Our latest publications

• Hermansen, Johanne Uthus; Kong, Weikaixin; Brodersen, Andrea M; Yanping, Yin; Urban, Aleksandra & Rein, Idun Dale [Show all 25 contributors for this article] (2026). A functional prognostic model predicts progression‐free survival in patients with relapsed chronic lymphocytic leukemia treated with ibrutinib + venetoclax. HemaSphere. 10(9). doi: 10.1002/hem3.70467. Full text in Research Archive Show summary

  Targeted therapies have improved outcomes for patients with chronic lymphocytic leukemia (CLL); yet, not all achieve durable responses. Existing prognostic models, developed for chemoimmunotherapy, have limited predictive value for newer targeted combinations. Here, we used data from a randomized phase II clinical trial to identify predictors of progression‐free survival (PFS) in response to Bruton tyrosine kinase (BTK) + B‐cell lymphoma 2 (Bcl‐2) inhibitor combination therapy. We studied the functional, genetic, and clinical characteristics of patients with relapsed/refractory (R/R) CLL treated with ibrutinib + venetoclax in the VISION/HO141 clinical trial (NCT03226301). Ex vivo drug sensitivity testing, (phospho)protein profiling, and immunophenotyping were performed on baseline peripheral blood mononuclear cells from 177 patients. Ex vivo drug sensitivity correlated with clinical responses to single‐agent therapies. (Phospho)protein patterns reflected intracellular pathway activity and predicted ex vivo drug sensitivity. Using stringent feature selection and regularized ridge regression to prevent overfitting, we integrated functional, genetic, and clinical data sets and identified six features—sensitivity to VEGFR‐2‐, JAK1/2 + Bcl‐2‐, and BTK + PI3K‐inhibitors; phosphorylation ratios of Bcl‐2:BTK and p90RSK:PLCγ2; and frequency of CD8 + T cells—that together outperformed the International Prognostic Index for CLL (CLL‐IPI) and other clinical scores in predicting PFS. These findings demonstrate that a prognostic model based on the functional features of CLL cell signaling and interaction with the microenvironment has the potential to improve upon conventional prognostic scores, warranting further clinical validation also in the frontline setting.

• Nothnagel, Patrizia Maria Christiane; Meslin, Paul-Arthur Richard; Wik, Jonas Aakre; Dufour, Damien; Strøm, Yunna Erika & Bjørås, Magnar [Show all 11 contributors for this article] (2026). Transient SUMOylation inhibition in human pre-adipocytes stably imprints a transcriptional beiging fate. Nucleic Acids Research (NAR). ISSN 0305-1048. 54(6), p. 1–22. doi: 10.1093/nar/gkag232. Full text in Research Archive Show summary

  SUMOylation regulates chromatin states and transcriptional programs that preserve cellular identity, yet how perturbation of the SUMOylation pathway impacts adipocyte plasticity remains unclear. Here, we show that brief pharmacologic inhibition of SUMOylation in human pre-adipocytes using TAK-981 primes stable de novo beige differentiation in the presence of the PPARG agonist rosiglitazone. Transient TAK-981 exposure produces changes in the transcriptome and metabolism of mature adipocytes, including robust induction of canonical beiging markers like UCP1 and increased mitochondrial respiration. Mechanistically, ATAC-seq and RNA-sequencing revealed immediate chromatin remodeling and early mobilization of CEBP family members, followed by stable activation of CEBPA and PPARG regulatory networks. ChIP experiments demonstrated loss of H3K27me3 and gain of H3K27ac at PPAR response elements at thermogenic enhancers, and increased PPARG occupancy across the UCP1 regulatory unit. This mechanism is enforced by enhanced cAMP-PKA-p38 signaling and stabilization of beiging transcription activators. We propose that transient relief of SUMO-mediated repression unlocks dominant regulatory units, notably the UCP1 enhancer cluster, producing a monomorphic reprogramming toward adaptive thermogenesis. These findings identify SUMOylation as a reversible epigenetic barrier to adipocyte beiging and suggest that temporally controlled SUMO pathway inhibition combined with PPARG activation could be exploited to modulate adipose tissue thermogenic capacity.

• Willoch-Allen, Katarina; Zhao, Zhi; Duran, Pilar Ayuda; Mikaeloff, Flora; Struyf, Nona & Österroos, Albin [Show all 13 contributors for this article] (2026). A multi-center study on the consistency of drug sensitivity testing in patients with acute myeloid leukemia. NPJ precision oncology. 10(1). doi: 10.1038/s41698-026-01480-4. Full text in Research Archive Show summary

  Clinical implementation of patient cell-based drug testing is hampered by inconsistencies and irreproducible results. Using linear mixed models and drug response data from four cohorts of patients with acute myeloid leukemia (n = 993 samples), we identified sources of variability in ex vivo drug sensitivity testing across the studies. We identified several experimental factors, including the time until sample was tested, the type of culture media and viability assay, which led to systematic differences in drug sensitivity profiles. We further investigated the effects of sample types (fresh or frozen) and disease status (diagnosis, relapse, or refractory), and observed that peripheral blood samples resulted in higher drug sensitivity levels, compared to those derived from bone marrow. Technical factors, such as the type of plate reader and centrifugation procedure, further contributed to the response variability depending on the chosen drug response metric. These factors need to be standardized for improved consistency in functional precision medicine studies.

• Rønneberg, Jørgen Dybevik; Hjeltnes, Martine; Krieger, Josephine; Elias, Joshua E.; Stensland, Maria & Holm, Kristian [Show all 15 contributors for this article] (2026). An epithelial GPR35 isoform supports tumor-associated transcriptional and metabolic phenotypes. FEBS Letters. ISSN 0014-5793. p. 1–23. doi: 10.1002/1873-3468.70423.
• Sargautis, Domantas & Thiede, Bernd (2026). LC-MS peptide mapping of monoclonal antibodies using the mirror proteases trypsin and Tryp-N. Journal of Pharmaceutical and Biomedical Analysis. ISSN 0731-7085. 272. doi: 10.1016/j.jpba.2026.117361. Full text in Research Archive
• Pati, Niladri Bhusan; Chung, Brian K.; Holm, Kristian; Sætre, Frank Ove; Reims, Henrik Mikael & Aasebø, Anders Theodor Michaelsen [Show all 12 contributors for this article] (2026). Ultra long-lived plasma cells in the human small intestine produce microbiota-reactive IgA antibodies. Beneficial Microbes. ISSN 1876-2883. p. 1–13. doi: 10.1163/18762891-bja00120. Full text in Research Archive
• Balaguera, Nathalia Chica; Andersen, Aram Nikolai; Munoz, Sara Orellana; Llorente, Ignacio Garcia; P, Aurélie Nguéa & Nakken, Sigve [Show all 14 contributors for this article] (2026). Time-resolved functional genomics using deep learning reveals global hierarchical control of autophagy. Nature Cell Biology. ISSN 1465-7392. 28(3), p. 465–479. doi: 10.1038/s41556-025-01837-0. Full text in Research Archive Show summary

  Recycling of cellular components through autophagy maintains homeostasis in changing nutrient environments. Although its core mechanisms are extensively studied, understanding of its systems-wide dynamic regulation remains limited, particularly regarding how autophagy is inactivated once nutrients are restored. Here we mapped the genetic network that controls activation and inactivation of autophagy during nitrogen changes by combining time-resolved high-content imaging, deep learning and latent feature analysis. This dataset, termed AutoDRY, categorizes 5,919 mutants based on nutrient response kinetics and their contributions to autophagosome formation and clearance. Integrating these profiles with functional and genetic network data uncovered hierarchical and multilayered control of autophagy and revealed multiple new regulatory pathways. Notably, we identified the retrograde pathway as a pivotal time-varying modulator that tunes the expression of core autophagy genes and plays a central role in autophagy inactivation. Together, this study establishes a systems-level resource to guide future investigations of autophagy.

• Kranjec, Christian; Eilertsen, Ina; Nunes, Luis; Moosavi, Hossein; Berg, Kaja Christine Graue & Eknæs, Mette [Show all 12 contributors for this article] (2026). Common gene mutations in 103 authenticated colorectal cancer cell lines. Oncogenesis. 15. doi: 10.1038/s41389-026-00599-0. Full text in Research Archive Show summary

  Colorectal cancer (CRC) cell lines represent the main molecular subtypes of tumors and are valuable models for preclinical investigations. However, cell lines can diverge over time and careful selection of models based on their molecular features is key. We have authenticated 103 commonly used CRC cell lines and present the mutation profiles of 20 CRC-relevant genes sequenced to an average depth of 575 times coverage. The cell lines reflected the distinct mutation patterns of hypermutation phenotypes associated with microsatellite instability and pathogenic POLE mutations. Hypermutated cell lines appeared to have a stronger mutational divergence and more frequent subclonal mutations, while mutations not associated with hypermutation were more frequently homozygous or hemizygous, classified as pathogenic, and subject to stronger selection pressure. Loss of heterozygosity at mutated loci was primarily observed in tumor suppressor genes. Genetic interactions based on co-occurring mutations identified cell lines representative of particularly aggressive subtypes of CRC, including concurrent BRAF p.V600 and truncating APC mutations, as well as APC/TP53/RAS triple mutations with double hits of APC. This study provides a resource to guide the selection of cell lines for functional studies of CRC, and detailed mutation data including classifications of pathogenicity, variant allele frequencies and illustrations of the mutation distribution along the length of encoded proteins are included.

• Iversen, Lars; Garrastacho, Manuel Ramirez; Hansen, Pernille May; Thiede, Bernd; Thyssen, Jacob Pontoppidan & Rasmussen, Marie Louise Roed [Show all 9 contributors for this article] (2026). Proteomic profiling of tear fluid in patients with atopic keratoconjunctivitis. The ocular surface. ISSN 1542-0124. 41, p. 66–75. doi: 10.1016/j.jtos.2026.05.005. Full text in Research Archive
• Falnes, Pål & Davydova, Erna (2026). The Protein Histidine Methyltransferase METTL9—From Mechanism to Biological Function. Life. 16(3). doi: 10.3390/life16030445. Full text in Research Archive
• Enserink, Jorrit Martijn & Balaguera, Nathalia Chica (2026). AutoDRY: a genome-wide resource revealing phase-specific control of autophagy activation, amplitude, and termination. Autophagy. ISSN 1554-8627. doi: 10.1080/15548627.2026.2660121. Full text in Research Archive
• Kärkkäinen, Virve; Saari, Toni; Rusanen, Minna; Uusitalo, Hannu; Leinonen, Ville & Thiede, Bernd [Show all 9 contributors for this article] (2026). Altered tear fluid protein expression in persons with mild cognitive impairment. Journal of Alzheimer's Disease. ISSN 1387-2877. 110(1), p. 109–119. doi: 10.1177/13872877251413291. Full text in Research Archive
• Nguyen, Xuan Thang; Martiensen, Emily Louise Beecher; Abdelhalim, Mohamed; Tran, Trung; Ledsaak, Marit & Rogne, Marie [Show all 8 contributors for this article] (2026). Proteome-wide multi-omics profiling of osteosarcoma transcription factor networks. bioRxiv (preprint article). doi: 10.64898/2026.03.29.714917. Full text in Research Archive Show summary

  Osteosarcoma (OS) is an aggressive bone cancer that most commonly affects children and young adults. OS exhibits a high degree of genomic complexity, as well as cellular plasticity, and dynamic transcriptional regulation is suggested to contribute to treatment resistance and metastasis. Cell lines are well characterized as models to advance our knowledge on OS biology. HOS and U2OS cells have increased invasiveness and higher migratory ability compared with MG63. In this study, we employed a tandem array of consensus transcription factor response elements (catTFREs) proteomic approach to characterize transcription factor (TF) regulatory networks related to OS aggressiveness. We mapped 7,594 proteins and enriched 352 transcription factors and coregulators. When we integrated proteomics with cell line specific gene expression and chromatin accessibility we classified the proteins into different OS cell line dependent sub-clusters and identified TFs and coregulators common for all cell lines and specific for individual cell lines. We demonstrate that RUNX2, MYBL2 and HMGA2 are specifically enriched in HOS and U2OS and may be linked to the cell aggressiveness. ETV5, JUNB, NFIX and ZEB1 were among TFs specific to MG63. Our analysis provides a more comprehensive understanding of the transcriptional drivers that shape OS regulatory landscapes and may have future therapeutic implications.

• Lauretis, Angela De; Wang, Qiang; Santacroce, Marco; Thiede, Bernd; Ma, Qianli & Lyngstadaas, Ståle Petter [Show all 9 contributors for this article] (2025). A Comparative Study of the Impact of Chemical Debridement Products on the Proteomic Profile of the Salivary Pellicle and Cell Adhesion on OsseoSpeed Titanium Dental Implant Surfaces. ACS Applied Materials & Interfaces. ISSN 1944-8244. 17(39), p. 54592–54592. doi: 10.1021/acsami.5c14570. Full text in Research Archive
• Tu, Yanyun; Cheng, Lina; Liu, Xianfeng; Hammerstad, Marta; Shi, Chunlin & Meng, Sida [Show all 9 contributors for this article] (2025). SlIDL6–SlHSL1/2/3 ligand-receptor pairs regulate tomato pedicel abscission. Journal of Integrative Agriculture (JIA). ISSN 2095-3119. 25(1), p. 118–126. doi: 10.1016/j.jia.2025.03.018. Full text in Research Archive Show summary

  Flower and fruit abscission reduce crop yield, so decreasing abscission is a significant agricultural issue. HAESA (HAE) and HAESA-like2 (HSL2) kinases and their ligand, INFLORESCENCE DEFICIENT IN ABSCISSION (IDA) peptide, have been confirmed to be the core elements regulating floral organ abscission in Arabidopsis thaliana. Our earlier research revealed that SlIDL6, a homolog of IDA in tomato, functions similarly to AtIDA, regulating the abscission of tomato flower organs. Here, we further isolated three HAESA-like homologs, SlHSL1/2/3, which are involved in tomato flower abscission. SlHSL1/2/3 are highly expressed in the abscission zone (AZ). The knockout mutant lines of Slhsl1, Slhsl2, and Slhsl3 showed lower flower pedicel abscission than wild type (WT). The double mutant of Slhsl1Slhsl2, Slhsl1Slhsl3, and Slhsl2Slhsl3 further depressed abscission than each of the single mutant lines, while triple mutants Slhsl1Slhsl2Slhsl3 exhibited the lowest abscission, indicating that SlHSL1/2/3 mediated abscission is non-redundancy, at least partially. Treating tomato pedicel explants with SlIDL6 peptide significantly accelerated pedicel abscission in WT. However, it had little effect on the abscission rate of SlHSL1/2/3 knockout lines, indicating that SlHSL1/2/3 are the receptors of SlIDL6 in pedicel abscission. Ethylene action inhibitor 1-methylcyclopropene (1-MCP) can significantly depress the expression of SlHSL1/2/3. Ethylene can significantly accelerate the abscission of WT, while less abscission was found in SlHSL1/2/3 knockout lines. Our findings indicate that SlHSL1/2/3 can act as receptors for SlIDL6 to positively regulate tomato pedicel abscission, and the abscission regulated by SlHSL1/2/3 was partially dependent on ethylene.

• Amono, Racheal; Rønning, Snöa Amanda Tora Nyx Fredlund; Chesnais, Morgane; Thiede, Bernd; Markussen, Turhan & Evensen, Øystein [Show all 7 contributors for this article] (2025). Defined domains and cleavage determine the diverse functions of piscine myocarditis virus p33 protein. Frontiers in Microbiology. 16. doi: 10.3389/fmicb.2025.1633241. Full text in Research Archive Show summary

  Piscine myocarditis virus (PMCV) causes chronic, necrotizing myocarditis in Atlantic salmon. Originally, PMCV was identified based on its genetic homology and genomic organization, indicating a relationship to viruses of the Ghabrivirales order, specifically the former Totiviridae family, whose members predominantly infect fungi or protozoans and lack an extracellular life cycle stage. However, PMCV was the first virus of this order found to infect a vertebrate host. Since then, other piscine viruses and viruses infecting terrestrial and aquatic arthropods have been described and recently assigned to new virus families within the order. PMCV is now classified in Pistolviridae. All these viruses infecting multicellular hosts encode proteins that are believed to be involved in extracellular transmission. In PMCV, this relates to a protein of size 33.4 kDa (p33) encoded by a unique third open reading frame. To investigate its characteristics and role, we expressed various recombinant variants of p33 in cultured cells. Our results demonstrate that p33 expression induces a cytotoxic phenotype in transfected cells. The full-length protein undergoes processing into smaller peptide variants. Previous in silico analysis predicted an N-terminal chemokine-like domain, and our present results show that this domain is secreted as peptides capable of inducing cytotoxicity when expressed alone. The C-terminal region includes sequence characteristics of a small hydrophobic domain, which appears crucial for the correct processing of the full-length protein into N- and C-terminal peptides and directing the C-terminal peptides to a high membrane concentration. Investigations into p33 function could elucidate how PMCV achieves extracellular transmission, a mechanism that may be conserved among viruses of Pistolviridae. The findings in this study provide evidence that p33 has structural and functional characteristics of a protein adapted to facilitate host cell membrane interaction and cell lysis, potentially enabling extracellular viral release. These insights may provide evolutionary evidence that pistolviruses have acquired the uncommon trait of viral transmission within the order Ghabrivirales, broadening our understanding of virus–host adaptation in vertebrates.

• Sagini, Krizia; Selvarajah, Mughilan; Romero, Silvana Angelica Troncoso; Bassols, Eva Citores; Gracia, Beatriz Martín & Ramirez, Manuel [Show all 16 contributors for this article] (2025). The Redox Enzyme Thioredoxin Is Increased in Plasma Extracellular Vesicles From Endurance‐Trained Females in Response to Acute Exercise. The FASEB Journal. ISSN 0892-6638. 39(16). doi: 10.1096/fj.202500678r. Full text in Research Archive Show summary

  The effects of exercise are mediated by molecules secreted into the circulation in response to exercise. Extracellular vesicles (EVs) have recently appeared as possible vehicles for some of these molecules, and as mediators of tissue crosstalk during exercise. This study aimed to identify changes in the proteome of EVs released into circulation by endurance running‐trained females after a bout of 60‐min aerobic exercise consisting of incremental treadmill running. Blood samples were collected immediately before and after exercise, and EVs were separated from plasma and characterized by Western blot and electron microscopy. Mass spectrometry‐based proteomics analysis of EV preparations was then performed and showed that the levels of 9 proteins were differentially expressed in response to exercise. Thioredoxin, an enzyme that plays an essential role in the maintenance of redox homeostasis, showed the largest change. The increase in the levels of thioredoxin after exercise was also observed by Western blot analysis. Interestingly, post‐exercise plasma EVs were able to protect cells against oxidative stress. These results support the effect of physical exercise in modulating the levels of molecules involved in the maintenance of redox homeostasis, such as thioredoxin and other antioxidant enzymes, in plasma EVs. Although further studies are needed to fully understand the functional role of these molecules, it is intriguing to consider the protective health effects that exercise‐induced EVs may have against oxidative stressors.

• Kärkkäinen, Virve; Saari, Toni; Rusanen, Minna; Uusitalo, Hannu; Leinonen, Ville & Thiede, Bernd [Show all 9 contributors for this article] (2025). Neuroinflammation Markers in Tear Fluid of Mild Alzheimer’s Disease. Journal of Molecular Neuroscience. ISSN 0895-8696. 75(2). doi: 10.1007/s12031-025-02368-x. Full text in Research Archive Show summary

  The protein composition of tear fluid (TF) reflects the severity and progression of many age-related diseases. Here, we evaluated TF proteins from patients with mild Alzheimer’s disease (AD) and cognitively healthy controls (CO) to explore potential new biomarker molecules. The aim of this study was to explore potential new biomarker molecules by examining the expression of TF proteins whose function is related to neuroinflammation. We examined 53 participants (34 COs, mean age 71 years, Mini-Mental State Examination (MMSE) score 28.9 ± 1.4; 19 with AD, Clinical Dementia Rating 0.5–1, mean age 72 years, MMSE 23.8 ± 2.8). All participants underwent neurological status examination, cognitive testing, and ophthalmological examination. TF was collected using Schirmer strips, and TF protein content was evaluated using mass spectrometry-based proteomics and label-free quantification. We report 14 TF proteins that showed altered protein expression in the AD group compared to the CO group. Twelve proteins were significantly upregulated (SERPINA3, FGA, SIAS, ORM1, ANXA3, G6PI/NLK, CH3L2, MSLN, CPPED1, JCHAIN, IGHV5-51, SPARCL1) and two were downregulated (PIP, SCGB2A1) (p ≤ 0.05). Observed altered expression of TF proteins in the AD group may have potential in AD pathology. Since inflammation is one of the earliest signs of neurodegeneration in AD, these proteins are putative new biomarker candidates of early AD.

• Roman, Giacomo; Lauritzen, Knut Husø; Harrison, Sean Philip; Bhattacharya, Anindita; Andresen, Marianne Seierstad & Mowinckel, Marie-Christine [Show all 16 contributors for this article] (2025). Ex vivo correction of severe coagulation Factor VII deficiency in patient-derived 3D liver organoids. Haematologica. ISSN 0390-6078. doi: 10.3324/haematol.2025.288046. Full text in Research Archive Show summary

  Coagulation factor (F) VII deficiency is the most frequent among the rare, inherited bleeding disorders and is predominantly caused by missense mutations in the F7 gene. The disease phenotype ranges from asymptomatic cases to extremely severe hemorrhagic forms, requiring prophylactic injections with plasma-derived or recombinant FVII concentrates. In response, we have developed an autologous cell-based approach that corrects the disease-causing mutation in patient-derived induced pluripotent stem cells (iPSCs) and generates therapeutic, three-dimensional hepatic organoids (HOs). We report the CRISPRmediated correction of homozygous c.718G>C (p.G240R), a missense mutation associated with a severe, life-threatening bleeding phenotype. The HOs contain all liver cell types and exhibit key liver functions, including coagulation factor production. After correction, our data indicate that the patient-derived HOs secrete consistent amounts of functional FVII protein, resulting in improved thrombin generation times. These results represent a significant milestone toward the establishment of an autologous cell-based therapy for patients with FVII- and other coagulation factor deficiencies.

• Nothnagel, Patrizia Maria Christiane; Meslin, Paul-Arthur Richard; Wik, Jonas Aakre; Strøm, Yunna Erika; Bjørås, Magnar & Enserink, Jorrit [Show all 10 contributors for this article] (2025). Transient SUMOylation Inhibition In Human Pre-adipocytes Stably Imprints a Transcriptional Beiging Fate. bioRxiv (preprint article). 54(6). doi: 10.1101/2025.03.07.642034. Full text in Research Archive Show summary

  Abstract SUMOylation regulates chromatin states and transcriptional programs that preserve cellular identity, yet how transient perturbation of the SUMO pathway impacts adipocyte plasticity remains unclear. Here we show that brief pharmacologic inhibition of SUMO conjugation in human pre-adipocytes using TAK-981 primes stable de novo beige differentiation in the presence of the PPARG agonist rosiglitazone. Transient TAK-981 exposure before adipogenic induction produces long-lasting changes in the transcriptome and metabolism of mature adipocytes, including robust induction of canonical beiging markers like UCP1 and increased mitochondrial respiration. Mechanistically, ATAC-seq and transcription factor footprinting revealed immediate and durable chromatin remodeling and early mobilization of CEBP family members, followed by stable activation of CEBPA and PPARG regulatory networks. ChIP experiments demonstrated loss of H3K27me3 and gain of H3K27ac at PPAR response elements within key thermogenic enhancers, with increased PPARG occupancy across the UCP1 regulatory unit. This mechanism is enforced by enhanced cAMP-PKA-p38 signaling, and stabilization of beiging transcriptional activators. Our data support a model in which transient relief of SUMO-mediated repression unlocks dominant regulatory units, notably the UCP1 enhancer cluster, producing a monomorphic, cell type specific reprogramming toward adaptive thermogenesis. These findings identify SUMOylation as a reversible epigenetic barrier to adipocyte beiging and suggest that temporally controlled SUMO pathway inhibition combined with PPARG activation could be exploited to modulate adipose tissue thermogenic capacity. Graphical abstract SUMOylation restricts de novo differentiation of pre-adipocytes into beige adipocytes by repressing cAMP signaling and preventing epigenetic and transcriptional reprogramming of white adipocytes.

• Deng, Ke; Pällmann, Nora; Livgård, Marte; Kildal, Wanja; Pradhan, Manohar & Fazli, Ladan [Show all 10 contributors for this article] (2025). Endoplasmic reticulum stress-driven nucleotide catabolism fuels prostate cancer. Cancer Letters. ISSN 0304-3835. 630. doi: 10.1016/j.canlet.2025.217888. Full text in Research Archive Show summary

  Endoplasmic reticulum (ER) stress is a critical regulator of cancer cell metabolism and survival. In this study, we elucidate the coordinated roles of two key ER stress mediators, Activating Transcription Factor 4 (ATF4) and X-box Binding Protein 1 spliced (XBP1s), in regulating purine homeostasis in prostate cancer (PCa) cells. We demonstrate that ATF4 directly upregulates Molybdenum Cofactor Sulfurase (MOCOS), a key enzyme in purine catabolism, while XBP1s induces the expression of xanthine dehydrogenase (XDH), the principal MOCOS target in this pathway. Knockdown of MOCOS significantly impairs PCa cell proliferation as well as prostatosphere and colony formation in vitro, and inhibits tumor growth in preclinical mouse models of PCa. Mechanistically, MOCOS suppression leads to purine accumulation, disrupts pyrimidine synthesis, and causes nucleotide imbalance, resulting in replication fork stalling. This imbalance is also accompanied by a compromised glutathione-mediated antioxidant response, rendering the cells more susceptible to DNA damage. Importantly, targeting XDH, either genetically or biochemically, also significantly hinders PCa cell growth. Collectively, our data highlight the pivotal role of ER stress-mediated purine homeostasis in sustaining PCa cell growth.

• Kuzu, Omer; Granerud, Lars Jørgen Tvenge & Saatcioglu, Fahri (2025). Navigating the landscape of protein folding and proteostasis: from molecular chaperones to therapeutic innovations. Signal Transduction and Targeted Therapy. ISSN 2095-9907. 10(1). doi: 10.1038/s41392-025-02439-w. Full text in Research Archive Show summary

  Abstract Protein folding is a fundamental process ensuring that polypeptide chains acquire the correct three-dimensional structures required for biological function. This complex journey from nascent polypeptides to mature proteins is tightly regulated by the cellular proteostasis network—an integrated system of molecular chaperones, folding enzymes, and degradation machineries. Disruptions in this network lead to dysproteostasis, a pathological state implicated in a growing list of human diseases, including neurodegenerative disorders, metabolic syndromes, and cancer. In this review, we provide a comprehensive and multidimensional analysis of protein folding biology, tracing its evolution from early theoretical foundations to cutting-edge biophysical and computational techniques that now permit near-atomic-resolution modeling of folding dynamics. We explore the historical progression of protein folding research, including landmark discoveries of secondary structure, chaperone biology, and energy landscape theory. We detail the roles of key molecular chaperones across cytosolic, mitochondrial, and endoplasmic reticulum compartments, emphasizing their collaborative actions in protein folding and quality control. We also discuss the multifactorial causes of protein misfolding—from genetic mutations to aging and oxidative stress—and examine the pathological consequences, paying special attention to diseases characterized by toxic protein aggregation and loss of proteome fidelity. We then examine therapeutic innovations targeting proteostasis, including chaperone modulators, proteostasis pathway inhibitors, and emerging strategies to increase proteome resilience. By consolidating insights at the molecular, cellular, and systems levels, this review underscores the central role of protein folding homeostasis in health and disease and highlights novel opportunities for therapeutic intervention through the modulation of the proteostasis network.

• Ünal, Bilal & Saatcioglu, Fahri (2025). Targeting the unfolded protein response for cancer therapy: mitigating tumor adaptation and immune suppression. Biomarker Research. 13. doi: 10.1186/s40364-025-00813-y. Full text in Research Archive Show summary

  There are significant stress factors within the tumor microenvironment (TME), such as hypoxia, oxidative stress, and nutrient deprivation. These disrupt endoplasmic reticulum (ER) function in cancer cells, as well as the infiltrating immune cells, leading to activation of the unfolded protein response (UPR) signaling, which the tumor uses to mitigate stress and survive. There are three canonical UPR pathways that are regulated by respective ER-resident transmembrane sensors: inositol-requiring protein 1α (IRE1α), PKR-like ER kinase (PERK), and activating transcription factor 6 (ATF6); activation of these pathways results in expression of cognate transcription factors that regulate gene expression to mitigate ER stress. Persistent UPR activation in the TME has been linked to aberrant tumor growth, progression, metastasis, angiogenesis, and therapy resistance in different cancer types. In addition, modulation of UPR activity significantly impacts immune cell function at different levels further impacting its role on the TME. Therefore, there is now significant interest to design novel therapies that target the UPR to kill cancer cells and simultaneously enhance protective anti-tumor immunity. Here we summarize recent findings as to how targeting UPR signaling can induce tumor regression and at the same time galvanize the immune response. We discuss the potential of integrating UPR targeting with other therapies, such as immune checkpoint inhibition, highlighting emerging strategies to improve therapeutic efficacy and overcome resistance. These recent insights underscore the importance of UPR as a novel therapeutic target for cancer treatment.

• Farhan, Hesso; Zahoor, Muhammad; Tauer, Josephine T. & Högler, Wolfgang (2025). The endoplasmic reticulum proteostasis network and bone disease. Trends in Molecular Medicine. ISSN 1471-4914. 2193, p. 1–14. doi: 10.1016/j.molmed.2025.06.005. Full text in Research Archive Show summary

  Bone homeostasis relies on the coordinated activity of osteoblasts and osteoclasts that balances bone formation and resorption, and of osteocytes for biomechanical sensing and hormone secretion. A key factor in the function of these cells is proteostasis, where the endoplasmic reticulum (ER) oversees protein synthesis, quality control, folding, and the secretion of proteins such as collagen type I. Emerging research links ER proteostasis defects to skeletal disorders caused by impaired bone development and mass. We explore the mechanisms of ER proteostasis, including the unfolded protein response (UPR), and discuss how genetic, metabolic, and environmental factors disrupt these pathways and contribute to bone pathology. We also highlight the need for further mechanistic insights which could pave the way for novel therapies that target ER–Golgi traffic and inhibit ER stress in bone diseases.

• Pharo, Heidi; Brandt-Winge, Sara; Thorkildsen, Evy Marie; Vedeld, Hege Marie & Lind, Guro Elisabeth (2025). DNA Methylation Analyses of Tissue Samples and Cell-Free DNA by Droplet Digital PCR. In Pavlovic, Guillaume (Eds.), Digital PCR. Springer Nature. ISSN 9781071647677. p. 179–194. doi: 10.1007/978-1-0716-4767-7_12. Full text in Research Archive
• Auxillos, Jamie; Stigliani, Arnaud; Vaagensø, Christian Skov; Garland, William; Niazi, Adnan Muhammad & Valen, Eivind [Show all 8 contributors for this article] (2025). True length of diverse capped RNA sequencing (TLDR-seq): 5′–3′-end sequencing of capped RNAs regardless of 3′-end status. Nucleic Acids Research (NAR). ISSN 0305-1048. 53(6). doi: 10.1093/nar/gkaf240. Full text in Research Archive Show summary

  Analysis of transcript function is greatly aided by knowledge of the full-length RNA sequence. New long-read sequencing enabled by Oxford Nanopore and PacBio devices have the potential to provide full-length transcript information; however, standard methods still lack the ability to capture true RNA 5′ ends and select for polyadenylated (pA+) transcripts only. Here, we present a method that, by utilizing cap trapping and 3′-end adapter ligation, sequences transcripts between their exact 5′ and 3′ ends regardless of polyadenylation status and without the need for ribosomal RNA depletion, with the ability to characterize polyadenylation length of RNAs, if any. The method shows high reproducibility, can faithfully detect 5′ ends, 3′ ends and splice junctions, and produces gene-expression estimates that are highly correlated to those of short-read sequencing techniques. We also demonstrate that the method can detect and sequence full-length nonadenylated (pA−) RNAs, including long noncoding RNAs, promoter upstream transcripts, and enhancer RNAs, and present cases where pA+ and pA− RNAs show preferences for different but closely located transcription start sites. Our method is therefore useful for the characterization of diverse capped RNA species and analysis of relationships between transcription initiation, termination, and RNA processing.

• Chothani, Sonia; Ruiz-Orera, Jorge; Tierney, Jack A. S.; Clauwaert, Jim; Deutsch, Eric W & Alba, M. Mar [Show all 46 contributors for this article] (2025). An expanded reference catalog of translated open reading frames for biomedical research. bioRxiv. ISSN 2692-8205. doi: 10.1101/2025.07.03.662928. Full text in Research Archive Show summary

  Non-canonical (i.e., unannotated) open reading frames (ncORFs) have until recently been omitted from reference genome annotations, despite evidence of their translation, limiting their incorporation into biomedical research. To address this, in 2022, we initiated the TransCODE consortium and built the first community-driven consensus catalog of human ncORFs, which was openly distributed to the research community via Ensembl-GENCODE. While this catalog represented a starting point for reference ncORF annotation, major technical and scientific issues remained. In particular, this initial catalogue had no standardized framework to judge the evidence of translation for individual ncORFs. Here, we present an expanded and refined catalog of the human reference annotation of ncORFs. By incorporating more datasets and by lifting constraints on ORF length and start-codon, we define a comprehensive set of 28,359 ncORFs that is nearly four times the size of the previous catalog. Furthermore, to aid users who wish to work with ncORFs with the strongest and most reproducible signals of translation, we utilized a data-driven framework (i.e. translation signature scores) to assess the accumulated evidence for any individual ncORF. Using this approach, we derive a subset of 7,888 ncORFs with translation evidence on par with canonical protein-coding genes, which we refer to as the Primary set. This set can serve as a reliable reference for downstream analyses and validation, with a particular emphasis on high quality. Overall, this update reflects continual community-driven efforts to make ncORFs accessible and actionable to the broader research public and further iterations of the catalog will continue to expand and refine this resource.

• Kute, Preeti; Pauzin, Francois Philippe; Labun, Kornel; Bramham, Clive Raymond Evjen & Valen, Eivind (2025). Transcriptome-wide mapping of small ribosomal subunits elucidates scanning mechanisms of translation initiation in the mammalian brain. Communications Biology. 8(1). doi: 10.1038/s42003-025-08804-3. Full text in Research Archive Show summary

  Abstract Neuronal protein synthesis is highly compartmentalised and regulated, with key roles for translation initiation and elongation factors. Ribosome profiling, the most widely used transcriptome-wide method for measuring translation, captures translation elongation, but not the initiation phase involving small ribosomal subunit (SSU) scanning. Here, we adapted ribosome complex profiling (RCP-seq) for mouse dentate gyrus and cerebral cortex, to characterize translation initiation. In both tissues, SSUs accumulate near the start codon on synaptically localised RNAs, and this ‘poised’ SSU configuration correlates with enhanced translational efficiency. Upstream open reading frames (uORFs) are associated with less poised SSUs, potentially by disassociating the SSUs. We further find that neuron-specific transcripts recruit more ribosomes and are more efficiently translated than glia-specific transcripts. For neuronal transcripts, monosome-preferring mRNAs show less SSU occupancy relative to polysome-preferring mRNAs, suggesting reduced recruitment of ribosomes. In summary, RCP-seq elucidates translation initiation dynamics and cell-type- and transcript-specific regulation in the mammalian brain.

• Engelfriet, Melanie Lianne; Guo, Yanwu; Arnold, Andreas; Valen, Eivind & Ciosk, Rafal (2025). Reprograming gene expression in ‘hibernating’ C. elegans involves the IRE-1/XBP-1 pathway. eLIFE. 13. doi: 10.7554/elife.101186.3. Full text in Research Archive Show summary

  In nature, many animals respond to cold by entering hibernation, while in clinical settings, controlled cooling is used in transplantation and emergency medicine. However, the molecular mechanisms that enable cells to survive severe cold are still not fully understood. One key aspect of cold adaptation is the global downregulation of protein synthesis. Studying it in the nematode Caenorhabditis elegans , we find that the translation of most mRNAs continues in the cold, albeit at a slower rate, and propose that cold-specific gene expression is regulated primarily at the transcription level. Supporting this idea, we found that the transcription of certain cold-induced genes is linked to the activation of unfolded protein response (UPR) through the conserved IRE-1/XBP-1 signaling pathway. Our findings suggest that this pathway is triggered by cold-induced perturbations in proteins and lipids within the endoplasmic reticulum, and that its activation is beneficial for cold survival.

• Pinto, Rita Isabel Morais; Sjurgard, Ingrid Vikan; Pharo, Heidi; Mala, Tom; Vedeld, Hege Marie & Lind, Guro Elisabeth (2025). Molecular stratification of esophageal adenocarcinoma: implications for prognosis and treatment strategy. Oncogene. ISSN 0950-9232. 45, p. 353–367. doi: 10.1038/s41388-025-03650-3. Full text in Research Archive Show summary

  Genome-wide molecular profiling has emerged as a promising approach for advancing the clinical management of esophageal adenocarcinoma (EAC), with the potential to improve prognostic accuracy and enable more personalized treatment strategies. In this review, we summarize current evidence from genomic and epigenomic EAC stratification studies, highlighting the proposed molecular subtypes and evaluating their clinical relevance. We discuss how these subclassifications may inform disease outcomes, refine patient selection for specific therapies and uncover new treatment opportunities aligned with tumor molecular profiles. Additionally, we explore molecular subtypes associated with Barrett’s esophagus, a precursor lesion of EAC, and consider how these insights can help elucidate the mechanisms underlying EAC development. Such understanding may inform improved strategies for early tumor detection, risk stratification and prevention, ultimately aiming to reduce the burden of EAC. We also address the current challenges limiting the clinical application of these molecular classifiers, including restricted sample availability, insufficient validation and the difficulty of translating genome-wide findings into practical and clinical useful biomarkers. Integrating molecular subtyping into clinical workflows is a key step toward precision medicine in EAC, with the goal of enhancing treatment response rates and patient outcomes. Future advances will require collaborative efforts and robust clinical validation in large prospective studies to ensure that molecular stratification strategies can be effectively translated into improved management of EAC.

• Kärkkäinen, Virve; Saari, Toni; Hannonen, Sanna; Rusanen, Minna; Lehtola, Juha Matti & Uusitalo, Hannu [Show all 11 contributors for this article] (2025). Altered tear fluid protein expression in persons with mild Alzheimer's disease in proteins involved in oxidative stress, protein synthesis, and energy metabolism. Journal of Alzheimer's Disease. ISSN 1387-2877. 105(1), p. 292–301. doi: 10.1177/13872877251326868. Full text in Research Archive Show summary

  Background Tear fluid (TF) is a protein-rich solution that reflects pathophysiological changes in Alzheimer's disease (AD). Objective In this study, we examined whether TF proteins were differently expressed in persons with mild AD dementia compared to cognitively healthy controls (CO). Methods We analyzed data from 53 study participants including 34 CO (mean age, 71 years; Mini-Mental State Examination [MMSE] score, 28.9 ± 1.4), and 19 patients with AD (Clinical Dementia Rating, 0.5–1; mean age, 72 years; MMSE score, 23.8 ± 2.8). All participants underwent cognitive testing, as well as neurological and ophthalmological examinations. TF was collected using Schirmer strips, and TF protein content was evaluated using mass spectrometry-based proteomics and label-free quantification. Results We found that 16 proteins exhibited significantly upregulated expression in the AD group compared to the CO group (p ≤ 0.05). These proteins were NP1L4, BBOX1, CYTC, RNAS4, PCD, RNT2, AL1A3, SYSC, TPIS, CLH1, PGAM1, EIF3L, 5NTC, HNRNPA2B1, PYGL, and ERO1α. No proteins were significantly downregulated in the AD group compared to the CO group. Conclusions Our results support the hypothesis that TF is a potential source of biomarkers for AD. Part of those proteins with altered expression have previously linked to increased oxidative stress, changed protein synthesis, and disturbed regulation of energy metabolism related to AD or neurodegenerative disease. The present results indicate the value of continued investigation of TF proteins in AD.

• Hammerstad, Marta; Steinvik, Erlend & Hersleth, Hans-Petter (2025). The methyltransferase NmbA methylates the low-molecular weight thiol bacillithiol, and displays a specific structural architecture. Redox Biology. 88. doi: 10.1016/j.redox.2025.103937. Full text in Research Archive Show summary

  Low-molecular-weight (LMW) thiols maintain the cellular redox balance and protect cells against reactive species, heavy metals, toxins, and antibiotics. Despite having similar metabolic functions, structurally distinct LMW thiols are widespread in nature, with bacillithiol (BSH) being the predominant LMW thiol in bacteria. The LMW thiol N-methyl-BSH (N-Me-BSH) has been identified in the green sulfur bacterium Chlorobaculum tepidum, revealing the presence of a putative S-adenosyl-l-methionine (SAM)-dependent methyltransferase (MT), NmbA, which could catalyze the final biosynthetic step of N-Me-BSH. In this study, we report biochemical evidence for NmbA's specific function as an MT of the N-atom of the BSH cysteine moiety. We also present the crystal structure of NmbA, confirming that NmbA is a Class I SAM-dependent MT, however, displaying a unique three-dimensional architecture that differs from those of other natural product MTs (NPMTs). The NmbA active site has a narrow molecular basket structure resulting from an unusual organization of the variable Cap domain, and our docking calculations suggests that it can specifically accommodate the BSH substrate. Our research provides a valuable overview of the phylogenetic distribution of N-Me-BSH in bacteria, alongside essential functional and structural insight into a new class of N-directed NPMTs. These findings contribute to the field of SAM-dependent MTs and may allow for targeting distinct bacterial defense mechanisms involving LMW thiols with potential environmental, biotechnological, and medical implications.

• Pharo, Heidi; Vedeld, Hege Marie; Sjurgard, Ingrid Vikan; Pinto, Rita Isabel Morais & Lind, Guro Elisabeth (2025). From concept to clinic: a roadmap for DNA methylation biomarkers in liquid biopsies. Oncogene. ISSN 0950-9232. 44, p. 4814–4831. doi: 10.1038/s41388-025-03624-5. Full text in Research Archive Show summary

  Global cancer incidence continues to rise, emphasizing the urgent need for improved diagnostics and management strategies. DNA methylation biomarkers in liquid biopsies offer a promising, minimally invasive solution. Despite their potential, only a few tests have successfully transitioned from research to clinical practice. This review addresses key aspects influencing successful biomarker development and clinical implementation—including liquid biopsy source selection, biomarker discovery workflow and targeted validation in clinical sample series—and provide strategies to improve accuracy, reproducibility and clinical utility. Altogether, these considerations could aid in bridging the translational gap from research to clinical application, and to increase the number of clinically implemented liquid biopsy tests.

• Andresen, Siri; Outa, Amani al; Formica, Miriam; Enserink, Jorrit & Knævelsrud, Helene (2025). Improved detection of lipidated Atg8a by immunoblotting in Drosophila melanogaster cells and tissues enables precise investigation of Atg8a flux and its termination. Autophagy. ISSN 1554-8627. 21(11), p. 2497–2517. doi: 10.1080/15548627.2025.2508551. Full text in Research Archive Show summary

  Macroautophagy/autophagy is an essential intracellular catabolic process for maintaining cellularhomeostasis. In Drosophila melanogaster, Atg8a lipidation serves as a key marker for autophagy, yettraditional methods often fail to effectively detect its lipidated state. To overcome this limitation, wedeveloped a refined approach that employs N-ethylmaleimide (NEM) to inhibit Atg4, thereby pre-serving Atg8a lipidation during sample preparation both in vitro and in vivo. We determined theoptimal concentration of the autophagic inhibitors bafilomycin A1 (BafA1) and chloroquine (CQ)required for inhibition of autolysosomal degradation. Furthermore, we investigated the effects ofprolonged nutrient deprivation on autophagic flux and TORC1 signaling. Our findings not onlyvalidate the effectiveness of this new approach to monitor lipidation of Atg8a but also provideinsights into selection of autolysosomal inhibitors and nutrient-dependent regulatory roles of TORC1in Drosophila.

• Aaen, Kristin Hovden; Testa, Maria Francesca; Nilsen, Jeannette; Tarantino, Rebecca; Canepari, Cesare & Benedusi, Mascia [Show all 16 contributors for this article] (2025). Tailored collagen binding of albumin-fused hyperactive coagulation factor IX dictates in vivo distribution and functional properties. Nature Communications. 16. doi: 10.1038/s41467-025-62955-9. Full text in Research Archive Show summary

  The efficacy of hemophilia B (HB) replacement therapy is evaluated by coagulation factor IX (FIX) activity in plasma, although FIX bound to extravascular type IV collagen (Col4) also contributes to efficient hemostasis. Here, we investigated the impact of engineering FIX for improved (K5R) or reduced (K5A) Col4 binding on the pharmacokinetic properties of FIX Padua, fused to human serum albumin (HSA QMP ) engineered for favorable neonatal Fc receptor (FcRn) engagement. Hyperactive features and extended plasma half-life in human FcRn expressing mice, attributed to FIX Padua and HSA QMP engineering, respectively, was confirmed. In HB mice, Padua KA -HSA QMP exhibited negligible extravascular distribution and the highest plasma levels at early time points followed by the steepest decay. Conversely, Padua KR -HSA QMP showed increased extravascular distribution and a 3-fold longer functional half-life (80 hours). These findings support the use of Padua KA -HSA QMP and Padua KR -HSA QMP as hyperactive short- or long-term therapeutics, respectively, with opportunities for tailored HB replacement therapy.

• Schroer, Lisa; Weirich, Sara; Hammerstad, Marta; Hersleth, Hans-Petter; Grønsberg, Ida Andrietta & Hagen, Lars [Show all 11 contributors for this article] (2025). Orthologues of the human protein histidine methyltransferase METTL9 display distinct substrate specificities. Journal of Biological Chemistry. ISSN 0021-9258. 301(7). doi: 10.1016/j.jbc.2025.110318. Full text in Research Archive Show summary

  The human (Homo sapiens; Hs) methyltransferase (MTase) METTL9 is the first enzyme shown to generate 1-methylhistidine (π-methylhistidine) in proteins. METTL9 preferentially methylates an alternating histidine (HxH) motif, where “x” is a small, uncharged amino acid, and multiple substrates have been identified. Putative METTL9 orthologues are found in most eukaryotes, and we have here investigated the activity of such enzymes from several species, representing all five eukaryotic supergroups. The majority of the tested enzymes demonstrated in vitro MTase activity on the prototype HsMETTL9 substrates ARMC6 and DNAJB12. We also detected protein methylation activity of the Caenorhabditis elegans METTL9 which had previously been suggested to be a DNA MTase. However, METTL9 from the fruit fly (Drosophila melanogaster; Dm) and the picoplankton Ostreococcus tauri (Ot) displayed distinct substrate specificities, differing from each other and from that of HsMETTL9. These differences were observed when recombinant proteins and short peptides were used as METTL9 substrates. To further analyze substrate specificity, we used peptide arrays to systematically replace the “x” residue and the residues flanking the HxH motif in a substrate peptide. This revealed varying degrees of tolerance among the METTL9 orthologues (Hs > Dm > Ot) for substitutions at these positions. Our results show that the METTL9 orthologues, although requiring an HxH target site, have evolved different substrate specificities, likely due to differing biologically relevant substrates in the respective organisms. Furthermore, we solved the X-ray structure of OtMETTL9, revealing several differences from the previously published HsMETTL9 structures that may explain its distinct substrate specificity.

• Lauretis, Angela de; Wang, Qiang; Santacroce, Marco; Thiede, Bernd; Ma, Qianli & Lyngstadaas, Ståle Petter [Show all 9 contributors for this article] (2025). A Comparative Study of the Impact of Chemical Debridement Products on the Proteomic Profile of the Salivary Pellicle and Cell Adhesion on OsseoSpeed Titanium Dental Implant Surfaces. ACS Applied Materials & Interfaces. ISSN 1944-8244. 17, p. 54579–54592. doi: 10.1021/acsami.5c14570. Full text in Research Archive Show summary

  Background and objectives: Dental implants are commonly used for tooth replacement, with excellent success rates over 10 years. However, exposure to oral cavity bacteria makes them susceptible to biofilm accumulation, which can lead to peri-implant diseases. The salivary pellicle, a protein-rich layer formed on implant surfaces, mediates bacterial adhesion and cell attachment. Effective decontamination is essential for managing bacterial infections, yet no consensus exists on the best chemical agent. Understanding how chemical agents affect the pellicle proteomic profile may guide the selection of optimal treatments. Methods: We investigate the impact of chemical decontamination agents on the salivary pellicle on OsseoSpeed-like titanium dental implant surfaces using a BCA assay and proteomics, and on cell adhesion using human gingival fibroblasts (HGF) and human bone marrow mesenchymal stem cells (hBMSC). We compare the effects of hydrogen peroxide (H2O2), Poloxamer 407 (P407), P407 + H2O2 and sodium hypochlorite + amino acids (NaOCl + AA) using the salivary pellicle and a clean titanium surface as controls. By quantifying and identifying proteins on titanium surfaces after decontamination, this study reveals how chemical agents affect the salivary pellicle. Results: All products reduced the total protein on the surfaces, with the P407 + H2O2 hydrogel demonstrating lower protein variety and superior effectiveness in preventing surface recontamination. Not all proteins were removed during decontamination procedures, and unique proteins were detected in each experimental condition, suggesting protein readsorption depends on surface chemistry. P407 + H2O2 shows the highest enrichment in integrin- and cadherin-binding proteins upon recontamination. P407 and P407 + H2O2 promoted HGFs and hBMSCs attachment. Conversely, NaOCl + AA and H2O2 showed lower HGFs and hBMSCs counts. Conclusion: The P407 + H2O2 hydrogel had the strongest decontamination effect and limited surface recontamination. Each chemical agent produced a distinct proteomic profile. P407 and P407 + H2O2 promoted initial cell adhesion.

• Labun, Kornel; Rio, Oline Eriksen; Tjeldnes, Håkon; Swirski, Michał; Komisarczuk, Anna Zofia & Haapaniemi, Emma Maria [Show all 7 contributors for this article] (2025). CHOPOFF: symbolic alignments enable fast and sensitive CRISPR off-target detection. bioRxiv - the preprint server for biology. doi: 10.1101/2025.01.06.603201. Full text in Research Archive Show summary

  ABSTRACT CRISPR/Cas systems offer powerful tools for genome editing, but their therapeutic application is hampered by the risk of unintended off-target mutations. Many molecular methods have been established to detect off-target editing, however, their sensitivity depends on first identifying potential sites using in silico methods. However, these in silico prediction methods are challenged by a trade-off between speed and sensitivity, and can fail to comprehensively detect all edited off-target sites. Here, we demonstrate that ignoring bulges has led to missing editing at off-target sites in previous studies and that continuing this practice can lead to inflated claims of fidelity. As a solution, we introduce the concept of symbolic alignments to efficiently identify off-targets without sacrificing sensitivity. We further present specialized data structures that enable rapid, alignment-free probabilistic ranking of guide RNAs based on their predicted off-target burden. Implemented in the tool CHOPOFF, these innovations accommodate mismatches, bulges (insertions/deletions), and incorporate genomic sequence variants for personalized off-target assessment. Benchmarking demonstrates that CHOPOFF significantly outperforms state-of-the-art tools in both prediction accuracy and computational speed. Availability CHOPOFF command line available at https://github.com/JokingHero/CHOPOFF.jl CHOPOFF web server available at https://crisprtools.org/chopoff

• Mamia, Katariina Aino Inkeri; Kolbeinsdottir, Solrun; Labun, Kornel; Li, Zhuokun; Komisarczuk, Anna Zofia & Keskitalo, Salla [Show all 32 contributors for this article] (2025). Precision T cell correction platform for Inborn Errors of Immunity. Molecular Therapy. ISSN 1525-0016. 33(11), p. 5624–5643. doi: 10.1016/j.ymthe.2025.08.018. Full text in Research Archive Show summary

  CRISPR/Cas9 gene editing is a promising tool to correct pathogenic variants for autologous cell therapies targeting Inborn errors of immunity (IEI). Current strategies, such as gene knockout or cDNA knock-in, address many single-gene defects but can disrupt gene expression, highlighting the need for precise correction platforms. While transplanting corrected autologous hematopoietic stem cells is a curative approach, it is unsuitable for patients with advanced disease, inflammation or acute infections. As correcting T cells is an alternative therapeutic strategy for lymphoid IEIs, we present an efficient T cell single nucleotide variant (SNV) correction platform based on homology-directed repair (HDR). By using STAT1 gain-of-function (STAT1-GOF), Cartilage Hair Hypoplasia (CHH), Deficiency of ADA2 (DADA2), and Autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) as IEI models, we demonstrate that our platform achieves up to 80% correction, with resultant functional correction of the disease phenotype in the selected models. Furthermore, we performed safety profiling using GUIDE-seq, single-cell RNA sequencing, long-read genome sequencing, and proteomics analysis and detected no genomic, transcriptomic, or proteomic aberrations. This study establishes HDR-based SNV editing as a portable method for developing clinical autologous T cell therapies and represents a promising step toward a broad-spectrum gene correction platform for treating diverse monogenic immune disorders.

• Nilsen, Jeannette; Aaen, Kristin Hovden; Benjakul, Sopisa; Ruso-Julve, Fulgencio; Greiner, Thomas Uwe & Bejan, Daniela [Show all 11 contributors for this article] (2025). Enhanced plasma half-life and efficacy of engineered human albumin-fused GLP-1 despite enzymatic cleavage of its C-terminal end. Communications Biology. 8(1). doi: 10.1038/s42003-025-08249-8. Full text in Research Archive Show summary

  Albumin has a long plasma half-life due to engagement of the neonatal Fc receptor (FcRn), which prevents intracellular degradation. However, its C-terminal end can be cleaved by carboxypeptidase A, and removal of the last leucine residue (L585) weakens receptor binding, reducing its half-life from 20 days to 3.5 days in humans. This biology has so far been overlooked when designing human albumin-fused biologics. Thus, there is a need for an engineering strategy to secure favorable FcRn binding and pharmacokinetic properties. Here, we show that a branched aliphatic amino acid or methionine at position 585 of albumin is required for optimal receptor binding, which cannot be replaced to prevent enzymatic cleavage without negatively affecting FcRn engagement. As a solution, we report that C-terminally cleaved albumin can be efficiently rescued from intracellular degradation by introducing amino acid substitutions that improve FcRn binding. This albumin-engineering strategy was also effective when applied with a therapeutic fusion partner, glucagon-like peptide 1 (GLP-1), resulting in a 2-fold increase in plasma half-life and prolonged efficacy in human FcRn transgenic mice. We demonstrate how human albumin fusions should be tailored to ensure a long plasma half-life and enhanced efficacy of fused biologics, despite potential C-terminal cleavage in vivo.

• Al-Rubaye, Noor; Mrsa, Ago; Ramirez, Manuel; Halvorsen, Trine Grønhaug & Reubsaet, Leon (2025). Smart proteolytic samplers for liquid chromatography-mass spectrometry protein quantification: Assessing trypsin immobilization via divinyl sulfone or periodate functionalization. Journal of Chromatography A. ISSN 0021-9673. 1747. doi: 10.1016/j.chroma.2025.465825. Full text in Research Archive Show summary

  In this report we try to answer the question if smart proteolytic samplers can be used for LC-MS based quantitative determination of proteins from paper. Production of smart proteolytic samplers is a two-stage process: In the first stage cellulose is functionalized while in the second stage trypsin is covalently bound. Two functionalization strategies, divinyl sulfone - and periodate functionalization, were compared under various conditions. The divinyl sulfone functionalization in combination with trypsin immobilization is here described for the first time. This functionalization chemistry shows to be equally successful as the periodate functionalization. Using Whatman Grade CF12 paper, it appears that smart proteolytic samplers with high tryptic activity can be produced. These have the capability to produce tryptic peptides with good linearity in buffered solutions: up to 0.998 for BSA (using HLVDEPQNLIK as signature peptide and IO4-functionalization) and 0.990 for Cytochrome C (using MIFAGIK as signature peptide and IO4-functionalization). The reproducibility in complex biological samples is acceptable (<20 % average RSD for divinyl sulfone functionalized paper) when internal correction is carried out. Overall, smart proteolytic samplers can be used for reliable LC-MS based determination of proteins when the correct proteotypic peptides are chosen.

• Dufour, Damien; Zhao, Xu; Chaleil, Florian; Nothnagel, Patrizia Maria Christiane; Bjørås, Magnar & Lefrançois-Martinez, Anne-Marie [Show all 8 contributors for this article] (2025). Pharmacological inhibition of SUMOylation with TAK-981 mimics genetic HypoSUMOylation in murine perigonadal white adipose tissue. Adipocyte. ISSN 2162-3945. 14(1). doi: 10.1080/21623945.2025.2474107. Full text in Research Archive Show summary

  Post-translational modification by the small ubiquitin-like modifier (SUMO) is essential for cellular differentiation and homeostasis. Here, we investigate the role of SUMOylation in adipose tissue development using TAK-981, a pharmacological inhibitor of SUMOylation. Administration of TAK-981 to mice resulted in significant defect in weight gain and adipocyte atrophy in perigonadal white adipose tissue (gWAT) depots. Gene expression analyses revealed a marked downregulation of adipogenic genes, including Pparg, Cebpa, and Fasn. Our data thus indicate that TAK-981 treatment impaired adipogenesis in gWAT, consistent with prior findings that SUMOylation supports transcriptional regulation of adipogenesis and lipid metabolism. We also found significant infiltration of immune cells and efferocytosis in gWAT. Our results thus indicate that SUMOylation inhibition using a small molecule phenocopies genetic hypoSUMOylation models, highlighting its critical role in maintaining adipocyte functionality and immune environment. These findings provide evidence that SUMOylation is essential for fat accumulation in vivo. Furthermore, given that TAK-981 is currently under clinical evaluation for the treatment of solid tumors, our results underscore the importance of considering the potential unintended effects of SUMOylation inhibition on adipose tissue in patients.

• Bjune, Jan-Inge; Laber, Samantha; Dyer, Laurence; Nothnagel, Patrizia Maria Christiane; Yamada, Shuntaro & Zhao, Xu [Show all 15 contributors for this article] (2025). IRX3 controls a SUMOylation-dependent differentiation switch in adipocyte precursor cells. Nature Communications. 16. doi: 10.1038/s41467-025-62361-1. Full text in Research Archive Show summary

  IRX3 is linked to predisposition to obesity through the FTO locus and is upregulated during early adipogenesis in risk-allele carriers, shifting adipocyte fate toward fat storage. However, how this elevated IRX3 expression influences later developmental stages remains unclear. Here we show that IRX3 regulates adipocyte fate by modulating epigenetic reprogramming. ChIP-sequencing in preadipocytes identifies over 300 IRX3 binding sites, predominantly at promoters of genes involved in SUMOylation and chromatin remodeling. IRX3 knockout alters expression of SUMO pathway genes, increases global SUMOylation, and inhibits PPARγ activity and adipogenesis. Pharmacological SUMOylation inhibition rescues these effects. IRX3 KO also reduces SUMO occupancy at Wnt-related genes, enhancing Wnt signaling and promoting osteogenic fate in 3D cultures. This fate switch is partially reversible by SUMOylation inhibition. We identify IRX3 as a key transcriptional regulator of epigenetic programs, acting upstream of SUMOylation to maintain mesenchymal identity and support adipogenesis while suppressing osteogenesis in mouse embryonic fibroblasts.

• Golnarnik, Golnaz; Thiede, Bernd; Søland, Tine Merete; Galtung, Hilde & Haug, Trude Marie (2025). Hydrogen peroxide-induced oxidative stress alters protein expression in two rat salivary acinar cell lines. Archives of Oral Biology. ISSN 0003-9969. 175. doi: 10.1016/j.archoralbio.2025.106254. Full text in Research Archive Show summary

  Objectives: This study aimed to investigate the impact of hydrogen peroxide-induced oxidative stress on the protein expression profiles of submandibular and parotid acinar cells using a proteomic approach. We sought to evaluate how oxidative stress might contribute to salivary gland dysfunction and whether the two glands respond differently.<p> <p>Design: Immortalized rat parotid gland (PG) and submandibular gland (SMG) acinar epithelial cell lines were exposed to 50 µM and 150 µM hydrogen peroxide for 24 hr, followed by protein identification and quantification via liquid chromatography-mass spectrometry. Immunofluorescence microscopy and western blot analysis validated selected protein expressions, and cell viability was assessed using trypan blue exclusion assays. <p>Results: Compared to controls, histone H4 expression increased in both cell types after hydrogen peroxide exposure, whereas voltage-dependent anion-selective channel 1, keratin 7, and keratin 8 increased only in parotid gland cells. Conversely, mitochondrial aldehyde dehydrogenase and kidney isoform glutaminase were downregulated in parotid gland cells. Basal expression of mitochondrial aldehyde dehydrogenase and catalase was higher in submandibular gland cells. At higher hydrogen peroxide concentrations, antioxidant proteins expression and cell viability were greater in submandibular gland cells compared to parotid gland cells. <p>Conclusions: Our results suggest that submandibular gland acinar cells exhibit greater resistance to oxidative stress compared to parotid gland cells, potentially due to distinct antioxidant and metabolic coping strategies. Understanding these gland-specific responses may contribute to future approaches to protect salivary glands from oxidative damage under pathological conditions.

• gjølberg, torleif; mester, simone; Calamera, Gaia; Telstad, Jenny Skjermo; Sandlie, Inger & Andersen, Jan Terje (2025). Targeting the Neonatal Fc Receptor in Autoimmune Diseases: Pipeline and Progress. BioDrugs. ISSN 1173-8804. 39(3), p. 373–409. doi: 10.1007/s40259-025-00708-2. Full text in Research Archive Show summary

  Autoimmune diseases are highly prevalent and affect people at all ages, women more often than men. The most prominent immunological manifestation is the production of antibodies directed against self-antigens. In many cases, these antibodies (Abs) drive the pathogenesis by attacking the body’s own healthy cells, causing serious health problems that may be life threatening. Most autoantibodies are of the immunoglobulin G (IgG) isotype, which has a long plasma half-life and potent effector functions. Thus, there is a need for specific treatment options that rapidly eliminate these pathogenic IgG auto-Abs. In this review, we discuss how the neonatal Fc receptor (FcRn) acts as a regulator of the high levels of not only IgG Abs, but also albumin, by rescuing both these soluble proteins from cellular catabolism, and how a molecular and cellular understanding of this complex biology has spurred an intense interest in the development of FcRn-targeting strategies for the treatment of IgG-driven autoimmune diseases. We find that this emerging therapeutic class demonstrates efficacy within several autoimmune diseases with distinct pathophysiology. This offers hope for both new therapeutic avenues for highly prevalent diseases currently treated by other means, and rare diseases with no approved therapies to date. In addition, we elaborate on studies that have led to approval of the first FcRn antagonists, the clinical progress and structural design of molecules in the pipeline, their position in the overall therapeutic landscape of autoimmunity, the design of next-generation antagonists as well as the use of this receptor-targeting principle for other therapeutic applications.

• Pati, Niladri Bhusan; Jin, Yixin; Kumar, Suman; Kyte, Jon A & Ciosk, Rafal (2025). Thermal potentiation improves IFN-γ production but not cytotoxicity in human CAR-T cells. BMC Research Notes. 18(1). doi: 10.1186/s13104-025-07249-5. Full text in Research Archive Show summary

  Objective Body temperature plays an important role in cancer, with febrile temperature generally associated with improved cancer resistance. In murine models, this resistance has been linked to the cytotoxic T cells, whose differentiation into cancer-killing effector cells is poor at lower but robust at elevated temperatures. If conserved, temperature-mediated potentiation of patient-derived T cells could be implemented to improve the existing cancer treatments, including the chimeric antigen receptor T-cell therapy (CAR T-cell therapy). Here, we tested this possibility using human STEAP1 CAR-T cells developed to target prostate cancer. Results In mice, transient exposure to febrile temperature (39–40 ºC) increases the production of IFN-γ and the cancer-killing ability of CD8 + T cells. Using a similar temperature treatment, we observed elevated levels of IFN-γ also in the human CAR-T cells. However, these cells displayed no improvement in their ability to kill cancer cells. Although we cannot discount the possibility that alternative protocols might lead to other outcomes, our findings suggest that incorporating thermal potentiation into existing protocols of CAR-T cell therapy may be more complicated than anticipated.

• Kresse, Stine Henrichson; Thorkildsen, Evy Marie; Brandt-Winge, Sara; Pharo, Heidi; Vedeld, Hege Marie & Lind, Guro Elisabeth (2025). Comparison of enzymatic and bisulfite conversion of circulating cell-free tumor DNA for DNA methylation analyses. Clinical Epigenetics. 1(17). doi: 10.1186/s13148-025-01901-4. Full text in Research Archive Show summary

  Background Detection of DNA methylation biomarkers in circulating cell-free DNA (cfDNA) has great clinical potential for cancer management, but there is a high need for method optimization and standardization. Bisulfite conversion of DNA is the gold-standard pre-treatment method for DNA methylation analyses, but causes also DNA fragmentation and loss. Enzymatic conversion of DNA represents a promising alternative due to the more gentle treatment minimizing damage to DNA. The aim of this study was to evaluate and compare enzymatic and bisulfite conversion to identify the best pre-treatment method for detecting DNA methylation biomarkers in cfDNA from plasma using droplet digital PCR (ddPCR). Results The performance of the NEBNext Enzymatic Methyl-seq Kit (both the full kit intended for sequencing and the sub-component NEBNext Enzymatic Methyl-seq Conversion Module) and the EpiTect Plus DNA Bisulfite Kit was evaluated and compared using normal cfDNA and tumor cfDNA samples from colorectal cancer patients. The cytosine conversion efficiency was 99–100% for both enzymatic and bisulfite conversion. Enzymatic conversion resulted in longer DNA fragments with higher peak fragment sizes compared to bisulfite conversion, but the DNA recovery was considerably lower after enzymatic conversion (34–47%) compared to bisulfite conversion (61–81%). For enzymatic conversion, the full kit gave slightly better DNA recovery than the conversion module. A comparison of five magnetic bead brands, as well as several different magnetic bead-to-sample ratios revealed no major improvements in DNA recovery for the enzymatic conversion. DNA methylation of the biomarker BCAT1 was detected at similar rates in parallel tumor cfDNA samples pre-treated with either enzymatic or bisulfite conversion. However, enzymatic conversion resulted in lower number of positive droplets for both target and control ddPCR assays, in line with the lower DNA recovery after conversion. Conclusions Based on a thorough evaluation of enzymatic and bisulfite conversion of cfDNA using ddPCR, bisulfite conversion emerges as the best pre-treatment method due to higher DNA recovery after conversion and higher number of positive droplets in the ddPCR reactions.

• Drevland, Ole Martin; Muinck, Eric de; Trosvik, Pål; Hammerstad, Marta; Kvitne, Kine Eide & Midtvedt, Karsten [Show all 8 contributors for this article] (2025). Microbiome-derived reactivation of mycophenolate explains variations in enterohepatic recirculation in kidney transplant recipients. Microbiome. 13(1). doi: 10.1186/s40168-025-02142-6. Full text in Research Archive Show summary

  Background The pivotal role of microbes in drug metabolism is increasingly recognized, as variation in the gut microbiome composition between individuals has been shown to impact systemic drug exposure, efficacy and toxicity. Mycophenolate mofetil (MMF) is a cornerstone in immunosuppressive therapy following solid organ transplantation. However, dosing and tolerance are challenged by significant pharmacokinetic variability among patients, largely due to variable degrees of enterohepatic recirculation of mycophenolic acid (MPA), the active moiety of MMF. It is hypothesized that the variability in MPA recirculation is driven by gut microbiome-derived β-glucuronidase (β-GUS) mediated cleavage of MPA-glucuronide (MPAG) excreted in the bile. Here, we investigated the bidirectional interaction between MPA and the gut microbiome in kidney transplant recipients, using a combination of in vivo and in vitro data. Results We compared the fecal microbiomes of kidney transplant recipients (n = 21) both pre- and post-transplantation to healthy individuals (n = 15) using shotgun metagenomic sequencing. We also determined the individual microbiome-derived reactivation rate of MPAG to MPA and show a strong positive correlation between this reactivation rate and the degree of MPA enterohepatic recirculation in vivo. Through metagenomic analysis, the reactivation rate of MPA was linked to specific gut microbial species. In particular, specific β-GUS gene variants associated with Faecalibacterium prausnitzii showed a strong impact on the conversion of MPAG to MPA. Furthermore, our study confirmed a significant shift in microbial composition post-transplantation and revealed notable fluctuations in species such as F. prausnitzii and Akkermansia muciniphila across different time points after transplantation. Lastly, we provide evidence that the microbiome-derived reactivation rate of MPA is linked to specific beta-glucuronidase alleles. Conclusions We highlight for the first time that the ex vivo determined reactivation rate of MPA explains the variation of enterohepatic recirculation, emphasizing the important role of F. prausnitzii in this process. More broadly, our findings suggest that the gut microbiome significantly influences the degree of enterohepatic recirculation of MPA, providing valuable insights that could be relevant for optimizing individualized immunosuppressive drug dosing in transplant patients.

• mester, simone; Chan, Chilam; Lustig, Marta; Foss, Stian; Jansen, J. H. Marco & Herigstad, Marie Leangen [Show all 17 contributors for this article] (2025). Engineering of anticancer human immunoglobulin A equipped with albumin for enhanced plasma half-life. PNAS Nexus. 4(2). doi: 10.1093/pnasnexus/pgaf042. Full text in Research Archive Show summary

  Most therapeutic antibodies are based on immunoglobulin G (IgG) due to their potent effector functions and long plasma half-life. However, also monomeric IgA has emerged as an attractive candidate for cancer treatment as, upon specific binding to tumor cells, it can activate myeloid cells, like polymorphonuclear leukocytes and macrophages, to kill the tumor cells by engaging the Fc α receptor I (FcαRI). Despite this favorable property, human IgA has a short plasma half-life in both mice and humans, which is clearly limiting preclinical studies in a translational perspective. Here, we report on albumin-equipped designs of human IgA antibodies that are long acting due to tailored binding to the human form of neonatal Fc receptor (FcRn), which is a natural plasma half-life regulator of albumin. Importantly, this was achieved without compromising the ability of IgA to engage and activate FcαRI-expressing effector cells for tumor cell killing in vitro and in vivo in a new mouse model transgenic for the human forms of FcRn and FcαRI. We further show that the potency of the engineered long-acting human IgA against tumor cells with intermediate target antigen expression levels could be enhanced by myeloid checkpoint inhibitors targeting the signal regulatory protein α-CD47 axis.

• Tierney, Jack A.S.; Świrski, Michał I.; Tjeldnes, Håkon; Kiran, Anmol M.; Carancini, Gionmattia & Kiniry, Stephen J. [Show all 10 contributors for this article] (2025). RiboSeq.Org: an integrated suite of resources for ribosome profiling data analysis and visualization. Nucleic Acids Research (NAR). ISSN 0305-1048. 53(1), p. D268–D274. doi: 10.1093/nar/gkae1020. Full text in Research Archive
• Deutsch, Eric W; Kok, Leron W; Mudge, Jonathan M; Ruiz-Orera, Jorge; Fierro-Monti, Ivo & Sun, Zhi [Show all 43 contributors for this article] (2024). High-quality peptide evidence for annotating non-canonical open reading frames as human proteins. bioRxiv. ISSN 2692-8205. doi: 10.1101/2024.09.09.612016. Full text in Research Archive Show summary

  Abstract A major scientific drive is to characterize the protein-coding genome as it provides the primary basis for the study of human health. But the fundamental question remains: what has been missed in prior genomic analyses? Over the past decade, the translation of non-canonical open reading frames (ncORFs) has been observed across human cell types and disease states, with major implications for proteomics, genomics, and clinical science. However, the impact of ncORFs has been limited by the absence of a large-scale understanding of their contribution to the human proteome. Here, we report the collaborative efforts of stakeholders in proteomics, immunopeptidomics, Ribo-seq ORF discovery, and gene annotation, to produce a consensus landscape of protein-level evidence for ncORFs. We show that at least 25% of a set of 7,264 ncORFs give rise to translated gene products, yielding over 3,000 peptides in a pan-proteome analysis encompassing 3.8 billion mass spectra from 95,520 experiments. With these data, we developed an annotation framework for ncORFs and created public tools for researchers through GENCODE and PeptideAtlas. This work will provide a platform to advance ncORF-derived proteins in biomedical discovery and, beyond humans, diverse animals and plants where ncORFs are similarly observed.

• Caulier, Benjamin William; Joaquina, Sandy; Gelebart, Pascal Francois; Dowling, Tara Helen Steinsland; Kaveh, Fatemeh & Thomas, Moritz [Show all 20 contributors for this article] (2024). CD37 is a safe chimeric antigen receptor target to treat acute myeloid leukemia. Cell Reports Medicine. 5(6). doi: 10.1016/j.xcrm.2024.101572. Full text in Research Archive Show summary

  Acute myeloid leukemia (AML) is characterized by the accumulation of immature myeloid cells in the bone marrow and the peripheral blood. Nearly half of the AML patients relapse after standard induction therapy, and new forms of therapy are urgently needed. Chimeric antigen receptor (CAR) T therapy has so far not been successful in AML due to lack of efficacy and safety. Indeed, the most attractive antigen targets are stem cell markers such as CD33 or CD123. We demonstrate that CD37, a mature B cell marker, is expressed in AML samples, and its presence correlates with the European LeukemiaNet (ELN) 2017 risk stratification. We repurpose the anti-lymphoma CD37CAR for the treatment of AML and show that CD37CAR T cells specifically kill AML cells, secrete proinflammatory cytokines, and control cancer progression in vivo. Importantly, CD37CAR T cells display no toxicity toward hematopoietic stem cells. Thus, CD37 is a promising and safe CAR T cell AML target.

• Hansen, Stian Bakke; Ünal, Bilal; Kuzu, Ömer Faruk & Saatcioglu, Fahri (2024). Immunological facets of prostate cancer and the potential of immune checkpoint inhibition in disease management. Theranostics. 14(18), p. 6913–6934. doi: 10.7150/thno.100555. Full text in Research Archive Show summary

  Prostate cancer (PCa) is the most common non-cutaneous cancer in men and a major cause of cancer-related deaths. Whereas localized PCa can be cured by surgery and radiotherapy, metastatic disease can be treated, but is not curable. Inhibition of androgen signaling remains the main therapeutic intervention for treatment of metastatic PCa, in addition to chemotherapy, radionuclide therapy and emerging targeted therapies. Although initial responses are favorable, resistance to these therapies invariably arise with development of castration resistant PCa (CRPC) and lethal phenotypes. Recent findings have implicated the crosstalk between PCa cells and the tumor microenvironment (TME) as a key factor for disease progression and metastasis, and the immune system is becoming an increasingly attractive target for therapy. Given the striking success of immune checkpoint inhibitors (ICIs) in various cancer types, preclinical and clinical studies have begun to explore their potential in PCa. It has become clear that the PCa TME is largely immunosuppressive, and ICI therapy does not have efficacy for PCa. Intense effort is therefore being made in the field to understand the mechanisms of suppression and to turn the immunosuppressive TME into an immune active one that would enable ICI efficacy. Herein we examine this recent body of knowledge and how the mutational landscape of PCa integrates with an immunosuppressive TME to circumvent ICI-mediated T-cell activity and tumor killing. We then review the emerging potential success of combinatorial ICI approaches, utility of careful patient selection, and potential novel strategies to improve the efficacy of ICI for PCa therapy.

• Foss, Stian; Sakya, Siri Aastedatter; Aguinagalde, Leire; Lustig, Marta; Shaughnessy, Jutamas & Cruz, Ana Rita [Show all 25 contributors for this article] (2024). Human IgG Fc-engineering for enhanced plasma half-life, mucosal distribution and killing of cancer cells and bacteria. Nature Communications. 15(1). doi: 10.1038/s41467-024-46321-9. Full text in Research Archive Show summary

  Monoclonal IgG antibodies constitute the fastest growing class of therapeutics. Thus, there is an intense interest to design more potent antibody formats, where long plasma half-life is a commercially competitive differentiator affecting dosing, frequency of administration and thereby potentially patient compliance. Here, we report on an Fc-engineered variant with three amino acid substitutions Q311R/M428E/N434W (REW), that enhances plasma half-life and mucosal distribution, as well as allows for needle-free delivery across respiratory epithelial barriers in human FcRn transgenic mice. In addition, the Fc-engineered variant improves on-target complement-mediated killing of cancer cells as well as both gram-positive and gram-negative bacteria. Hence, this versatile Fc technology should be broadly applicable in antibody design aiming for long-acting prophylactic or therapeutic interventions.

• Falnes, Pål (2024). Closing in on human methylation-the versatile family of seven-β-strand (METTL) methyltransferases. Nucleic Acids Research (NAR). ISSN 0305-1048. 52(19), p. 11423–11441. doi: 10.1093/nar/gkae816. Full text in Research Archive Show summary

  Methylation is a common biochemical reaction, and a number of methyltransferase (MTase) enzymes mediate the various methylation events occurring in living cells. Almost all MTases use the methyl donor S-adenosylmethionine (AdoMet), and, in humans, the largest group of AdoMet-dependent MTases are the so-called seven-β-strand (7BS) MTases. Collectively, the 7BS MTases target a wide range of biomolecules, i.e. nucleic acids and proteins, as well as several small metabolites and signaling molecules. They play essential roles in key processes such as gene regulation, protein synthesis and metabolism, as well as neurotransmitter synthesis and clearance. A decade ago, roughly half of the human 7BS MTases had been characterized experimentally, whereas the remaining ones merely represented hypothetical enzymes predicted from bioinformatics analysis, many of which were denoted METTLs (METhylTransferase-Like). Since then, considerable progress has been made, and the function of > 80% of the human 7BS MTases has been uncovered. In this review, I provide an overview of the (estimated) 120 human 7BS MTases, grouping them according to substrate specificities and sequence similarity. I also elaborate on the challenges faced when studying these enzymes and describe recent major advances in the field.

• Wu, Chenyu; Gunnarsson, Einar Bjarki; Myklebust, Even Moa; Luque, Alvaro Kohn; Tadele, Dagim Shiferaw & Enserink, Jorrit [Show all 9 contributors for this article] (2024). Using birth-death processes to infer tumor subpopulation structure from live-cell imaging drug screening data. PLoS Computational Biology. ISSN 1553-734X. 20(3). doi: 10.1371/journal.pcbi.1011888. Full text in Research Archive Show summary

  Tumor heterogeneity is a complex and widely recognized trait that poses significant challenges in developing effective cancer therapies. In particular, many tumors harbor a variety of subpopulations with distinct therapeutic response characteristics. Characterizing this heterogeneity by determining the subpopulation structure within a tumor enables more precise and successful treatment strategies. In our prior work, we developed PhenoPop, a computational framework for unravelling the drug-response subpopulation structure within a tumor from bulk high-throughput drug screening data. However, the deterministic nature of the underlying models driving PhenoPop restricts the model fit and the information it can extract from the data. As an advancement, we propose a stochastic model based on the linear birth-death process to address this limitation. Our model can formulate a dynamic variance along the horizon of the experiment so that the model uses more information from the data to provide a more robust estimation. In addition, the newly proposed model can be readily adapted to situations where the experimental data exhibits a positive time correlation. We test our model on simulated data (in silico) and experimental data (in vitro), which supports our argument about its advantages.

• Zhang, Tingting; Zhao, Faming; Hu, Yanxia; Wei, Jinlan; Cui, Fengzhen & Lin, Yahang [Show all 8 contributors for this article] (2024). Environmental monobutyl phthalate exposure promotes liver cancer via reprogrammed cholesterol metabolism and activation of the IRE1α-XBP1s pathway. Oncogene. ISSN 0950-9232. 43, p. 2355–2370. doi: 10.1038/s41388-024-03086-1. Full text in Research Archive
• Ma, Qianli; Piaia, Lya; Loča, Dagnija; Rubenis, Kristaps; Locs, Janis & Thiede, Bernd [Show all 8 contributors for this article] (2024). Soluble Proteins From Conventional and Organic Eggshell Membranes With Different Proteomic Profiles Show Similar In Vitro Biofunctions. Journal of Biomedical Materials Research. Part A. ISSN 1549-3296. 113(1). doi: 10.1002/jbm.a.37848. Full text in Research Archive Show summary

  The eggshell membrane (ESM), resembling the extracellular matrix, acts as a protective barrier against bacterial invasion and offers various biofunctions due to its porous structure and protein-rich composition, such as ovalbumin, ovotransferrin, collagen, soluble protein, and antimicrobial proteins. However, ESM's structure primarily comprises disulfide bonds and heterochains, challenging protein solubilization/extraction. Therefore, the method of dissolving and extracting bioactive protein components in ESM has extremely high potential value and importance for exploring the reuse of egg waste and environmental protection. In this study we extracted soluble ESM proteins (SEPs) from conventional (industrial-fed) and organic (free-grounded) using an acidic 3-mercaptopropionic acid (3-MPA) extraction strategy. FTIR was used to monitor the chemical changes in the ESM, and LC-MS/MS was employed to conduct the proteomic analysis. The biocompatibility and the effects of SEPs cocktails on extracellular matrix (ECM) synthesis were also investigated. The results indicated that the acidic 3-MPA strategy could effectively alter the ESM chemical composition, thereby facilitating SEPs extraction. The SEPs from conventional and organic eggs have different protein profiles but with partial overlapping. SEPs from different sources exhibited similarly desirable biosafety profiles and similar dose-dependent promotion of osteoblastic (ECM) component synthesis, suggesting that different egg sources may contribute to consistent core biological functions of protein products but may also introduce different functional priorities

• Mrsa, Ago; Nardini, Greta; Halvorsen, Trine Grønhaug; Thiede, Bernd & Reubsaet, Leon (2024). One-step functionalization of paper and simplified antibody immobilization for on-the-spot immunocapture from dried serum in liquid chromatography-tandem mass spectrometry based targeted protein determination. Journal of Mass Spectrometry. ISSN 1076-5174. 59(1). doi: 10.1002/jms.4989. Full text in Research Archive Show summary

  This work aimed to simplify and improve the process of binding monoclonal antibodies (mAbs) covalently to filter paper for use in dried blood spot sampling, enabling instant capture of protein biomarkers for targeted protein determination. Incorporating the necessary immunocapture sample preparation step in the initial sampling stage saves time and reduces the workload. The biomarker human chorionic gonadotropin (hCG) was used as the model analyte. The antibody-based paper samplers were prepared by functionalizing paper discs (6 mm) through a simple reaction using divinyl sulfone (DVS). After DVS activation, the paper discs were incubated with E27 hCG mAbs, followed by 0.05% tween/phosphate buffer saline to block the surface. After sample application and drying, the discs only needed to be washed before tryptic digestion and finally analysed on a nanoliquid chromatography–tandem mass spectrometry system. The finished DVS-mAbs samplers could selectively capture hCG (100 ng/mL) from human serum, with a recovery of 50%. Sample clean-up reduced the number of identified proteins from 132 to 82 before and after wash, respectively, with a 70% reduction in serum albumin signal while still retaining hCG on the sampler during the washing protocol. An evaluation of the samplers revealed excellent linearity (R2 = 0.9995) for hCG in serum with relative standard deviations below 15%. This work has presented the first ever reported paper samplers immobilized with antibodies utilizing DVS chemistry, showing promise in the future of paper-based sampling.

• Saatcioglu, Fahri & Kuzu, Omer Faruk (2024). GeneSetR: A web server for gene set analysis based on genome-wide Perturb-Seq data. BioRxiv. ISSN 0362-4331. doi: 10.1101/2023.09.18.558211. Full text in Research Archive
• Fernandez, Javier Gutierrez; Hersleth, Hans-Petter & Hammerstad, Marta (2024). The crystal structure of myco­thiol di­sulfide reductase (Mtr) provides mechanistic insight into the specific low-molecular-weight thiol reductase activity of Actinobacteria. Acta Crystallographica Section D: Structural Biology. 80(3), p. 181–193. doi: 10.1107/S205979832400113X. Full text in Research Archive Show summary

  Low-molecular-weight (LMW) thiols are involved in many processes in all organisms, playing a protective role against reactive species, heavy metals, toxins and antibiotics. Actinobacteria, such as Mycobacterium tuberculosis, use the LMW thiol mycothiol (MSH) to buffer the intracellular redox environment. The NADPH-dependent FAD-containing oxidoreductase mycothiol disulfide reductase (Mtr) is known to reduce oxidized mycothiol disulfide (MSSM) to MSH, which is crucial to maintain the cellular redox balance. In this work, the first crystal structures of Mtr are presented, expanding the structural knowledge and understanding of LMW thiol reductases. The structural analyses and docking calculations provide insight into the nature of Mtrs, with regard to the binding and reduction of the MSSM substrate, in the context of related oxidoreductases. The putative binding site for MSSM suggests a similar binding to that described for the homologous glutathione reductase and its respective substrate glutathione disulfide, but with distinct structural differences shaped to fit the bulkier MSSM substrate, assigning Mtrs as uniquely functioning reduc- tases. As MSH has been acknowledged as an attractive antitubercular target, the structural findings presented in this work may contribute towards future anti- tuberculosis drug development.

• Johansen, Anette; Thiede, Bernd; Anonsen, Jan Haug & Nilsson, Göran Erik (2024). Phosphoproteomic changes in response to anoxia are tissue-specific in the anoxia-tolerant crucian carp (Carassius carassius). Frontiers in Physiology. 15. doi: 10.3389/fphys.2024.1407834. Full text in Research Archive Show summary

  Crucian carp (Carassius carassius), a freshwater fish, can survive chronic anoxia for several months at low temperatures. Consequently, anoxia-related physiological and biochemical adaptations in this species have been studied for more than half a century. Still, despite for the well-known role of protein phosphorylation in regulating cellular processes, no studies have comprehensively characterized the phosphoproteome in crucian carp. In this study, we report the global phosphoproteome in crucian carp brain and liver during anoxia and reoxygenation. By applying a bottom-up proteomic approach on enriched phosphopeptides we found that the brain phosphoproteome shows surprisingly few changes during anoxia-reoxygenation exposure with only 109 out of 4200 phosphopeptides being differentially changed compared to normoxic controls. By contrast, in the liver 395 out of 1287 phosphopeptides changed. Although most changes occurred in the liver phosphoproteome, the pattern of changes indicated metabolic depression and decreased translation in both brain and liver. We also found changes in phosphoproteins involved in apoptotic regulation and reactive oxygen species handling in both tissues. In the brain, some of the most changed phosphopeptides belonged to proteins involved in central nervous system development and neuronal activity at the synaptic cleft. Changed phosphoproteins specific for liver tissue were related to glucose metabolism, such as glycolytic flux and glycogenolysis. In conclusion, protein phosphorylation in response to anoxia and reoxygenation showed both common and tissue-specific changes related to the functional differences between brain and liver.

• Corrales, Jeanne Gaspar; Alonso, Lucia Ramos; González-Sabín, Javier; Ríos-Lombardía, Nicolás; Trevijano-Contador, Nuria & Berg, Henriette Engen [Show all 12 contributors for this article] (2024). Characterization of a selective, iron-chelating antifungal compound that disrupts fungal metabolism and synergizes with fluconazole. Microbiology spectrum. 12(2). doi: 10.1128/spectrum.02594-23. Full text in Research Archive Show summary

  Fungal infections are a growing global health concern due to the limited number of available antifungal therapies as well as the emergence of fungi that are resistant to first-line antimicrobials, particularly azoles and echinocandins. Development of novel, selective antifungal therapies is challenging due to similarities between fungal and mammalian cells. An attractive source of potential antifungal treatments is provided by ecological niches co-inhabited by bacteria, fungi, and multicellular organisms, where complex relationships between multiple organisms have resulted in evolution of a wide variety of selective antimicrobials. Here, we characterized several analogs of one such natural compound, collismycin A. We show that NR-6226C has antifungal activity against several pathogenic Candida species, including C. albicans and C. glabrata, whereas it only has little toxicity against mammalian cells. Mechanistically, NR-6226C selectively chelates iron, which is a limiting factor for pathogenic fungi during infection. As a result, NR-6226C treatment causes severe mitochondrial dysfunction, leading to formation of reactive oxygen species, metabolic reprogramming, and a severe reduction in ATP levels. Using an in vivo model for fungal infections, we show that NR-6226C significantly increases survival of Candida-infected Galleria mellonella larvae. Finally, our data indicate that NR-6226C synergizes strongly with fluconazole in inhibition of C. albicans. Taken together, NR-6226C is a promising antifungal compound that acts by chelating iron and disrupting mitochondrial functions.

• Vedeld, Hege Marie; Pharo, Heidi Dietrichson; Sørbø, Anne; Brandt-Winge, Sara; Five, May-Britt & Jeanmougin, Marine Anne-Lise Francoise [Show all 9 contributors for this article] (2024). Distinct longitudinal patterns of urine tumor DNA in patients undergoing surveillance for bladder cancer. Molecular Oncology. ISSN 1574-7891. 18, p. 2684–2695. doi: 10.1002/1878-0261.13639. Full text in Research Archive Show summary

  Cystoscopy is the gold standard for surveillance of non-muscle invasive bladder cancer (NMIBC), but the procedure is invasive and has suboptimal accuracy. The aim of this study was to investigate the potential of analyzing urine samples for the presence of urine tumor DNA (utDNA) to replace cystoscopy for surveillance of bladder cancer recurrence. In this longitudinal, prospective, and observational study, 47 patients were followed for recurrence for 2 years, involving analysis of utDNA using the BladMetrix DNA methylation biomarker test at each cystoscopy. In total, utDNA was detected in 21/23 recurrences (91% sensitivity), including 5/5 T1, T2, and carcinoma in situ (CIS) tumors (100%) and 10/12 Ta tumors (83%), with < 1% false-negative test results. Importantly, utDNA analysis showed the potential to reduce the number of cystoscopies by 55%, benefitting 79% of the patients. Eleven of 23 recurrences (48%) were detected earlier with utDNA than with cystoscopy, and distinct patterns of residual utDNA post-surgery indicated minimal residual disease (MRD) or field effect in 6% and 15% of the patients, respectively. In conclusion, utDNA analysis shows high sensitivity to detect tumor recurrence, potential to reduce the number of cystoscopies, and promise to guide patient-specific surveillance regimens.

• Bjørnestad, Synne Arstad; Solbakken, Monica Hongrø; Krokene, Pia; Thiede, Bernd; Hylland, Ketil & Jakobsen, Kjetill Sigurd [Show all 9 contributors for this article] (2024). The Atlantic Cod MHC I compartment has the properties needed for cross-presentation in the absence of MHC II. Scientific Reports. 14(1). doi: 10.1038/s41598-024-76225-z. Full text in Research Archive Show summary

  Atlantic cod has a peculiar immune system, characterized by the loss of Major Histocompatibility Complex (MHC) class II pathway, and an extreme expansion of the MHC class I gene repertoire. This has led to the hypothesis that some of the MHC I variants have replaced MHC II by presenting exogenous-peptides in a process similar to cross-presentation. In mammals, MHC I loads endogenous antigens in the endoplasmic reticulum, but we recently found that different Atlantic cod MHC I gene variants traffic to endolysosomes. There, they colocalize with Tapasin and other components of the peptide-loading complex, indicating a plausible peptide-loading system outside the endoplasmic reticulum. In this study, we further characterize the identity of the Atlantic cod MHC I compartment (cMIC). We found that, similarly to mammalian MHC II compartment, cMIC contains late endosomal markers such as Rab7, LAMP1 and CD63. Furthermore, we identified Hsp90b1 (also known as grp94) and LRP1 (also known as CD91) as interactors of MHC I by mass spectrometry. As these two proteins are involved in cross-presentation in mammals, this further suggests that Atlantic cod MHC I might use a similar mechanism to present exogenous peptides, thus, compensating for the absence of MHC II.

• Fuentes-Martín, Raúl; Duran, Pilar Ayuda; Hanes, Robert; Gallego-Yerga, Laura; Wolterinck, Lisanne & Enserink, Jorrit [Show all 8 contributors for this article] (2024). Promising anti-proliferative indolic benzenesulfonamides alter mechanisms with sulfonamide nitrogen substituents. European Journal of Medicinal Chemistry. ISSN 0223-5234. 275, p. 1–20. doi: 10.1016/j.ejmech.2024.116617. Full text in Research Archive Show summary

  Agents that cause apoptotic cell death by interfering with tubulin dynamics, such as vinblastine and paclitaxel, are an important class of chemotherapeutics. Unfortunately, these compounds are substrates for multidrug resistance (MDR) pumps, allowing cancer cells to gain resistance to these chemotherapeutics. The indolesulfonamide family of tubulin inhibitors are not excluded by MDR pumps and have a promising activity profile, although their high lipophilicity is a pharmacokinetic limitation for their clinical use. Here we present a new family of N-indolyl-3,4,5-trimethoxybenzenesulfonamide derivatives with modifications on the indole system at positions 1 and 3 and on the sulfonamide nitrogen. We synthesized and screened against HeLa cells 34 novel indolic benzenesulfonamides. The most potent derivatives (1.7–109 nM) were tested against a broad panel of cancer cell lines, which revealed that substituted benzenesulfonamides analogs had highest potency. Importantly, these compounds were only moderately toxic to non-tumorigenic cells, suggesting the presence of a therapeutic index. Consistent with known clinical anti-tubulin agents, these compounds arrested the cell cycle at G2/M phase. Mechanistically, they induced apoptosis via caspase 3/7 activation, which occurred during M arrest. The substituents on the sulfonamide nitrogen appeared to determine different mechanistic results and cell fates. These results suggest that the compounds act differently depending on the bridge substituents, thus making them very interesting as mechanistic probes as well as potential drugs for further development.

• Tierney, Jack A.S.; Sẃirski, Michał; Tjeldnes, Håkon; Mudge, Jonathan M.; Kufel, Joanna & Whiffin, Nicola [Show all 8 contributors for this article] (2024). Ribosome decision graphs for the representation of eukaryotic RNA translation complexity. Genome Research. ISSN 1088-9051. 34(4), p. 530–538. doi: 10.1101/gr.278810.123. Full text in Research Archive
• Korkmaz, Asli; Bernhardsen, Guro Pauck; Cirit, Burcu; Suzer, Gayem Koprucu; Kayan, Hale & Biçmen, Hülya [Show all 11 contributors for this article] (2024). Sudarshan Kriya Yoga Breathing and a Meditation Program for Burnout Among Physicians: A Randomized Clinical Trial. JAMA Network Open. 7(1). doi: 10.1001/jamanetworkopen.2023.53978. Full text in Research Archive Show summary

  Importance Physicians are exposed to high stress and strain that results in burnout, which affects them, their families, their patients, and the entire health care system; thus, there is an urgent need to develop methods to increase the resiliency of physicians. Sudarshan Kriya Yoga (SKY) is a comprehensive yoga breathing and meditation-based program that is a potential approach to mitigate physician burnout. Objective To determine whether SKY can reduce psychological distress and improve wellness in physicians. Design, Setting, and Participants This randomized clinical trial assessed the potential efficacy of SKY compared with a stress management education (SME) training as control. This study was conducted online from November 11, 2021, to March 14, 2022, and included physicians from Turkey, Germany, and Dubai. Both the SKY and the SME control groups received 1.5 hours of training for 3 consecutive days via a group video conference call. Participants were physicians willing to do some form of relaxation exercise everyday for 2 months. Exclusion criteria included presence of major illness and maintaining a regular mind-body program practice. Statistical analysis took place from March to November 2023. Interventions Participants were randomly assigned 1:1 into 2 groups—the SKY group or the SME (control) group—using a computer algorithm. After the 3-day instruction period, the participants in the SKY group practiced for approximately 30 minutes per day on their own and participated in a weekly 1-hour, group-based online follow-up practice. After the 3-day instruction period, participants in the SME group reviewed and applied the notes from stress management education training at their initiative and had a weekly 1-hour group-based online follow-up session. Main Outcomes and Measures The primary outcomes were stress and depression (measured by the 42-item Depression, Anxiety, and Stress Scale [DASS-42]) and insomnia measured by the Regensburg Insomnia Scale (RIS) with primary end point at 8 weeks. Secondary outcomes included anxiety (DASS-42); optimism (Life Orientation Test-Revised [LOT-R]); professional fulfillment, work exhaustion, interpersonal disengagement, and overall burnout (Professional Fulfillment Index [PFI]); and self-reported professional errors (Self-Reported Professional Error Questionnaire). Results This study included 129 participants (SME, 63 participants [48.9%]; SKY, 66 participants [51.1%]; 115 females [89.2%]; 14 males [10.8%]; mean [SD] age, 46.2 [9.0] years). Compared with the SME control group, participants in the SKY group had significantly decreased stress on the DASS-42 at posttraining (difference, −6.8 points; 95% CI, −9.6 to −4.1 points; P = .006) and at postintervention (difference, −6.0 points; 95% CI, −8.8 to −3.3 points; P = .03), significantly decreased depression at posttraining (difference, −5.7 points; 95% CI, −8.6 to −2.8 points; P < .001) and postintervention (difference, −5.4 points; 95% CI, −8.3 to −2.5 points; P < .001), and significantly decreased anxiety at postintervention. In addition, there was a significant decrease in insomnia from baseline to postintervention in the SKY group (difference, −0.3 points; 95% CI, −2.3 to 1.7 points; P = .01). The SKY group also showed significantly increased professional fulfillment as well as significant decreases in work exhaustion, interpersonal disengagement, and burnout. There was no effect on self-reported medical errors. Conclusions and Relevance In this randomized clinical trial, physicians who regularly practiced SKY throughout a 2-month period experienced improvements in wellness and decreased burnout. These data suggest that SKY may be an effective, practical, and safe strategy to increase wellness and mitigate burnout in physicians. Trial Registration ClinicalTrials.gov Identifier: NCT05956470

• Seybold, Anna Cosima; Kumar, Suman; Tumu, Sharat Chandra & Hausen, Harald (2024). Neurons with larval synaptic targets pioneer the later nervous system in the annelid Malacoceros fuliginosus. Frontiers in Neuroscience. ISSN 1662-4548. 18. doi: 10.3389/fnins.2024.1439897. Full text in Research Archive Show summary

  Comparative studies on the development of nervous systems have a significant impact on understanding animal nervous system evolution. Nevertheless, an important question is to what degree neuronal structures, which play an important role in early stages, become part of the adult nervous system or are relevant for its formation. This is likely in many direct developers, but it is not the case in forms with catastrophic metamorphosis. It is not clear in many forms with gradual metamorphosis. This introduces uncertainty in tracing the evolution of nervous systems and of larval forms. One of the prominent larval characteristics of numerous planktonic marine organisms is the epidermal ciliation used for swimming and steering, which disappears during metamorphosis. Therefore, the neuronal elements controlling the ciliary beating are often assumed to vanish with the cilia and regarded as purely larval specializations. With volume EM, we followed the neuronal targets of the very first pioneer neurons at the apical and posterior ends of the larva of the annelid Malacoceros fuliginosus. We observed that all of these pioneers appear to have a dual function. Although they are laying the paths for the later adult nervous system, they also make synaptic contact with the main ciliated ring of the larva. We propose that the formation of the later adult nervous system and the innervation of the larval locomotory organ are indeed closely linked to each other. This has implications for understanding the early nervous system development of marine larvae and for existing hypotheses on nervous system evolution.

• Ünal, Bilal; Kuzu, Omer; Yang, Jin; Osorio, Daniel; Kildal, Wanja & Pradhan, Manohar [Show all 14 contributors for this article] (2024). Targeting IRE1α reprograms the tumor microenvironment and enhances anti-tumor immunity in prostate cancer. Nature Communications. 15(1), p. 1–21. doi: 10.1038/s41467-024-53039-1. Full text in Research Archive Show summary

  Unfolded protein response (UPR) is a central stress response pathway that is hijacked by tumor cells for their survival. Here, we find that IRE1α signaling, one of the canonical UPR arms, is increased in prostate cancer (PCa) patient tumors. Genetic or small molecule inhibition of IRE1α in syngeneic mouse PCa models and an orthotopic model decreases tumor growth. IRE1α ablation in cancer cells potentiates interferon responses and activates immune system related pathways in the tumor microenvironment (TME). Single-cell RNA-sequencing analysis reveals that targeting IRE1α in cancer cells reduces tumor-associated macrophage abundance. Consistently, the small molecule IRE1α inhibitor MKC8866, currently in clinical trials, reprograms the TME and enhances anti-PD-1 therapy. Our findings show that IRE1α signaling not only promotes cancer cell growth and survival but also interferes with anti-tumor immunity in the TME. Thus, targeting IRE1α can be a promising approach for improving anti-PD-1 immunotherapy in PCa.

• Winiarczyk, Mateusz; Thiede, Bernd; Utheim, Tor Paaske; Kaarniranta, Kai; Winiarczyk, Dagmara & Michalak, Katarzyna [Show all 7 contributors for this article] (2024). Oxidative Stress, Persistent Inflammation and Blood Coagulation Alterations in Serum Proteome of Patients with Neovascular Age-Related Macular Degeneration. Life. 14(5). doi: 10.3390/life14050624. Full text in Research Archive Show summary

  Neovascular age-related macular degeneration (AMD) is a major cause of irreversible blindness in elderly populations in developed countries. AMD’s etiopathology is multifactorial, with strong environmental and genetic components, but the exact molecular pathomechanisms underlying the disease are still unknown. In this study, we analyzed blood serum collected from 74 neovascular AMD patients and 58 healthy controls to identify proteins that may serve as potential biomarkers and expand our knowledge about the etiopathogenesis of the disease. The study revealed 17 differentially expressed proteins—11 up-regulated and 6 down-regulated—in neovascular AMD, which are involved in the biological processes previously linked with the disease—oxidative stress and persistent inflammation, impaired cellular transport, lipid metabolism and blood coagulation. In conclusion, the differences in the expressions of the proteins identified in this study may contribute to our understanding of the mechanisms underlying AMD and possibly serve in future as promising biomarkers.

• Kärkkäinen, Virve; Hannonen, Sanna; Rusanen, Minna; Lehtola, Juha-Matti; Saari, Toni & Uusitalo, Hannu [Show all 11 contributors for this article] (2024). Tear fluid reflects the altered protein expressions of Alzheimer's disease patients in proteins involved in protein repair and clearance system or the regulation of cytoskeleton. Journal of Alzheimer's Disease. ISSN 1387-2877. doi: 10.1177/13872877241295315. Full text in Research Archive
• Antewi, Milton Boaheng; Dumitriu, Gianina Aurica; Simon, Jaione Santamaria; Romano, Javier Sanchez; Li, Ruomei & Smedsrød, Bård [Show all 9 contributors for this article] (2023). Liver sinusoidal endothelial cells show reduced scavenger function and downregulation of Fc gamma receptor IIb, yet maintain a preserved fenestration in the Glmp gt/gt mouse model of slowly progressing liver fibrosis. PLOS ONE. 18(11), p. 1–29. doi: 10.1371/journal.pone.0293526. Full text in Research Archive Show summary

  Liver sinusoidal endothelial cells (LSECs) are fenestrated endothelial cells with a unique, high endocytic clearance capacity for blood-borne waste macromolecules and colloids. This LSEC scavenger function has been insufficiently characterized in liver disease. The Glmpgt/gt mouse lacks expression of a subunit of the MFSD1/GLMP lysosomal membrane protein transporter complex, is born normal, but soon develops chronic, mild hepatocyte injury, leading to slowly progressing periportal liver fibrosis, and splenomegaly. This study examined how LSEC scavenger function and morphology are affected in the Glmpgt/gt model. FITC-labelled formaldehyde-treated serum albumin (FITC-FSA), a model ligand for LSEC scavenger receptors was administered intravenously into Glmpgt/gt mice, aged 4 months (peak of liver inflammation), 9–10 month, and age-matched Glmpwt/wt mice. Organs were harvested for light and electron microscopy, quantitative image analysis of ligand uptake, collagen accumulation, LSEC ultrastructure, and endocytosis receptor expression (also examined by qPCR and western blot). In both age groups, the Glmpgt/gt mice showed multifocal liver injury and fibrosis. The uptake of FITC-FSA in LSECs was significantly reduced in Glmpgt/gt compared to wild-type mice. Expression of LSEC receptors stabilin-1 (Stab1), and mannose receptor (Mcr1) was almost similar in liver of Glmpgt/gt mice and age-matched controls. At the same time, immunostaining revealed differences in the stabilin-1 expression pattern in sinusoids and accumulation of stabilin-1-positive macrophages in Glmpgt/gt liver. FcγRIIb (Fcgr2b), which mediates LSEC endocytosis of soluble immune complexes was widely and significantly downregulated in Glmpgt/gt liver. Despite increased collagen in space of Disse, LSECs of Glmpgt/gt mice showed well-preserved fenestrae organized in sieve plates but the frequency of holes >400 nm in diameter was increased, especially in areas with hepatocyte damage. In both genotypes, FITC-FSA also distributed to endothelial cells of spleen and bone marrow sinusoids, suggesting that these locations may function as possible compensatory sites of clearance of blood-borne scavenger receptor ligands in liver fibrosis.

• Willemen, Hanneke L.D.M.; Ribeiro, Patrícia Silva Santos; Broeks, Melissa; Meijer, Nils; Versteeg, Sabine & Tiggeler, Annefien [Show all 11 contributors for this article] (2023). Inflammation-induced mitochondrial and metabolic disturbances in sensory neurons control the switch from acute to chronic pain. Cell Reports Medicine. 4(11), p. 1–24. doi: 10.1016/j.xcrm.2023.101265. Full text in Research Archive Show summary

  Pain often persists in patients with an inflammatory disease, even when inflammation has subsided. The molecular mechanisms leading to this failure in pain resolution and the transition to chronic pain are poorly understood. Mitochondrial dysfunction in sensory neurons links to chronic pain, but its role in resolution of inflammatory pain is unclear. Transient inflammation causes neuronal plasticity, called hyperalgesic priming, which impairs resolution of pain induced by a subsequent inflammatory stimulus. We identify that hyperalgesic priming in mice increases the expression of a mitochondrial protein (ATPSc-KMT) and causes mitochondrial and metabolic disturbances in sensory neurons. Inhibition of mitochondrial respiration, knockdown of ATPSCKMT expression, or supplementation of the affected metabolite is sufficient to restore resolution of inflammatory pain and prevents chronic pain development. Thus, inflammation-induced mitochondrial-dependent disturbances in sensory neurons predispose to a failure in resolution of inflammatory pain and development of chronic pain.

• Kumari, Pooja; Thuestad, Lars Harald & Ciosk, Rafal (2023). Post-transcriptional repression of CFP-1 expands the regulatory repertoire of LIN-41/TRIM71. Nucleic Acids Research (NAR). ISSN 0305-1048. 51(19), p. 10668–10680. doi: 10.1093/nar/gkad729. Full text in Research Archive Show summary

  Abstract The Caenorhabditis elegans LIN-41/TRIM71 is a well-studied example of a versatile regulator of mRNA fate, which plays different biological functions involving distinct post-transcriptional mechanisms. In the soma, LIN-41 determines the timing of developmental transitions between larval stages. The somatic LIN-41 recognizes specific mRNAs via LREs (LIN-41 Recognition Elements) and elicits either mRNA decay or translational repression. In the germline, LIN-41 controls the oocyte-to-embryo transition (OET), although the relevant targets and regulatory mechanisms are poorly understood. The germline LIN-41 was suggested to regulate mRNAs indirectly by associating with another RNA-binding protein. We show here that LIN-41 can also regulate germline mRNAs via the LREs. Through a computational-experimental analysis, we identified the germline mRNAs potentially controlled via LREs and validated one target, the cfp-1 mRNA, encoding a conserved chromatin modifier. Our analysis suggests that cfp-1 may be a long-sought target whose LIN-41-mediated regulation during OET facilitates the transcriptional reprogramming underlying the switch from germ- to somatic cell identity.

• Johannessen, Julie Aarmo; Formica, Miriam; Haukeland, Aina Louise C.; Bråthen, Nora Rojahn; Outa, Amani al & Larsen, Miriam Aarsund [Show all 9 contributors for this article] (2023). The human leukemic oncogene MLL-AF4 promotes hyperplastic growth of hematopoietic tissues in Drosophila larvae. iScience. 26(10). doi: 10.1016/j.isci.2023.107726. Full text in Research Archive Show summary

  MLL-rearranged (MLL-r) leukemias are among the leukemic subtypes with poorest survival, and treatment options have barely improved over the last decades. Despite increasing molecular understanding of the mechanisms behind these hematopoietic malignancies, this knowledge has had poor translation into the clinic. Here, we report a Drosophila melanogaster model system to explore the pathways affected in MLL-r leukemia. We show that expression of the human leukemic oncogene MLL-AF4 in the Drosophila hematopoietic system resulted in increased levels of circulating hemocytes and an enlargement of the larval hematopoietic organ, the lymph gland. Strikingly, depletion of Drosophila orthologs of known interactors of MLL-AF4, such as DOT1L, rescued the leukemic phenotype. In agreement, treatment with small-molecule inhibitors of DOT1L also prevented the MLL-AF4-induced leukemia-like phenotype. Taken together, this model provides an in vivo system to unravel the genetic interactors involved in leukemogenesis and offers a system for improved biological understanding of MLL-r leukemia.

• Storås, Andrea; Magnø, Morten Schjerven; Fineide, Fredrik; Thiede, Bernd; Chen, Xiangjun & Strumke, Inga [Show all 9 contributors for this article] (2023). Identifying Important Proteins in Meibomian Gland Dysfunction with Explainable Artificial Intelligence. IEEE International Symposium on Computer-Based Medical Systems. doi: 10.1109/CBMS58004.2023.00217. Full text in Research Archive Show summary

  Meibomian gland dysfunction is the most common cause of dry eye disease and leads to significantly reduced quality of life and social burdens. Because meibomian gland dysfunction results in impaired function of the tear film lipid layer, studying the expression of tear proteins might increase the understanding of the etiology of the condition. Machine learning is able to detect patterns in complex data. This study applied machine learning to classify levels of meibomian gland dysfunction from tear proteins. The aim was to investigate proteomic changes between groups with different severity levels of meibomian gland dysfunction, as opposed to only separating patients with and without this condition. An established feature importance method was used to identify the most important proteins for the resulting models. Moreover, a new method that can take the uncertainty of the models into account when creating explanations was proposed. By examining the identified proteins, potential biomarkers for meibomian gland dysfunction were discovered. The overall findings are largely conirmatory, indicating that the presented machine learning approaches are promising for detecting clinically relevant proteins. While this study provides valuable insights into proteomic changes associated with varying severity levels of meibomian gland dysfunction, it should be noted that it was conducted without a healthy control group. Future research could benefit from including such a comparison to further validate and extend the findings presented here.

• Alonso, Lucia Ramos; Holland, Petter; Gras, Stéphanie Le; Zhao, Xu; Jost, Bernard & Bjørås, Magnar [Show all 9 contributors for this article] (2023). Mitotic chromosome condensation resets chromatin to safeguard transcriptional homeostasis during interphase. Proceedings of the National Academy of Sciences of the United States of America (PNAS). ISSN 0027-8424. 120(4). doi: 10.1073/pnas.2210593120. Full text in Research Archive Show summary

  Mitotic entry correlates with the condensation of the chromosomes, changes in histone modifications, exclusion of transcription factors from DNA, and the broad downregulation of transcription. However, whether mitotic condensation influences transcription in the subsequent interphase is unknown. Here, we show that preventing one chromosome to condense during mitosis causes it to fail resetting of transcription. Rather, in the following interphase, the affected chromosome contains unusually high levels of the transcription machinery, resulting in abnormally high expression levels of genes in cis, including various transcription factors. This subsequently causes the activation of inducible transcriptional programs in trans, such as the GAL genes, even in the absence of the relevant stimuli. Thus, mitotic chromosome condensation exerts stringent control on interphase gene expression to ensure the maintenance of basic cellular functions and cell identity across cell divisions. Together, our study identifies the maintenance of transcriptional homeostasis during interphase as an unexpected function of mitosis and mitotic chromosome condensation.

• Malecka, Agnieszka Maria; Østlie, Ingunn; Trøen, Gunhild; Malecki, Jedrzej Mieczyslaw; Delabie, Jan & Tierens, Anne [Show all 9 contributors for this article] (2023). Gene expression analysis revealed downregulation of complement receptor 1 in clonal B cells in cold agglutinin disease. Clinical and Experimental Immunology. ISSN 0009-9104. 216(1), p. 45–54. doi: 10.1093/cei/uxad135. Full text in Research Archive Show summary

  Abstract Cold agglutinin disease (CAD) is a rare B-cell lymphoproliferative disorder of the bone marrow, manifested by autoimmune hemolytic anemia caused by binding of monoclonal IgM autoantibodies to the I antigen. Underlying genetic changes have previously been reported, but their impact on gene expression profile has been unknown. Here, we define differentially expressed genes in CAD B cells. To unravel downstream alteration in cellular pathways, gene expression by RNA sequencing was undertaken. Clonal B-cell samples from 12 CAD patients and IgM-expressing memory B cells from 4 healthy individuals were analyzed. Differential expression analysis and filtering resulted in 93 genes with significant differential expression. Top upregulated genes included SLC4A1, SPTA1, YBX3, TESC, HBD, AHSP, TRAF1, HBA2, RHAG, CA1, SPTB, IL10, UBASH3B, ALAS2, HBA1, CRYM, RGCC, KANK2, and IGHV4-34. They were upregulated at least 8-fold, while complement receptor 1 (CR1/CD35) was downregulated 11-fold in clonal CAD B cells compared to control B cells. Flow cytometry analyses further confirmed reduced CR1 (CD35) protein expression by clonal CAD IgM+ B cells compared to IgM+ memory B cells in controls. CR1 (CD35) is an important negative regulator of B-cell activation and differentiation. Therefore, reduced CR1 (CD35) expression may increase activation, proliferation, and antibody production in CAD-associated clonal B cells.

• Juvkam, Inga Solgård; Zlygosteva, Olga; Sitarz, Mateusz; Thiede, Bernd; Sørensen, Brita Singers & Malinen, Eirik [Show all 9 contributors for this article] (2023). Proton Compared to X-Irradiation Induces Different Protein Profiles in Oral Cancer Cells and Their Derived Extracellular Vesicles. International Journal of Molecular Sciences. ISSN 1661-6596. 24(23), p. 1–17. doi: 10.3390/ijms242316983. Full text in Research Archive Show summary

  Extracellular vesicles (EVs) are membrane-bound particles released from cells, and their cargo can alter the function of recipient cells. EVs from X-irradiated cells have been shown to play a likely role in non-targeted effects. However, EVs derived from proton irradiated cells have not yet been studied. We aimed to investigate the proteome of EVs and their cell of origin after proton or X-irradiation. The EVs were derived from a human oral squamous cell carcinoma (OSCC) cell line exposed to 0, 4, or 8 Gy from either protons or X-rays. The EVs and irradiated OSCC cells underwent liquid chromatography–mass spectrometry for protein identification. Interestingly, we found different protein profiles both in the EVs and in the OSCC cells after proton irradiation compared to X-irradiation. In the EVs, we found that protons cause a downregulation of proteins involved in cell growth and DNA damage response compared to X-rays. In the OSCC cells, proton and X-irradiation induced dissimilar cell death pathways and distinct DNA damage repair systems. These results are of potential importance for understanding how non-targeted effects in normal tissue can be limited and for future implementation of proton therapy in the clinic.

• Kresse, Stine Henrichson; Brandt-Winge, Sara; Pharo, Heidi Dietrichson; Flatin, Bjørnar Tovson Bae; Jeanmougin, Marine Anne-Lise Francoise & Vedeld, Hege Marie [Show all 7 contributors for this article] (2023). Evaluation of commercial kits for isolation and bisulfite conversion of circulating cell-free tumor DNA from blood. Clinical Epigenetics. 15(1). doi: 10.1186/s13148-023-01563-0. Full text in Research Archive Show summary

  Background DNA methylation biomarkers in circulating cell-free DNA (cfDNA) have great clinical potential for cancer management. Most methods for DNA methylation analysis require bisulfite conversion, causing DNA degradation and loss. This is particularly challenging for cfDNA, which is naturally fragmented and normally present in low amounts. The aim of the present study was to identify an optimal combination of cfDNA isolation and bisulfite conversion kits for downstream analysis of DNA methylation biomarkers in plasma. Results Of the five tested bisulfite conversion kits (EpiJET Bisulfite Conversion Kit, EpiTect Plus DNA Bisulfite Kit (EpiTect), EZ DNA Methylation-Direct Kit, Imprint DNA Modification Kit (Imprint) and Premium Bisulfite Kit), the highest and lowest DNA yield and recovery were achieved using the EpiTect kit and the Imprint kit, respectively, with more than double the amount of DNA for the EpiTect kit. Of the three tested cfDNA isolation kits (Maxwell RSC ccfDNA Plasma Kit, QIAamp Circulating Nucleic Acid Kit (CNA) and QIAamp MinElute ccfDNA Mini Kit), the CNA kit yielded around twice as much cfDNA compared to the two others kits, although with more high molecular weight DNA present. When comparing various combinations of cfDNA isolation kits and bisulfite conversion kits, the CNA kit and the EpiTect kit were identified as the best-performing combination, resulting in the highest yield of bisulfite converted cfDNA from normal plasma, as measured by droplet digital PCR (ddPCR). As a proof of principle, this kit combination was used to process plasma samples from 13 colorectal cancer patients for subsequent ddPCR methylation analysis of BCAT1 and IKZF1. Methylation of BCAT1 and/or IKZF1 was identified in 6/10 (60%) stage IV patients and 1/3 (33%) stage III patients. Conclusions Based on a thorough evaluation of five bisulfite conversion kits and three cfDNA isolation kits, both individually and in combination, the CNA kit and the EpiTect kit were identified as the best-performing kit combination, with highest DNA yield and recovery across a range of DNA input amounts. The combination was successfully used for detection of clinically relevant DNA methylation biomarkers in plasma from cancer patients.

• Hammerstad, Marta; Rugtveit, Anne Kristine; Dahlen, Sondov Åsmundson Braathen; Andersen, Hilde Kristin & Hersleth, Hans-Petter (2023). Functional Diversity of Homologous Oxidoreductases—Tuning of Substrate Specificity by a FAD-Stacking Residue for Iron Acquisition and Flavodoxin Reduction. Antioxidants. 12(6). doi: 10.3390/antiox12061224. Full text in Research Archive Show summary

  Although bacterial thioredoxin reductase-like ferredoxin/flavodoxin NAD(P)+ oxidoreduc- tases (FNRs) are similar in terms of primary sequences and structures, they participate in diverse biological processes by catalyzing a range of different redox reactions. Many of the reactions are critical for the growth, survival of, and infection by pathogens, and insight into the structural basis for substrate preference, specificity, and reaction kinetics is crucial for the detailed understanding of these redox pathways. Bacillus cereus (Bc) encodes three FNR paralogs, two of which have assigned distinct biological functions in bacillithiol disulfide reduction and flavodoxin (Fld) reduction. Bc FNR2, the endogenous reductase of the Fld-like protein NrdI, belongs to a distinct phylogenetic cluster of homologous oxidoreductases containing a conserved His residue stacking the FAD cofactor. In this study, we have assigned a function to FNR1, in which the His residue is replaced by a conserved Val, in the reduction of the heme-degrading monooxygenase IsdG, ultimately facilitating the release of iron in an important iron acquisition pathway. The Bc IsdG structure was solved, and IsdG-FNR1 interactions were proposed through protein–protein docking. Mutational studies and bioinformatics analyses confirmed the importance of the conserved FAD-stacking residues on the respective reaction rates, proposing a division of FNRs into four functionally unique sequence similarity clusters likely related to the nature of this residue.

• Outa, Amani al; Hicks, Steven; Thambawita, Vajira L B; Andresen, Siri; Enserink, Jorrit Martijn & Halvorsen, Pål [Show all 8 contributors for this article] (2023). CELLULAR, A Cell Autophagy Imaging Dataset. Scientific Data. doi: 10.1038/s41597-023-02687-x. Full text in Research Archive Show summary

  Cells in living organisms are dynamic compartments that continuously respond to changes in their environment to maintain physiological homeostasis. While basal autophagy exists in cells to aid in the regular turnover of intracellular material, autophagy is also a critical cellular response to stress, such as nutritional depletion. Conversely, the deregulation of autophagy is linked to several diseases, such as cancer, and hence, autophagy constitutes a potential therapeutic target. Image analysis to follow autophagy in cells, especially on high-content screens, has proven to be a bottleneck. Machine learning (ML) algorithms have recently emerged as crucial in analyzing images to efficiently extract information, thus contributing to a better understanding of the questions at hand. This paper presents CELLULAR, an open dataset consisting of images of cells expressing the autophagy reporter mRFP-EGFP-Atg8a with cell-specific segmentation masks. Each cell is annotated into either basal autophagy, activated autophagy, or unknown. Furthermore, we introduce some preliminary experiments using the dataset that can be used as a baseline for future research.

• Chen, Yingjia; He, Liye; Ianevski, Aleksandr; Duran, Pilar Ayuda; Potdar, Swapnil & Saarela, Jani [Show all 14 contributors for this article] (2023). Robust scoring of selective drug responses for patient-tailored therapy selection. Nature Protocols. ISSN 1754-2189. p. 1–30. doi: 10.1038/s41596-023-00903-x. Full text in Research Archive Show summary

  Most patients with advanced malignancies are treated with severely toxic, first-line chemotherapies. Personalized treatment strategies have led to improved patient outcomes and could replace one-size-fits-all therapies, yet they need to be tailored by testing of a range of targeted drugs in primary patient cells. Most functional precision medicine studies use simple drug-response metrics, which cannot quantify the selective effects of drugs (i.e., the differential responses of cancer cells and normal cells). We developed a computational method for selective drug-sensitivity scoring (DSS), which enables normalization of the individual patient’s responses against normal cell responses. The selective response scoring uses the inhibition of noncancerous cells as a proxy for potential drug toxicity, which can in turn be used to identify effective and safer treatment options. Here, we explain how to apply the selective DSS calculation for guiding precision medicine in patients with leukemia treated across three cancer centers in Europe and the USA; the generic methods are also widely applicable to other malignancies that are amenable to drug testing. The open-source and extendable R-codes provide a robust means to tailor personalized treatment strategies on the basis of increasingly available ex vivo drug-testing data from patients in real-world and clinical trial settings. We also make available drug-response profiles to 527 anticancer compounds tested in 10 healthy bone marrow samples as reference data for selective scoring and de-prioritization of drugs that show broadly toxic effects. The procedure takes <60 min and requires basic skills in R.

• Falnes, Pål; Malecki, Jedrzej Mieczyslaw; Gonzalez-Molina, Maria del Carmen Herrera; Bengtsen, Mads & Davydova, Erna (2023). Human seven-β-strand (METTL) methyltransferases - conquering the universe of protein lysine methylation. Journal of Biological Chemistry. ISSN 0021-9258. 299(6), p. 1–15. doi: 10.1016/j.jbc.2023.104661. Full text in Research Archive Show summary

  Lysine methylation is an abundant posttranslational modification, which has been most intensively studied in the context of histone proteins, where it represents an important epigenetic mark. Lysine methylation of histone proteins is primarily catalyzed by SET-domain methyltransferases (MTases). However, it has recently become evident that also another MTase family, the so-called seven-β-strand (7BS) MTases, often denoted METTLs (methyltransferase-like), contains several lysine (K)-specific MTases (KMTs). These enzymes catalyze the attachment of up to three methyl groups to lysine residues in specific substrate proteins, using S-adenosylmethionine (AdoMet) as methyl donor. About a decade ago, only a single human 7BS KMT was known, namely the histone-specific DOT1L, but 15 additional 7BS KMTs have now been discovered and characterized. These KMTs typically target a single nonhistone substrate that, in most cases, belongs to one of the following three protein groups: components of the cellular protein synthesis machinery, mitochondrial proteins, and molecular chaperones. This article provides an extensive overview and discussion of the human 7BS KMTs and their biochemical and biological roles.

• Benjakul, Sopisa; Anthi, Aina Karen; Kolderup, Heidi Anette; Vaysburd, Marina; Lode, Heidrun Elisabeth & Mallery, D [Show all 20 contributors for this article] (2023). A pan-SARS-CoV-2-specific soluble angiotensin-converting enzyme 2-albumin fusion engineered for enhanced plasma half-life and needle-free mucosal delivery. PNAS Nexus. 2(12), p. 1–15. doi: 10.1093/pnasnexus/pgad403. Full text in Research Archive Show summary

  Immunocompromised patients often fail to raise protective vaccine-induced immunity against the global emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants. Although monoclonal antibodies have been authorized for clinical use, most have lost their ability to potently neutralize the evolving Omicron subvariants. Thus, there is an urgent need for treatment strategies that can provide protection against these and emerging SARS-CoV-2 variants to prevent the development of severe coronavirus disease 2019. Here, we report on the design and characterization of a long-acting viral entry-blocking angiotensin-converting enzyme 2 (ACE2) dimeric fusion molecule. Specifically, a soluble truncated human dimeric ACE2 variant, engineered for improved binding to the receptor-binding domain of SARS-CoV-2, was fused with human albumin tailored for favorable engagement of the neonatal fragment crystallizable receptor (FcRn), which resulted in enhanced plasma half-life and allowed for needle-free transmucosal delivery upon nasal administration in human FcRn-expressing transgenic mice. Importantly, the dimeric ACE2-fused albumin demonstrated potent neutralization of SARS-CoV-2 immune escape variants.

• Storås, Andrea; Fineide, Fredrik; Magnø, Morten Schjerven; Thiede, Bernd; Chen, Xiangjun & Strumke, Inga [Show all 11 contributors for this article] (2023). Using machine learning model explanations to identify proteins related to severity of meibomian gland dysfunction. Scientific Reports. 13(1). doi: 10.1038/s41598-023-50342-7. Full text in Research Archive Show summary

  Meibomian gland dysfunction is the most common cause of dry eye disease and leads to significantly reduced quality of life and social burdens. Because meibomian gland dysfunction results in impaired function of the tear film lipid layer, studying the expression of tear proteins might increase the understanding of the etiology of the condition. Machine learning is able to detect patterns in complex data. This study applied machine learning to classify levels of meibomian gland dysfunction from tear proteins. The aim was to investigate proteomic changes between groups with different severity levels of meibomian gland dysfunction, as opposed to only separating patients with and without this condition. An established feature importance method was used to identify the most important proteins for the resulting models. Moreover, a new method that can take the uncertainty of the models into account when creating explanations was proposed. By examining the identified proteins, potential biomarkers for meibomian gland dysfunction were discovered. The overall findings are largely confirmatory, indicating that the presented machine learning approaches are promising for detecting clinically relevant proteins. While this study provides valuable insights into proteomic changes associated with varying severity levels of meibomian gland dysfunction, it should be noted that it was conducted without a healthy control group. Future research could benefit from including such a comparison to further validate and extend the findings presented here.

• Andersen, Aram Nikolai; Brodersen, Andrea M; Duran, Pilar Ayuda; Piechaczyk, Laure Isabelle; Tadele, Dagim Shiferaw & Baken, Lizet [Show all 12 contributors for this article] (2023). Clinical forecasting of acute myeloid leukemia using ex vivo drug-sensitivity profiling. Cell Reports Methods. 3(12), p. 1–16. doi: 10.1016/j.crmeth.2023.100654. Full text in Research Archive Show summary

  Current treatment selection for acute myeloid leukemia (AML) patients depends on risk stratification based on cytogenetic and genomic markers. However, the forecasting accuracy of treatment response remains modest, with most patients receiving intensive chemotherapy. Recently, ex vivo drug screening has gained traction in personalized treatment selection and as a tool for mapping patient groups based on relevant cancer dependencies. Here, we systematically evaluated the use of drug sensitivity profiling for predicting patient survival and clinical response to chemotherapy in a cohort of AML patients. We compared computational methodologies for scoring drug efficacy and characterized tools to counter noise and batch-related confounders pervasive in high-throughput drug testing. We show that ex vivo drug sensitivity profiling is a robust and versatile approach to patient prognostics that comprehensively maps functional signatures of treatment response and disease progression. In conclusion, ex vivo drug profiling can assess risk for individual AML patients and may guide clinical decision-making.

• Jeanmougin, Marine Anne-Lise Francoise; Brodal, Hans Petter; Pharo, Heidi Dietrichson; Vedeld, Hege Marie & Lind, Guro Elisabeth (2023). PoDCall: positive droplet calling and normalization of droplet digital PCR DNA methylation data. Bioinformatics. ISSN 1367-4803. 39(1). doi: 10.1093/bioinformatics/btac766. Full text in Research Archive Show summary

  Abstract Motivation Droplet digital PCR (ddPCR) holds great promises for investigating DNA methylation with high sensitivity. Yet, the lack of methods for analyzing ddPCR DNA methylation data has resulted in users processing the data manually at the expense of standardization. Results PoDCall is an R package performing automated calling of positive droplets, quantification and normalization of methylation levels in ddPCR experiments. A Shiny application provides users with an intuitive and interactive interface to access PoDCall functionalities. Availability and implementation The PoDCall R package is freely available on Bioconductor at https://bioconductor.org/packages/PoDCall/. The Shiny application can be executed from the R console using the wrapper function PoDCall::podcallShiny(). Supplementary information Supplementary data are available at Bioinformatics online.

• Duran, Pilar Ayuda; Hermansen, Johanne Uthus; Giliberto, Mariaserena; Yin, Yanping; Hanes, Robert & Gordon, Sandra [Show all 13 contributors for this article] (2023). Standardized assays to monitor drug sensitivity in hematologic cancers. Cell death discovery. 9(1), p. 1–13. doi: 10.1038/s41420-023-01722-5. Full text in Research Archive Show summary

  Abstract The principle of drug sensitivity testing is to expose cancer cells to a library of different drugs and measure its effects on cell viability. Recent technological advances, continuous approval of targeted therapies, and improved cell culture protocols have enhanced the precision and clinical relevance of such screens. Indeed, drug sensitivity testing has proven diagnostically valuable for patients with advanced hematologic cancers. However, different cell types behave differently in culture and therefore require optimized drug screening protocols to ensure that their ex vivo drug sensitivity accurately reflects in vivo drug responses. For example, primary chronic lymphocytic leukemia (CLL) and multiple myeloma (MM) cells require unique microenvironmental stimuli to survive in culture, while this is less the case for acute myeloid leukemia (AML) cells. Here, we present our optimized and validated protocols for culturing and drug screening of primary cells from AML, CLL, and MM patients, and a generic protocol for cell line models. We also discuss drug library designs, reproducibility, and quality controls. We envision that these protocols may serve as community guidelines for the use and interpretation of assays to monitor drug sensitivity in hematologic cancers and thus contribute to standardization. The read-outs may provide insight into tumor biology, identify or confirm treatment resistance and sensitivity in real time, and ultimately guide clinical decision-making.

• Meuskens, Ina; Kristiansen, Per Eugen; Bardiaux, Benjamin; Koynarev, Vladimir Rosenov; Hatlem, Daniel & Prydz, Kristian [Show all 9 contributors for this article] (2023). A poly-proline II helix in YadA from Yersinia enterocolitica serotype O:9 facilitates heparin binding through electrostatic interactions. The FEBS Journal. ISSN 1742-464X. 291(4), p. 761–777. doi: 10.1111/febs.17001. Full text in Research Archive Show summary

  Poly‐proline II helices are secondary structure motifs frequently found in ligand‐binding sites. They exhibit increased flexibility and solvent exposure compared to the strongly hydrogen‐bonded α‐helices or β‐strands and can therefore easily be misinterpreted as completely unstructured regions with an extremely high rotational freedom. Here, we show that the adhesin YadA of Yersinia enterocolitica serotype O:9 contains a poly‐proline II helix interaction motif in the N‐terminal region. The motif is involved in the interaction of YadA O:9 with heparin, a host glycosaminoglycan. We show that the basic residues within the N‐terminal motif of YadA are required for electrostatic interactions with the sulfate groups of heparin. Biophysical methods including CD spectroscopy, solution‐state NMR and SAXS all independently support the presence of a poly‐proline helix allowing YadA O:9 binding to the rigid heparin. Lastly, we show that host cells deficient in sulfation of heparin and heparan sulfate are not targeted by YadA O:9 ‐mediated adhesion. We speculate that the YadA O:9 –heparin interaction plays an important and highly strain‐specific role in the pathogenicity of Yersinia enterocolitica serotype O:9.

• Engelfriet, Melanie Lianne; Malecki, Jedrzej Mieczyslaw; Forsberg, Frida; Falnes, Pål & Ciosk, Rafal (2023). Characterization of the biochemical activity and tumor-promoting role of the dual protein methyltransferase METL-13/METTL13 in Caenorhabditis elegans. PLOS ONE. 18(6), p. 1–16. doi: 10.1371/journal.pone.0287558. Full text in Research Archive Show summary

  The methyltransferase-like protein 13 (METTL13) methylates the eukaryotic elongation factor 1 alpha (eEF1A) on two locations: the N-terminal amino group and lysine 55. The absence of this methylation leads to reduced protein synthesis and cell proliferation in human cancer cells. Previous studies showed that METTL13 is dispensable in non-transformed cells, making it potentially interesting for cancer therapy. However, METTL13 has not been examined yet in whole animals. Here, we used the nematode Caenorhabditis elegans as a simple model to assess the functions of METTL13. Using methyltransferase assays and mass spectrometry, we show that the C . elegans METTL13 (METL-13) methylates eEF1A (EEF-1A) in the same way as the human protein. Crucially, the cancer-promoting role of METL-13 is also conserved and depends on the methylation of EEF-1A, like in human cells. At the same time, METL-13 appears dispensable for animal growth, development, and stress responses. This makes C . elegans a convenient whole-animal model for studying METL13-dependent carcinogenesis without the complications of interfering with essential wild-type functions.

• Meyer, Saskia; Blaas, Isaac; Bollineni, Ravi Chand; Delic-Sarac, Marina; Tran, Trung & Knetter, Cathrine [Show all 20 contributors for this article] (2023). Prevalent and immunodominant CD8 T cell epitopes are conserved in SARS-CoV-2 variants. Cell reports. 42(1). doi: 10.1016/j.celrep.2023.111995. Full text in Research Archive Show summary

  The emergence of SARS-CoV-2 variants of concern (VOC) is driven by mutations that mediate escape from neutralizing antibodies. There is also evidence that mutations can cause loss of T cell epitopes. However, studies on viral escape from T cell immunity have been hampered by uncertain estimates of epitope prevalence. Here, we map and quantify CD8 T cell responses to SARS-CoV-2-specific minimal epitopes in blood drawn from April to June 2020 from 83 COVID-19 convalescents. Among 37 HLA ligands eluted from five prevalent alleles and an additional 86 predicted binders, we identify 29 epitopes with an immunoprevalence ranging from 3% to 100% among individuals expressing the relevant HLA allele. Mutations in VOC are reported in 10.3% of the epitopes, while 20.6% of the non-immunogenic peptides are mutated in VOC. The nine most prevalent epitopes are conserved in VOC. Thus, comprehensive mapping of epitope prevalence does not provide evidence that mutations in VOC are driven by escape of T cell immunity.

• Alonso, Lucia Ramos & Chymkowitch, Pierre (2023). Maintaining transcriptional homeostasis during cell cycle. Transcription. ISSN 2154-1264. doi: 10.1080/21541264.2023.2246868. Full text in Research Archive Show summary

  The preservation of gene expression patterns that define cellular identity throughout the cell division cycle is essential to perpetuate cellular lineages. However, the progression of cells through different phases of the cell cycle severely disrupts chromatin accessibility, epigenetic marks, and the recruitment of transcriptional regulators. Notably, chromatin is transiently disassembled during S-phase and undergoes drastic condensation during mitosis, which is a significant challenge to the preservation of gene expression patterns between cell generations. This article delves into the specific gene expression and chromatin regulatory mechanisms that facilitate the preservation of transcriptional identity during replication and mitosis. Furthermore, we emphasize our recent findings revealing the unconventional role of yeast centromeres and mitotic chromosomes in maintaining transcriptional fidelity beyond mitosis.

• Luque, Alvaro Kohn; Myklebust, Even Moa; Tadele, Dagim Shiferaw; Giliberto, Mariaserena; Schmiester, Leonard & Noory, Jasmine [Show all 15 contributors for this article] (2023). Phenotypic deconvolution in heterogeneous cancer cell populations using drug-screening data. Cell Reports Methods. 3(3). doi: 10.1016/j.crmeth.2023.100417. Full text in Research Archive Show summary

  Tumor heterogeneity is an important driver of treatment failure in cancer since therapies often select for drug-tolerant or drug-resistant cellular subpopulations that drive tumor growth and recurrence. Profiling the drug-response heterogeneity of tumor samples using traditional genomic deconvolution methods has yielded limited results, due in part to the imperfect mapping between genomic variation and functional characteristics. Here, we leverage mechanistic population modeling to develop a statistical framework for profiling phenotypic heterogeneity from standard drug-screen data on bulk tumor samples. This method, called PhenoPop, reliably identifies tumor subpopulations exhibiting differential drug responses and estimates their drug sensitivities and frequencies within the bulk population. We apply PhenoPop to synthetically generated cell populations, mixed cell-line experiments, and multiple myeloma patient samples and demonstrate how it can provide individualized predictions of tumor growth under candidate therapies. This methodology can also be applied to deconvolution problems in a variety of biological settings beyond cancer drug response.

• Bogucka-Janczi, Katarzyna; Harms, Gregory; Coissieux, Marie-May; Bentires-Alj, Mohamed; Thiede, Bernd & Rajalingam, Krishnaraj (2023). ERK3/MAPK6 dictates CDC42/RAC1 activity and ARP2/3-dependent actin polymerization. eLIFE. 12, p. 1–40. doi: 10.7554/eLife.85167. Full text in Research Archive Show summary

  The actin cytoskeleton is tightly controlled by RhoGTPases, actin binding-proteins and nucleation-promoting factors to perform fundamental cellular functions. We have previously shown that ERK3, an atypical MAPK, controls IL-8 production and chemotaxis (Bogueka et al., 2020). Here, we show in human cells that ERK3 directly acts as a guanine nucleotide exchange factor for CDC42 and phosphorylates the ARP3 subunit of the ARP2/3 complex at S418 to promote filopodia formation and actin polymerization, respectively. Consistently, depletion of ERK3 prevented both basal and EGF-dependent RAC1 and CDC42 activation, maintenance of F-actin content, filopodia formation, and epithelial cell migration. Further, ERK3 protein bound directly to the purified ARP2/3 complex and augmented polymerization of actin in vitro. ERK3 kinase activity was required for the formation of actin-rich protrusions in mammalian cells. These findings unveil a fundamentally unique pathway employed by cells to control actin-dependent cellular functions.

• Reubsaet, Leon; Thiede, Bernd & Halvorsen, Trine Grønhaug (2022). Next generation VAMS®–Trypsin immobilization for instant proteolysis in bottom-up protein determination. Advances in Sample Preparation. 3. doi: 10.1016/j.sampre.2022.100027. Full text in Research Archive Show summary

  The purpose of this study was to develop a smart proteolysis sampler based on commercially available VAMS® tips employing Volumetric Absorptive Microsampling. In this proof-of-concept paper we show that covalently bound trypsin to VAMS exhibits sufficient activity for proteomic purposes without altering physical conditions. Alkaline pH was needed for the covalent binding of trypsin and at least 50 μg trypsin was required to obtain maximal activity. Prolonged digestion times (up to overnight) of a sample containing 7 model proteins on the VAMS yielded higher peak intensities for signature peptides which are beneficial for quantitative analysis. In-solution digestions of three proteins (carbonic anhydrase 2, catalase and BSA) yielded higher intensities of their signature peptides whereas the other four proteins (cytochrome C, myoglobin, β-lactoglobulin, and phosphorylase B) showed higher intensities for their signature peptides from digestions performed on trypsin modified VAMS. In comparison with conventional in-solution digestions, approximately half of the proteins were identified from diluted serum using trypsin on VAMS. For shotgun serum proteomics, diluted samples collected on trypsin modified VAMS followed by immediate drying yielded similar peptide- (942 vs 875) and protein (96 vs 99) identifications compared to those collected on trypsin modified VAMS followed by overnight digestions. This result shows the potential of trypsin on VAMS digestions.

• Cui, Xueyan; Stavik, Benedicte; Thiede, Bernd; Sandset, Per Morten & Kanse, Sandip (2022). FSAP Protects against Histone-Mediated Increase in Endothelial Permeability In Vitro. International Journal of Molecular Sciences. ISSN 1661-6596. 23(22). doi: 10.3390/ijms232213706. Full text in Research Archive Show summary

  Factor-VII-activating protease (FSAP) is involved in the regulation of hemostasis and inflammation. Extracellular histones play a role in inflammation and the conversion of latent pro-FSAP into active FSAP. FSAP has been shown to regulate endothelial permeability, but the mechanisms are not clear. Here, we have investigated the effects of FSAP on endothelial permeability in vitro. A mixture of histones from calf thymus stimulated permeability, and the wild-type (WT) serine protease domain (SPD) of FSAP blocked this effect. WT–SPD–FSAP did not influence permeability on its own, nor that stimulated by thrombin or vascular endothelial growth factor (VEGF)-A165. Histones induced a large-scale rearrangement of the junction proteins VE-cadherin and zona occludens-1 from a clear junctional distribution to a diffuse pattern. The presence of WT–SPD–FSAP inhibited these changes. Permeability changes by histones were blocked by both TLR-2 and TLR4 blocking antibodies. Histones upregulated the expression of TLR-2, but not TLR-4, in HUVEC cells, and WT–SPD–FSAP abolished the upregulation of TLR-2 expression. An inactive variant, Marburg I (MI)–SPD–FSAP, did not have any of these effects. The inhibition of histone-mediated permeability may be an important function of FSAP with relevance to sepsis, trauma, and stroke and the need to be investigated further in in vivo experiments.

• Nguyen, Minh Thao; Halvorsen, Trine Grønhaug; Thiede, Bernd & Reubsaet, Leon (2022). Smart proteolysis samplers for pre-lab bottom-up protein analysis – Performance of on-paper digestion compared to conventional digestion. Separation Science Plus (SSC plus). 5(5), p. 171–183. doi: 10.1002/sscp.202100062. Full text in Research Archive Show summary

  Here the relation between digestion of proteins by trypsin covalently bound to paper and trypsin in-solution is investigated. The trypsin acting on paper is covalently bound. A trypsin concentration of 0.5% (w/v) results in the highest digestion activity of all concentrations tested. Additionally, it can be seen that trypsin on-paper has retained approx. 50% of its activity. Unlike trypsin in-solution, the stability of the smart proteolysis samplers was regarded to be stable for at least four months when kept refrigerated. Autolysis was very small for covalently bound trypsin: less than 2% compared to in-solution trypsin. Proteomic analysis of diluted human serum showed more protein identifications (214) in-solution digestions than on-paper digestions (76). Also, higher coverage for the in-solution digestion was obtained. Those proteins identified after on-paper digestion with no or few disulfide bonds seem to have more similar sequence coverages compared to those identified after in-solution digestion. Smart samplers allow the determination of at least 70–75 proteins without performing the overnight digestion. All in all, trypsin covalently bound to paper shows to retain high proteolytic activity and is a stable alternative for conventional digestions. In this way, smart proteolytic samplers show their feasibility in pre-lab sample preparation.

• Saatcioglu, Fahri; Cirit, Burcu & Suzer, Gayem Koprucu (2022). The Promise of Well-Being Interventions to Mitigate Physician Burnout During the COVID-19 Pandemic and Beyond. JCO Oncology Practice (JCO OP). ISSN 2688-1527. 18(12), p. 808–814. doi: 10.1200/OP.22.00108. Full text in Research Archive Show summary

  The high degree of burnout in physicians, including oncologists, is detrimental to physicians themselves, their families, patients, health care organizations, and the health care systems as a whole. This dire situation has significantly worsened during the COVID-19 pandemic. It is well established that both organizational and individual measures are urgently needed to mitigate the negative consequences of physician burnout. Here, we review the research that has begun to indicate potential evidence-based individual approaches to promote physician well-being. We give an overview of these emerging programs and their importance, provide an example from our own experience, and enumerate considerations for future research. We also discuss the need for developing new approaches that are evidence-based and the best ways in which they can be incorporated in the health care setting. When judiciously combined with organizational approaches, preferentially as an integral part of them, individual wellness programs for physicians are poised to contribute significantly toward the much needed relief from physician burnout.

• Chica-Balaguera, Nathalia; Portantier, Marina; Nyquist-Andersen, Mari; Burriel, Silvia Espada & Aviles, Sandra Lopez (2022). Uncoupling of Mitosis and Cytokinesis Upon a Prolonged Arrest in Metaphase Is Influenced by Protein Phosphatases and Mitotic Transcription in Fission Yeast. Frontiers in Cell and Developmental Biology. 10. doi: 10.3389/fcell.2022.876810. Full text in Research Archive Show summary

  Depletion of the Anaphase-Promoting Complex/Cyclosome (APC/C) activator Cdc20 arrests cells in metaphase with high levels of the mitotic cyclin (Cyclin B) and the Separase inhibitor Securin. In mammalian cells this arrest has been exploited for the treatment of cancer with drugs that engage the spindle assembly checkpoint and, recently, with chemical inhibitors of the APC/C. While most cells arrested in mitosis for prolonged periods undergo apoptosis, others skip cytokinesis and enter G1 with unsegregated chromosomes. This process, known as mitotic slippage, generates aneuploidy and increases genomic instability in the cancer cell. Here, we analyze the behavior of fission yeast cells arrested in mitosis through the transcriptional silencing of the Cdc20 homolog slp1 . While depletion of slp1 readily halts cells in metaphase, this arrest is only transient and a majority of cells eventually undergo cytokinesis and show steady mitotic dephosphorylation. Notably, this occurs in the absence of Cyclin B (Cdc13) degradation. We investigate the involvement of phosphatase activity in these events and demonstrate that PP2A-B55 Pab1 is required to prevent septation and, during the arrest, its CDK-mediated inhibition facilitates the induction of cytokinesis. In contrast, deletion of PP2A-B56 Par1 completely abrogates septation. We show that this effect is partly due to this mutant entering mitosis with reduced CDK activity. Interestingly, both PP2A-B55 Pab1 and PP2A-B56 Par1 , as well as Clp1 (the homolog of the budding yeast mitotic phosphatase Cdc14) are required for the dephosphorylation of mitotic substrates during the escape. Finally, we show that the mitotic transcriptional wave controlled by the RFX transcription factor Sak1 facilitates the induction of cytokinesis and also requires the activity of PP2A-B56 Par1 in a mechanism independent of CDK.

• Nissen-Meyer, Jon; Skotland, Tore & Boye, Erik (2022). Are doping tests in sports trustworthy?: Athletes suffer from insufficiently defined criteria for doping tests: Athletes suffer from insufficiently defined criteria for doping tests. EMBO Reports. ISSN 1469-221X. 23. doi: 10.15252/embr.202154431. Full text in Research Archive
• Sikkeland, Jørgen; Ng, Matthew; Nenseth, Hatice Zeynep; Ünal, Bilal; Qu, Su & Jin, Yang [Show all 8 contributors for this article] (2022). STAMP2 suppresses autophagy in prostate cancer cells by modulating the integrated stress response pathway. American Journal of Cancer Research. 12(1), p. 327–336. Full text in Research Archive Show summary

  Six Transmembrane Protein of Prostate 2 (STAMP2) is critical for prostate cancer (PCa) growth. We previously showed that STAMP2 regulates the expression of stress induced transcription factor ATF4, which is implicated in starvation-induced autophagy. We therefore investigated whether STAMP2 is involved in the regulation of autophagy in PCa cells. Here we show that STAMP2 suppresses autophagy in PCa cells through modulation of the integrated stress response axis. We also find that STAMP2 regulates mitochondrial respiration. These findings suggest that STAMP2 has significant metabolic effects through mitochondrial function and autophagy, both of which support PCa growth.

• Pharo, Heidi Dietrichson; Jeanmougin, Marine; Ager-Wick, Eirill; Vedeld, Hege Marie; Sørbø, Anne & Dahl, Christina [Show all 17 contributors for this article] (2022). BladMetrix: a novel urine DNA methylation test with high accuracy for detection of bladder cancer in hematuria patients. Clinical Epigenetics. 14(1). doi: 10.1186/s13148-022-01335-2. Full text in Research Archive Show summary

  Background Cystoscopy is the gold standard for bladder cancer detection, but is costly, invasive and has imperfect diagnostic accuracy. We aimed to identify novel and accurate DNA methylation biomarkers for non-invasive detection of bladder cancer in urine, with the potential to reduce the number of cystoscopies among hematuria patients. Results Biomarker candidates (n = 32) were identified from methylome sequencing of urological cancer cell lines (n = 16) and subjected to targeted methylation analysis in tissue samples (n = 60). The most promising biomarkers (n = 8) were combined into a panel named BladMetrix. The performance of BladMetrix in urine was assessed in a discovery series (n = 112), consisting of bladder cancer patients, patients with other urological cancers and healthy individuals, resulting in 95.7% sensitivity and 94.7% specificity. BladMetrix was furthermore evaluated in an independent prospective and blinded series of urine from patients with gross hematuria (n = 273), achieving 92.1% sensitivity, 93.3% specificity and a negative predictive value of 98.1%, with the potential to reduce the number of cystoscopies by 56.4%. Conclusions We here present BladMetrix, a novel DNA methylation urine test for non-invasive detection of bladder cancer, with high accuracy across tumor grades and stages, and the ability to spare a significant number of cystoscopies among patients with gross hematuria.

• Xiao, Rourou; You, Lixin; Zhang, Li; Guo, Xichen; Guo, Ensong & Zhao, Faming [Show all 29 contributors for this article] (2022). Inhibiting the IRE1α Axis of the Unfolded Protein Response Enhances the Antitumor Effect of AZD1775 in TP53 Mutant Ovarian Cancer. Advanced Science. 9(21). doi: 10.1002/advs.202105469. Full text in Research Archive Show summary

  Targeting the G2/M checkpoint mediator WEE1 has been explored as a novel treatment strategy in ovarian cancer, but mechanisms underlying its efficacy and resistance remains to be understood. Here, it is demonstrated that the WEE1 inhibitor AZD1775 induces endoplasmic reticulum stress and activates the protein kinase RNA-like ER kinase (PERK) and inositol-required enzyme 1α (IRE1α) branches of the unfolded protein response (UPR) in TP53 mutant (mtTP53) ovarian cancer models. This is facilitated through NF-κB mediated senescence-associated secretory phenotype. Upon AZD1775 treatment, activated PERK promotes apoptotic signaling via C/EBP-homologous protein (CHOP), while IRE1α-induced splicing of XBP1 (XBP1s) maintains cell survival by repressing apoptosis. This leads to an encouraging synergistic antitumor effect of combining AZD1775 and an IRE1α inhibitor MKC8866 in multiple cell lines and preclinical models of ovarian cancers. Taken together, the data reveal an important dual role of the UPR signaling network in mtTP53 ovarian cancer models in response to AZD1775 and suggest that inhibition of the IRE1α-XBP1s pathway may enhance the efficacy of AZD1775 in the clinics.

• Berge-Seidl, Sebastian; Nielsen, Nis Valentin; Alfonso, Armando A. Rodriguez; Etscheid, Michael; Kandanur, Sai Priya Sarma & Haug, Bengt Erik [Show all 15 contributors for this article] (2022). Identification of a phage display-derived peptide interacting with the N-terminal region of factor VII activating protease (FSAP) enables characterization of zymogen activation. ACS Chemical Biology. ISSN 1554-8929. 17(9), p. 2631–2642. doi: 10.1021/acschembio.2c00538. Full text in Research Archive Show summary

  Factor VII Activating protease (FSAP) has a protective effect in diverse disease conditions as inferred from studies in FSAP–/– mice and humans deficient in FSAP activity due to single-nucleotide polymorphism. The zymogen form of FSAP in plasma is activated by extracellular histones that are released during tissue injury or inflammation or by positively charged surfaces. However, it is not clear whether this activation mechanism is specific and amenable to manipulation. Using a phage display approach, we have identified a Cys-constrained 11 amino acid peptide, NNKC9/41, that activates pro-FSAP in plasma. The synthetic linear peptide has a propensity to cyclize through the terminal Cys groups, of which the antiparallel cyclic dimer, but not the monocyclic peptide, is the active component. Other commonly found zymogens in the plasma, related to the hemostasis system, were not activated. Binding studies with FSAP domain deletion mutants indicate that the N-terminus of FSAP is the key interaction site of this peptide. In a monoclonal antibody screen, we identified MA-FSAP-38C7 that prevented the activation of pro-FSAP by the peptide. This antibody bound to the LESLDP sequence (amino acids 30–35) in an intrinsically disordered stretch in the N-terminus of FSAP. The plasma clotting time was shortened by NNKC9/41, and this was reversed by MA-FSAP-38C7, demonstrating the utility of this peptide. Peptide NNKC9/41 will be useful as a tool to delineate the molecular mechanism of activation of pro-FSAP, elucidate its biological role, and provide a starting point for the pharmacological manipulation of FSAP activity.

• Berg, Henriette Engen; Halldórsso, Skarphéðinn; Bakketeig, Eris Aas; Thiede, Bernd; Sandberg, Cecilie & Lundanes, Elsa [Show all 8 contributors for this article] (2022). Micro‐pillar array columns (µPAC): An efficient tool for comparing tissue and cultured cells of glioblastoma. Journal of chromatography open. ISSN 2772-3917. 2. doi: 10.1016/j.jcoa.2022.100047. Full text in Research Archive
• Hanes, Robert; Ayuda-Duran, Maria del Pilar; Rønneberg, Leiv; Nakken, Sigve; Hovig, Eivind & Zucknick, Manuela [Show all 7 contributors for this article] (2022). screenwerk: a modular tool for the design and analysis of drug combination screens. Bioinformatics. ISSN 1367-4803. 39(1). doi: 10.1093/bioinformatics/btac840. Full text in Research Archive Show summary

  Abstract Motivation There is a rapidly growing interest in high-throughput drug combination screening to identify synergizing drug interactions for treatment of various maladies, such as cancer and infectious disease. This creates the need for pipelines that can be used to design such screens, perform quality control on the data and generate data files that can be analyzed by synergy-finding bioinformatics applications. Results screenwerk is an open-source, end-to-end modular tool available as an R-package for the design and analysis of drug combination screens. The tool allows for a customized build of pipelines through its modularity and provides a flexible approach to quality control and data analysis. screenwerk is adaptable to various experimental requirements with an emphasis on precision medicine. It can be coupled to other R packages, such as bayesynergy, to identify synergistic and antagonistic drug interactions in cell lines or patient samples. screenwerk is scalable and provides a complete solution for setting up drug sensitivity screens, read raw measurements and consolidate different datasets, perform various types of quality control and analyze, report and visualize the results of drug sensitivity screens. Availability and implementation The R-package and technical documentation is available at https://github.com/Enserink-lab/screenwerk; the R source code is publicly available at https://github.com/Enserink-lab/screenwerk under GNU General Public License v3.0; bayesynergy is accessible at https://github.com/ocbe-uio/bayesynergy. Selected modules are available through Galaxy, an open-source platform for FAIR data analysis at https://oncotools.elixir.no Supplementary information Supplementary data are available at Bioinformatics online.

• Jin, Yixin; Lorvik, Kristina Berg; Jin, Yang; Beck, Carole; Sike, Adam & Persiconi, Irene [Show all 10 contributors for this article] (2022). Development of STEAP1 targeting chimeric antigen receptor for adoptive cell therapy against cancer. Molecular Therapy Oncolytics. 26, p. 189–206. doi: 10.1016/j.omto.2022.06.007. Full text in Research Archive
• Zhao, Xu; Hendriks, Ivo A; Gras, Stephanie Le; Ye, Tao; Alonso, Lucia Ramos & P, Aurélie Nguéa [Show all 13 contributors for this article] (2022). Waves of sumoylation support transcription dynamics during adipocyte differentiation. Nucleic Acids Research (NAR). ISSN 0305-1048. 50(3), p. 1351–1369. doi: 10.1093/nar/gkac027. Full text in Research Archive Show summary

  Abstract Tight control of gene expression networks required for adipose tissue formation and plasticity is essential for adaptation to energy needs and environmental cues. However, the mechanisms that orchestrate the global and dramatic transcriptional changes leading to adipocyte differentiation remain to be fully unraveled. We investigated the regulation of nascent transcription by the sumoylation pathway during adipocyte differentiation using SLAMseq and ChIPseq. We discovered that the sumoylation pathway has a dual function in differentiation; it supports the initial downregulation of pre-adipocyte-specific genes, while it promotes the establishment of the mature adipocyte transcriptional program. By characterizing endogenous sumoylome dynamics in differentiating adipocytes by mass spectrometry, we found that sumoylation of specific transcription factors like PPARγ/RXR and their co-factors are associated with the transcription of adipogenic genes. Finally, using RXR as a model, we found that sumoylation may regulate adipogenic transcription by supporting the chromatin occurrence of transcription factors. Our data demonstrate that the sumoylation pathway supports the rewiring of transcriptional networks required for formation of functional adipocytes. This study also provides the scientists in the field of cellular differentiation and development with an in-depth resource of the dynamics of the SUMO-chromatin landscape, SUMO-regulated transcription and endogenous sumoylation sites during adipocyte differentiation.

• Marquina, Maribel; Lambea, Eva; Carmona, Mercé; Sánchez-Marinas, Marta; Aviles, Sandra Lopez & Ayte, José [Show all 8 contributors for this article] (2022). A new negative feedback mechanism for MAPK pathway inactivation through Srk1 MAPKAP kinase. Scientific Reports. 12(1). doi: 10.1038/s41598-022-23970-8. Full text in Research Archive Show summary

  Abstract The fission yeast mitogen-activated kinase (MAPK) Sty1 is essential for cell survival in response to different environmental insults. In unstimulated cells, Sty1 forms an inactive ternary cytoplasmatic complex with the MAPKK Wis1 and the MAPKAP kinase Srk1. Wis1 phosphorylates and activates Sty1, inducing the nuclear translocation of the complex. Once in the nucleus, Sty1 phosphorylates and activates Srk1, which in turns inhibits Cdc25 and cell cycle progression, before being degraded in a proteasome-dependent manner. In parallel, active nuclear Sty1 activates the transcription factor Atf1, which results in the expression of stress response genes including pyp2 (a MAPK phosphatase) and srk1 . Despite its essentiality in response to stress, persistent activation of the MAPK pathway can be deleterious and induces cell death. Thus, timely pathway inactivation is essential to ensure an appropriate response and cell viability. Here, uncover a role for the MAPKAP kinase Srk1 as an essential component of a negative feedback loop regulating the Sty1 pathway through phosphorylation and inhibition of the Wis1 MAPKK. This feedback regulation by a downstream kinase in the pathway highlights an additional mechanism for fine-tuning of MAPK signaling. Thus, our results indicate that Srk1 not only facilitates the adaptation to stress conditions by preventing cell cycle progression, but also plays an instrumental role regulating the upstream kinases in the stress MAPK pathway.

• Enserink, Jorrit & Chymkowitch, Pierre (2022). Cell Cycle-Dependent Transcription: The Cyclin Dependent Kinase Cdk1 Is a Direct Regulator of Basal Transcription Machineries. International Journal of Molecular Sciences. ISSN 1661-6596. 23(3), p. 1–23. doi: 10.3390/ijms23031293. Full text in Research Archive Show summary

  The cyclin-dependent kinase Cdk1 is best known for its function as master regulator of the cell cycle. It phosphorylates several key proteins to control progression through the different phases of the cell cycle. However, studies conducted several decades ago with mammalian cells revealed that Cdk1 also directly regulates the basal transcription machinery, most notably RNA polymerase II. More recent studies in the budding yeast Saccharomyces cerevisiae have revisited this function of Cdk1 and also revealed that Cdk1 directly controls RNA polymerase III activity. These studies have also provided novel insight into the physiological relevance of this process. For instance, cell cycle-stage-dependent activity of these complexes may be important for meeting the increased demand for various proteins involved in housekeeping, metabolism, and protein synthesis. Recent work also indicates that direct regulation of the RNA polymerase II machinery promotes cell cycle entry. Here, we provide an overview of the regulation of basal transcription by Cdk1, and we hypothesize that the original function of the primordial cell-cycle CDK was to regulate RNAPII and that it later evolved into specialized kinases that govern various aspects of the transcription machinery and the cell cycle.

• Kogler, Stian; Aizenshtadt, Aleksandra; Harrison, Sean; Skottvoll, Frøydis Sved; Berg, Henriette Engen & Abadpour, Shadab [Show all 14 contributors for this article] (2022). "Organ-in-a-Column" Coupled On-line with Liquid Chromatography-Mass Spectrometry. Analytical Chemistry. ISSN 0003-2700. 94(50), p. 17677–17684. doi: 10.1021/acs.analchem.2c04530. Full text in Research Archive Show summary

  Organoids, i.e., laboratory-grown organ models developed from stem cells, are emerging tools for studying organ physiology, disease modeling, and drug development. On-line analysis of organoids with mass spectrometry would provide analytical versatility and automation. To achieve these features with robust hardware, we have loaded liquid chromatography column housings with induced pluripotent stem cell (iPSC) derived liver organoids and coupled the “organ-in-a-column” units on-line with liquid chromatography-mass spectrometry (LC-MS). Liver organoids were coloaded with glass beads to achieve an even distribution of organoids throughout the column while preventing clogging. The liver organoids were interrogated “on column” with heroin, followed by on-line monitoring of the drug’s phase 1 metabolism. Enzymatic metabolism of heroin produced in the “organ-in-a-column” units was detected and monitored using a triple quadrupole MS instrument, serving as a proof-of-concept for on-line coupling of liver organoids and mass spectrometry. Taken together, the technology allows direct integration of liver organoids with LC-MS, allowing selective and automated tracking of drug metabolism over time.

• Pekec, Tina; Lewandowski, Jarosław; Komur, Alicja; Sobańska, Daria; Guo, Yanwu & Świtońska-Kurkowska, Karolina [Show all 12 contributors for this article] (2022). Ferritin-mediated iron detoxification promotes hypothermia survival in Caenorhabditis elegans and murine neurons. Nature Communications. 13(1). doi: 10.1038/s41467-022-32500-z. Full text in Research Archive Show summary

  Abstract How animals rewire cellular programs to survive cold is a fascinating problem with potential biomedical implications, ranging from emergency medicine to space travel. Studying a hibernation-like response in the free-living nematode Caenorhabditis elegans , we uncovered a regulatory axis that enhances the natural resistance of nematodes to severe cold. This axis involves conserved transcription factors, DAF-16/FoxO and PQM-1, which jointly promote cold survival by upregulating FTN-1, a protein related to mammalian ferritin heavy chain (FTH1). Moreover, we show that inducing expression of FTH1 also promotes cold survival of mammalian neurons, a cell type particularly sensitive to deterioration in hypothermia. Our findings in both animals and cells suggest that FTN-1/FTH1 facilitates cold survival by detoxifying ROS-generating iron species. We finally show that mimicking the effects of FTN-1/FTH1 with drugs protects neurons from cold-induced degeneration, opening a potential avenue to improved treatments of hypothermia.

• Sobańska, Daria; Komur, Alicja; Chabowska-Kita, Agnieszka; Gumna, Julita; Kumari, Pooja & Pachulska-Wieczorek, Katarzyna [Show all 7 contributors for this article] (2022). The silencing of ets-4 mRNA relies on the functional cooperation between REGE-1/Regnase-1 and RLE-1/Roquin-1. Nucleic Acids Research (NAR). ISSN 0305-1048. 50(14), p. 8226–8239. doi: 10.1093/nar/gkac609. Full text in Research Archive Show summary

  Abstract Regnase-1 is an evolutionarily conserved endoribonuclease. It degrades diverse mRNAs important for many biological processes including immune homeostasis, development and cancer. There are two competing models of Regnase-1-mediated mRNA silencing. One model postulates that Regnase-1 works together with another RNA-binding protein, Roquin-1, which recruits Regnase-1 to specific mRNAs. The other model proposes that the two proteins function separately. Studying REGE-1, the Caenorhabditis elegans ortholog of Regnase-1, we have uncovered its functional relationship with RLE-1, the nematode counterpart of Roquin-1. While both proteins are essential for mRNA silencing, REGE-1 and RLE-1 appear to associate with target mRNA independently of each other. Thus, although the functional interdependence between REGE-1/Regnase-1 and RLE-1/Roquin-1 is conserved, the underlying mechanisms may display species-specific variation, providing a rare perspective on the evolution of this important post-transcriptional regulatory mechanism.

• Alonso, Lucia Ramos; Llorente, Ignacio Garcia; Enserink, Jorrit & Chymkowitch, Pierre (2022). Analysis of the pheromone signaling pathway by RT-qPCR in the budding yeast Saccharomyces cerevisiae. STAR Protocols. 3(1). doi: 10.1016/j.xpro.2022.101210. Full text in Research Archive Show summary

  FUS3 and STE2 expression levels can be used as reporters for signaling through the pheromone pathway in the budding yeast Saccharomyces cerevisiae. Here, we describe an optimized protocol to measure the expression levels of FUS3 and STE2 using quantitative reverse transcription PCR (RT-qPCR). We describe the steps for comparing untreated and pheromone-treated yeast cells and how to quantify the changes in various deletion strains. The protocol can be applied to determine potential regulators of the pheromone pathway. For complete details on the use and execution of this protocol, please refer to Garcia et al. (2021).

• Grapotte, Mathys; Saraswat, Manu; Bessière, Chloé; Menichelli, Christophe; Ramilowski, Jordan A. & Severin, Jessica [Show all 490 contributors for this article] (2021). Discovery of widespread transcription initiation at microsatellites predictable by sequence-based deep neural network. Nature Communications. 12(1). doi: 10.1038/s41467-021-23143-7. Full text in Research Archive Show summary

  Abstract Using the Cap Analysis of Gene Expression (CAGE) technology, the FANTOM5 consortium provided one of the most comprehensive maps of transcription start sites (TSSs) in several species. Strikingly, ~72% of them could not be assigned to a specific gene and initiate at unconventional regions, outside promoters or enhancers. Here, we probe these unassigned TSSs and show that, in all species studied, a significant fraction of CAGE peaks initiate at microsatellites, also called short tandem repeats (STRs). To confirm this transcription, we develop Cap Trap RNA-seq, a technology which combines cap trapping and long read MinION sequencing. We train sequence-based deep learning models able to predict CAGE signal at STRs with high accuracy. These models unveil the importance of STR surrounding sequences not only to distinguish STR classes, but also to predict the level of transcription initiation. Importantly, genetic variants linked to human diseases are preferentially found at STRs with high transcription initiation level, supporting the biological and clinical relevance of transcription initiation at STRs. Together, our results extend the repertoire of non-coding transcription associated with DNA tandem repeats and complexify STR polymorphism.

• Nenseth, Hatice Zeynep; Sahu, Aparna; Saatcioglu, Fahri & Osguthorpe, Steven (2021). A Nutraceutical Formula Is Effective in Raising the Circulating Vitamin and Mineral Levels in Healthy Subjects: A Randomized Trial. Frontiers in Nutrition. 8. doi: 10.3389/fnut.2021.703394. Full text in Research Archive Show summary

  Low levels of nutrient intake are common in industrialized countries. This has negative implications on health and is associated with chronic diseases. Supplementation of vitamins, minerals, and key nutrients to optimal levels may, therefore, be beneficial for individual health and for the health economy. Although the use of supplements has become very common, due to a lack of monitoring, there is very limited data on the efficacy of supplementation with different formulas. In this study, we present the results of a randomized controlled study on the efficacy of a novel formulated nutraceutical, N247, in 250 healthy volunteers aged 26–75 years and a placebo control group (n = 35). The broad-spectrum formulation of N247 includes essential vitamins, minerals, and trace elements that are adequately balanced in regard to synergies and related metabolic functions. Moreover, tolerance, safety, and nutrient availability is an important aspect of daily, long-term use of N247. After 3 months of regular N247 use, levels of vitamins and minerals in serum were significantly increased in the N247 group compared with the control group and a placebo group, with excellent compliance rates. Coupled with additional natural ingredients that aim to increase the potency of the nutrients, N247 may represent a novel and beneficial supplement for individuals with nutritional deficiencies.

• Mester, Simone; Evers, Mitchell; Meyer, Saskia; Nilsen, Jeannette; Greiff, Victor & Sandlie, Inger [Show all 8 contributors for this article] (2021). Extended plasma half-life of albumin-binding domain fused human IgA upon pH-dependent albumin engagement of human FcRn in vitro and in vivo. mAbs. ISSN 1942-0862. 13:1893888(1), p. 1–14. doi: 10.1080/19420862.2021.1893888. Full text in Research Archive Show summary

  Albumin has a serum half-life of 3 weeks in humans. This feature can be used to improve the pharmacokinetics of shorter-lived biologics. For instance, an albumin-binding domain (ABD) can be used to recruit albumin. A prerequisite for such design is that the ABD-albumin interaction does not interfere with pH-dependent binding of albumin to the human neonatal Fc receptor (FcRn), as FcRn acts as the principal regulator of the half-life of albumin. Thus, there is a need to know how ABDs act in the context of fusion partners and human FcRn. Here, we studied the binding and transport properties of human immunoglobulin A1 (IgA1), fused to a Streptococcus protein G-derived engineered ABD, in in vitro and in vivo systems harboring human FcRn. IgA has great potential as a therapeutic protein, but its short half-life is a major drawback. We demonstrate that ABD-fused IgA1 binds human FcRn pH-dependently and is rescued from cellular degradation in a receptor-specific manner in the presence of albumin. This occurs when ABD is fused to either the light or the heavy chain. In human FcRn transgenic mice, IgA1-ABD in complex with human albumin, gave 4-6-fold extended half-life compared to unmodified IgA1, where the light chain fusion showed the longest half-life. When the heavy chain-fused protein was pre-incubated with an engineered human albumin with improved FcRn binding, cellular rescue and half-life was further enhanced. Our study reveals how an ABD, which does not interfere with albumin binding to human FcRn, may be used to extend the half-life of IgA.

• Shoor, Marita; Gudim, Ingvild; Hersleth, Hans-Petter & Hammerstad, Marta (2021). Thioredoxin reductase from Bacillus cereus exhibits distinct reduction and NADPH-binding properties. FEBS Open Bio. 11(11), p. 3019–3031. doi: 10.1002/2211-5463.13289. Full text in Research Archive Show summary

  Low molecular weight (low Mr) thioredoxin reductases (TrxRs) are homodimeric NADPH-dependent dithiol flavoenzymes that reduce thioredoxins (Trxs) or Trx-like proteins involved in the activation networks of enzymes, such as the bacterial class Ib ribonucleotide reductase (RNR). During the last few decades, TrxR-like ferredoxin/flavodoxin NADP+ oxidoreductases (FNRs) have been discovered and characterized in several types of bacteria, including those not encoding the canonical plant-type FNR. In Bacillus cereus, a TrxR-like FNR has been shown to reduce the flavodoxin-like protein NrdI in the activation of class Ib RNR. However, some species only encode TrxR, and lack the homologous TrxR-like FNR. Due to the structural similarity between TrxRs and TrxR-like FNRs, as well as variations in their occurrence in different microorganisms, we hypothesized that low Mr TrxR may be able to replace TrxR-like FNR in, for example, the reduction of NrdI. In this study, characterization of TrxR from B. cereus has revealed a weak FNR activity towards NrdI reduction. Additionally, the crystal structure shows that only one out of two binding sites of the B. cereus TrxR homodimer is occupied with NADPH, indicating a possible asymmetric co-substrate binding in TrxR.

• Caulier, Benjamin William; Enserink, Jorrit & Wälchli, Sébastien (2021). Pharmacologic control of CAR T cells. International Journal of Molecular Sciences. ISSN 1661-6596. 22(9), p. 1–23. doi: 10.3390/ijms22094320. Full text in Research Archive Show summary

  Chimeric antigen receptor (CAR) therapy is a promising modality for the treatment of advanced cancers that are otherwise incurable. During the last decade, different centers worldwide have tested the anti-CD19 CAR T cells and shown clinical benefits in the treatment of B cell tumors. However, despite these encouraging results, CAR treatment has also been found to lead to serious side effects and capricious response profiles in patients. In addition, the CD19 CAR success has been difficult to reproduce for other types of malignancy. The appearance of resistant tumor variants, the lack of antigen specificity, and the occurrence of severe adverse effects due to over-stimulation of the therapeutic cells have been identified as the major impediments. This has motivated a growing interest in developing strategies to overcome these hurdles through CAR control. Among them, the combination of small molecules and approved drugs with CAR T cells has been investigated. These have been exploited to induce a synergistic anti-cancer effect but also to control the presence of the CAR T cells or tune the therapeutic activity. In the present review, we discuss opportunistic and rational approaches involving drugs featuring anti-cancer efficacy and CAR-adjustable effect.

• Dalum, Alf Seljenes; Kraus, Aurora; Khan, Shanawaz; Davydova, Erna; Rigaudeau, Dimitri & Bjørgen, Håvard [Show all 13 contributors for this article] (2021). High-Resolution, 3D Imaging of the Zebrafish Gill-Associated Lymphoid Tissue (GIALT) Reveals a Novel Lymphoid Structure, the Amphibranchial Lymphoid Tissue. Frontiers in Immunology. 12. doi: 10.3389/fimmu.2021.769901. Full text in Research Archive Show summary

  The zebrafish is extensively used as an animal model for human and fish diseases. However, our understanding of the structural organization of its immune system remains incomplete, especially the mucosa-associated lymphoid tissues (MALTs). Teleost MALTs are commonly perceived as diffuse and scattered populations of immune cells throughout the mucosa. Yet, structured MALTs have been recently discovered in Atlantic salmon (Salmo salar L.), including the interbranchial lymphoid tissue (ILT) in the gills. The existence of the ILT was only recently identified in zebrafish and other fish species, highlighting the need for in-depth characterizations of the gill-associated lymphoid tissue (GIALT) in teleosts. Here, using 3-D high-resolution microscopy, we analyze the GIALT of adult zebrafish with an immuno-histology approach that reveals the organization of lymphoid tissues via the labeling of T/NK cells with an antibody directed to a highly conserved epitope on the kinase ZAP70. We show that the GIALT in zebrafish is distributed over at least five distinct sub-regions, an organization found in all pairs of gill arches. The GIALT is diffuse in the pharyngeal part of the gill arch, the interbranchial septum and the filaments/ lamellae, and structured in two sub-regions: the ILT, and a newly discovered lymphoid structure located along each side of the gill arch, which we named the Amphibranchial Lymphoid Tissue (ALT). Based on RAG2 expression, neither the ILT nor the ALT constitute additional thymi. The ALT shares several features with the ILT such as presence of abundant lymphoid cells and myeloid cells embedded in a network of reticulated epithelial cells. Further, the ILT and the ALT are also a site for T/NK cell proliferation. Both ILT and ALT show structural changes after infection with Spring Viraemia of Carp Virus (SVCV). Together, these data suggest that ALT and ILT play an active role in immune responses. Comparative studies show that whereas the ILT seems absent in most neoteleosts (“Percomorphs”), the ALT is widely present in cyprinids, salmonids and neoteleosts, suggesting that it constitutes a conserved tissue involved in the protection of teleosts via the gills.

• Grad, Iwona; Hanes, Robert; Ayuda-Duran, Maria del Pilar; Kuijjer, Marieke Lydia; Enserink, Jorrit & Meza, Leonardo Zepeda [Show all 7 contributors for this article] (2021). Discovery of novel candidates for anti-liposarcoma therapies by medium-scale high-throughput drug screening. PLOS ONE. 16(3), p. 1–23. doi: 10.1371/journal.pone.0248140. Full text in Research Archive Show summary

  Sarcomas are a heterogeneous group of mesenchymal orphan cancers and new treatment alternatives beyond traditional chemotherapeutic regimes are much needed. So far, tumor mutation analysis has not led to significant treatment advances, and we have attempted to bypass this limitation by performing direct drug testing of a library of 353 anti-cancer compounds that are either FDA-approved, in clinical trial, or in advanced stages of preclinical development on a panel of 13 liposarcoma cell lines. We identified and validated six drugs, targeting different mechanisms and with good efficiency across the cell lines: MLN2238 –a proteasome inhibitor, GSK2126458 –a PI3K/mTOR inhibitor, JNJ-26481585 –a histone deacetylase inhibitor, triptolide–a multi-target drug, YM155 –a survivin inhibitor, and APO866 (FK866)–a nicotinamide phosphoribosyl transferase inhibitor. GR50s for those drugs were mostly in the nanomolar range, and in many cases below 10 nM. These drugs had long-lasting effect upon drug withdrawal, limited toxicity to normal cells and good efficacy also against tumor explants. Finally, we identified potential genomic biomarkers of their efficacy. Being approved or in clinical trials, these drugs are promising candidates for liposarcoma treatment.

• Cokan, Kaja Blagotinšek; Urlep, Žiga; Moškon, Miha; Mraz, Miha; Kong, Xiang Yi & Eskild, Winnie [Show all 9 contributors for this article] (2021). Common Transcriptional Program of Liver Fibrosis in Mouse Genetic Models and Humans. International Journal of Molecular Sciences. ISSN 1661-6596. 22(2), p. 1–21. doi: 10.3390/ijms22020832. Full text in Research Archive Show summary

  Multifactorial metabolic diseases, such as non-alcoholic fatty liver disease, are a major burden to modern societies, and frequently present with no clearly defined molecular biomarkers. Herein we used system medicine approaches to decipher signatures of liver fibrosis in mouse models with malfunction in genes from unrelated biological pathways: cholesterol synthesis—Cyp51, notch signaling—Rbpj, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling—Ikbkg, and unknown lysosomal pathway—Glmp. Enrichment analyses of Kyoto Encyclopedia of Genes and Genomes (KEGG), Reactome and TRANScription FACtor (TRANSFAC) databases complemented with genome-scale metabolic modeling revealed fibrotic signatures highly similar to liver pathologies in humans. The diverse genetic models of liver fibrosis exposed a common transcriptional program with activated estrogen receptor alpha (ERα) signaling, and a network of interactions between regulators of lipid metabolism and transcription factors from cancer pathways and the immune system. The novel hallmarks of fibrosis are downregulated lipid pathways, including fatty acid, bile acid, and steroid hormone metabolism. Moreover, distinct metabolic subtypes of liver fibrosis were proposed, supported by unique enrichment of transcription factors based on the type of insult, disease stage, or potentially, also sex. The discovered novel features of multifactorial liver fibrotic pathologies could aid also in improved stratification of other fibrosis related pathologies.

• Furumizu, Chihiro; Krabberød, Anders Kristian; Hammerstad, Marta; Alling, Renate; Wildhagen, Mari & Sawa, Shinichiro [Show all 7 contributors for this article] (2021). The sequenced genomes of non-flowering land plants reveal the innovative evolutionary history of peptide signaling. The Plant Cell. ISSN 1040-4651. 33(9), p. 2915–2934. doi: 10.1093/plcell/koab173. Full text in Research Archive Show summary

  Abstract An understanding of land plant evolution is a prerequisite for in-depth knowledge of plant biology. Here we extract and explore information hidden in the increasing number of sequenced plant genomes, from bryophytes to angiosperms, to elucidate a specific biological question—how peptide signaling evolved. To conquer land and cope with changing environmental conditions, plants have gone through transformations that must have required innovations in cell-to-cell communication. We discuss peptides mediating endogenous and exogenous changes by interaction with receptors activating intracellular molecular signaling. Signaling peptides were discovered in angiosperms and operate in tissues and organs such as flowers, seeds, vasculature, and 3D meristems that are not universally conserved across land plants. Nevertheless, orthologs of angiosperm peptides and receptors have been identified in nonangiosperms. These discoveries provoke questions regarding coevolution of ligands and their receptors, and whether de novo interactions in peptide signaling pathways may have contributed to generate novel traits in land plants. The answers to such questions will have profound implications for the understanding of the evolution of cell-to-cell communication and the wealth of diversified terrestrial plants. Under this perspective, we have generated, analyzed, and reviewed phylogenetic, genomic, structural, and functional data to elucidate the evolution of peptide signaling.

• Guo, Yanwu; Tocchini, Cristina & Ciosk, Rafal (2021). CLK-2/TEL2 is a conserved component of the nonsense-mediated mRNA decay pathway. PLOS ONE. 16(1), p. 1–16. doi: 10.1371/journal.pone.0244505. Full text in Research Archive Show summary

  Nonsense-mediated mRNA decay (NMD) controls eukaryotic mRNA quality, inducing the degradation of faulty transcripts. Key players in the NMD pathway were originally identified, through genetics, in Caenorhabditis elegans as smg (suppressor with morphological effect on genitalia) genes. Using forward genetics and fluorescence-based NMD reporters, we reexamined the genetic landscape underlying NMD. Employing a novel strategy for mapping sterile mutations, Het-Map, we identified clk-2 , a conserved gene previously implicated in DNA damage signaling, as a player in the nematode NMD. We find that CLK-2 is expressed predominantly in the germline, highlighting the importance of auxiliary factors in tissue-specific mRNA decay. Importantly, the human counterpart of CLK-2/TEL2, TELO2, has been also implicated in the NMD, suggesting a conserved role of CLK-2/TEL2 proteins in mRNA surveillance. Recently, variants of TELO2 have been linked to an intellectual disability disorder, the You-Hoover-Fong syndrome, which could be related to its function in the NMD.

• Bengtsen, Mads; Winje, Ivan Myhre; Eftestøl, Einar; Landskron, Johannes; Sun, Chengyi & Nygård, Kamilla Børve [Show all 10 contributors for this article] (2021). Comparing the epigenetic landscape in myonuclei purified with a PCM1 antibody from a fast/glycolytic and a slow/oxidative muscle. PLoS Genetics. ISSN 1553-7390. 17:e1009907(11). doi: 10.1371/journal.pgen.1009907. Full text in Research Archive Show summary

  Muscle cells have different phenotypes adapted to different usage, and can be grossly divided into fast/glycolytic and slow/oxidative types. While most muscles contain a mixture of such fiber types, we aimed at providing a genome-wide analysis of the epigenetic landscape by ChIP-Seq in two muscle extremes, the fast/glycolytic extensor digitorum longus (EDL) and slow/oxidative soleus muscles. Muscle is a heterogeneous tissue where up to 60% of the nuclei can be of a different origin. Since cellular homogeneity is critical in epigenome-wide association studies we developed a new method for purifying skeletal muscle nuclei from whole tissue, based on the nuclear envelope protein Pericentriolar material 1 (PCM1) being a specific marker for myonuclei. Using antibody labelling and a magnetic-assisted sorting approach, we were able to sort out myonuclei with 95% purity in muscles from mice, rats and humans. The sorting eliminated influence from the other cell types in the tissue and improved the myo-specific signal. A genome-wide comparison of the epigenetic landscape in EDL and soleus reflected the differences in the functional properties of the two muscles, and revealed distinct regulatory programs involving distal enhancers, including a glycolytic super-enhancer in the EDL. The two muscles were also regulated by different sets of transcription factors; e.g. in soleus, binding sites for MEF2C, NFATC2 and PPARA were enriched, while in EDL MYOD1 and SIX1 binding sites were found to be overrepresented. In addition, more novel transcription factors for muscle regulation such as members of the MAF family, ZFX and ZBTB14 were identified.

• Cui, Fengzhen; Pan, Qingfei; Wang, Siyi; Zhao, Faming; Wang, Runxin & Zhang, Tingting [Show all 19 contributors for this article] (2021). Maternal Benzophenone Exposure Impairs Hippocampus Development and Cognitive Function in Mouse Offspring. Advanced Science. 8, p. 1–16. doi: 10.1002/advs.202102686. Full text in Research Archive Show summary

  Benzophenones are widely supplemented in personal care products, but little is known about its neurodevelopmental toxicity. The previous epidemiological study discovered a negative correlation between maternal exposure to a benzophenone metabolite 4-hydroxybenzophenone (4HBP) and child's neurodevelopment, yet the causal relationship and detailed mechanism remain to be defined. Here, it is reported that prenatal, but not postnatal, exposure to environmentally relevant level of 4HBP impairs hippocampus development and causes cognitive dysfunction in offspring mice. Transcriptomic analyses reveal that 4HBP induces the endoplasmic reticulum stress-induced apoptotic signaling and inflammatory response in hippocampal neural stem cells. Mechanistically, 4HBP exposure activates protein kinase R-like ER kinase (PERK) signaling, which induces CHOP, inhibits IκB translation, and transactivates p65, thereby promoting inflammation and apoptosis on multiple levels. Importantly, genetic or pharmacological inhibition of PERK pathway significantly attenuates 4HBP-induced NFκB signaling and neurodevelopmental abnormalities in mice and in a human brain organoid model. The study uncovers the neurodevelopmental toxicity of BP and cautions its exposure during pregnancy.

• Johannsen, Christina; Koehler, Christian & Thiede, Bernd (2021). Comparison of LFQ and IPTL for Protein Identification and Relative Quantification. Molecules. ISSN 1431-5157. 26(17). doi: 10.3390/molecules26175330. Full text in Research Archive Show summary

  (1) Background: Mass spectrometry-based quantitative proteome profiling is most commonly performed by label-free quantification (LFQ), stable isotopic labeling with amino acids in cell culture (SILAC), and reporter ion-based isobaric labeling methods (TMT and iTRAQ). Isobaric peptide termini labeling (IPTL) was described as an alternative to these methods and is based on crosswise labeling of both peptide termini and MS2 quantification. High quantification accuracy was assumed for IPTL because multiple quantification points are obtained per identified MS2 spectrum. A direct comparison of IPTL with other quantification methods has not been performed yet because IPTL commonly requires digestion with endoproteinase Lys-C. (2) Methods: To enable tryptic digestion of IPTL samples, a novel labeling for IPTL was developed that combines metabolic labeling (Arg-0/Lys-0 and Arg-d4/Lys-d4, respectively) with crosswise N-terminal dimethylation (d4 and d0, respectively). (3) Results: The comparison of IPTL with LFQ revealed significantly more protein identifications for LFQ above homology ion scores but not above identity ion scores. (4) Conclusions: The quantification accuracy was superior for LFQ despite the many quantification points obtained with IPTL.

• Venz, Richard; Pekec, Tina; Katic, Iskra; Ciosk, Rafal & Ewald, Collin Yves (2021). End-of-life targeted auxin-mediated degradation of DAF-2 Insulin/IGF-1 receptor promotes longevity free from growth-related pathologies. eLIFE. 10. doi: 10.7554/eLife.71335. Full text in Research Archive Show summary

  Preferably, lifespan-extending therapies should work when applied late in life without causing undesired pathologies. Reducing insulin/insulin-like growth factor (IGF)-1 signaling (IIS) increases lifespan across species, but the effects of reduced IIS interventions in extreme geriatric ages remains unknown. Using the nematode Caenorhabditis elegans , we engineered the conditional depletion of the DAF-2/insulin/IGF-1 transmembrane receptor using an auxin-inducible degradation (AID) system. This allowed for the temporal and spatial reduction in DAF-2 protein levels at time points after which interventions such as RNAi become ineffective. Using this system, we found that AID-mediated depletion of DAF-2 protein surpasses the longevity of daf-2 mutants. Depletion of DAF-2 during early adulthood resulted in multiple adverse phenotypes, including growth retardation, germline shrinkage, egg retention, and reduced brood size. By contrast, AID-mediated depletion of DAF-2 post-reproduction, or specifically in the intestine in early adulthood, resulted in an extension of lifespan without these deleterious effects. Strikingly, at geriatric ages, when 75% of the population had died, AID-mediated depletion of DAF-2 protein resulted in a doubling in lifespan. Thus, we provide a proof-of-concept that even close to the end of an individual’s lifespan, it is possible to slow aging and promote longevity.

• Pällmann, Nora; Ke, Deng; Livgård, Marte; Tesikova, Martina; Jin, Yixin & Frengen, Nicolai Sebastian [Show all 18 contributors for this article] (2021). Stress-mediated reprogramming of prostate cancer one-carbon cycle drives disease progression. Cancer Research. ISSN 0008-5472. 81(15), p. 4066–4078. doi: 10.1158/0008-5472.CAN-20-3956. Full text in Research Archive Show summary

  One-carbon (1C) metabolism has a key role in metabolic programming with both mitochondrial (m1C) and cytoplasmic (c1C) components. Here we show that activating transcription factor 4 (ATF4) exclusively activates gene expression involved in m1C, but not the c1C cycle in prostate cancer cells. This includes activation of methylenetetrahydrofolate dehydrogenase 2 (MTHFD2) expression, the central player in the m1C cycle. Consistent with the key role of m1C cycle in prostate cancer, MTHFD2 knockdown inhibited prostate cancer cell growth, prostatosphere formation, and growth of patient-derived xenograft organoids. In addition, therapeutic silencing of MTHFD2 by systemically administered nanoliposomal siRNA profoundly inhibited tumor growth in preclinical prostate cancer mouse models. Consistently, MTHFD2 expression is significantly increased in human prostate cancer, and a gene expression signature based on the m1C cycle has significant prognostic value. Furthermore, MTHFD2 expression is coordinately regulated by ATF4 and the oncoprotein c-MYC, which has been implicated in prostate cancer. These data suggest that the m1C cycle is essential for prostate cancer progression and may serve as a novel biomarker and therapeutic target. Significance: These findings demonstrate that the mitochondrial, but not cytoplasmic, one-carbon cycle has a key role in prostate cancer cell growth and survival and may serve as a biomarker and/or therapeutic target.

• Thiede, Bernd & Koehler, Christian (2021). An Approach for Triplex-IPTL. In Marcus, Katrin; Eisenacher, Martin & Sitek, Barbara (Ed.), Quantitative Methods in Proteomics. Humana Press. ISSN 9781071610237. p. 133–144. doi: 10.1007/978-1-0716-1024-4_10. Full text in Research Archive Show summary

  Isobaric peptide termini labeling (IPTL) is an approach for quantitative proteomics based on crosswise isotopic labeling of peptides at the N- and C-terminus. The labeling reagents are chosen in isotopic variations that the resulting mass of all labels per peptide is isobaric, but the individual label on each peptide terminus is different. Therefore, the quantitative difference of the peptide signal can be determined by the fragment ions of the corresponding MS2 spectra. Here, we describe an approach for triplex-IPTL to allow the comparison of three proteomes. This approach is based on digestion of the proteins by endoproteinase Lys-C, followed by three combinations of selective dimethylation of the peptide N-termini and subsequent dimethylation of the lysine residues at the C-termini. Data analysis is performed using Mascot for database searches and the freely available software package IsobariQ for quantification.

• Andersen, Elisabeth; Dugarte, Maria Eugenia Chollet; Sletten, Marit; Stavik, Benedicte; Skarpen, Ellen & Backe, Paul Hoff [Show all 9 contributors for this article] (2021). Molecular Characterization of Two Homozygous Factor VII Variants Associated with Intracranial Bleeding. Thrombosis and Haemostasis. ISSN 0340-6245. 121(12), p. 1588–1598. doi: 10.1055/a-1450-8568. Full text in Research Archive
• Bruun, Jarle; Eide, Peter Andreas Wold; Bergsland, Christian Holst; Brück, Oscar; Svindland, Aud & Arjama, Mariliina [Show all 13 contributors for this article] (2021). E-cadherin is a robust prognostic biomarker in colorectal cancer and low expression is associated with sensitivity to inhibitors of topoisomerase, aurora, and HSP90 in preclinical models. Molecular Oncology. ISSN 1574-7891. 00, p. 1–18. doi: 10.1002/1878-0261.13159. Full text in Research Archive Show summary

  Cell–cell and cell–matrix adhesion proteins that have been implicated in colorectal epithelial integrity and epithelial-to-mesenchymal transition could be robust prognostic and potential predictive biomarkers for standard and novel therapies. We analyzed in situ protein expression of E-cadherin (ECAD), integrin β4 (ITGB4), zonula occludens 1 (ZO-1), and cytokeratins in a single-hospital series of Norwegian patients with colorectal cancer (CRC) stages I–IV (n = 922) using multiplex fluorescence-based immunohistochemistry (mfIHC) on tissue microarrays. Pharmacoproteomic associations were explored in 35 CRC cell lines annotated with drug sensitivity data on > 400 approved and investigational drugs. ECAD, ITGB4, and ZO-1 were positively associated with survival, while cytokeratins were negatively associated with survival. Only ECAD showed independent prognostic value in multivariable Cox models. Clinical and molecular associations for ECAD were technically validated on a different mfIHC platform, and the prognostic value was validated in another Norwegian series (n = 798). In preclinical models, low and high ECAD expression differentially associated with sensitivity to topoisomerase, aurora, and HSP90 inhibitors, and EGFR inhibitors. E-cadherin protein expression is a robust prognostic biomarker with potential clinical utility in CRC.

• Malecki, Jedrzej Mieczyslaw; O'Donohue, Marie-Francoise; Kim, Yeji; Jakobsson, Magnus; Gessa, Luca & Pinto, Rita [Show all 14 contributors for this article] (2021). Human METTL18 is a histidine-specific methyltransferase that targets RPL3 and affects ribosome biogenesis and function. Nucleic Acids Research (NAR). ISSN 0305-1048. 49(6), p. 3185–3203. doi: 10.1093/nar/gkab088. Full text in Research Archive Show summary

  Abstract Protein methylation occurs primarily on lysine and arginine, but also on some other residues, such as histidine. METTL18 is the last uncharacterized member of a group of human methyltransferases (MTases) that mainly exert lysine methylation, and here we set out to elucidate its function. We found METTL18 to be a nuclear protein that contains a functional nuclear localization signal and accumulates in nucleoli. Recombinant METTL18 methylated a single protein in nuclear extracts and in isolated ribosomes from METTL18 knockout (KO) cells, identified as 60S ribosomal protein L3 (RPL3). We also performed an RPL3 interactomics screen and identified METTL18 as the most significantly enriched MTase. We found that His-245 in RPL3 carries a 3-methylhistidine (3MH; τ-methylhistidine) modification, which was absent in METTL18 KO cells. In addition, both recombinant and endogenous METTL18 were found to be automethylated at His-154, thus further corroborating METTL18 as a histidine-specific MTase. Finally, METTL18 KO cells displayed altered pre-rRNA processing, decreased polysome formation and codon-specific changes in mRNA translation, indicating that METTL18-mediated methylation of RPL3 is important for optimal ribosome biogenesis and function. In conclusion, we have here established METTL18 as the second human histidine-specific protein MTase, and demonstrated its functional relevance.

• Høydahl, Lene; Frigstad, Terje; Rasmussen, Ingunn B; Øynebråten, Inger; Schjetne, Karoline W & Andersen, Jan Terje [Show all 10 contributors for this article] (2021). Antibody-mediated delivery of T-cell epitopes to antigen-presenting cells induce strong CD4 and CD8 T-cell responses. Vaccine. ISSN 0264-410X. 39(11), p. 1583–1592. doi: 10.1016/j.vaccine.2021.02.012. Full text in Research Archive Show summary

  Targeted delivery of antigen to antigen-presenting cells (APCs) enhances antigen presentation and thus, is a potent strategy for making more efficacious vaccines. This can be achieved by use of antibodies with specificity for endocytic surface molecules expressed on the APC. We aimed to compare two different antibody-antigen fusion modes in their ability to induce T-cell responses; first, exchange of immunoglobulin (Ig) constant domain loops with a T-cell epitope (Troybody), and second, fusion of T-cell epitope or whole antigen to the antibody C-terminus. Although both strategies are well-established, they have not previously been compared using the same system. We found that both antibody-antigen fusion modes led to presentation of the T-cell epitope. The strength of the T-cell responses varied, however, with the most efficient Troybody inducing CD4 T-cell proliferation and cytokine secretion at 10–100-fold lower concentration than the antibodies carrying antigen fused to the C-terminus, both in vitro and after intravenous injection in mice. Furthermore, we exchanged this loop with an MHCI-restricted T-cell epitope, and the resulting antibody enabled efficient cross-presentation to CD8 T cells in vivo. Targeting of antigen to APCs by use of such antibody-antigen fusions is thus an attractive vaccination strategy for increased activation of both CD4 and CD8 peptide-specific T cells.

• Stepanenko, Iryna; Babak, Maria V.; Spengler, Gabriella; Hammerstad, Marta; Popovic-Bijelic, Ana & Shova, Sergiu [Show all 10 contributors for this article] (2021). Coumarin-Based Triapine Derivatives and Their Copper(II) Complexes: Synthesis, Cytotoxicity and mR2 RNR Inhibition Activity. Biomolecules. 11(862). doi: 10.3390/biom11060862. Full text in Research Archive Show summary

  A series of thiosemicarbazone-coumarin hybrids (HL1-HL3 and H2L4) has been synthesised in 12 steps and used for the preparation of mono- and dinuclear copper(II) complexes, namely Cu(HL1)Cl2 (1), Cu(HL2)Cl2 (2), Cu(HL3)Cl2 (3) and Cu2(H2L4)Cl4 (4), isolated in hydrated or solvated forms. Both the organic hybrids and their copper(II) and dicopper(II) complexes were comprehensively characterised by analytical and spectroscopic techniques, i.e., elemental analysis, ESI mass spectrometry, 1D and 2D NMR, IR and UV–vis spectroscopies, cyclic voltammetry (CV) and spectroelectrochemistry (SEC). Re-crystallisation of 1 from methanol afforded single crystals of copper(II) complex with monoanionic ligand Cu(L1)Cl, which could be studied by single crystal X-ray diffraction (SC-XRD). The prepared copper(II) complexes and their metal-free ligands revealed antiproliferative activity against highly resistant cancer cell lines, including triple negative breast cancer cells MDA-MB-231, sensitive COLO-205 and multidrug resistant COLO-320 colorectal adenocarcinoma cell lines, as well as in healthy human lung fibroblasts MRC-5 and compared to those for triapine and doxorubicin. In addition, their ability to reduce the tyrosyl radical in mouse R2 protein of ribonucleotide reductase has been ascertained by EPR spectroscopy and the results were compared with those for triapine.

• Besleaga, Iuliana; Stepanenko, Iryna; Petrasheuskaya, Tatsiana V.; Darvasiova, Denisa; Breza, Martin & Hammerstad, Marta [Show all 14 contributors for this article] (2021). Triapine Analogues and Their Copper(II) Complexes: Synthesis, Characterization, Solution Speciation, Redox Activity, Cytotoxicity, and mR2 RNR Inhibition. Inorganic Chemistry. ISSN 0020-1669. 60, p. 11297–11319. doi: 10.1021/acs.inorgchem.1c01275. Full text in Research Archive Show summary

  Three new thiosemicarbazones (TSCs) HL1–HL3 as triapine analogues bearing a redox-active phenolic moiety at the terminal nitrogen atom were prepared. Reactions of HL1–HL3 with CuCl2·2H2O in anoxic methanol afforded three copper(II) complexes, namely, Cu(HL1)Cl2 (1), [Cu(L2)Cl] (2′), and Cu(HL3)Cl2 (3), in good yields. Solution speciation studies revealed that the metal-free ligands are stable as HL1–HL3 at pH 7.4, while being air-sensitive in the basic pH range. In dimethyl sulfoxide they exist as a mixture of E and Z isomers. A mechanism of the E/Z isomerization with an inversion at the nitrogen atom of the Schiff base imine bond is proposed. The monocationic complexes [Cu(L1–3)]+ are the most abundant species in aqueous solutions at pH 7.4. Electrochemical and spectroelectrochemical studies of 1, 2′, and 3 confirmed their redox activity in both the cathodic and the anodic region of potentials. The one-electron reduction was identified as metal-centered by electron paramagnetic resonance spectroelectrochemistry. An electrochemical oxidation pointed out the ligand-centered oxidation, while chemical oxidations of HL1 and HL2 as well as 1 and 2′ afforded several two-electron and four-electron oxidation products, which were isolated and comprehensively characterized. Complexes 1 and 2′ showed an antiproliferative activity in Colo205 and Colo320 cancer cell lines with half-maximal inhibitory concentration values in the low micromolar concentration range, while 3 with the most closely related ligand to triapine displayed the best selectivity for cancer cells versus normal fibroblast cells (MRC-5). HL1 and 1 in the presence of 1,4-dithiothreitol are as potent inhibitors of mR2 ribonucleotide reductase as triapine.

• Davydova, Erna; Shimazu, Tadahiro; Schuhmacher, Maren Kirstin; Jakobsson, Magnus E.; Willemen, Hanneke L.D.M. & Liu, Tongri [Show all 28 contributors for this article] (2021). The methyltransferase METTL9 mediates pervasive 1-methylhistidine modification in mammalian proteomes. Nature Communications. 12, p. 1–14. doi: 10.1038/s41467-020-20670-7. Full text in Research Archive Show summary

  Abstract Post-translational methylation plays a crucial role in regulating and optimizing protein function. Protein histidine methylation, occurring as the two isomers 1- and 3-methylhistidine (1MH and 3MH), was first reported five decades ago, but remains largely unexplored. Here we report that METTL9 is a broad-specificity methyltransferase that mediates the formation of the majority of 1MH present in mouse and human proteomes. METTL9-catalyzed methylation requires a His-x-His (HxH) motif, where “x” is preferably a small amino acid, allowing METTL9 to methylate a number of HxH-containing proteins, including the immunomodulatory protein S100A9 and the NDUFB3 subunit of mitochondrial respiratory Complex I. Notably, METTL9-mediated methylation enhances respiration via Complex I, and the presence of 1MH in an HxH-containing peptide reduced its zinc binding affinity. Our results establish METTL9-mediated 1MH as a pervasive protein modification, thus setting the stage for further functional studies on protein histidine methylation.

• Pizzi, Eleonora; Halvorsen, Trine Grønhaug; Koehler, Christian & Reubsaet, Leon (2021). Next-Generation Dried Blood Spot Samplers for Protein Analysis: Describing Trypsin-Modified Smart Sampling Paper. Separations. 8(5). doi: 10.3390/separations8050066. Full text in Research Archive Show summary

  This paper describes smart sampling paper to be used for bottom-up protein analysis. Four different manners to immobilize trypsin on cellulose were evaluated. Untreated paper, potassium-periodate-functionalized paper (with and without post-immobilization reduction) and 2-hydroxyethyl methacrylate (HEMA)/2-vinyl-4,4-dimethylazlactone (VDM)-functionalized paper were all used to immobilize trypsin. For the evaluation, Coomassie Brilliant Blue staining of proteins on paper and the BAEE trypsin activity assay needed to be modified. These methods allowed, together with data from mass spectrometric analysis of cytochrome C digestions, us to acquire fundamental insight into protein binding, and trypsin action and activity on paper. All functionalized discs bind more protein than the untreated discs. Protein binding to functionalized discs is based on both adsorption and covalent binding. Trypsin immobilized on potassium-periodate-functionalized discs exhibits the highest trypsin activity when using cytochrome C as substrate. It is proven that it is trypsin attached to paper (and not desorbed trypsin) which is responsible for the enzyme activity. The use of discs on complex biological samples shows that all functionalized discs are able to digest diluted serum; for the best-performing disc, HEMA-VDM functionalized, up to 200 high-confidence proteins are qualified, showing its potential.

• Eskild, Winnie; May, Gerhard; Michelangeli, Frank; Minchin, Steve; Roberts, Mark & Saðýn, Ferhan [Show all 9 contributors for this article] (2021). Meeting reports: Evolving Molecular Bioscience Education 2021. Biochemist. ISSN 0954-982X. 43(4), p. 49–49. doi: 10.1042/bio_2021_160. Full text in Research Archive
• Frick, Rahel; Høydahl, Lene Støkken; Petersen, Jan; Pré, Fleur du; Kumari, Shraddha & Berntsen, Grete [Show all 20 contributors for this article] (2021). A high-affinity human TCR-like antibody detects celiac disease gluten peptide-MHC complexes and inhibits T cell activation. Science immunology. 6:eabg4925(62), p. 1–16. doi: 10.1126/sciimmunol.abg4925. Full text in Research Archive Show summary

  Antibodies specific for peptides bound to human leukocyte antigen (HLA) molecules are valuable tools for studies of antigen presentation and may have therapeutic potential. Here, we generated human T cell receptor (TCR)–like antibodies toward the immunodominant signature gluten epitope DQ2.5-glia-α2 in celiac disease (CeD). Phage display selection combined with secondary targeted engineering was used to obtain highly specific antibodies with picomolar affinity. The crystal structure of a Fab fragment of the lead antibody 3.C11 in complex with HLA-DQ2.5:DQ2.5-glia-α2 revealed a binding geometry and interaction mode highly similar to prototypic TCRs specific for the same complex. Assessment of CeD biopsy material confirmed disease specificity and reinforced the notion that abundant plasma cells present antigen in the inflamed CeD gut. Furthermore, 3.C11 specifically inhibited activation and proliferation of gluten-specific CD4+ T cells in vitro and in HLA-DQ2.5 humanized mice, suggesting a potential for targeted intervention without compromising systemic immunity.

• Klionsky, Daniel J.; Abdel-Aziz, Amal Kamal; Abdelfatah, Sara; Abdellatif, Mahmoud Khaled; Abdoli, Asghar & Abudu, Yakubu Princely [Show all 50 contributors for this article] (2021). Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition). Autophagy. ISSN 1554-8627. 17(1), p. 1–382. doi: https:/www.tandfonline.com/doi/full/10.1080/15548627.2020.1797280. Full text in Research Archive
• Rønneberg, Leiv; Cremaschi, Andrea; Hanes, Robert; Enserink, Jorrit & Zucknick, Manuela (2021). bayesynergy: flexible Bayesian modelling of synergistic interaction effects in in vitro drug combination experiments. Briefings in Bioinformatics. ISSN 1467-5463. 22(6), p. 1–12. doi: 10.1093/bib/bbab251. Full text in Research Archive Show summary

  Abstract The effect of cancer therapies is often tested pre-clinically via in vitro experiments, where the post-treatment viability of the cancer cell population is measured through assays estimating the number of viable cells. In this way, large libraries of compounds can be tested, comparing the efficacy of each treatment. Drug interaction studies focus on the quantification of the additional effect encountered when two drugs are combined, as opposed to using the treatments separately. In the bayesynergy R package, we implement a probabilistic approach for the description of the drug combination experiment, where the observed dose response curve is modelled as a sum of the expected response under a zero-interaction model and an additional interaction effect (synergistic or antagonistic). Although the model formulation makes use of the Bliss independence assumption, we note that the posterior estimates of the dose–response surface can also be used to extract synergy scores based on other reference models, which we illustrate for the Highest Single Agent model. The interaction is modelled in a flexible manner, using a Gaussian process formulation. Since the proposed approach is based on a statistical model, it allows the natural inclusion of replicates, handles missing data and uneven concentration grids, and provides uncertainty quantification around the results. The model is implemented in the open-source Stan programming language providing a computationally efficient sampler, a fast approximation of the posterior through variational inference, and features parallel processing for working with large drug combination screens.

• Llorente, Ignacio Garcia; Munoz, Sara Orellana; Alonso, Lucia Ramos; Andersen, Aram Nikolai; Zimmermann, Christine & Eriksson, Jens [Show all 11 contributors for this article] (2021). Kel1 is a phosphorylation-regulated noise suppressor of the pheromone signaling pathway. Cell reports. 37(13), p. 1–20. doi: 10.1016/j.celrep.2021.110186. Full text in Research Archive Show summary

  Mechanisms have evolved that allow cells to detect signals and generate an appropriate response. The accuracy of these responses relies on the ability of cells to discriminate between signal and noise. How cells filter noise in signaling pathways is not well understood. Here, we analyze noise suppression in the yeast pheromone signaling pathway and show that the poorly characterized protein Kel1 serves as a major noise suppressor and prevents cell death. At the molecular level, Kel1 prevents spontaneous activation of the pheromone response by inhibiting membrane recruitment of Ste5 and Far1. Only a hypophosphorylated form of Kel1 suppresses signaling, reduces noise, and prevents pheromone-associated cell death, and our data indicate that the MAPK Fus3 contributes to Kel1 phosphorylation. Taken together, Kel1 serves as a phospho-regulated suppressor of the pheromone pathway to reduce noise, inhibit spontaneous activation of the pathway, regulate mating efficiency, and prevent pheromone-associated cell death.

• Hammerstad, Marta & Hersleth, Hans-Petter (2021). Overview of structurally homologous flavoprotein oxidoreductases containing the low Mr thioredoxin reductase-like fold – A functionally diverse group. Archives of Biochemistry and Biophysics. ISSN 0003-9861. 702, p. 1–19. doi: 10.1016/j.abb.2021.108826. Full text in Research Archive Show summary

  Structural studies show that enzymes have a limited number of unique folds, although structurally related enzymes have evolved to perform a large variety of functions. In this review, we have focused on enzymes containing the low molecular weight thioredoxin reductase (low Mr TrxR) fold. This fold consists of two domains, both containing a three-layer ββα sandwich Rossmann-like fold, serving as flavin adenine dinucleotide (FAD) and, in most cases, pyridine nucleotide (NAD(P)H) binding-domains. Based on a search of the Protein Data Bank for all published structures containing the low Mr TrxR-like fold, we here present a comprehensive overview of enzymes with this structural architecture. These range from TrxR-like ferredoxin/flavodoxin NAD(P)+ oxidoreductases, through glutathione reductase, to NADH peroxidase. Some enzymes are solely composed of the low Mr TrxR-like fold, while others contain one or two additional domains. In this review, we give a detailed description of selected enzymes containing only the low Mr TrxR-like fold, however, catalyzing a diversity of chemical reactions. Our overview of this structurally similar, yet functionally distinct group of flavoprotein oxidoreductases highlights the fascinating and increasing number of studies describing the diversity among these enzymes, especially during the last decade(s).

• Guerreiro, Eduarda Mazagão; Øvstebø, Reidun; Thiede, Bernd; Costea, Daniela Elena; Søland, Tine Merete & Galtung, Hilde (2020). Cancer cell line-specific protein profiles in extracellular vesicles identified by proteomics. PLOS ONE. 15(9), p. 1–20. doi: 10.1371/journal.pone.0238591. Full text in Research Archive Show summary

  Extracellular vesicles (EVs), are important for intercellular communication in both physiological and pathological processes. To explore the potential of cancer derived EVs as disease biomarkers for diagnosis, monitoring, and treatment decision, it is necessary to thoroughly characterize their biomolecular content. The aim of the study was to characterize and compare the protein content of EVs derived from three different cancer cell lines in search of a specific molecular signature, with emphasis on proteins related to the carcinogenic process. Oral squamous cell carcinoma (OSCC), pancreatic ductal adenocarcinoma (PDAC) and melanoma brain metastasis cell lines were cultured in CELLine AD1000 flasks. EVs were isolated by ultrafiltration and size-exclusion chromatography and characterized. Next, the isolated EVs underwent liquid chromatography-mass spectrometry (LC-MS) analysis for protein identification. Functional enrichment analysis was performed for a more general overview of the biological processes involved. More than 600 different proteins were identified in EVs from each particular cell line. Here, 14%, 10%, and 24% of the identified proteins were unique in OSCC, PDAC, and melanoma vesicles, respectively. A specific protein profile was discovered for each cell line, e.g., EGFR in OSCC, Muc5AC in PDAC, and FN1 in melanoma vesicles. Nevertheless, 25% of all the identified proteins were common to all cell lines. Functional enrichment analysis linked the proteins in each data set to biological processes such as “biological adhesion”, “cell motility”, and “cellular component biogenesis”. EV proteomics discovered cancer-specific protein profiles, with proteins involved in processes promoting tumor progression. In addition, the biological processes associated to the melanoma-derived EVs were distinct from the ones linked to the EVs isolated from OSCC and PDAC. The malignancy specific biomolecular cues in EVs may have potential applications as diagnostic biomarkers and in therapy.

• Hammerstad, Marta; Gudim, Ingvild & Hersleth, Hans-Petter (2020). The Crystal Structures of Bacillithiol Disulfide Reductase Bdr (YpdA) Provide Structural and Functional Insight into a New Type of FAD-Containing NADPH-Dependent Oxidoreductase. Biochemistry. ISSN 0006-2960. 59(51), p. 4793–4798. doi: 10.1021/acs.biochem.0c00745. Full text in Research Archive Show summary

  Low G+C Gram-positive Firmicutes, such as the clinically important pathogens Staphylococcus aureus and Bacillus cereus, use the low-molecular weight thiol bacillithiol (BSH) as a defense mechanism to buffer the intracellular redox environment and counteract oxidative stress encountered by human neutrophils during infections. The protein YpdA has recently been shown to function as an essential NADPH-dependent reductase of oxidized bacillithiol disulfide (BSSB) resulting from stress responses and is crucial for maintaining the reduced pool of BSH and cellular redox balance. In this work, we present the first crystallographic structures of YpdAs, namely, those from S. aureus and B. cereus. Our analyses reveal a uniquely organized biological tetramer; however, the structure of the monomeric subunit is highly similar to those of other flavoprotein disulfide reductases. The absence of a redox active cysteine in the vicinity of the FAD isoalloxazine ring implies a new direct disulfide reduction mechanism, which is backed by the presence of a potentially gated channel, serving as a putative binding site for BSSB in the proximity of the FAD cofactor. We also report enzymatic activities for both YpdAs, which along with the structures presented in this work provide important structural and functional insight into a new class of FAD-containing NADPH-dependent oxidoreductases, related to the emerging fight against pathogenic bacteria.

• Guldvik, Ingrid Jenny; Zuber, Verena; Braadland, Peder Rustøen; Grytli, Helene Hartvedt; Ramberg, Håkon Andre & Lilleby, Wolfgang [Show all 18 contributors for this article] (2020). Identification and Validation of Leucine-rich alpha-2-glycoprotein 1 as a Noninvasive Biomarker for Improved Precision in Prostate Cancer Risk Stratification. European Urology Open Science. ISSN 2666-1691. 21, p. 51–61. doi: 10.1016/j.euros.2020.08.007. Full text in Research Archive Show summary

  Background More accurate risk assessments are needed to improve prostate cancer management. Objective To identify blood-based protein biomarkers that provided prognostic information for risk stratification. Design, setting, and participants Mass spectrometry was used to identify biomarker candidates from blood, and validation studies were performed in four independent cohorts retrospectively collected between 1988 and 2015. Outcome measurements and statistical analysis The primary outcome objectives were progression-free survival, prostate cancer–specific survival (PCSS), and overall survival. Statistical analyses to assess survival and model performance were performed. Results and limitation Serum leucine-rich α-2-glycoprotein 1 (LRG1) was found to be elevated in fatal prostate cancer. LRG1 provided prognostic information independent of metastasis and increased the accuracy in predicting PCSS, particularly in the first 3 yr. A high LRG1 level is associated with an average of two-fold higher risk of disease-progression and mortality in both high-risk and metastatic patients. However, our study design, with a retrospective analysis of samples spanning several decades back, limits the assessment of the clinical utility of LRG1 in today’s clinical practice. Thus, independent prospective studies are needed to establish LRG1 as a clinically useful biomarker for patient management. Conclusions High blood levels of LRG1 are unfavourable in newly diagnosed high-risk and metastatic prostate cancer, and LRG1 increased the accuracy of risk stratification of prostate cancer patients. Patient summary High blood levels of leucine-rich α-2-glycoprotein 1 are unfavourable in newly diagnosed high-risk and metastatic prostate cancer.

• Kara, Emrah; Nielsen, Nis Valentin; Eggertsdottir, Bergrun; Thiede, Bernd; Kanse, Sandip & Løset, Geir Åge (2020). Design and characterization of a new pVII combinatorial phage display peptide library for protease substrate mining using Factor VII activating protease (FSAP) as model. ChemBioChem. ISSN 1439-4227. 21(13), p. 1875–1884. doi: 10.1002/cbic.201900705. Full text in Research Archive Show summary

  We describe a novel, easy and efficient combinatorial phage display peptide substrate‐mining method to map the substrate specificity of proteases. The peptide library is displayed on the pVII capsid of the M13 bacteriophage, which renders pIII necessary for infectivity and efficient retrieval, in an unmodified state. As capture module, the 3XFLAG was chosen due to its very high binding efficiency to anti‐FLAG mAbs and its independency of any post‐translational modification. This library was tested with Factor‐VII activating protease (WT‐FSAP) and its single‐nucleotide polymorphism variant Marburg‐I (MI)‐FSAP. The WT‐FSAP results confirmed the previously reported Arg/Lys centered FSAP cleavage site consensus as dominant, as well as reinforcing MI‐FSAP as a loss‐of‐function mutant. Surprisingly, rare substrate clones devoid of basic amino acids were also identified. Indeed one of these peptides was cleaved as free peptide, thus suggesting a broader range of WT‐FSAP substrates than previously anticipated.

• Krishnan, Aswini; Berthelet, Jean; Renaud, Emilie; Rosigkeit, Sebastian; Distler, Ute & Stawiski, Eric [Show all 17 contributors for this article] (2020). Proteogenomics analysis unveils a TFG-RET gene fusion and druggable targets in papillary thyroid carcinomas. Nature Communications. 11. doi: 10.1038/s41467-020-15955-w. Full text in Research Archive Show summary

  Papillary thyroid cancer (PTC) is the most common type of endocrine malignancy. By RNA-seq analysis, we identify a RET rearrangement in the tumour material of a patient who does not harbour any known RAS or BRAF mutations. This new gene fusion involves exons 1–4 from the 5′ end of the Trk fused Gene (TFG) fused to the 3′ end of RET tyrosine kinase leading to a TFG-RET fusion which transforms immortalized human thyroid cells in a kinase-dependent manner. TFG-RET oligomerises in a PB1 domain-dependent manner and oligomerisation of TFG-RET is required for oncogenic transformation. Quantitative proteomic analysis reveals the upregulation of E3 Ubiquitin ligase HUWE1 and DUBs like USP9X and UBP7 in both tumor and metastatic lesions, which is further confirmed in additional patients. Expression of TFG-RET leads to the upregulation of HUWE1 and inhibition of HUWE1 significantly reduces RET-mediated oncogenesis.

• Khan, Ayyad Ahmad Zartasht; Jackson, Catherine; Utheim, Tor Paaske; Reppe, Sjur; Sapkota, Dipak & Olstad, Ole Kristoffer [Show all 8 contributors for this article] (2020). Sericin-induced melanogenesis in cultured retinal pigment epithelial cells is associated with elevated levels of hydrogen peroxide and inflammatory proteins. Molecules. ISSN 1431-5157. 25(19), p. 1–20. doi: 10.3390/molecules25194395. Full text in Research Archive Show summary

  We previously demonstrated that the silk protein sericin promotes pigmentation of retinal pigment epithelium (RPE) by activating the NF-κB pathway. Among numerous agents, NF-κB can be activated by hydrogen peroxide. In the present study, we explored possible associations between reactive oxygen species and sericin-induced melanogenesis in RPE. The proteome of human fetal RPE cultured for seven days with or without 1% sericin was analyzed using ingenuity pathway analysis (IPA). The proteomic data was verified by immunofluorescence and immunoblotting. Light microscopy and scanning electron microscopy were used to assess morphology. Dihydroethidium (DHE) and dihydrorhodamine (DHR) assays were used to measure superoxide and hydrogen peroxide species. Expression levels of proteins related to inflammation, differentiation, cell survival and cell adhesion were higher in cells cultured in Dulbecco’s Modified Eagle Medium (DMEM) with 1% sericin, whereas cells cultured in DMEM alone showed higher expression levels of proteins associated with Bruch’s membrane and cytoskeleton. Despite upregulation of inflammatory proteins, sericin co-cultured RPE yielded significantly higher cell viability compared to cells cultured without sericin. Addition of sericin to culture media significantly increased hydrogen peroxide-levels without significantly affecting superoxide-levels. We suggest that sericin-induced melanogenesis in cultured RPE is associated with elevated levels of superoxide dismutase, hydrogen peroxide and inflammatory proteins.

• Flatin, Bjørnar Tovson Bae; Vedeld, Hege Marie; Pinto, Rita; Langerud, Jonas; Lind, Guro Elisabeth & Lothe, Ragnhild A [Show all 8 contributors for this article] (2020). Multiregional assessment of CIMP in primary colorectal cancers: Phenotype concordance but marker variability. International Journal of Cancer. ISSN 0020-7136. p. 1–6. doi: 10.1002/ijc.33425. Full text in Research Archive Show summary

  Intratumor heterogeneity of colorectal cancers (CRCs) is manifested both at the genomic and epigenomic levels. Early genetic aberrations in carcinogenesis are clonal and present throughout the tumors, but less is known about the heterogeneity of the epigenetic CpG island methylator phenotype (CIMP). CIMP characterizes a subgroup of CRCs thought to originate from specific precursor lesions, and it is defined by widespread DNA methylation within promoter regions. In this work, we investigated CIMP in two to four multiregional samples from 30 primary tumors (n = 86 samples) using the consensus Weisenberger gene panel (CACNA1G, IGF2, NEUROG1, RUNX3 and SOCS1). Twenty‐nine of 30 tumors (97%) showed concordant CIMP status in all samples, and percent methylated reference (PMR) values of all five markers had higher intertumor than intratumor variation (P value = 1.5e−09). However, a third of the CIMP+ tumors exhibited discrepancies in methylation status in at least one of the five gene markers. To conclude, CIMP status was consistent within primary CRCs, and it is likely a clonal phenotype. However, spatial discordances of the individual genes suggest that large‐scale analysis of multiregional samples could be of interest for identifying CIMP markers that are robust to intratumor heterogeneity.

• Jin, Yang & Saatcioglu, Fahri (2020). Targeting the unfolded protein response in hormone-regulated cancers. Trends in Cancer. ISSN 2405-8033. 6(2), p. 160–171. doi: 10.1016/j.trecan.2019.12.001. Full text in Research Archive
• Haabeth, Ole Audun; Hennig, Kjartan; Fauskanger, Marte; Løset, Geir Åge; Bogen, Bjarne & Tveita, Anders Aune (2020). CD4+ T-cell killing of multiple myeloma cells is mediated by resident bone marrow macrophages. Blood Advances. ISSN 2473-9529. 4(12), p. 2595–2605. doi: 10.1182/bloodadvances.2020001434. Full text in Research Archive Show summary

  Abstract CD4+ T cells may induce potent antitumor immune responses through interaction with antigen-presenting cells within the tumor microenvironment. Using a murine model of multiple myeloma, we demonstrated that adoptive transfer of idiotype-specific CD4+ T cells may elicit curative responses against established multifocal myeloma in bone marrow. This finding indicates that the myeloma bone marrow niche contains antigen-presenting cells that may be rendered tumoricidal. Given the complexity of the bone marrow microenvironment, the mechanistic basis of such immunotherapeutic responses is not known. Through a functional characterization of antitumor CD4+ T-cell responses within the bone marrow microenvironment, we found that killing of myeloma cells is orchestrated by a population of bone marrow–resident CD11b+F4/80+MHC-IIHigh macrophages that have taken up and present secreted myeloma protein. The present results demonstrate the potential of resident macrophages as powerful mediators of tumor killing within the bone marrow and provide a basis for novel therapeutic strategies against multiple myeloma and other malignancies that affect the bone marrow.

• Tadele, Dagim Shiferaw; Robertson, Joseph; Crispin, Richard; Herrera, Carmen; Chlubnová, Markéta & Piechaczyk, Laure Isabelle [Show all 14 contributors for this article] (2020). A cell competition-based small molecule screen identifies a novel compound that induces dual c-Myc depletion and p53 activation. Journal of Biological Chemistry. ISSN 0021-9258. 296. doi: 10.1074/jbc.RA120.015285. Full text in Research Archive Show summary

  Breakpoint Cluster Region-Abelson kinase (BCR–Abl) is a driver oncogene that causes chronic myeloid leukemia and a subset of acute lymphoid leukemias. Although tyrosine kinase inhibitors provide an effective treatment for these diseases, they generally do not kill leukemic stem cells (LSCs), the cancer-initiating cells that compete with normal hematopoietic stem cells for the bone marrow niche. New strategies to target cancers driven by BCR–Abl are therefore urgently needed. We performed a small molecule screen based on competition between isogenic untransformed cells and BCR–Abl-transformed cells and identified several compounds that selectively impair the fitness of BCR–Abl-transformed cells. Interestingly, systems-level analysis of one of these novel compounds, DJ34, revealed that it induced depletion of c-Myc and activation of p53. DJ34-mediated c-Myc depletion occurred in a wide range of tumor cell types, including lymphoma, lung, glioblastoma, breast cancer, and several forms of leukemia, with primary LSCs being particularly sensitive to DJ34. Further analyses revealed that DJ34 interferes with c-Myc synthesis at the level of transcription, and we provide data showing that DJ34 is a DNA intercalator and topoisomerase II inhibitor. Physiologically, DJ34 induced apoptosis, cell cycle arrest, and cell differentiation. Taken together, we have identified a novel compound that dually targets c-Myc and p53 in a wide variety of cancers, and with particularly strong activity against LSCs.

• Solhaug, Anita; Torgersen, Maria Lyngaas; Holme, Jørn Andreas; Wiik-Nielsen, Jannicke; Thiede, Bernd & Eriksen, Gunnar Sundstøl (2020). The Fusarium mycotoxin, 2-Amino-14,16-dimethyloctadecan-3-ol (AOD) induces vacuolization in HepG2 cells. Toxicology. ISSN 0300-483X. p. 433–434. doi: 10.1016/j.tox.2020.152405. Full text in Research Archive Show summary

  The mycotoxin 2-Amino-14,16-dimethyloctadecan-3-ol (AOD) has been isolated from cultures of the fungus Fusarium avenaceum, one of the most prevalent Fusarium species. AOD is an analogue of sphinganine and 1-deoxysphinganine, important intermediates in the de novo biosynthesis of cellular sphingolipids. Here we studied cellular effects of AOD using the human liver cell line HepG2 as a model system. AOD (10 μM) induced a transient accumulation of vacuoles in the cells. The effect was observed at non-cytotoxic concentrations and was not linked to cell death processes. Proteomic analyses indicated that protein degradation and/or vesicular transport may be a target for AOD. Further studies revealed that AOD had only minor effects on the initiation rate of macropinocytosis and autophagy. However, the AOD-induced vacuoles were lysosomal-associated membrane protein-1 (LAMP-1) positive, suggesting that they most likely originate from lysosomes or late endosomes. Accordingly, both endosomal and autophagic protein degradation were inhibited. Further studies revealed that treatment with concanamycin A or chloroquine completely blocked the AOD-induced vacuolization, suggesting that the vacuolization is dependent of acidic lysosomes. Overall, the results strongly suggest that the increased vacuolization is due to an accumulation of AOD in lysosomes or late endosomes thereby disturbing the later stages of the endolysosomal process.

• Perdreau-Dahl, Harmonie; Progida, Cinzia; Barfeld, Stefan Jörg; Guldsten, Hanne; Thiede, Bernd & Arntzen, Magnus Øverlie [Show all 10 contributors for this article] (2020). Sjögren syndrome/scleroderma autoantigen 1 is a direct Tankyrase binding partner in cancer cells. Communications Biology. 3(1). doi: 10.1038/s42003-020-0851-2. Full text in Research Archive
• Bern, Malin C.; Nilsen, Jeannette; Ferrarese, Mattia; Sand, Kine Marita Knudsen; Gjølberg, Torleif Tollefsrud & Lode, Heidrun Elisabeth [Show all 24 contributors for this article] (2020). An engineered human albumin enhances half-life and transmucosal delivery when fused to protein-based biologics. Science Translational Medicine. ISSN 1946-6234. 12(565), p. 1–13. doi: 10.1126/scitranslmed.abb0580. Full text in Research Archive Show summary

  Needle-free uptake across mucosal barriers is a preferred route for delivery of biologics, but the efficiency of unassisted transmucosal transport is poor. To make administration and therapy efficient and convenient, strategies for the delivery of biologics must enhance both transcellular delivery and plasma half-life. We found that human albumin was transcytosed efficiently across polarized human epithelial cells by a mechanism that depends on the neonatal Fc receptor (FcRn). FcRn also transported immunoglobulin G, but twofold less than albumin. We therefore designed a human albumin variant, E505Q/T527M/K573P (QMP), with improved FcRn binding, resulting in enhanced transcellular transport upon intranasal delivery and extended plasma half-life of albumin in transgenic mice expressing human FcRn. When QMP was fused to recombinant activated coagulation factor VII, the half-life of the fusion molecule increased 3.6-fold compared with the wild-type human albumin fusion, without compromising the therapeutic properties of activated factor VII. Our findings highlight QMP as a suitable carrier of protein-based biologics that may enhance plasma half-life and delivery across mucosal barriers.

• Næs, Guro; Storesund, Jan Ove; Udayakumar, Priyanga-Dina; Ledsaak, Marit & Gabrielsen, Odd Stokke (2020). Dissecting the transactivation domain (tAD) of the transcription factor c‐Myb to assess recent models of tAD function. FEBS Open Bio. 10(11), p. 2329–2342. doi: 10.1002/2211-5463.12978. Full text in Research Archive Show summary

  Transcription factors use a DNA‐binding domain to localize their action and a transactivation domain (tAD) to stimulate activation of the associated gene. Recent work has renewed interest in how tADs activate genes, which remains poorly understood. Key features in the new models are exposure of short linear motifs (SLMs) and liquid–liquid phase separation (LLPS). Inspired by the new models for tAD function, we decided to revisit the tAD of the haematopoietic transcription factor c‐Myb by performing a mutational analysis to see how these new models fit and potentially explain the tAD behaviour of this master regulator. We know that c‐Myb has an acidic tAD, which contains a well‐characterized SLM in the form of a LxxLL motif. By testing 12 alanine‐scanning mutants and three mutants with major reorganization of its tAD in two mammalian reporter systems, we found a pattern of effects very close to what would be expected from the SLM‐exposure model, with strong effects exerted by both acidic replacements and SLM mutation. When the same mutants were tested in a yeast system, the pattern of effects was dramatically different, with the SLM mutation exerting no effect, and tAD behaviour was much less affected by small alterations, as would be expected from a LLPS model. These observations are discussed in the light of the two new tAD models, and a two‐step hypothesis for transactivation, combining both models, is proposed.

• Fan, Qiong; Nørgaard, Rikke Christine; Grytten, Ivar; Ness, Cecilie; Lucas, Christin & Vekterud, Kristin [Show all 15 contributors for this article] (2020). LXRα Regulates ChREBPα Transactivity in a Target Gene-Specific Manner through an Agonist-Modulated LBD-LID Interaction. Cells. 9(5), p. 1–26. doi: 10.3390/cells9051214. Full text in Research Archive Show summary

  The cholesterol-sensing nuclear receptor liver X receptor (LXR) and the glucose-sensing transcription factor carbohydrate responsive element-binding protein (ChREBP) are central players in regulating glucose and lipid metabolism in the liver. More knowledge of their mechanistic interplay is needed to understand their role in pathological conditions like fatty liver disease and insulin resistance. In the current study, LXR and ChREBP co-occupancy was examined by analyzing ChIP-seq datasets from mice livers. LXR and ChREBP interaction was determined by Co-immunoprecipitation (CoIP) and their transactivity was assessed by real-time quantitative polymerase chain reaction (qPCR) of target genes and gene reporter assays. Chromatin binding capacity was determined by ChIP-qPCR assays. Our data show that LXRα and ChREBPα interact physically and show a high co-occupancy at regulatory regions in the mouse genome. LXRα co-activates ChREBPα and regulates ChREBP-specific target genes in vitro and in vivo. This co-activation is dependent on functional recognition elements for ChREBP but not for LXR, indicating that ChREBPα recruits LXRα to chromatin in trans. The two factors interact via their key activation domains; the low glucose inhibitory domain (LID) of ChREBPα and the ligand-binding domain (LBD) of LXRα. While unliganded LXRα co-activates ChREBPα, ligand-bound LXRα surprisingly represses ChREBPα activity on ChREBP-specific target genes. Mechanistically, this is due to a destabilized LXRα:ChREBPα interaction, leading to reduced ChREBP-binding to chromatin and restricted activation of glycolytic and lipogenic target genes. This ligand-driven molecular switch highlights an unappreciated role of LXRα in responding to nutritional cues that was overlooked due to LXR lipogenesis-promoting function.

• Hammerstad, Marta; Kjendseth, Åsmund Røhr & Hersleth, Hans-Petter (2019). A Research-Inspired Biochemistry Laboratory Module - Combining Expression, Purification, Crystallization, Structure-Solving, and Characterization of a Flavodoxin-like Protein. Biochemistry and Molecular Biology Education. ISSN 1470-8175. 47(3), p. 318–332. doi: 10.1002/bmb.21218. Full text in Research Archive Show summary

  Many laboratory courses consist of short and seemingly unconnected individual laboratory exercises. To increase the course consistency, relevance, and student engagement, we havedevelopedaresearch-inspiredandproject-basedmodule, “From Gene to Structure and Function”. This 2.5-week full-day biochemistry and structural biology module covers protein expression, purification, structure solving, and characterization.Themoduleiscenteredaroundthe flavodoxin-likeprotein NrdI, involved in the activation of the bacterial ribonucleotide reductase enzyme system. Through an in-depth focus on one specific protein, the students will learn the basic laboratory skills needed in order to generate a broader knowledge and breadth within the field. With respect to generic skills, the studentsreporttheir findingsasascientificarticle,withtheaim to learn to present concise research results and write scientific papers. The current research-inspired project has the potential ofbeingfurtherdevelopedintoamorediscovery-drivenproject and extended to include other molecular biological techniques orbiochemical/biophysicalcharacterizations.Instudentevaluations, this research-inspired laboratory course has received very high ratings and been highly appreciated, where the students have gained research experience for more independent future work in the laboratory. © 2019 The Authors. Biochemistry and Molecular Biology Education published by Wiley Periodicals, Inc. on behalf of International Union of Biochemistry andMolecularBiology,47(3):318–332,2019.

• Aqrawi, Lara Adnan; Galtung, Hilde; Guerreiro, Eduarda; Øvstebø, Reidun; Thiede, Bernd & Utheim, Tor Paaske [Show all 11 contributors for this article] (2019). Proteomic and histopathological characterisation of sicca subjects and primary Sjögren’s syndrome patients reveals promising tear, saliva and extracellular vesicle disease biomarkers. Arthritis Research & Therapy. ISSN 1478-6354. 21, p. 1–14. doi: 10.1186/s13075-019-1961-4. Full text in Research Archive Show summary

  Background Mononuclear cell infiltration of exocrine glands, production of Ro/SSA and La/SSB autoantibodies, along with oral and ocular dryness, are characteristic features of primary Sjögren’s syndrome (pSS). Non-SS sicca subjects, an underexplored group in relation to pSS, display similar sicca symptoms, with possible mild signs of inflammation in their salivary glands, yet with no serological detection of autoantibody production. In this study, we investigated inflammatory manifestations in the salivary gland tissue, tear fluid and saliva of non-SS subjects, as compared to pSS patients and healthy individuals. Methods Fifteen non-SS, 10 pSS and 10 healthy subjects were included in the analyses. Histological evaluation of salivary gland biopsies was performed. Liquid chromatography-mass spectrometry (LC-MS) was conducted on tear fluid and stimulated whole saliva, and proteomic biomarker profiles were generated. Extracellular vesicle (EVs) isolation and characterisation from both fluids were also combined with LC-MS. The LC-MS data were analysed for quantitative differences between patient and control groups using Scaffold. Database for Annotation, Visualization and Integrated Discovery (DAVID) and Functional Enrichment Analysis Tool (FunRich) were applied for functional analyses. Results Histopathological evaluation of salivary gland biopsies showed implications of milder inflammation in non-SS subjects through mononuclear cell infiltration, fibrosis and fatty replacement, as compared to pSS patients. Although unaffected in the non-SS group, upregulation of proinflammatory pathways and proteins involved in ubiquitination (LMO7 and HUWE1) and B cell differentiation (TPD52) were detected in tear fluid of pSS patients. Moreover, overexpression of proteins STOM, ANXA4 and ANXA1, regulating cellular innate and adaptive immunological pathways, were further identified in EVs from tear fluid of pSS patients. Finally, whole saliva and EVs isolated from whole saliva of pSS patients expressed proteins vital for innate MHC class I cellular regulation (NGAL) and T cell activation (CD44). Conclusions Non-SS sicca subjects may show implications of mild inflammation in their glandular tissue, while their protein profile was strikingly more similar to healthy controls than to pSS patients. Hence, the tear and salivary biomarkers identified could be implemented as potential non-invasive diagnostic tools that may aid in increasing diagnostic accuracy when evaluating non-SS subjects and pSS patients and monitoring disease progression.

• Malecki, Jedrzej Mieczyslaw; Willemen, Hanneke L.D.M.; Pinto, Rita; Ho, Angela Yeuan Yen; Moen, Anders & Eijkelkamp, Niels [Show all 7 contributors for this article] (2019). Human FAM173A is a mitochondrial lysine-specific methyltransferase that targets adenine nucleotide translocase and affects mitochondrial respiration. Journal of Biological Chemistry. ISSN 0021-9258. 294(31), p. 11654–11664. doi: 10.1074/jbc.RA119.009045. Full text in Research Archive
• Wagner, Fabienne; Kunz, Tobias C.; Chowdhury, Suvagata R.; Thiede, Bernd; Fraunholz, Martin & Eger, Debora [Show all 7 contributors for this article] (2019). Armadillo repeat-containing protein 1 is a dual localization protein associated with mitochondrial intermembrane space bridging complex. PLOS ONE. 14(10), p. 1–19. doi: 10.1371/journal.pone.0218303. Full text in Research Archive Show summary

  Cristae architecture is important for the function of mitochondria, the organelles that play the central role in many cellular processes. The mitochondrial contact site and cristae organizing system (MICOS) together with the sorting and assembly machinery (SAM) forms the mitochondrial intermembrane space bridging complex (MIB), a large protein complex present in mammalian mitochondria that partakes in the formation and maintenance of cristae. We report here a new subunit of the mammalian MICOS/MIB complex, an armadillo repeat-containing protein 1 (ArmC1). ArmC1 localizes both to cytosol and mitochondria, where it associates with the outer mitochondrial membrane through its carboxy-terminus. ArmC1 interacts with other constituents of the MICOS/MIB complex and its amounts are reduced upon MICOS/MIB complex depletion. Mitochondria lacking ArmC1 do not show defects in cristae structure, respiration or protein content, but appear fragmented and with reduced motility. ArmC1 represents therefore a peripheral MICOS/MIB component that appears to play a role in mitochondrial distribution in the cell.

• Zadraznik, Tanja; Moen, Anders & Sustar-Vozlic, Jelka (2019). Chloroplast proteins involved in drought stress response in selected cultivars of common bean (Phaseolus vulgaris L.). 3 Biotech. ISSN 2190-572X. 9(9), p. 1–15. doi: 10.1007/s13205-019-1862-x. Full text in Research Archive
• Riegel, Kristina; Schlöder, Janine; Sobczak, Marco; Jonuleit, Helmut; Thiede, Bernd & Schild, Hansjörg [Show all 7 contributors for this article] (2019). RAF kinases are stabilized and required for dendritic cell differentiation and function. Cell Death and Differentiation. ISSN 1350-9047. 27, p. 1300–1315. doi: 10.1038/s41418-019-0416-4. Full text in Research Archive Show summary

  RAF kinases (ARAF, BRAF, and CRAF) are highly conserved enzymes that trigger the RAF-MEK1/2-ERK1/2 (MAPK) pathway upon activation of RAS. Despite enormous clinical interest, relatively little is known on the role of RAFs in mediating immune responses. Here, we investigated the role of RAF kinases and MEK1/2 in dendritic cells (DCs), the central regulators of T cell-mediated antitumor immune responses and the adaptive immune system. We demonstrate that RAF kinases are active and stabilized at their protein levels during DC differentiation. Inhibition of RAF kinases but not MEK1/2 impaired the activation of DCs in both mice and human. As expected, DCs treated with RAF inhibitors show defects in activating T cells. Further, RAF and MEK1/2 kinases are directly required for the activation and proliferation of CD4+ T cells. Our observations suggest that RAF and MEK1/2 have independent roles in regulating DC function that has important implications for administering RAF–MAPK inhibitors in the clinics.

• Itkonen, Harri; Urbanucci, Alfonso; ES, Martin Sara; Khan, Aziz; Mathelier, Anthony & Thiede, Bernd [Show all 8 contributors for this article] (2019). High OGT activity is essential for MYC-driven proliferation of prostate cancer cells. Theranostics. 9(8), p. 2183–2197. doi: 10.7150/thno.30834. Full text in Research Archive Show summary

  O-GlcNAc transferase (OGT) is overexpressed in aggressive prostate cancer. OGT modifies intra-cellular proteins via single sugar conjugation (O-GlcNAcylation) to alter their activity. We recently discovered the first fast-acting OGT inhibitor OSMI-2. Here, we probe the stability and function of the chromatin O-GlcNAc and identify transcription factors that coordinate with OGT to promote proliferation of prostate cancer cells. Methods: Chromatin immunoprecipitation (ChIP) coupled to sequencing (seq), formaldehyde-assisted isolation of regulatory elements, RNA-seq and reverse-phase protein arrays (RPPA) were used to study the importance of OGT for chromatin structure and transcription. Mass spectrometry, western blot, RT-qPCR, cell cycle analysis and viability assays were used to establish the role of OGT for MYC-related processes. Prostate cancer patient data profiled for both mRNA and protein levels were used to validate findings. Results: We show for the first time that OGT inhibition leads to a rapid loss of O-GlcNAc chromatin mark. O-GlcNAc ChIP-seq regions overlap with super-enhancers (SE) and MYC binding sites. OGT inhibition leads to down-regulation of SE-dependent genes. We establish the first O-GlcNAc chromatin consensus motif, which we use as a bait for mass spectrometry. By combining the proteomic data from oligonucleotide enrichment with O-GlcNAc and MYC ChIP-mass spectrometry, we identify host cell factor 1 (HCF-1) as an interaction partner of MYC. Inhibition of OGT disrupts this interaction and compromises MYC's ability to confer androgen-independent proliferation to prostate cancer cells. We show that OGT is required for MYC-mediated stabilization of mitotic proteins, including Cyclin B1, and/or the increased translation of their coding transcripts. This implies that increased expression of mRNA is not always required to achieve increased protein expression and confer aggressive phenotype. Indeed, high expression of Cyclin B1 protein has strong predictive value in prostate cancer patients (p=0.000014) while mRNA does not. Conclusions: OGT promotes SE-dependent gene expression. OGT activity is required for the interaction between MYC and HCF-1 and expression of MYC-regulated mitotic proteins. These features render OGT essential for the androgen-independent, MYC-driven proliferation of prostate cancer cells. Androgen-independency is the major mechanism of prostate cancer progression, and our study identifies OGT as an essential mediator in this process.

• Thakur, Avinash; Wong, Jasper C.H.; Wang, Evan Y.; Lotto, Jeremy; Kim, Donghwan & Cheng, Jung-Chien [Show all 20 contributors for this article] (2019). Hepatocyte nuclear factor 4-alpha is essential for the active epigenetic state at enhancers in mouse liver. Hepatology. ISSN 0270-9139. 70(4), p. 1360–1376. doi: 10.1002/hep.30631. Full text in Research Archive
• Pällmann, Nora; Livgård, Marte; Tesikova, Martina; Nenseth, Hatice Zeynep; Akkus, Erman & Sikkeland, Jørgen [Show all 20 contributors for this article] (2019). Regulation of the unfolded protein response through ATF4 and FAM129A in prostate cancer. Oncogene. ISSN 0950-9232. 38, p. 6301–6318. doi: 10.1038/s41388-019-0879-2. Full text in Research Archive Show summary

  Cancer cells exploit many of the cellular adaptive responses to support their survival needs. One such critical pathway in eukaryotic cells is the unfolded protein response (UPR) that is important in normal physiology as well as disease states, including cancer. Since UPR can serve as a lever between survival and death, regulated control of its activity is critical for tumor formation and growth although the underlying mechanisms are poorly understood. Here we show that one of the main transcriptional effectors of UPR, activating transcription factor 4 (ATF4), is essential for prostate cancer (PCa) growth and survival. Using systemic unbiased gene expression and proteomic analyses, we identified a novel direct ATF4 target gene, family with sequence similarity 129 member A (FAM129A), which is critical in mediating ATF4 effects on prostate tumorigenesis. Interestingly, FAM129A regulated both PERK and eIF2α in a feedback loop that differentially channeled the UPR output. ATF4 and FAM129A protein expression is increased in patient PCa samples compared with benign prostate. Importantly, in vivo therapeutic silencing of ATF4-FAM129A axis profoundly inhibited tumor growth in a preclinical PCa model. These data support that one of the canonical UPR branches, through ATF4 and its target gene FAM129A, is required for PCa growth and thus may serve as a novel therapeutic target.

• Zaucker, Andreas; Kumari, Pooja & Sampath, Karuna (2019). Zebrafish embryogenesis – A framework to study regulatory RNA elements in development and disease. Developmental Biology. ISSN 0012-1606. 457, p. 172–180. doi: 10.1016/j.ydbio.2019.01.008. Full text in Research Archive
• Abudu, Yakubu Princely; Pankiv, Serhiy; Mathai, Benan John; Lystad, Alf Håkon; Bindesbøll, Christian & Brenne, Hanne Britt [Show all 14 contributors for this article] (2019). NIPSNAP1 and NIPSNAP2 act as “eat-me signal” for mitophagy. Developmental Cell. ISSN 1534-5807. 49(4), p. 509–525.e12. doi: 10.1016/j.devcel.2019.03.013. Full text in Research Archive Show summary

  The clearance of damaged or dysfunctional mitochondria by selective autophagy (mitophagy) is important for cellular homeostasis and prevention of disease. Our understanding of the mitochondrial signals that trigger their recognition and targeting by mitophagy is limited. Here, we show that the mitochondrial matrix proteins 4-Nitrophenylphosphatase domain and non-neuronal SNAP25-like protein homolog 1 (NIPSNAP1) and NIPSNAP2 accumulate on the mitochondria surface upon mitochondrial depolarization. There, they recruit proteins involved in selective autophagy, including autophagy receptors and ATG8 proteins, thereby functioning as an “eat me” signal for mitophagy. NIPSNAP1 and NIPSNAP2 have a redundant function in mitophagy and are predominantly expressed in different tissues. Zebrafish lacking a functional Nipsnap1 display reduced mitophagy in the brain and parkinsonian phenotypes, including loss of tyrosine hydroxylase (Th1)-positive dopaminergic (DA) neurons, reduced motor activity, and increased oxidative stress.

• Pyzik, Michal; Sand, Kine Marita Knudsen; Hubbard, Jonathan J.; Andersen, Jan Terje; Sandlie, Inger & Blumberg, Richard S. (2019). The neonatal Fc Receptor (FcRn): A misnomer? Frontiers in Immunology. 10:1540(JULY), p. 1–24. doi: 10.3389/fimmu.2019.01540. Full text in Research Archive Show summary

  Antibodies are essential components of an adaptive immune response. Immunoglobulin G (IgG) is the most common type of antibody found in circulation and extracellular fluids. Although IgG alone can directly protect the body from infection through the activities of its antigen binding region, the majority of IgG immune functions are mediated via proteins and receptors expressed by specialized cell subsets that bind to the fragment crystallizable (Fc) region of IgG. Fc gamma (γ) receptors (FcγR) belong to a broad family of proteins that presently include classical membrane-bound surface receptors as well as atypical intracellular receptors and cytoplasmic glycoproteins. Among the atypical FcγRs, the neonatal Fc receptor (FcRn) has increasingly gained notoriety given its intimate influence on IgG biology and its ability to also bind to albumin. FcRn functions as a recycling or transcytosis receptor that is responsible for maintaining IgG and albumin in the circulation, and bidirectionally transporting these two ligands across polarized cellular barriers. More recently, it has been appreciated that FcRn acts as an immune receptor by interacting with and facilitating antigen presentation of peptides derived from IgG immune complexes (IC). Here we review FcRn biology and focus on newer advances including how emerging FcRn-targeted therapies may affect the immune responses to IgG and IgG IC.

• Sikkeland, Jørgen; Lindstad, Torstein; Nenseth, Hatice Zeynep; Dezitter, Xavier Gabriel M; Qu, Su & Muhumed, Ridhwan Mohamed [Show all 9 contributors for this article] (2019). Inflammation and ER stress differentially regulate STAMP2 expression and localization in adipocytes. Metabolism: Clinical and Experimental. ISSN 0026-0495. 93, p. 75–85. doi: 10.1016/j.metabol.2019.01.014. Full text in Research Archive Show summary

  Background Chronic ER stress and dysfunction is a hallmark of obesity and a critical contributor to metaflammation, abnormal hormone action and altered substrate metabolism in metabolic tissues, such as liver and adipocytes. Lack of STAMP2 in lean mice induces inflammation and insulin resistance on a regular diet, and it is dysregulated in the adipose tissue of obese mice and humans. We hypothesized that the regulation of STAMP2 is disrupted by ER stress. Methods 3T3-L1 and MEF adipocytes were treated with ER stress inducers thapsigargin and tunicamycin, and inflammation inducer TNFα. The treatments effect on STAMP2 expression and enzymatic function was assessed. In addition, 3T3-L1 adipocytes and HEK cells were utilized for Stamp2 promoter activity investigation performed with luciferase and ChIP assays. Results ER stress significantly reduced both STAMP2 mRNA and protein expression in cultured adipocytes whereas TNFα had the opposite effect. Concomitant with loss of STAMP2 expression during ER stress, intracellular localization of STAMP2 was altered and total iron reductase activity was reduced. Stamp2 promoter analysis by reporter assays and chromatin immunoprecipitation, showed that induction of ER stress disrupts C/EBPα-mediated STAMP2 expression. Conclusion These data suggest a clear link between ER stress and quantitative and functional STAMP2-deficiency.

• Eerde, André van; Varnai, Aniko; Jameson, John-Kristian; Paruch, Lisa; Moen, Anders & Anonsen, Jan Haug [Show all 12 contributors for this article] (2019). In-depth characterization of Trichoderma reesei cellobiohydrolase TrCel7A produced in Nicotiana benthamiana reveals limitations of cellulase production in plants by host-specific post-translational modifications. Plant Biotechnology Journal. ISSN 1467-7644. 18(3), p. 631–643. doi: 10.1111/pbi.13227. Full text in Research Archive Show summary

  Sustainable production of biofuels from lignocellulose feedstocks depends on cheap enzymes for degradation of such biomass. Plants offer a safe and cost‐effective production platform for biopharmaceuticals, vaccines and industrial enzymes boosting biomass conversion to biofuels. Production of intact and functional protein is a prerequisite for large‐scale protein production, and extensive host‐specific post‐translational modifications (PTMs) often affect the catalytic properties and stability of recombinant enzymes. Here we investigated the impact of plant PTMs on enzyme performance and stability of the major cellobiohydrolase TrCel7A from Trichoderma reesei, an industrially relevant enzyme. TrCel7A was produced in Nicotiana benthamiana using a vacuum‐based transient expression technology, and this recombinant enzyme (TrCel7Arec) was compared with the native fungal enzyme (TrCel7Anat) in terms of PTMs and catalytic activity on commercial and industrial substrates. We show that the N‐terminal glutamate of TrCel7Arec was correctly processed by N. benthamiana to a pyroglutamate, critical for protein structure, while the linker region of TrCel7Arec was vulnerable to proteolytic digestion during protein production due to the absence of O‐mannosylation in the plant host as compared with the native protein. In general, the purified full‐length TrCel7Arec had 25% lower catalytic activity than TrCel7Anat and impaired substrate‐binding properties, which can be attributed to larger N‐glycans and lack of O‐glycans in TrCel7Arec. All in all, our study reveals that the glycosylation machinery of N. benthamiana needs tailoring to optimize the production of efficient cellulases.

• Sheng, Xia; Nenseth, Hatice Zeynep; Qu, Su; Kuzu, Ömer Faruk; Frahnow, Turid & Simon, Lukas [Show all 16 contributors for this article] (2019). IRE1α-XBP1s pathway promotes prostate cancer by activating c-MYC signaling. Nature Communications. 10:323, p. 1–12. doi: 10.1038/s41467-018-08152-3. Full text in Research Archive Show summary

  Activation of endoplasmic reticulum (ER) stress/the unfolded protein response (UPR) has been linked to cancer, but the molecular mechanisms are poorly understood and there is a paucity of reagents to translate this for cancer therapy. Here, we report that an IRE1α RNase-specific inhibitor, MKC8866, strongly inhibits prostate cancer (PCa) tumor growth as monotherapy in multiple preclinical models in mice and shows synergistic antitumor effects with current PCa drugs. Interestingly, global transcriptomic analysis reveal that IRE1α-XBP1s pathway activity is required for c-MYC signaling, one of the most highly activated oncogenic pathways in PCa. XBP1s is necessary for optimal c-MYC mRNA and protein expression, establishing, for the first time, a direct link between UPR and oncogene activation. In addition, an XBP1-specific gene expression signature is strongly associated with PCa prognosis. Our data establish IRE1α-XBP1s signaling as a central pathway in PCa and indicate that its targeting may offer novel treatment strategies.

• Fan, Qiong; Nørgaard, Rikke Christine; Grytten, Ivar; Ness, Cecilie Maria; Lucas, Christin & Vekterud, Kristin [Show all 15 contributors for this article] (2019). Open the LID: LXRα regulates ChREBPα transactivity in a target gene-specific manner through an agonist-modulated LBD-LID interaction. BioRxiv. ISSN 0362-4331. doi: 10.1101/2019.12.20.869974. Full text in Research Archive Show summary

  The cholesterol-sensing nuclear receptor liver X receptor (LXR) and the glucose-sensing transcription factor carbohydrate responsive element-binding protein (ChREBP) are central players in regulating glucose and lipid metabolism in liver. We have previously shown that LXR regulates ChREBP transcription and activity; however, the underlying mechanisms are unclear. In the current study, we demonstrate that LXRα and ChREBPα interact physically, and show a high co-occupancy at regulatory regions in the mouse genome. LXRα co-activates ChREBPα, and regulates ChREBP-specific target genes in vitro and in vivo. This co-activation is dependent on functional recognition elements for ChREBP, but not for LXR, indicating that ChREBPα recruits LXRα to chromatin in trans. The two factors interact via their key activation domains; ChREBPα’s low glucose inhibitory domain (LID) and the ligand-binding domain (LBD) of LXRα. While unliganded LXRα co-activates ChREBPα, ligand-bound LXRα surprisingly represses ChREBPα activity on ChREBP-specific target genes. Mechanistically, this is due to a destabilized LXRα:ChREBPα interaction, leading to reduced ChREBP-binding to chromatin and restricted activation of glycolytic and lipogenic target genes. This ligand-driven molecular switch highlights an unappreciated role of LXRα that was overlooked due to LXR lipogenesis-promoting function.

• Shi, Jihua; Guo, Wen-Zhi; Jin, Yang; Zhang, Hua-Peng; Pang, Chun & Li, Jie [Show all 8 contributors for this article] (2019). Recognition of HER2 expression in hepatocellular carcinoma and its significance in postoperative tumor recurrence. Cancer Medicine. 8(3), p. 1269–1278. doi: 10.1002/cam4.2006. Full text in Research Archive Show summary

  Background The ERBB2 oncogene hypothesis is challenged in hepatocellular carcinoma (HCC) with the conflicting evidences of human epidermal growth factor receptor 2 (HER2) overexpression. HER2 could be a new target as a treatment option for HCC as well as tumor recurrence after surgery. HER2 in HCC biology needs further explorations. Methods Clinical and mRNA data of HCC patients were obtained from TCGA HCC cohort, GSE89377 and GSE115018. Western Blotting and immunohistochemistry were employed to test expression of HER2, E‐cadherin, and Vimentin. In HepG2, JM1, HER2‐transfected McA cells, and TGF‐β cocultured JM1 cells, HCC biology, including cell survival, proliferation, and epithelial‐to‐mesenchymal transition (EMT) phenotypes were evaluated. Results ERBB2 mRNA amplification was found in HCC datasets, and its expression was downregulated in high grade HCC with a worse overall survival. HER2 overexpression was identified in H4IIE, HepG2, JM1 cells, and 82% (14/17) HCC samples, and tumor stage was correlated with expression of HER2, E‐cadherin, and Vimentin (P < 0.05). Trastuzumab with the high concentrations suppressed proliferation of HER2‐positive hepatoma cells (P < 0.05); in the coculture model to induce EMT of JM1 cells, HER2 expression increased with downregulated E‐cadherin and upregulated Vimentin. Trastuzumab intravenous injection inhibited in vivo tumor size and metastases (P < 0.05). Signal analysis revealed that HER2 functioned through upregulation of β‐catenin and inhibition of SMAD3. Conclusion HER2 expression pattern is linked with tumor stage and overall survival; the transforming function of HER2 is found more relevant through β‐catenin and SMAD3. HER2‐targeted treatment is recommended to suppress the HER2‐mediated tumor growth during postoperative liver regeneration.

• Koehler, Christian & Thiede, Bernd (2019). Predominant cleavage of proteins N-terminal to serines and threonines using scandium(III) triflate. JBIC Journal of Biological Inorganic Chemistry. ISSN 0949-8257. 25, p. 61–66. doi: 10.1007/s00775-019-01733-7. Full text in Research Archive Show summary

  Proteolytic digestion prior to LC–MS analysis is a key step for the identification of proteins. Digestion of proteins is typically performed with trypsin, but certain proteins or important protein sequence regions might be missed using this endoproteinase. Only few alternative endoproteinases are available and chemical cleavage of proteins is rarely used. Recently, it has been reported that some metal complexes can act as artificial proteases. In particular, the Lewis acid scandium(III) triflate has been shown to catalyze the cleavage of peptide bonds to serine and threonine residues. Therefore, we investigated if this compound can also be used for the cleavage of proteins. For this purpose, several single proteins, the 20S immune-proteasome (17 proteins), and the Universal Proteomics Standard UPS1 (48 proteins) were analyzed by MALDI–MS and/or LC–MS. A high cleavage specificity N-terminal to serine and threonine residues was observed, but also additional peptides with deviating cleavage specificity were found. Scandium(III) triflate can be a useful tool in protein analysis as no other reagent has been reported yet which showed cleavage specificity within proteins to serines and threonines.

• P, Aurélie Nguéa; Robertson, Joseph; Herrera, Maria Carmen; Chymkowitch, Pierre & Enserink, Jorrit (2019). Desumoylation of RNA polymerase III lies at the core of the Sumo stress response in yeast. Journal of Biological Chemistry. ISSN 0021-9258. 294(49), p. 18784–18795. doi: 10.1074/jbc.RA119.009721. Full text in Research Archive Show summary

  Post-translational modification by small ubiquitin-like modifier (Sumo) regulates many cellular processes, including the adaptive response to various types of stress, referred to as the Sumo stress response (SSR). However, it remains unclear whether the SSR involves a common set of core proteins regardless of the type of stress or whether each particular type of stress induces a stress-specific SSR that targets a unique, largely nonoverlapping set of Sumo substrates. In this study, we used MS and a Gene Ontology approach to identify differentially sumoylated proteins during heat stress, hyperosmotic stress, oxidative stress, nitrogen starvation, and DNA alkylation in Saccharomyces cerevisiae cells. Our results indicate that each stress triggers a specific SSR signature centered on proteins involved in transcription, translation, and chromatin regulation. Strikingly, whereas the various stress-specific SSRs were largely nonoverlapping, all types of stress tested here resulted in desumoylation of subunits of RNA polymerase III, which correlated with a decrease in tRNA synthesis. We conclude that desumoylation and subsequent inhibition of RNA polymerase III constitutes the core of all stress-specific SSRs in yeast.

• Bottermann, Maria; Foss, Stian; Caddy, Sarah L.; Clift, Dean; Tienen, Laurens M. van & Vaysburd, Marina [Show all 16 contributors for this article] (2019). Complement C4 prevents viral infection through capsid inactivation. Cell Host and Microbe. ISSN 1931-3128. 25(4), p. 617–629. doi: 10.1016/j.chom.2019.02.016. Full text in Research Archive Show summary

  The complement system is vital for anti-microbial defense. In the classical pathway, pathogen-bound antibody recruits the C1 complex (C1qC1r2C1s2) that initiates a cleavage cascade involving C2, C3, C4, and C5 and triggering microbial clearance. We demonstrate a C4-dependent antiviral mechanism that is independent of downstream complement components. C4 inhibits human adenovirus infection by directly inactivating the virus capsid. Rapid C4 activation and capsid deposition of cleaved C4b are catalyzed by antibodies via the classical pathway. Capsid-deposited C4b neutralizes infection independent of C2 and C3 but requires C1q antibody engagement. C4b inhibits capsid disassembly, preventing endosomal escape and cytosolic access. C4-deficient mice exhibit heightened viral burdens. Additionally, complement synergizes with the Fc receptor TRIM21 to block transduction by an adenovirus gene therapy vector but is partially restored by Fab virus shielding. These results suggest that the complement system could be altered to prevent virus infection and enhance virus gene therapy efficacy.

• Mousavi, Seyed Ali; Skjeldal, Frode Miltzow; Fønhus, Marita Sporstøl; Haugen, Linda Hofstad; Eskild, Winnie & Berg, Trond [Show all 7 contributors for this article] (2019). Receptor-mediated endocytosis of VEGF-A in rat liver sinusoidal endothelial cells. BioMed Research International. ISSN 2314-6133. 2019, p. 1–12. doi: 10.1155/2019/5496197. Full text in Research Archive Show summary

  Background and Aims. Vascular endothelial growth factor (VEGF) receptors (VEGFR1 and VEGFR2) bind VEGF-A with high affinity. This study sought to determine the relative contributions of these two receptors to receptor-mediated endocytosis of VEGF-A and to clarify their endocytic itineraries in rat liver sinusoidal endothelial cells (LSECs). Methods. Isolated LSECs and radiolabeled VEGF-A were used to examine surface binding and receptor-mediated endocytosis. Quantitative real time RT-PCR (Q-RT-PCR) and Western blotting were applied to demonstrate receptor expression. Results. Q-RT-PCR analysis showed that VEGFR1 and VEGFR2 mRNA were expressed in LSECs. Ligand saturation analysis at 4°C indicated two different classes of [125I]-VEGFA binding sites on LSECs with apparent dissociation constants of 8 and 210 pM. At 37°C, LSECs efficiently took up and degraded [125I]-VEGF-A for at least 2 hours. Uptake of [125I]-VEGF-A by LSECs was blocked by dynasore that inhibits dynamin-dependent internalization, whereas inhibition of cysteine proteases by leupeptin inhibited degradation without affecting the uptake of [125I]-VEGF-A, suggesting that it is degraded following transport to lysosomes. Incubation of LSECs in the continued presence of a saturating concentration of unlabeled VEGF-A at 37°C was associated with a loss of as much as 75% of the total VEGFR2 within 30 min as shown by Western blot analysis, whereas there was no appreciable decrease in protein levels for VEGFR1 after 120 min incubation, suggesting that VEGF-A stimulation downregulates VEGFR2, but not VEGFR1, in LSECs. This possibility was supported by the observation that a hexapeptide that specifically blocks VEGF-A binding to VEGFR1 caused a marked reduction in the uptake of [125I]-VEGF-A, whereas a control peptide had no effect. Finally, live cell imaging studies using a fluorescently labeled anti-VEGFR2 antibody showed that VEGFR2 was transported via early and late endosomes to reach endolysosomes where degradation of the VEGFR2 takes place. Conclusion. Our studies suggest that, subsequent to VEGF-A binding and internalization, the unoccupied VEGFR1 may recycle to the cell surface allowing its reutilization, whereas the majority of the internalized VEGFR2 is targeted for degradation.

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• Rio, Oline Eriksen; Labun, Kornel; Dahal-Koirala, Shiva; Komisarczuk, Anna Zofia; Valen, Eivind & Haapaniemi, Emma Maria (2026). SNIPSNP: precision design of CRISPR reagents. Full text in Research Archive
• Hammerstad, Marta; Gudim, Ingvild; Andersen, Hilde Kristin; Aracena-Solem, Matias & Hersleth, Hans-Petter (2026). Metals and thiols in bacterial defense and survival. Full text in Research Archive
• Aarnes, Halvor & Jørgensen, Marte Holten (2026). blomsterformel – Store norske leksikon. Store Norske Leksikon (Nettutgaven). Full text in Research Archive Show summary

  Blomsterformel er en tekst som viser strukturen og oppbygningen av en blomst. Når man beskriver en blomst på blomsterformel så angis de ulike delene av blomstene med en fast kode og deretter antall. Det er også koder for sammenvoksing, symmetri og plassering.

• Kleven, Tina; Grødem, Sverre; Ciosk, Rafal & Fyhn, Marianne (2025). Temperature-dependent neuronal plasticity in a mouse model of induced hypothermia. Full text in Research Archive
• Brodal, Hans Petter; Pharo, Heidi; Vedeld, Hege Marie & Lind, Guro Elisabeth (2025). PoDCall: a robust tool for automated droplet classification in DNA‐methylation droplet digital PCR. Molecular Oncology. ISSN 1574-7891. 19(9), p. 2472–2473. doi: 10.1002/1878-0261.70105. Full text in Research Archive
• Pharo, Heidi; Vedeld, Hege Marie; Wahlqvist, Rolf & Lind, Guro Elisabeth (2025). Re: Thomas Dreyer, Simone Brandt, Knud Fabrin, et al. Use of the Xpert Bladder Cancer Monitor Urinary Biomarker Test for Guiding Cystoscopy in High-grade Non-muscle-invasive Bladder Cancer: Results from the Randomized Controlled DaBlaCa-15 Trial. Eur Urol 2025;88:23–30. European Urology. ISSN 0302-2838. 88(6), p. e121–e121. doi: 10.1016/j.eururo.2025.07.028. Full text in Research Archive
• Świrski, Michał I.; Tierney, Jack A. S.; Albà, M. Mar; Andreev, Dmitry E.; Aspden, Julie L. & Atkins, John F. [Show all 170 contributors for this article] (2025). Translon: a single term for translated regions. Nature Methods. ISSN 1548-7091. 22(10), p. 2002–2006. doi: 10.1038/s41592-025-02810-3. Full text in Research Archive
• Haugen, Frida Loe; Mamia, Katariina Aino Inkeri; Rio, Oline Eriksen; Sollano, Anniken Solveig Matheson; Li, Zhuokun & Labun, Kornel [Show all 20 contributors for this article] (2025). CRISPR/Cas9-based pipeline to introduce custom edits in autologous T cells. Full text in Research Archive
• Haugen, Frida Loe; Mamia, Katariina Aino Inkeri; Rio, Oline Eriksen; Sollano, Anniken Solveig Matheson; Li, Zhuokun & Labun, Kornel [Show all 20 contributors for this article] (2025). CRISPR-Cas9 pipeline for custom editing in autologous T cells. doi: https:/event.fourwaves.com/crisprmed25/abstracts/b5911e8e-de8e-4ee4-b8e2-9f08cc2f31d8. Full text in Research Archive Show summary

  CRISPR/Cas9 gene editing technology is a promising tool for correcting pathogenic variants in Inborn Errors of Immunity (IEIs). Current IEI correction strategies rely on cDNA knock-in or gene knockout, which can disrupt gene expression and need extensive optimization. To address this, we developed a safe, efficient CRISPR/Cas9-based T cell editing pipeline using HDR for SNV correction. We observe a rapid and selective expansion of CD3+ T cells from patient peripheral blood following an 8-day cytokine stimulation. To enhance HDR outcomes, we explored multiple strategies and developed an optimized T cell pipeline capable of achieving up to 80% HDR. We found no pre-malignant off-target effects or karyotypic, transcriptomic, or proteomic abnormalities in patient T cells analysed with GUIDE-seq, scRNA-seq, long-read WGS, and proteomics. We observed functional correction of the disease phenotype in patient T cells demonstrated on transcriptomic level by single cell RNA sequencing, on proteomic level by Mass Spectrometry and in cell culture-based assays. So far, we have implemented the pipeline and corrected 12 unique SNV mutations causing 8 diverse IEIs. This platform is designed to streamline CRISPR tool discovery/optimization and can be further developed into a GMP-compatible, scaled-up process for salvage therapy in T cell-related IEIs.

• Hammerstad, Marta & Hersleth, Hans-Petter (2025). Structural studies of flavodoxins from Bacillus cereus in different redox states. Full text in Research Archive
• Hammerstad, Marta; Karlsen, Marthe Lindahl; Gudim, Ingvild; Gutiérrez-Fernández, Javier & Hersleth, Hans-Petter (2025). Structural and functional studies of enzymes involved in thiol-based redox pathways. Full text in Research Archive
• Karlsen, Marthe Lindahl; Hersleth, Hans-Petter & Hammerstad, Marta (2025). Structural and functional insights into bacillithiol disulfide reductase – A potential antimicrobial target. Full text in Research Archive
• Hammerstad, Marta; Karlsen, Marthe Lindahl; Gutiérrez-Fernández, Javier; Gudim, Ingvild & Hersleth, Hans-Petter (2024). Structures of bacilli­thiol and myco­thiol di­sulfide reductases of bacilli and mycobacteria. Acta Crystallographica Section A: Foundations of Crystallography. ISSN 0108-7673. 80. doi: 10.1107/S2053273324099200. Full text in Research Archive
• Ruso-Julve, Fulgencio; Sakya, Siri A; Kolderup, Heidi Anette; Tjärnhage, Elias; Benjakul, Sopisa & Nilsen, Jeannette [Show all 10 contributors for this article] (2024). Engineered human albumin as an effector negative scaffold for tailored design of long-acting antibody fragment fusions. Full text in Research Archive
• Falnes, Pål (2024). The versatile seven-beta-strand methyltransferase family. Full text in Research Archive
• Falnes, Pål (2024). A tour of the landscape of human seven-β-strand methyltransferases. Full text in Research Archive
• Sheng, Xia; Nenseth, Hatice Zeynep; Qu, Su; Kuzu, Omer; Frahnow, Turid & Simon, Lukas [Show all 16 contributors for this article] (2024). Correction to: IRE1α-XBP1s pathway promotes prostate cancer by activating c-MYC signaling (Nature Communications, (2019), 10, 1, (323), 10.1038/s41467-018-08152-3). Nature Communications. 15(1). doi: 10.1038/s41467-024-50645-x. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick (2024). Dear readers of NBS-Nytt, dear friends. NBS-nytt. ISSN 0801-3535. 1. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick (2024). Dear readers of NBS-Nytt, dear friends. NBS-nytt. ISSN 0801-3535. 4. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick (2024). Dear readers of NBS-Nytt, dear friends. NBS-nytt. ISSN 0801-3535. 3. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick (2024). Dear readers of NBS-Nytt, dear friends. NBS-nytt. ISSN 0801-3535. 2. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick (2024). Experience of a presenter at the NBS contact meeting. NBS-nytt. ISSN 0801-3535. 4. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick (2024). NBS Winter meeting 2024. NBS-nytt. ISSN 0801-3535. 1. Full text in Research Archive
• Hersleth, Hans-Petter (2024). Structures of bacillithiol and mycothiol disulfide reductases of bacilli and mycobacteria. Full text in Research Archive
• Crnic, Irene Fausk; Saragliadis, Athanasios; Cortés, Natalia Mojica; Kristiansen, Per Eugen & Krengel, Ute (2024). Molecular interactions between LPS and bacterial toxin. Full text in Research Archive
• Hammerstad, Marta; Karlsen, Marthe Lindahl; Fernandez, Javier Gutierrez; Gudim, Ingvild & Hersleth, Hans-Petter (2024). Structures of bacillithiol and mycothiol disulfide reductases of Firmicutes and Actinobacteria. Full text in Research Archive
• Karlsen, Marthe Lindahl; Hersleth, Hans-Petter & Hammerstad, Marta (2024). Structural and functional investigation of bacillithiol disulfide reductases. Full text in Research Archive
• Andersson-Rusch, Clara; Quist-Løkken, Ingrid; Sætrom, Pål; Hanif, Md Abu; Rolinski, Milosz & Møen, Johanna [Show all 12 contributors for this article] (2024). Whole genome CRISPR knockout screen reveals key function for ID3 in regulating myeloma cell survival via TCF3 and c-MYC. Full text in Research Archive
• Quist-Løkken, Ingrid; Tilseth, Mira; Hanif, Md Abu; Andersson-Rusch, Clara; Walz, Michael & Hausch, Felix [Show all 7 contributors for this article] (2024). Novel FKBP12 targeting PROTACs enhances BMP-induced apoptosis in multiple myeloma. Full text in Research Archive
• Mamia, Katariina Aino Inkeri; Kolbeinsdottir, Solrun; Li, Zhuokun; Reint, Ganna; Komisarczuk, Anna Zofia & Haugen, Frida Høsøien [Show all 21 contributors for this article] (2024). CRISPR/Cas9 T cell editing pipeline for monogenic primary immunodeficiency disorders. Full text in Research Archive
• Andersen, Jan Terje; Benjakul, Sopisa; Anthi, Aina Karen; Kolderup, Heidi Anette & Sandlie, Inger (2023). Et skreddersydd protein - designet av forskere i OUS - jakter koronaviruset. doi: https:/www.oslo-universitetssykehus.no/om-oss/innsikt/et-skredersydd-protein---designet-av-forskere-i-ous---jakter-coronaviruset/. Full text in Research Archive
• Montero, Ruth; Blihovde, Veronica Fitzpatrick; Klau, Leesa Jane; Hooft, Jamie Marie; Morales-Lange, Byron & Aarstad, Olav Andreas [Show all 11 contributors for this article] (2023). IMMUNOMODULATORY EFFECTS OF LAMINARIN FROM Laminaria hyperborea ON ATLANTIC SALMON LEUKOCYTES– FIRST STEPS TOWARDS NOVEL FUNCTIONAL FEED APPLICATION FOR SALMON. Full text in Research Archive
• Montero, Ruth; Blihovde, Veronica Fitzpatrick; Klau, Leesa Jane; Hooft, Jamie Marie; Morales-Lange, Byron & Aarstad, Olav Andreas [Show all 11 contributors for this article] (2023). First insights into the immunomodulatory effects of the b-glucan Laminarin on Atlantic salmon leukocytes. Full text in Research Archive
• Anthi, Aina Karen; Kolderup, Heidi Anette; Vaage, Eline Benno; Bern, Malin C.; Benjakul, Sopisa & Tjärnhage, Elias [Show all 30 contributors for this article] (2023). A novel non-invasive FcRn-targeted mucosal vaccine strategy. Full text in Research Archive
• Anthi, Aina Karen; mester, simone; Bern, Malin C.; Sandlie, Inger & Andersen, Jan Terje (2023). HemoMagic: Anemia Meets Unleashed Innovation. Full text in Research Archive
• Lind, Guro Elisabeth & Korsvoll, Nils Hallvard (2023). Postdoktorene fortjener bedre betingelser. Full text in Research Archive
• Anthi, Aina Karen; Kolderup, Heidi Anette; Vaage, Eline Benno; Bern, Malin C.; Benjakul, Sopisa & Tjärnhage, Elias [Show all 30 contributors for this article] (2023). A non-invasive subunit vaccine technology targeting FcRn induces protective mucosal and systemic antibody immunity in mouse models. Full text in Research Archive
• Anthi, Aina Karen; Kolderup, Heidi Anette; Vaage, Eline Benno; Bern, Malin C.; Benjakul, Sopisa & Tjärnhage, Elias [Show all 30 contributors for this article] (2023). A novel non-invasive FcRn-targeted mucosal vaccine strategy. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick (2023). NBS Winter meeting 2023. NBS-nytt. ISSN 0801-3535. 1. Full text in Research Archive
• Andreasen, Johan Kristian; Andreassen, Unni Soltun; Fossland, Trine; Høgestøl, Sofie Alexandra Engli; Lund, Ingrid & Lind, Guro Elisabeth [Show all 9 contributors for this article] (2023). Mangelfulle systemer for konfliktutsatte stipendiater. Full text in Research Archive
• Anthi, Aina Karen; Vaage, Eline Benno; Bern, Malin C.; Benjakul, Sopisa; Tjärnhage, Elias & Lode, Heidrun Elisabeth [Show all 29 contributors for this article] (2023). A non-invasive subunit vaccine technology targeting FcRn induces protective mucosal and systemic antibody immunity in mouse models. Full text in Research Archive
• Hammerstad, Marta; Andersen, Hilde Kristin; Rugtveit, Anne Kristine; Dahlen, Sondov Åsmundson Braathen & Hersleth, Hans-Petter (2023). Functional diversity among flavin-containing oxidoreductases involved in redox protection, iron homeostasis, and ribonucleotide reduction. Full text in Research Archive
• Fernandez, Javier Gutierrez; Hammerstad, Marta & Hersleth, Hans-Petter (2023). Structural determination of Mycobacterium smegmatis and Rhodococcus erythropolis mycothiol disulphide reductases. Full text in Research Archive
• Karlsen, Marthe Lindahl; Hersleth, Hans-Petter & Hammerstad, Marta (2023). Diversity among bacillithiol disulfide reductases – a structural and functional investigation. Full text in Research Archive
• Meuskens, Ina; Kristiansen, Per Eugen; Bardiaux, Benjamin; Koynarev, Vladimir Rosenov; Hatlem, Daniel & Prydz, Kristian [Show all 9 contributors for this article] (2023). Strain variations in a bacterial adhesin lead to different binding partners in the pathogen Yersinia enterocolitica. Full text in Research Archive
• Falnes, Pål (2023). Unravelling novel methyltransferases involved in protein and nucleic acid methylation. Full text in Research Archive
• Tislevoll, Benedicte Sjo; Hellesøy, Monica; Fagerholt, Oda Helen Eck; Gullaksen, Stein-Erik; Srivastava, Aashish & Birkeland, Even [Show all 24 contributors for this article] (2023). Author Correction: Early response evaluation by single cell signaling profiling in acute myeloid leukemia (Nature Communications, (2023), 14, 1, (115), 10.1038/s41467-022-35624-4). Nature Communications. 14(1). doi: 10.1038/s41467-023-37488-8. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick & Montero_Meza, Ruth Tamara (2023). Dear readers of NBS-Nytt, dear friends. NBS-nytt. ISSN 0801-3535. 2. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick (2023). Dear readers of NBS-Nytt, dear friends. NBS-nytt. ISSN 0801-3535. 1. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick & Montero_Meza, Ruth Tamara (2023). Dear readers of NBS-Nytt, dear friends. NBS-nytt. ISSN 0801-3535. 4. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick & Montero_Meza, Ruth Tamara (2023). Dear readers of NBS-Nytt, dear friends. NBS-nytt. ISSN 0801-3535. 3. Full text in Research Archive
• Fernandez, Javier Gutierrez; Hammerstad, Marta & Hersleth, Hans-Petter (2023). Structural determination of Mycobacterium smegmatis and Rhodococcus erythropolis mycothiol disulphide reductases. Full text in Research Archive
• Fernandez, Javier Gutierrez; Hammerstad, Marta & Hersleth, Hans-Petter (2023). Structural determination of Mycobacterium smegmatis and Rhodococcus erythropolis mycothiol disulphide reductases. Full text in Research Archive
• Hammerstad, Marta; Kjendseth, Åsmund Røhr & Hersleth, Hans-Petter (2023). A Research-inspired Laboratory Module on a Redox Protein Combining Biochemistry and Structural Biology. Full text in Research Archive
• Dugarte, Maria Eugenia Chollet; Harrison, Sean; Baumgarten, Saphira Felicitas; Morena-Barrio, Maria Eugenia de la; Bravo-Pérez, Carlos & Mowinckel, Marie-Christine [Show all 12 contributors for this article] (2022). Human iPSC-derived hepatic organoids – a model for blood coagulation research and therapy. Human Gene Therapy. ISSN 1043-0342. doi: 10.1089/hum.2022.29225.abstracts. Full text in Research Archive
• Fernandez, Javier Gutierrez; Hammerstad, Marta & Hersleth, Hans-Petter (2022). Structural determination of Mycobacterium tuberculosis and Rhodococcus erythropolis mycothiol disulphide reductases. Full text in Research Archive
• Hammerstad, Marta; Gudim, Ingvild & Hersleth, Hans-Petter (2022). Redox protection of pathogens by low molecular weight thiols. Full text in Research Archive
• Rese, Morten; Hammerstad, Marta & Hersleth, Hans-Petter (2022). Diversity in myoglobin function. Full text in Research Archive
• Andersen, Hilde Kristin; Hersleth, Hans-Petter & Hammerstad, Marta (2022). Structure-function studies of ferredoxin-flavodoxin NADP+ oxidoreductase 1 from Bacillus cereus as an iron-uptake oxidoreductase. Full text in Research Archive
• Falnes, Pål (2022). Discovering novel lysine-specific protein methyltransferases. Full text in Research Archive
• Falnes, Pål (2022). Discovering novel human histidine methyltransferases. Full text in Research Archive
• Bengtsen, Mads (2022). Epigenetiske endringer ved bruk av anabole steroider. Full text in Research Archive
• Ruso-Julve, Fulgencio; Sakya, Siri A; Lode, Heidrun Elisabeth; Foss, Stian; Benjakul, Sopisa & Nilsen, Jeannette [Show all 10 contributors for this article] (2022). Engineered albumin as a scaffold for improving the pharmacokinetic properties of antibody fragments. Full text in Research Archive
• Falnes, Pål (2022). Using mass spectrometry to study methylation of nucleic acids and proteins. Full text in Research Archive
• Enserink, Jorrit (2022). Brukermedvirkning i Kreftforskning. Full text in Research Archive
• Enserink, Jorrit (2022). Brukermedvirkning - Forskernes perspektiv. Full text in Research Archive
• Enserink, Jorrit (2022). Novel Methods for Stratification of Acute Myeloid Leukemia. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick (2022). Dear readers of NBS-Nytt, dear friends. NBS-nytt. ISSN 0801-3535. 2. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick (2022). Dear readers of NBS-Nytt, dear friends. NBS-nytt. ISSN 0801-3535. 3. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick (2022). Dear readers of NBS-Nytt, dear friends. NBS-nytt. ISSN 0801-3535. 4. Full text in Research Archive
• Malecka, Agnieszka Maria; Østlie, Ingunn; Trøen, Gunhild; Malecki, Jedrzej Mieczyslaw; Delabie, Jan & Tierens, Anne [Show all 8 contributors for this article] (2022). Gene Expression Analysis Reveals Downregulation of Complement Receptor 1 in Cold Agglutinin Disease. Blood. ISSN 0006-4971. 140(1), p. 6419–6420. doi: 10.1182/blood-2022-156489. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick (2022). Dear readers of NBS-Nytt, dear friends. NBS-nytt. ISSN 0801-3535. 1. Full text in Research Archive
• Rese, Morten; Hammerstad, Marta & Hersleth, Hans-Petter (2022). Diversity in myoglobin function. Full text in Research Archive
• Hammerstad, Marta; Gudim, Ingvild & Hersleth, Hans-Petter (2022). Redox protection of pathogens by low molecular weight thiols. Full text in Research Archive
• Bengtsen, Mads (2022). How does testosterone induced hypertrophy affects the muscle epigenome. Full text in Research Archive
• Bjune, Jan-Inge; Laber, Samantha; Dyer, Laurence; Xu, Zhao; Claussnitzer, Melina & Njølstad, Pål Rasmus [Show all 11 contributors for this article] (2021). Epigenetic control of adipogenesis by Irx3. Full text in Research Archive
• Malecka, Agnieszka Maria; Trøen, Gunhild; Delabie, Jan; Malecki, Jedrzej Mieczyslaw; Østlie, Ingunn & Tierens, Anne [Show all 9 contributors for this article] (2021). The mutational landscape of cold agglutinin disease: CARD11 and CXCR4 mutations are correlated with lower hemoglobin levels. American Journal of Hematology. ISSN 0361-8609. doi: 10.1002/ajh.26205. Full text in Research Archive
• Aders, Eirin; Nygård, Kamilla Børve; Bengtsen, Mads & Gundersen, Kristian (2021). Uncovering epigenetic mechanisms behind muscle memory. Full text in Research Archive
• Dyhre, Benjamin; Bengtsen, Mads; Nygård, Kamilla Børve; Millay, Douglas P. & Eftestøl, Einar (2021). How does a reduced myonuclear number affect epigenetic regulation of muscle function? Full text in Research Archive
• Bengtsen, Mads (2021). Anabolic steroid-induced alterations in the muscle epigenome. Full text in Research Archive
• Hanson, Ingunn; Pitman, Kathinka Elinor; Thiede, Bernd; Cai, Yiqing & Edin, Nina Frederike Jeppesen (2021). Proteomic Analysis of Low Dose-Rate Primed Cells and Extracellular Exosomes. Full text in Research Archive
• Lind, Guro Elisabeth & Falkum, Ingrid Lossius (2021). En ny modell for postdoktorstillingen. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick (2021). Dear readers of NBS-Nytt, dear friends. NBS-nytt. ISSN 0801-3535. 4. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick (2021). Dear readers of NBS-Nytt, dear friends. NBS-nytt. ISSN 0801-3535. 3. Full text in Research Archive
• Blihovde, Veronica Fitzpatrick (2021). Dear readers of NBS-Nytt, dear friends. NBS-nytt. ISSN 0801-3535. 2. Full text in Research Archive
• Bakketeig, Eris Aas; Olsen, Christine; Berg, Henriette Engen; Nyman, Tuula Anneli; Thiede, Bernd & Lundanes, Elsa [Show all 8 contributors for this article] (2021). Optimization of concatenation of fractions in comprehensive two dimensional liquid chromatography bottom-up proteomics. Full text in Research Archive Show summary

  Two dimensional liquid chromatography (2D-LC) has long been a commonly used technique in bottom-up proteomics, and is as relevant today as ever [1, 2]. To maximize the number of protein identifications, analytical chemists have strived to obtain 2D-LC systems with a high degree of orthogonality, which is needed to reach the required high peak capacities. A system utilizing reversed-phase chromatography (RP) in both column dimensions with a large difference in pH has proven to yield a higher peak capacity for peptides, even with a lower degree of orthogonality compared to other combinations [3]. The present study is based on Reubsaet, et al. [4]. The study is a comprehensive offline RPRP 2D-LC system with high pH (>9) in the first dimension and low pH (3) in the second dimension. The method’s purpose is to maximise protein identifications from one trypsinated protein sample. To achieve maximum identifications, Reubsaet incorporated the concept of concatenation of fractions. In practice, concatenation is applied through collecting the column output from the first dimension in eight numbered vials, where the 1st, 9th, 17th, … fractions are collected in the first vial, the 2nd, 10th, 18th, … are collected in the second, etc. [4]. In the present study, the fractionation scheme is examined by changing the total number of vials in which samples are concatenated, the number of fractions in each vial, and the collection time for each fraction. As the method demands high precision and is very timeconsuming, automatization is practically essential. Automatization is achieved with the spider fractionation scheme, proposed and shown by Kulak, et al.[5]. Here, a 10-port injector connects the first dimension column to each sample vial, and switches between them automatically, reducing the strain on the operator drastically, while removing human errors. The optimization of the concatenation will lead to higher numbers of identified proteins in proteomics.

• Dugarte, Maria Eugenia Chollet; andersen, elisabeth; Morena-Barrio, Maria Eugenia de la; Bravo-Perez, Carlos; Mowinckel, Marie-Christine & Myklebust, Christiane Fillion [Show all 12 contributors for this article] (2020). Characterizing Coagulation FVII from iPSC-Hepatocytes-like Cells: Setting the Basis for Cell Therapy Development. Blood. ISSN 0006-4971. 136. doi: 10.1182/blood-2020-139211. Full text in Research Archive
• Hanson, Ingunn; Edin, Nina Frederike J; Thiede, Bernd & Altanerova, Ursula (2020). Communication of radioprotective efects by TGF-β3. Full text in Research Archive
• Pesci, Giacomo; Johannesen, Hedda; Løset, Geir Åge & Krengel, Ute (2020). A promising antibody to treat cancer. How does it work? Full text in Research Archive
• Søreide, Øystein; Olsen, Dag Rune; Lind, Guro Elisabeth & Holden, Lars (2020). Ingen forskning, ingen omstilling. Dagens næringsliv. ISSN 0803-9372. Full text in Research Archive
• Johannesen, Hedda; Bjerregaard-Andersen, Kaare; Grzybek, Michal; Løset, Geir Åge & Krengel, Ute (2020). Molecular basis of specificity of a promising anti-tumour antibody. Full text in Research Archive
• Falnes, Pål Øystein (2020). Novel human methyltransferases - roles in mitochondrial respiration, protein synthesis and beyond. Full text in Research Archive
• Hammerstad, Marta; Gudim, Ingvild & Hersleth, Hans-Petter (2020). The Crystal Structures of Bacillithiol Disulfide Reductase YpdA Reveal Structural and Functional Insight into a New Type of FAD-Containing NADPH-Dependent Oxidoreductases. Full text in Research Archive
• Berg, Henriette Engen; Halldórsson, Skarphéðinn; Thiede, Bernd; Vik-Mo, Einar O.; Lundanes, Elsa & Wilson, Steven Ray Haakon (2020). Micro pillar array columns for comprehensive proteomics. Full text in Research Archive Show summary

  Pillar array nano liquid chromatography columns can potentially replace packed nano liquid chromatography (nLC) columns for analyses of complex samples due to high chromatographic performance and low backpressure. In our study, pillar array columns are used for the detection of potential biomarkers in Glioblastoma Multiforme (GBM), which is the most aggressive form of brain cancer in adults. Pillar array columns consist of several micropillars etched into silicon wafers using lithographic micromachining. The interest of these columns in nLC format has increased as an option to packed nLC columns as the chromatographic efficiency could outperform nLC columns packed with sub-2 µm particles while maintaining the sensitivity [1]. In contrast to packed columns, where eddy dispersion is the main source of band broadening due to irregular flow paths between the particles, the perfectly ordered pillars in pillar array columns eliminate the eddy dispersion to a minimum. The low backpressure of pillar array columns compared to that of packed columns also permits the use of exceptionally long columns, increasing the chromatographic performance further. Using pillar array columns, plate numbers over 1 million and reduced plate heights (h) even <1 µm have been achieved [2-4]. High chromatographic performance is especially important in comprehensive proteomics. Pre-fractionation (using e.g. gel electrophoresis) is often added in proteomic protocols to increase the throughput [5]. However, performing pre-fractionation can be time-consuming in addition to the highly increased instrument time required. Recently, a micro-Chip pillar array nLC column (µPACTM) compatible with standard nLC instrumentation has been commercialized (Figure 1) [6]. Our question was; could the use of µPAC nLC columns reduce the need for pre-fractionation in comprehensive proteomics of GBM biopsies?

• Hanson, Ingunn; Altanerova, Ursula; Thiede, Bernd & Edin, Nina Frederike J (2020). Radioprotector TGF-β3 is secreted in exosomes and activated by LDR priming. Full text in Research Archive Show summary

  The biological effects of ionizing radiation in the low-dose region is important for diagnostic imaging, healthy tissue in radiation therapy and environmental radiation. For a majority of tested cell lines, the low dose hyper-radiosensitive (HRS) response has been shown to be default. We have previously shown that cells primed with a low dose-rate (⁓0.3 Gy/h) ionizing radiation permanently loses the HRS response. We have also shown that the transforming growth factor (TGF)-β3 is responsible for this effect. Here, we investigated the secretion mechanism of TGF-β3 from LDR primed cells, and theorize regarding the way it is activated. Exosomes were isolated from LDR primed cells, and were shown to remove HRS in reporter cells when co-cultured with these. Adding a TGF-β3 inhibitor to the medium restored the HRS response in reporter cells, confirming that TGF-β3 was the factor in the exosomes responsible for removing HRS. Mass spectrometry was performed on the content of exosomes from primed and control cells. Proteomic analysis confirmed the presence of exosome-specific proteins in exosomes from primed and control groups. TGF-β3 along with latency-associated protein (LAP), the protein that TGF-β3 is bound to in its inactive form, was present in similar amounts in both groups, indicating that TGF-β3 was secreted in its inactive form, regardless of LDR priming. Three proteins were significantly upregulated in the primed exosomes. These proteins will be investigated further in future experiments. We conclude that TGF-β3 is secreted in its inactive form in exosomes from LDR primed and unprimed cells, and that the priming is responsible for its activation. We will further investigate three significantly upregulated proteins with regards to their role in the activation of TGF-β3.

• Hammerstad, Marta; Gudim, Ingvild; Lofstad, Marie; Røhr, Åsmund Kjendseth & Hersleth, Hans-Petter (2019). Enzyme Activation by a Flavoprotein Redox Network. Full text in Research Archive
• Falnes, Pål Øystein (2019). Regulation of protein synthesis through methylation of human eukaryotic elongation factor 1 alpha (eEF1A). Full text in Research Archive
• Gudim, Ingvild; Lofstad, Marie; Hammerstad, Marta; Beek, Wouter van & Hersleth, Hans-Petter (2019). New structural insight into the well known peptide flip observed in flavodoxins. Acta Crystallographica Section A: Foundations and Advances. A75, p. e123–e123. doi: 10.1107/S2053273319094336. Full text in Research Archive
• Hammerstad, Marta; Gudim, Ingvild; Lofstad, Marie; Kjendseth, Åsmund Røhr & Hersleth, Hans-Petter (2019). Enzyme activation by a flavoprotein redox network. Acta Crystallographica Section A: Foundations and Advances. A75, p. e128–e128. doi: 10.1107/S2053273319094282. Full text in Research Archive
• Kendall-Price, Sophie; Evans, Rhiannon M; Rowbotham, Jack S; Reeve, Holly A; Carr, Stephen B & Frogley, Mark D [Show all 11 contributors for this article] (2019). Generating Single Metalloprotein Crystals in Well-Defined Redox States – Electrochemical Microspectroscopy As a Tool for Mechanistic Studies. Meeting Abstracts - The Electrochemical Society [ECS]. ISSN 1091-8213. 9, p. 425–425. Full text in Research Archive
• Shaw, Alan; Hoffecker, Ian T.; Smyrlaki, Ioanna; Rosa, João; Grevys, Algirdas & Bryant-Bratlie, Diane Lynn [Show all 10 contributors for this article] (2019). Publisher Correction: Binding to nanopatterned antigens is dominated by the spatial tolerance of antibodies. Nature Nanotechnology. ISSN 1748-3387. 14(4), p. 398–398. doi: 10.1038/s41565-019-0404-3. Full text in Research Archive
• Falnes, Pål Øystein (2019). Modulation of human mitochondrial metabolism by novel lysine-specific methyltransferases. Full text in Research Archive
• Zhang, Beibei; Ayuda-Duran, Maria del Pilar; Piechaczyk, Laure Isabelle; Fløisand, Yngvar; Safont, Mireia Mayoral & Karlsen, Ida Tveit [Show all 14 contributors for this article] (2019). GRP94 rewires and buffers the FLT3-ITD signaling network and promotes survival of acute myeloid leukemic stem cells. Haematologica. ISSN 0390-6078. 104(5). doi: 10.3324/haematol.2019.220533. Full text in Research Archive
• Fan, Qiong; Nørgaard, Rikke Christine; Grytten, Ivar; Ness, Cecilie Maria; Lucas, Christin & Vekterud, Kristin [Show all 15 contributors for this article] (2019). LXRα interacts with the glucose-sensing transcription factor ChREBPα to regulate its transcriptional activity. Full text in Research Archive Show summary

  The cholesterol-sensing nuclear receptor Liver X Receptor (LXR) and the glucose-sensing transcription factor carbohydrate responsive element-binding protein (ChREBP) are central players in the regulation of glucose and lipid metabolism. LXR does this job in part by regulating the expression of ChREBP. We have previously shown that LXR also regulates ChREBP activity. To get a better understanding of mechanisms at play, we asked if LXR and ChREBP interact physically. Interestingly, LXRα binds to ChREBPα, but not the shorter isoform ChREBPβ. Co-immunoprecipitation (CoIP) of different LXR and ChRBEP domains shows that it is ChREBPα’s low glucose inhibitory domain (LID), which is lacking in ChREBPβ, that interacts with the ligand-binding domain (LBD) of LXRα. In line with this, we see a surprisingly high co-occupancy of LXR and ChREBP on regulatory regions in the mouse genome when re-analysing two independently published chromatin immunoprecipitation-sequencing (ChIP-seq) datasets. Moreover, Functional studies show that LXRα is able to co-activate together with ChREBPα, but not ChREBPβ, and increase ChREBP-specific target gene expression in vitro and in vivo. Unexpectedly however, ligand-engaged LXR exhibits a repressive effect on the expression of the same genes in primary mouse hepatocytes, in contrast to what we observe with target genes that are common to LXR and ChREBP. Performing CoIP and ChIP on selected target genes, we demonstrate mechanistically that the repressive effect most likely is due to a weakened ChREBPα:LXRα interaction and reduced binding of ChREBP to chromatin. Altogether, the novel transcriptional complex comprising ChREBPα and LXRα adds to the intricate integration of nutrient signals in glucose and lipid metabolism.

• Gudim, Ingvild; Lofstad, Marie; Hammerstad, Marta; Røhr, Åsmund Kjendseth & Hersleth, Hans-Petter (2019). Activation of the Class Ib Ribonucleotide Reductase by a Flavodoxin Reductase in Bacillus cereus. Full text in Research Archive
• Falnes, Pål Øystein (2019). Identification and characterisation of novel human methyltransferases. Full text in Research Archive
• Sørensen, Karen Kristine; Antwi, Milton Boaheng; Dumitriu, Gianina; Li, Ruomei; Smedsrød, Bård & Eskild, Winnie (2019). Scavenger function of liver sinusoidal endothelial cells in a transgenic mouse model of liver fibrosis is reduced. Full text in Research Archive
• Johannesen, Hedda; Bjerregaard-Andersen, Kaare; Grzybek, Michal; Ünal, Coskun; Løset, Geir Åge & Krengel, Ute (2019). Crystal structure of an anti-tumour antibody in complex with its tumour-specific antigen. Full text in Research Archive
• Andersen, Jan Terje; Sakya, Siri Aastedatter; Foss, Stian; Sandlie, Inger & Voets, Ardi (2019). Is an HIV vaccine within reach? Vaksinebloggen. Full text in Research Archive
• Naseri, Nafasgol; Dahal-Koirala, Shiva; Haapaniemi, Emma Maria & Falnes, Pål (2026). Investigation of on-target Chromosomal deletions and loss during HDR-enhanced CRISPR-Cas9 correction of a CARMIL2 point mutation in patient-derived T cells. University of Oslo (UiO). Full text in Research Archive
• Borg, Nora Helene Hammersten; Komisarczuk, Anna Zofia; Falnes, Pål & Haapaniemi, Emma Maria (2026). Development of a Multimodal Screening Platform for Comparative Evaluation of Prime Editing Efficiency. Establishing an experimental framework through reporter cell line generation, prime editing vector construction, and detection assay development. Universitetet i Oslo. Full text in Research Archive Show summary

  --- Abstract will be published: 2027-05-19

• Andersen, Hilde Kristin; Hammerstad, Marta & Hersleth, Hans-Petter (2022). Struktur-funksjonsstudier av ferredoksin-flavodoksin NADP+ oksidoreduktase 1 fra Bacillus cereus som en jern-opptak oksidoreduktase. Universitetet i Oslo. Full text in Research Archive Show summary

  Bacillus cereus har tre enzymer som er annotert som ferredoksin-flavodoksin NADP+ oksidoreduktaser (FNRer). FNR3 er vist å redusere disulfidformen av bacillitiol, FNR2 er vist å effektivt redusere flavodoksiner, men det er enda ikke funnet noen god redokspartner til FNR1. FNR1 er vist å skille seg fra FNR2 ved å være del av en fylogenetisk undergruppe av tioredoksinreduktase-aktige FNRer hvor en konservert aromatisk residu i C-terminalen, som stabiliserer FAD-kofaktoren er erstattet med valin sammenlignet med FNR2-gruppen. Staphylococcus aureus har en nær homolog til FNR1 som deler den konserverte valinresiduen, som har blitt vist å kunne fungere som en jern-opptak-oksidoreduktase (IruO). IruO kan tilgjengeliggjøre jern for bakterier ved å redusere hemoksygenasene iron-regulated surface determinant (Isd)G og IsdI som degraderer hem, eller sideroforer. B. cereus har kun en homolog til IsdG, men ikke IsdI. I dette prosjektet har det blitt utført aktivitetsstudier for å undersøke om B. cereus FNR1 kan fungere som en IruO, og redusere IsdG for å degradere hem, ved bruk av UV-vis-spektroskopi. FNR1 og IsdG har blitt uttrykt og renset. Strukturen til IsdG er blitt løst ved bruk av proteinkrystallografi. IsdG viste å ha svært lik struktur i B. cereus og S. aureus, med ferredoksinfolding og de konserverte residuene Asn7, Trp67 og His77 i bindingssetet til hem. Aktivitetsstudiene viste at FNR1 kunne redusere IsdG for å degradere hem og tilgjengeliggjøre jern. FNR1 er anslått å ha evner i samme størrelsesorden til å redusere IsdG som til å redusere flavodoksiner. Aktivitetsstudiene tyder på at hem blir degradert ved først å redusere Fe(III) til Fe(II) og at det går via Fe(II)-O2 som i kanoniske hemoksygenaser.

• Rese, Morten; Hersleth, Hans-Petter; Hammerstad, Marta & Andersen, Niels Højmark (2022). Functional diversity of myoglobin isozymes from Carassius auratus - Biophysical study of myoglobin as a peroxidase. Universitetet i Oslo. Full text in Research Archive Show summary

  Myoglobin (Mb) is a heme protein expressed in oxidative muscle tissue and functions in oxygen storage and transport. Mb can also display complex redox chemistry involved in several important physiological processes including scavenging of reactive oxygen species and nitric oxide metabolism. Goldfish and the common carp are the only vertebrates known today that express two paralogs of Mb (Mb1 and Mb2) as a result of a recent whole genome duplication. Mb1 is expressed in gills, liver, and skeletal muscle tissue, whereas Mb2 is exclusively expressed in the brain. The tissue-specific expression led to work showing that Mb2 in carp fish is a more effective peroxidase. In this study, work has been done to gain detailed structural and functional insight into the differences of the goldfish Mb1 and Mb2 isozymes by biophysical characterization and 3D structure analysis. The intensively studied Mb from horse heart (hMb) has been studied together with Mb1 and Mb2 as a reference. The reaction of Mb with hydrogen peroxide (H2O2) has been of particular interest. In addition, studies on how radiation damage during diffraction data collection affects redox states of Mb have been performed. This study shows that goldfish Mb2 reacts with H2O2 significantly faster than Mb1. Structural modeling of Mb1 and Mb2 indicates that no differences around the active site are responsible for the difference in reactivity. The structural basis of this functional difference is therefore complex in nature and might be explained by altered dynamics as a consequence of differing amino acid composition towards the surface of the proteins. The ferric and ferryl redox states of Mb are damaged by X-ray radiation during diffraction data collection. The ferric state is significantly more susceptible to radiation damage compared to the ferryl state. This study shows the importance of performing a time/dose calculation on the amount of radiation a specific redox state can handle when collecting diffraction data.

• Berg, Henriette Engen; Wilson, Steven Ray Haakon; Lundanes, Elsa; Thiede, Bernd & Vik-Mo, Einar Osland (2022). Exploring miniaturized chromatography formats and sample preparation methods for shotgun proteomics in limited biological samples. Department of Chemistry, Faculty of Mathematics and Natural Sciences, University of Oslo. Full text in Research Archive Show summary

  A significant challenge in medical diagnostics is the development of simple but efficient tools for the detection/quantification of several biomarkers simultaneously using non-invasive sampling techniques. In this regard, the analysis of proteins (proteomics) is essential for understanding cellular processes and biomarker discovery. However, proteins vary greatly in terms of concentration levels and chemical properties in biological materials. Further, low sample sizes of modern biological models (e.g., patient-derived cell cultures, exosomes, and organoids) remain a big analytical challenge. The present work has focused on the brain cancer glioblastoma, which is in great need of increased knowledge and non-invasive sampling techniques. In addition, human organoids, which could act as a future in vitro model for disease modeling and personalized medicine, have been investigated. We have used high-resolution mass spectrometry for protein identification, exploring a selection of miniaturized liquid chromatography formats (for separation) and sample preparation techniques. By implementing these techniques, we have been able to study exosomes, 2D/3D cell cultures, and organoids, identifying over 6300 proteins in a single run using less than 5 µg of protein. The work has provided important insight into the possibilities and challenges of several novel models. It represents a development toward deeper proteomic profiling focusing on maintaining a high protein yield and time efficiency.

• Bakketeig, Eris Aas; Wilson, Steven Ray Haakon; Lundanes, Elsa; Berg, Henriette Engen; Olsen, Christine & Reubsaet, Leon [Show all 8 contributors for this article] (2021). The setup and optimization of a complete global proteomics two-dimensional liquid chromatography system, using Spider Fractionation. Universitetet i Oslo. Full text in Research Archive
• Hersleth, Hans-Petter; Hammerstad, Marta & Dahlen, Sondov Åsmundson Braathen (2021). Structure and function of FNR1 from Bacillus cereus undergoing a mutation. Universitetet i Oslo. Full text in Research Archive
• Rugtveit, Anne Kristine; Hammerstad, Marta & Hersleth, Hans-Petter (2021). Structural and functional studies of a ferredoxin/flavodoxin NADP+-oxidoreductase mutant from Bacillus cereus. Universitetet i Oslo. Full text in Research Archive
• Solbakke, Kristoffer Gudesen; Krengel, Ute; Johannesen, Hedda; Løset, Geir Åge & Bjerregaard-Andersen, Kaare (2019). 4G9: An antibody that potentially imitates a cancer-related carbohydrate. Universitetet i Oslo. Full text in Research Archive

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