BMB Section Seminar: PML protein organizes heterochromatin domains and regulates histone H3.3 loading by ATRX
Erwan Delbarre, PhD, Collas lab, Institute of Basic Medical Sciences, UiO
H3.3 is targeted to PML bodies by DAXX. (Photos: E. Delbarre)
Histone variant H3.3 is emerging as a key player in the formation and maintenance of heterochromatin. Its incorporation in chromatin is mostly ensured by two specific histone chaperone complexes. While the HIRA complex deposits H3.3 mainly in open chromatin including regulatory regions and bodies of active genes, the DAXX/ATRX complex allows H3.3 incorporation in heterochromatin including telomeric, pericentric and other repeats-rich regions. We showed earlier in human cells that H3.3 is targeted to promyelocytic (PML) bodies by DAXX prior to its deposition in chromatin. This process was further showed to critical for H3.3 incorporation into pericentric heterochromatin, suggesting that PML is involved in the deposition of H3.3 more specifically into heterochromatic regions. In the present work, we directly investigate in mouse cells the role of PML in the distribution of H3.3 in chromatin. Using ChIP-sequencing, we show that PML associates with heterochromatic domains that we name PML-associated domains (PADs). PADs are enriched in H3K9m3 and H3K27me3 and with a limited access for neo-synthesized H3.3. Strikingly in PADs, incorporation of H3.3 is mainly ensured by ATRX. In absence of PML, these domains display reduced H3K9me3 level and enrichment in H3K27me3, suggesting the importance of maintaining a heterochromatic configuration in these domains. Loss of PML also results in increased accessibility for H3.3 deposition by HIRA in PADs, albeit with a loss of effect of ATRX. Moreover, live cell imaging and chromatin fractionation experiments using micrococcal nuclease show that in absence of PML, H3.3 is incorporated faster in chromatin and a fraction of H3.3 is redistributed from a loose MNase-accessible to a more compact, MNase-resistant chromatin compartment. Altogether, our results reveal new roles of PML in (1) maintaining heterochromatin domains (PADs) with specific epigenetic signatures and (2) in the regulation of H3.3 deposition in different chromatin compartments by modulating the activity of ATRX in specific regions. The data further unveil a hitherto unappreciated role of PML in the large-scale organization of chromatin domains.