H3K27me3 and the PRC1-H2AK119ub pathway cooperatively maintain heterochromatin and transcriptional silencing after the loss of H3K9 methylation.

BACKGROUND: Heterochromatin is a fundamental component of eukaryotic chromosome architecture, crucial for genome stability and cell type-specific gene regulation. In mammalian nuclei, heterochromatin forms condensed B compartments, distinct from the transcriptionally active euchromatic A compartments. Histone H3 lysine 9 and lysine 27 trimethylation (H3K9me3 and H3K27me3) are two major epigenetic modifications that enrich constitutive and facultative heterochromatin, respectively. Previously, we found that the redistribution of H3K27me3 following the loss of H3K9 methylation contributes to heterochromatin maintenance, while the simultaneous loss of both H3K27me3 and H3K9 methylation induces heterochromatin decondensation in mouse embryonic fibroblasts. However, the spatial positioning of B compartments largely persists, suggesting additional mechanisms are involved. RESULTS: In this study, we investigated the role of H2AK119 monoubiquitylation (uH2A), a repressive chromatin mark deposited by Polycomb Repressive Complex 1 (PRC1), in maintaining heterochromatin structure following the loss of H3K9 and H3K27 methylation. We observed that uH2A and H3K27me3 are independently enriched in B compartments after H3K9 methylation loss. Despite the absence of H3K9me3 and H3K27me3, uH2A remained localized and contributed to heterochromatin retention. These results suggest that PRC1-mediated uH2A functions independently and cooperatively with H3K27me3 to maintain heterochromatin organization originally created by H3K9 methylation. CONCLUSION: Our findings highlight a compensatory role for uH2A in preserving heterochromatin structure after the loss of other repressive chromatin modifications. The PRC1-uH2A pathway plays a critical role in maintaining the integrity of B compartments and suggests that heterochromatin architecture is supported by a network of redundant epigenetic mechanisms in mammalian cells.