P1.158: Histone H1 Redistribution During Mitosis and Its Impact on Chromatin Organization

Background: Histone H1 linker histones are the most abundant chromatin-bound proteins, essential for DNA compaction, genomic organization, and shaping the epigenome by influencing histone modifications. Although significant progress has been made in understanding their roles during interphase, the precise mechanisms by which H1 linker histones regulate chromatin structure throughout the cell cycle, especially during mitosis, remain unclear. Clarifying these mechanisms is crucial, as mutations in H1 genes are associated with developmental disorders such as overgrowth syndromes, characterized by abnormal cellular proliferation.

Methods: Accurately measuring H1 occupancy has been challenging due to multiple H1 subtypes and their broad genomic distribution. To address this, we used CRISPR-mediated gene editing to generate murine erythroid cell lines expressing high-affinity epitope tags on the three major subtypes: H1c, H1d, and H1e. Leveraging these engineered cell lines, we performed ChIP-seq with ultra-deep sequencing with more than one billion reads, generating unprecedentedly high-resolution maps of H1 distribution across the genome during interphase and mitosis. We analyzed H1 occupancy across 25 distinct chromatin states and assessed correlations with chromatin accessibility and gene transcription to determine how transcriptional activity and chromatin structure influence H1 dynamics.

Results: Total H1 protein levels were unchanged between interphase and mitotic phases, and individual H1 subtypes displayed highly similar binding patterns throughout the genome. In line with previous work, H1 occupancy was greatest in heterochromatic regions and lowest in active regions such as enhancers, promoters, and actively transcribed genes during interphase. However, during mitosis, the typical depletion of H1 in these active regions was largely reversed, resulting in increased H1 binding at enhancers, promoters, and gene bodies. This increase in H1 occupancy did not correspond with changes in chromatin accessibility in mitosis but correlated inversely with changes in RNA Polymerase II binding, suggesting that resumption of active transcription in the subsequent interphase causes formation of H1 depleted regions.

Conclusion: Our findings indicate a substantial redistribution of H1 linker histones during mitosis, with increased occupancy at typically active chromatin regions coinciding with global transcriptional silencing. This redistribution highlights the dynamic interplay between transcriptional activity and H1 chromatin binding, providing novel insights into chromatin organization during cell division. These results represent important first steps toward understanding developmental disorders caused by mutations in H1 histones, like overgrowth syndromes.