GSE97947
H3S10ph broadly marks early-replicating domains in interphase ESCs and shows reciprocal antagonism with H3K9me2
Pervasive phosphorylation of histone H3 at serine 10 (H3S10ph) by Aurora B/C plays an important role in mitosis; however, this mark has also been observed at specific genic promoters and enhancers in interphase, implicating mitosis-independent functions. Using the FUCCI cell cycle reporter, we found that 30% of the genome is persistently marked with H3S10ph in interphase mouse embryonic stem cells (ESCs). H3S10ph demarcates broad gene-rich euchromatic regions in G1 and shows remarkable correlation with domains of early DNA replication timing (RT). Consistent with mitosis-independent H3S10 kinase activity, this pattern was preserved in ESCs treated with hesperidin, a potent inhibitor of Aurora B/C. Disruption of H3S10ph by expression of non-phosphorylatable H3.3S10A results in ectopic spreading of H3K9me2 into adjacent H3S10ph-enriched euchromatic regions, mimicking the phenotype observed in Drosophila JIL-1 kinase mutants. Conversely, H3S10ph domains expand in interphase Glp-/- ESCs, revealing that H3S10ph expansion is restricted by H3K9me2. Strikingly, spreading of H3S10ph at RT transition regions (TTRs) is accompanied by aberrant strand-biased transcription initiation of genes and repetitive elements co-oriented with the replication fork, indicating that H3K9me2 plays a critical role in establishing repressive chromatin on the leading strand. Finally, we show that H3S10ph is also present in interphase murine embryonic fibroblasts (MEFs), but is restricted to intragenic regions of actively transcribing genes. Our study provides a detailed map of interphase H3S10ph in ESCs and MEFs, uncovering a previously unappreciated crosstalk between the opposing marks H3S10ph and H3K9me2, and a role for the latter in ensuring appropriate transcription at TTRs in ESCs. Overall design: H3S10ph ChIP & Inputs in asynchnous and FUCCI-sorted ESCs H3S10ph ChIP & Inputs in DMSO and Hesperadin-treated ESCs YFP & H3K9me2 ChIP in H3.3-YFP, 2 H3.3S10A-YFP overexpressing ESC clonal lines H3K9me2 & H3S10ph in Glp-/- and corresponding WT ESCs H3K36me3, H3K4me3, RNA-seq in G9a-/-, Glp-/- and corresponding WT ESCs H3S10ph ChIP & Inputs in DMSO and Hesperadin-treated WT and G9a-/- MEFs.