Dual function of H2AK119ub1 on gene regulation via modulating canonical-PRC1 and H1-dependent chromatin compaction H2AK119ub1 deferentially fine-tunes gene expression by modulating canonical-PRC1 and H1-dependent chromatin compaction
First, we applied ChIP-seq (Chromatin Immunoprecipitation) to measure the enrichment of Cbx7, Ring1B, and H3K27me3 on DNA in WT (wild-type), Ring1bI53A, and Rybp/Yaf2-DKO ESCs. The results suggest that accumulation of cPRC1 on chromatin isn't caused by the mis-regulation of H3K27me3 in Ring1bI53A mESCs and Rybp/Yaf2-DKO impairs the accumulation of ncPRC1 on chromatin. Next, here we adopt ChIP-seq to investigate the dynamic of Ring1B and Cbx7 on DNA during Rybp/Yaf2 degradation after IAA treatment. We found that that H2AK119ub1 plays an important role in restricting the binding of cPRC1 on chromatin and thereby compromises cPRC1-mediated chromatin compaction. And then, we carried out mRNA-seq and identified differential expressed genes among WT and Ring1bI53A mESCs. Next, we applied ChIP-seq (Chromatin Immunoprecipitation) to measure the enrichment of Cbx7, Ring1B, Jarid2, PHC1, and PCGF2 on DNA in WT (wild-type), Ring1bI53A, and Bap1-KO ESCs . The results suggest that the binding of “all cPRC1” with nucleosomes is blocked by H2AK119ub1. Finally, here we performed MNase-seq in WT and H1-QKO mESCs. We found that H1-QKO lead to a stronger decondensation of H2Aub1_enriched TSSs compared with residual TSS regions. The result of ChIP-seq for H1 and ATAC-seq (Assay for Transposase Accessible Chromatin with high-throughput sequencing) of WT and Ring1BI53A, and mRNA-seq of WT and H1-QKO mESCs suggest H2AK119ub1 represses genes via promoting H1-mediated chromatin compaction. Overall design: RNA-seq, ATAC-seq, ChIP-seq for WT, I53A, Rybp/Yaf2 DKO, H1QKO cells. And MNase-seq for WT and H1QKO cells.