A conserved H3K14ub-driven H3K9me3 for chromatin compartmentalization

To delineate the relationship between G2E3-catalyzed H3K14ub and H3K9me3 in genome-wide, we performed chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) for H3K14ub, H3K9me3 and H3K4me3 in Parental (sgG2E3) control HeLa cells and G2E3 KO-10 HeLa cells. Our data demonstrated that G2E3-catalyzed H3K14ub potentiates H3K9 trimethylation (H3K9me3) by SUV39H and is specifically required for SUV39H compartmentalization and H3K9me3 in pericentromeric heterochromatin. Loss ofG2E3 severely impairs not only pericentromeric heterochromatin organization, but also results in aberrant accumulation of SUV39H and H3K9me3 in numerous euchromatin regions and a widespread transcriptional repression (combined with RNA-seq data). Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for histone modifications H3K4me3, H3K9me3 and H3K14ub in Parental （sgG2E3）control HeLa cells and G2E3 knockout HeLa cells. sgG2E3 control HeLa cells is a sgRNA stable cell line that has not been cut by Cas9 protein. In our paper, it is marked as Parental control HeLa cells.

为在全基因组范围内解析G2E3催化的H3K14泛素化（H3K14ub）与H3K9三甲基化（H3K9me3）之间的调控关系，我们分别在亲本（sgG2E3）对照海拉（HeLa）细胞以及G2E3敲除-10（G2E3 KO-10）海拉细胞中，针对H3K14ub、H3K9me3与H3K4me3开展了染色质免疫共沉淀结合高通量测序（ChIP-seq）实验。本研究数据证实，G2E3催化的H3K14ub可通过SUV39H增强H3K9三甲基化（H3K9me3），且其对SUV39H的区域化定位以及近着丝粒异染色质中的H3K9me3修饰具有特异性调控作用。G2E3的缺失不仅会严重破坏近着丝粒异染色质的结构组织，还会导致SUV39H与H3K9me3在大量常染色质区域异常富集，并引发广泛的转录抑制（结合RNA测序（RNA-seq）数据进行分析）。
本研究同时针对亲本（sgG2E3）对照海拉细胞与G2E3敲除海拉细胞中的组蛋白修饰H3K4me3、H3K9me3与H3K14ub开展了染色质免疫共沉淀测序（ChIP-seq）实验。其中sgG2E3对照海拉细胞为未被Cas9蛋白切割的单导RNA（single guide RNA，sgRNA）稳定细胞系，在本论文中被标注为亲本对照海拉细胞。