Histone marks enable formation of immiscible phase-separated chromatin compartments

Eukaryotic chromatin is organized into compartments for gene expression regulation, but the underlying mechanisms remain unclear. Here, we demonstrate that multivalent H3K27me3 and its reader, CBX7-PRC1, regulate facultative heterochromatin via a phase separation mechanism, similar to constitutive heterochromatin. Facultative and constitutive heterochromatin represent distinct, coexisting repressive condensates in nuclei. H3K27me3- and H3K9me3-marked nucleosomal arrays and their reader complexes phase separate into immiscible condensates, analogous to the relationship between facultative and constitutive heterochromatin in vivo. Overexpression of CBX7-PRC1 causes aberrant chromatin compartmentalization in vivo and up-regulation of cancer-related genes. CBX7 inhibitor effectively inhibits cancer cell proliferation possibly through phase separation-mediated compartment reorganization. Our data suggest that histone marks mediate chromatin compartmentalization via phase separation, and offer potential epigenetic mechanistic insights into tumor development and cancer therapy. By employing RNA-seq, ChIP-seq, CUT&Tag, and Hi-C, we systematically examined the change in chromatin features and transcriptome upon overexpression of CBX7 in NIH3T3 cells.

真核染色质会被组装为参与基因表达调控的染色质区室，但其背后的潜在机制仍未阐明。本研究证实，多价H3K27三甲基化（H3K27me3）及其组蛋白阅读器CBX7-PRC1，可通过相分离机制调控兼性异染色质，这一过程与组成型异染色质的调控类似。兼性与组成型异染色质是细胞核内两类共存且功能迥异的抑制性凝聚体。携带H3K27me3与H3K9me3修饰的核小体阵列及其对应的阅读器复合物，会相分离形成互不相溶的凝聚体，这与体内兼性与组成型异染色质的相互关系类似。过表达CBX7-PRC1会在体内引发异常染色质区室化，并上调癌症相关基因的表达。CBX7抑制剂可有效抑制癌细胞增殖，这一效应可能通过相分离介导的区室重塑实现。本研究数据表明，组蛋白修饰可通过相分离介导染色质区室化，为肿瘤发生及癌症治疗提供了潜在的表观遗传机制视角。本研究通过RNA测序（RNA-seq）、染色质免疫共沉淀测序（ChIP-seq）、CUT&Tag技术及Hi-C染色体构象捕获技术，系统检测了NIH3T3细胞中CBX7过表达后染色质特征与转录组的变化。