PR-DUB preserves Polycomb repression by preventing excessive accumulation of H2Aub1, an antagonist of nucleosome stacking (ATAC-Seq)

The Polycomb repression machinery in Drosophila comprises PRC1 that monoubiquitinates histone H2A at lysine 118 (H2Aub1) and PR-DUB, a major H2Aub1 deubiquitinase, but how H2Aub1 levels must be balanced for Polycomb repression remains enigmatic. We show that H2Aub1 is enriched at Polycomb target genes in early embryos but depleted from these genes during developmental stages when PRC1 represses their transcription. Accordingly, Polycomb targets remain repressed in H2Aub1-deficient animals. In PR-DUB catalytic mutants, high-level H2Aub1 accumulation at Polycomb targets increases chromatin accessibility, consistent with disruption of chromatin fiber folding by H2Aub1 in vitro. Consequently, PR-DUB mutants show defective Polycomb repression, while general transcription is largely unperturbed by the genome-wide, low-level H2Aub1 increase. Changes in H2Aub1 levels alter H3K27 methylation-kinetics but PRC2 nevertheless generates canonical H3K27me3 domains in PRC1 or PR-DUB catalytic mutants. PR-DUB therefore acts as a rheostat that removes excessive H2Aub1 that, though deposited by PRC1, antagonizes PRC1-mediated chromatin compaction. Analysis of ATAC-seq profiles generated from wild-type and mutant Drosophila embryos.

果蝇（Drosophila）中的多梳抑制系统（Polycomb repression machinery）由PRC1与PR-DUB组成：PRC1可在组蛋白H2A的赖氨酸118位点介导单泛素化修饰（H2Aub1），而PR-DUB是主要的H2Aub1去泛素化酶。然而，多梳抑制过程中H2Aub1水平需如何维持平衡，这一问题至今仍未阐明。本研究发现，早期胚胎中H2Aub1在多梳靶基因区域富集，但在PRC1抑制这些基因转录的发育阶段，靶基因区域的H2Aub1会发生耗竭。相应地，在H2Aub1缺失的动物体内，多梳靶基因仍可维持抑制状态。在PR-DUB催化功能缺失突变体中，多梳靶基因区域的H2Aub1高水平积累会提升染色质可及性，这与体外实验中H2Aub1破坏染色质纤维折叠的结果相符。因此，PR-DUB突变体表现出多梳抑制功能缺陷，而全基因组范围内小幅升高的H2Aub1水平基本不会干扰常规转录过程。H2Aub1水平的变化会改变H3K27甲基化动力学，但即便在PRC1或PR-DUB催化突变体中，PRC2仍能生成经典的H3K27me3结构域。由此可见，PR-DUB发挥着类似变阻器的调控作用，可清除过量的H2Aub1：尽管这类修饰由PRC1介导沉积，但过量的H2Aub1会拮抗PRC1所介导的染色质压缩。本研究同时对野生型与突变体果蝇胚胎的转座酶可及性测序（ATAC-seq）图谱进行了分析。