Dynamics of de novo heterochromatin assembly and disassembly at replication forks ensures fork stability

Chromatin is dynamically reorganized when DNA replication forks are challenged. However, the process of epigenetic reorganization and its implication for fork stability is poorly understood. Here, we discover a checkpoint regulated cascade of chromatin signaling that activates 5 the histone methyltransferase EHMT2/G9a to catalyze heterochromatin assembly at stressed replication forks. Using biochemical and single molecule chromatin fiber approaches, we show that G9a together with SUV39h1 induces chromatin compaction by accumulating the repressive modifications, H3K9me1/me2/me3, in the vicinity of stressed replication forks. This closed conformation is also favored by the G9a-dependent exclusion of the H3K9-demethylase 10 JMJD1A/KDM3A, which facilitates heterochromatin disassembly upon fork restart. Untimely heterochromatin disassembly from stressed forks by KDM3A enables PRIMPOL access, triggering ssDNA gap formation and sensitizing cells towards chemotherapeutic drugs. These findings may help explaining chemotherapy resistance and poor prognosis observed in cancer patients displaying elevated level of G9a/H3K9me3.

当DNA复制叉受到应激时，染色质会发生动态重塑。然而，表观遗传重塑的过程及其对复制叉稳定性的影响尚不明确。本研究发现一条受检验点调控的染色质信号级联反应，该通路可激活组蛋白甲基转移酶EHMT2（G9a），在受应激的复制叉处催化异染色质组装。研究人员通过生化技术与单分子染色质纤维实验手段证实，G9a与SUV39h1可通过在受应激的复制叉附近富集组蛋白H3赖氨酸9单甲基化（H3K9me1）、二甲基化（H3K9me2）与三甲基化（H3K9me3）这类抑制性修饰，诱导染色质紧缩。此外，G9a依赖性排除H3K9去甲基化酶JMJD1A（KDM3A）也可促进这一紧缩构象——而JMJD1A原本可在复制叉重启时促进异染色质解聚。若KDM3A过早介导受应激复制叉处的异染色质解聚，将使PRIMPOL得以结合，触发单链DNA间隙形成，并使细胞对化疗药物更加敏感。上述发现或可解释G9a/H3K9me3水平升高的癌症患者中观察到的化疗耐药性与不良预后。