Guangdong Key Laboratory of Genome Instability and Human Disease Prevention, Department of Biochemistry and Molecular Biology, Shenzhen University School of Medicine, 518055 Shenzhen, China.
Key Laboratory of Carcinogenesis and Translational Research, Ministry of Education, Department of Biochemistry and Molecular Biology, Peking University Health Science Center, 100191 Beijing, China.
Proc Natl Acad Sci U S A. 2021 Jun 8;118(23). doi: 10.1073/pnas.2011278118.
Ataxia telangiectasia and Rad3 related (ATR) activation after replication stress involves a cascade of reactions, including replication protein A (RPA) complex loading onto single-stranded DNA and ATR activator loading onto chromatin. The contribution of histone modifications to ATR activation, however, is unclear. Here, we report that H3K14 trimethylation responds to replication stress by enhancing ATR activation. First, we confirmed that H3K14 monomethylation, dimethylation, and trimethylation all exist in mammalian cells, and that both SUV39H1 and SETD2 methyltransferases can catalyze H3K14 trimethylation in vivo and in vitro. Interestingly, SETD2-mediated H3K14 trimethylation markedly increases in response to replication stress induced with hydroxyurea, a replication stress inducer. Under these conditions, SETD2-mediated H3K14me3 recruited the RPA complex to chromatin via a direct interaction with RPA70. The increase in H3K14me3 levels was abolished, and RPA loading was attenuated when SETD2 was depleted or H3K14 was mutated. Rather, the cells were sensitive to replication stress such that the replication forks failed to restart, and cell-cycle progression was delayed. These findings help us understand how H3K14 trimethylation links replication stress with ATR activation.
共济失调毛细血管扩张症和 Rad3 相关（ATR）在复制应激后被激活涉及一系列反应，包括复制蛋白 A（RPA）复合物加载到单链 DNA 上以及 ATR 激活剂加载到染色质上。然而，组蛋白修饰对 ATR 激活的贡献尚不清楚。在这里，我们报告 H3K14 三甲基化通过增强 ATR 激活来响应复制应激。首先，我们证实 H3K14 单甲基化、二甲基化和三甲基化都存在于哺乳动物细胞中，并且 SUV39H1 和 SETD2 甲基转移酶都可以在体内和体外催化 H3K14 三甲基化。有趣的是，SETD2 介导的 H3K14 三甲基化在羟脲诱导的复制应激下显著增加，羟脲是一种复制应激诱导剂。在这些条件下，SETD2 介导的 H3K14me3 通过与 RPA70 的直接相互作用将 RPA 复合物募集到染色质上。当 SETD2 耗尽或 H3K14 发生突变时，H3K14me3 水平的增加被消除，并且 RPA 加载减弱。相反，细胞对复制应激敏感，使得复制叉无法重新启动，细胞周期进程被延迟。这些发现有助于我们理解 H3K14 三甲基化如何将复制应激与 ATR 激活联系起来。
