Hubei Key Laboratory of Cell Homeostasis, the Department of Genetics, College of Life Sciences and the Institute for Advanced Studies, Wuhan University, Wuhan, Hubei 430072, China.
National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
Nucleic Acids Res. 2018 Nov 30;46(21):11326-11339. doi: 10.1093/nar/gky918.
Repair of DNA double-strand breaks (DSBs) requires eviction of the histones around DNA breaks to allow the loading of numerous repair and checkpoint proteins. However, the mechanism and regulation of this process remain poorly understood. Here, we show that histone H2B ubiquitination (uH2B) promotes histone eviction at DSBs independent of resection or ATP-dependent chromatin remodelers. Cells lacking uH2B or its E3 ubiquitin ligase Bre1 exhibit hyper-resection due to the loss of H3K79 methylation that recruits Rad9, a known negative regulator of resection. Unexpectedly, despite excessive single-strand DNA being produced, bre1Δ cells show defective RPA and Rad51 recruitment and impaired repair by homologous recombination and response to DNA damage. The HR defect in bre1Δ cells correlates with impaired histone loss at DSBs and can be largely rescued by depletion of CAF-1, a histone chaperone depositing histones H3-H4. Overexpression of Rad51 stimulates histone eviction and partially suppresses the recombination defects of bre1Δ mutant. Thus, we propose that Bre1 mediated-uH2B promotes DSB repair through facilitating histone eviction and subsequent loading of repair proteins.
DNA 双链断裂 (DSB) 的修复需要将 DNA 断裂周围的组蛋白驱逐出去，以允许众多修复和检查点蛋白的加载。然而，这个过程的机制和调控仍知之甚少。在这里，我们表明组蛋白 H2B 的泛素化 (uH2B) 促进了 DSB 处的组蛋白驱逐，这独立于核酸酶切割或 ATP 依赖性染色质重塑酶。缺乏 uH2B 或其 E3 泛素连接酶 Bre1 的细胞由于 H3K79 甲基化的丧失而导致过度核酸酶切割，H3K79 甲基化招募了 Rad9，Rad9 是核酸酶切割的已知负调控因子。出乎意料的是，尽管产生了大量的单链 DNA，但 bre1Δ 细胞显示出 RPA 和 Rad51 募集的缺陷以及同源重组修复和对 DNA 损伤的反应受损。bre1Δ 细胞中的 HR 缺陷与 DSB 处的组蛋白丢失缺陷相关，并且可以通过耗尽组蛋白伴侣 CAF-1（其沉积组蛋白 H3-H4）来很大程度上挽救。Rad51 的过表达刺激了组蛋白驱逐，并部分抑制了 bre1Δ 突变体的重组缺陷。因此，我们提出 Bre1 介导的 uH2B 通过促进组蛋白驱逐和随后修复蛋白的加载来促进 DSB 修复。
