State Key Laboratory of Genetic Engineering, Institute of Genetics, School of Life Sciences, Fudan University, Shanghai, 200433, China.
J Cell Sci. 2013 May 1;126(Pt 9):2042-51. doi: 10.1242/jcs.122945. Epub 2013 Mar 22.
Protein ubiquitination plays an important role in initiating the DNA damage response. Following DNA damage, E2 ubiquitin conjugating enzymes are crucial for catalyzing substrate ubiquitination that recruits downstream DNA repair factors to DNA lesions. To identify novel E2 conjugating enzymes important for initiating the DNA-damage-induced ubiquitination cascade, we screened most of the known E2 enzymes and found that RAD6A and RAD6B function together with RNF168 in the ionizing radiation (IR)-induced DNA damage response. Similarly to RNF168-deficient cells, RAD6A- or RAD6B-deficient cells exhibit a reduction in DNA-damage-induced protein ubiquitination. Correspondingly, DNA-damage-induced foci formation of DNA damage repair proteins, such as BRCA1 and 53BP1, is impaired in the absence of RAD6A or RAD6B. Moreover, the RNF168-RAD6 complex targeted histone H1.2 for ubiquitination in vitro and regulated DNA-damage-induced histone H1.2 ubiquitination in vivo. Collectively, these data demonstrate that RNF168, in complex with RAD6A or RAD6B, is activated in the DNA-damage-induced protein ubiquitination cascade.
蛋白质泛素化在启动 DNA 损伤反应中起着重要作用。在 DNA 损伤后，E2 泛素连接酶对于催化底物泛素化至关重要，该过程可以募集下游的 DNA 修复因子到 DNA 损伤部位。为了鉴定在起始 DNA 损伤诱导的泛素化级联反应中起重要作用的新的 E2 连接酶，我们筛选了大部分已知的 E2 酶，发现 RAD6A 和 RAD6B 与 RNF168 一起在电离辐射（IR）诱导的 DNA 损伤反应中发挥作用。与 RNF168 缺陷细胞相似，RAD6A 或 RAD6B 缺陷细胞中 DNA 损伤诱导的蛋白质泛素化减少。相应地，RAD6A 或 RAD6B 的缺失会损害 DNA 损伤修复蛋白如 BRCA1 和 53BP1 的 DNA 损伤诱导焦点形成。此外，RNF168-RAD6 复合物在体外将组蛋白 H1.2 靶向泛素化，并调节体内 DNA 损伤诱导的组蛋白 H1.2 泛素化。总之，这些数据表明，RNF168 与 RAD6A 或 RAD6B 形成复合物，在 DNA 损伤诱导的蛋白质泛素化级联反应中被激活。
