Department of Biology, University of Copenhagen, DK-2200, Copenhagen N, Denmark.
DNA Repair (Amst). 2012 Nov 1;11(11):892-905. doi: 10.1016/j.dnarep.2012.08.005. Epub 2012 Sep 24.
The ubiquitylation cascade plays an important role in the recruitment of repair factors at DNA double-strand breaks. The involvement of a growing number of ubiquitin E3 ligases adds to the complexity of the DNA damage-induced ubiquitin signaling. Here we use the genetically tractable avian cell line DT40 to investigate the role of HERC2, RNF8 and RNF168 in the DNA damage-induced ubiquitylation pathway. We show that formation of ubiquitin foci as well as cell survival after DNA damage depends on both RNF8 and RNF168. However, we find that RNF8 and RNF168 knockout cell lines respond differently to treatment with camptothecin indicating that they do not function in a strictly linear manner. Surprisingly, we show that HERC2 is required neither for survival nor for ubiquitin foci formation after DNA damage in DT40. Moreover, the E3 ubiquitin ligase activity of HERC2 is not redundant to that of RNF8 or RNF168.
泛素化级联反应在 DNA 双链断裂处修复因子的招募中起着重要作用。越来越多的泛素 E3 连接酶的参与增加了 DNA 损伤诱导的泛素信号的复杂性。在这里，我们使用遗传上可操纵的禽类细胞系 DT40 来研究 HERC2、RNF8 和 RNF168 在 DNA 损伤诱导的泛素化途径中的作用。我们表明，泛素焦点的形成以及 DNA 损伤后的细胞存活都依赖于 RNF8 和 RNF168。然而，我们发现 RNF8 和 RNF168 敲除细胞系对喜树碱的处理反应不同，这表明它们不是以严格的线性方式发挥作用。令人惊讶的是，我们表明 HERC2 既不是 DT40 中 DNA 损伤后细胞存活所必需的，也不是形成泛素焦点所必需的。此外，HERC2 的 E3 泛素连接酶活性与 RNF8 或 RNF168 没有冗余。
