Live imaging of induced and controlled DNA double-strand break formation reveals extremely low repair by homologous recombination in human cells
Live imaging of induced and controlled DNA double-strand break formation reveals extremely low repair by homologous recombination in human cells
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DOI：
10.1038/onc.2011.516
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发表时间：
2012-07-01
期刊：
影响因子：
8
通讯作者：
Goldberg, M.
中科院分区：
文献类型：
--
作者：
Shahar, O. D.;Ram, E. V. S. Raghu;Goldberg, M.
DNA双链断裂（DSBs）是最危险的DNA损伤，可能导致基因组不稳定，这是癌细胞的标志。DSB的主要修复途径为非同源末端连接（NHEJ）和同源重组（HR）。在哺乳动物细胞中，NHEJ占主导地位，它会导致不准确的修复。HR修复（HRR）被认为是准确的，并且仅限于细胞周期的S、G2和M期。尽管它很重要，但有关人权的许多方面仍然未知。在这里，我们开发了一种新的可诱导开/关开关细胞系统，首次能够在人类细胞中以快速和可逆的方式诱导DSB。通过限制DSB诱导的持续时间，我们发现非持续性核酸内切酶诱导的DSB很少被HR修复，而持续性DSB导致已公布的HRR频率（非显著HR频率与相似的频率分别为10%）。我们证明，这些dsb通过精确的修复机制进行修复，这与HRR（最有可能是无错误的NHEJ）不同。值得注意的是，我们的数据显示，在人类细胞中，内切酶诱导的DSB的HRR频率比以前用流行方法估计的低10倍，这导致了反复形成DSB。我们的研究结果表明，与NHEJ经常修复的简单病变相比，HRR主要在修复具有挑战性的dsb中起作用。阻止HR修复复杂和重复的人类基因组中的dsb可能对维持基因组稳定性具有重要作用。中华肿瘤杂志，2012,31 (3):394 - 394；doi: 10.1038 / onc.2011.516;2011年11月21日在线发布
DNA double-strand breaks (DSBs), the most hazardous DNA lesions, may result in genomic instability, a hallmark of cancer cells. The main DSB repair pathways are non-homologous end joining (NHEJ) and homologous recombination (HR). In mammalian cells, NHEJ, which can lead to inaccurate repair, predominates. HR repair (HRR) is considered accurate and is restricted to S, G2 and M phases of the cell cycle. Despite its importance, many aspects regarding HRR remain unknown. Here, we developed a novel inducible on/off switch cell system that enables, for the first time, to induce a DSB in a rapid and reversible manner in human cells. By limiting the duration of DSB induction, we found that non-persistent endonuclease-induced DSBs are rarely repaired by HR, whereas persistent DSBs result in the published HRR frequencies (non-significant HR frequency versus frequency of similar to 10%, respectively). We demonstrate that these DSBs are repaired by an accurate repair mechanism, which is distinguished from HRR (most likely, error-free NHEJ). Notably, our data reveal that HRR frequencies of endonuclease-induced DSBs in human cells are >10-fold lower than what was previously estimated by prevailing methods, which resulted in recurrent DSB formation. Our findings suggest a role for HRR mainly in repairing challenging DSBs, in contrast to uncomplicated lesions that are frequently repaired by NHEJ. Preventing HR from repairing DSBs in the complex and repetitive human genome probably has an essential role in maintaining genomic stability. Oncogene (2012) 31, 3495-3504; doi:10.1038/onc.2011.516; published online 21 November 2011