Bidirectional resection of DNA double-strand breaks by Mre11 and Exo1
Bidirectional resection of DNA double-strand breaks by Mre11 and Exo1
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DOI：
10.1038/nature10515
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发表时间：
2011-11-10
期刊：
影响因子：
64.8
通讯作者：
Neale, Matthew J.
中科院分区：
文献类型：
--
作者：
Garcia, Valerie;Phelps, Sarah E. L.;Neale, Matthew J.
通过同源重组修复DNA双链断裂(DSB)需要切除5‘末端以产生3’单链DNA尾巴(1)。该反应的关键成分是核酸外切酶1和双功能内切/外切核酸酶Mre11(参考文献2-4)。当DSB末端被结合蛋白(如DNA末端连接复合体(5)、拓扑异构酶(6)或减数分裂转酯酶Spo11)阻断时，Mre11内切酶活性是至关重要的，但Mre11 3‘-5’外切酶活性的特定功能仍不清楚。在这里，我们使用酿酒酵母来揭示Mre11外切酶在体内切除Spo11连接的5‘-DNA末端过程中的作用。我们表明，在Exo1突变细胞中观察到的残余切除依赖于Mre11，并且这两种核酸外切酶活性都是有效的DSB修复所必需的。以前的工作已经表明，切除是单向遍历的(1)。使用物理分析的组合进行5‘端加工，我们的结果表明了一种涉及双向切除的替代机制。首先，mre11从DSB的5‘末端切下了多达300个核苷酸--比之前假设的要远得多。其次，该缺口实现双向切除，在远离DSB的5‘-3’方向使用Exo1，在朝向DSB末端的3‘-5’方向使用Mre11。Mre11核酸外切酶的活性也使细胞对DNA损伤具有抵抗力，提示Mre11催化的切除可能是各种DNA修复途径的一个共同特征。
Repair of DNA double-strand breaks (DSBs) by homologous recombination requires resection of 5'-termini to generate 3'-single-strand DNA tails(1). Key components of this reaction are exonuclease 1 and the bifunctional endo/exonuclease, Mre11 (refs 2-4). Mre11 endonuclease activity is critical when DSB termini are blocked by bound protein-such as by the DNA end-joining complex(5), topoisomerases(6) or the meiotic transesterase Spo11 (refs 7-13)-but a specific function for the Mre11 3'-5' exonuclease activity has remained elusive. Here we use Saccharomyces cerevisiae to reveal a role for the Mre11 exonuclease during the resection of Spo11-linked 5'-DNA termini in vivo. We show that the residual resection observed in Exo1-mutant cells is dependent on Mre11, and that both exonuclease activities are required for efficient DSB repair. Previous work has indicated that resection traverses unidirectionally(1). Using a combination of physical assays for 5'-end processing, our results indicate an alternative mechanism involving bidirectional resection. First, Mre11 nicks the strand to be resected up to 300 nucleotides from the 5'-terminus of the DSB-much further away than previously assumed. Second, this nick enables resection in a bidirectional manner, using Exo1 in the 5'-3' direction away from the DSB, and Mre11 in the 3'-5' direction towards the DSB end. Mre11 exonuclease activity also confers resistance to DNA damage in cycling cells, suggesting that Mre11-catalysed resection may be a general feature of various DNA repair pathways.