A role for human homologous recombination factors in suppressing microhomology-mediated end joining.
A role for human homologous recombination factors in suppressing microhomology-mediated end joining.
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DOI：
10.1093/nar/gkw326
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发表时间：
2016-07-08
影响因子：
14.9
通讯作者：
Humphrey TC
中科院分区：
文献类型：
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作者：
Ahrabi S;Sarkar S;Pfister SX;Pirovano G;Higgins GS;Porter AC;Humphrey TC
DNA双链断裂(DSB)是一种毒性损伤，如果修复不当，可能导致细胞死亡或基因组不稳定。DSB修复通常通过经典的非同源末端连接(C-NHEJ)或同源重组(HR)途径来促进。然而，一种诱变的替代NHEJ途径，微同源介导的末端连接(MMEJ)，也可以部署。当MMEJ被C-NHEJ抑制时，HR和MMEJ之间的关系不是很清楚。在这里，我们描述了HR基因在抑制人类细胞MMEJ中的作用。通过使用敏感的HPRT方法监测DSB错误修复，我们发现HR蛋白，包括BRCA2，BRCA1或RPA的耗尽，导致了明显的突变特征，伴随着断裂诱导的突变频率、缺失长度和断裂位点上微同源短区域(2-6bp)的退火。这一特征依赖于CtIP、Mre11、POLQ和PARP，因此表明MMEJ。与CtIP或Mre11相比，BRCA1缺失导致部分切除和MMEJ增加，从而揭示了这些早期作用的HR因子之间的功能差异。综上所述，这些发现表明HR因素抑制了DSB切除后MMEJ的突变。
DNA double-strand breaks (DSBs) are toxic lesions, which if improperly repaired can result in cell death or genomic instability. DSB repair is usually facilitated by the classical non-homologous end joining (C-NHEJ), or homologous recombination (HR) pathways. However, a mutagenic alternative NHEJ pathway, microhomology-mediated end joining (MMEJ), can also be deployed. While MMEJ is suppressed by C-NHEJ, the relationship between HR and MMEJ is less clear. Here, we describe a role for HR genes in suppressing MMEJ in human cells. By monitoring DSB mis-repair using a sensitive HPRT assay, we found that depletion of HR proteins, including BRCA2, BRCA1 or RPA, resulted in a distinct mutational signature associated with significant increases in break-induced mutation frequencies, deletion lengths and the annealing of short regions of microhomology (2–6 bp) across the break-site. This signature was dependent on CtIP, MRE11, POLQ and PARP, and thus indicative of MMEJ. In contrast to CtIP or MRE11, depletion of BRCA1 resulted in increased partial resection and MMEJ, thus revealing a functional distinction between these early acting HR factors. Together these findings indicate that HR factors suppress mutagenic MMEJ following DSB resection.