Lyndaker Amy M, Goldfarb Tamara, Alani Eric
Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York 14853-2703, USA.
Genetics. 2008 Aug;179(4):1807-21. doi: 10.1534/genetics.108.090654. Epub 2008 Jun 24.
Efficient repair of DNA double-strand breaks (DSBs) requires the coordination of checkpoint signaling and enzymatic repair functions. To study these processes during gene conversion at a single chromosomal break, we monitored mating-type switching in Saccharomyces cerevisiae strains defective in the Rad1-Rad10-Slx4 complex. Rad1-Rad10 is a structure-specific endonuclease that removes 3' nonhomologous single-stranded ends that are generated during many recombination events. Slx4 is a known target of the DNA damage response that forms a complex with Rad1-Rad10 and is critical for 3'-end processing during repair of DSBs by single-strand annealing. We found that mutants lacking an intact Rad1-Rad10-Slx4 complex displayed RAD9- and MAD2-dependent cell cycle delays and decreased viability during mating-type switching. In particular, these mutants exhibited a unique pattern of dead and switched daughter cells arising from the same DSB-containing cell. Furthermore, we observed that mutations in post-replicative lesion bypass factors (mms2Delta, mph1Delta) resulted in decreased viability during mating-type switching and conferred shorter cell cycle delays in rad1Delta mutants. We conclude that Rad1-Rad10-Slx4 promotes efficient repair during gene conversion events involving a single 3' nonhomologous tail and propose that the rad1Delta and slx4Delta mutant phenotypes result from inefficient repair of a lesion at the MAT locus that is bypassed by replication-mediated repair.
DNA双链断裂（DSB）的有效修复需要检查点信号传导和酶促修复功能的协调。为了在单个染色体断裂处的基因转换过程中研究这些过程，我们监测了缺乏Rad1-Rad10-Slx4复合物的酿酒酵母菌株中的交配型转换。Rad1-Rad10是一种结构特异性核酸内切酶，可去除在许多重组事件中产生的3'非同源单链末端。Slx4是DNA损伤反应的已知靶点，它与Rad1-Rad10形成复合物，并且对于通过单链退火修复DSB期间的3'末端加工至关重要。我们发现，缺乏完整Rad1-Rad10-Slx4复合物的突变体在交配型转换期间表现出RAD9和MAD2依赖性的细胞周期延迟以及活力下降。特别是，这些突变体表现出一种独特的模式，即来自同一个含DSB细胞的死亡和转换后的子细胞。此外，我们观察到复制后损伤旁路因子（mms2Delta，mph1Delta）中的突变导致交配型转换期间活力下降，并在rad1Delta突变体中导致更短的细胞周期延迟。我们得出结论，Rad1-Rad10-Slx4在涉及单个3'非同源尾巴的基因转换事件中促进有效修复，并提出rad1Delta和slx4Delta突变体表型是由于MAT位点处的损伤修复效率低下所致，该损伤被复制介导的修复绕过。
