PDF(Pubmed)
Abstract：
Homologous recombination (HR) is essential for the maintenance of genome stability. During HR, Replication Protein A (RPA) rapidly coats the 3\'-tailed single-strand DNA (ssDNA) generated by end resection. Then, the ssDNA-bound RPA must be timely replaced by Rad51 recombinase to form Rad51 nucleoprotein filaments that drive homology search and HR repair. How cells regulate Rad51 assembly dynamics and coordinate RPA and Rad51 actions to ensure proper HR remains poorly understood. Here, we identified that Rtt105, a Ty1 transposon regulator, acts to stimulate Rad51 assembly and orchestrate RPA and Rad51 actions during HR. We found that Rtt105 interacts with Rad51 in vitro and in vivo and restrains the adenosine 5\' triphosphate (ATP) hydrolysis activity of Rad51. We showed that Rtt105 directly stimulates dynamic Rad51-ssDNA assembly, strand exchange, and D-loop formation in vitro. Notably, we found that Rtt105 physically regulates the binding of Rad51 and RPA to ssDNA via different motifs and that both regulations are necessary and epistatic in promoting Rad51 nucleation, strand exchange, and HR repair. Consequently, disrupting either of the interactions impaired HR and conferred DNA damage sensitivity, underscoring the importance of Rtt105 in orchestrating the actions of Rad51 and RPA. Our work reveals additional layers of mechanisms regulating Rad51 filament dynamics and the coordination of HR.
摘要：
同源重组（HR）对于维持基因组稳定性至关重要。HR期间,复制蛋白A（RPA）快速包被末端切除产生的3'尾单链DNA（ssDNA）。然后，ssDNA结合的RPA必须及时用Rad51重组酶取代,以形成驱动同源性搜索和HR修复的Rad51核蛋白丝。细胞如何调节Rad51组装动力学并协调RPA和Rad51作用以确保适当的HR仍然知之甚少。这里,我们确定了Rtt105，一个Ty1转座子调节因子，在HR期间采取行动刺激Rad51组装并协调RPA和Rad51行动。我们发现Rtt105在体外和体内与Rad51相互作用，并抑制Rad51的腺苷5'三磷酸（ATP）水解活性。我们表明Rtt105直接刺激动态Rad51-ssDNA组装，链交换，和体外D环形成。值得注意的是,我们发现Rtt105通过不同的基序物理调节Rad51和RPA与ssDNA的结合，并且这两种调节在促进Rad51成核中是必要的和上位的，链交换，HR修复因此,破坏任何一种相互作用，损害HR并赋予DNA损伤敏感性，强调Rtt105在协调Rad51和RPA行动中的重要性。我们的工作揭示了调节Rad51细丝动力学和HR协调的其他机制层。
