Mechanisms of Homologous Recombination in Yeast

这项研究计划的长期目标是了解同源的分子机制， 在模式真核生物，芽殖酵母酿酒酵母中的重组。同源 重组（HR）在大多数生物的生命周期中起着两个重要的作用。需要修复 DNA中的致命损伤，如双链断裂，它是必不可少的配对和分离， 减数分裂时的同源染色体。这些职能的重要性体现在： 诱变，以及在不存在重组的情况下的有丝分裂和减数分裂非整倍性。由于许多人 癌症易感综合征与基因组不稳定性增加有关， 重组机制可能对理解这些疾病很重要。同源 重组起始于双链断裂处5 '-3'切除形成的单链DNA（ssDNA） （DSB），或在停止的复制分叉上。Rad 51催化ssDNA之间的突触和链交换 结合在Rad 51核蛋白丝和同源双链体DNA内，在需要RPA的反应中， Rad 52、Rad 54、Rad 55和Rad 57。Rad 51催化链的研究已经取得了很大进展 在某些情况下，切除核酸酶的身份和侵入后步骤所需的因子在很大程度上是不确定的。 未知我们已经开发了一种简单的基因检测单端链入侵，需要广泛的 DNA合成以完成重组修复。该试验将用于表征入侵后的 使用物理监测方法的人力资源的步骤，并确定涉及的复制因素。的作用 还将研究总染色体重排中的BIR。第二个目标是针对 切除核酸酶的鉴定。因为在断裂位点形成ssDNA被认为是 作为Rad 51促进链侵入的必要先决条件，令人惊讶的是， 基因编码切除核酸酶。这表明这一步是多余的，或者切除 核酸酶对于生存力是必需的。提出了几种遗传筛选来鉴定编码 核酸酶最后，我们展示了重复的高频R/AD 57独立组合， 序列;建议进行遗传研究以确定R/\D57独立重组的组分 通路

The long-term objective of this research program is to understand the molecular mechanisms of homologous recombination in a model eukaryote, the budding yeast Saccharomyces cerevisiae. Homologous recombination (HR) plays two essential roles during the life cycle of most organisms. It is required to repair lethal lesions in DMA, such as double-strand breaks, and it is essential for the pairing and segregation of homologous chromosomes during meiosis. The importance of these functions is evidenced by increased mutagenesis, and mitotic and meiotic aneuploidy in the absence of recombination. Since many human cancer-prone syndromes are associated with increased genome instability, an understanding of the mechanisms of recombination is likely to be important in understanding these diseases. Homologous recombination initiates at single-stranded DNA (ssDNA) formed by 5'-3' resection at double-strand breaks (DSBs), or at stalled replication forks. Rad51 catalyzes synapsis and strand exchange between ssDNA bound within the Rad51 nucleoprotein filament and homologous duplex DNA, in a reaction requiring RPA, Rad52, Rad54, Rad55 and Rad57. Much progress has been made in understanding Rad51-catalyzedstrand invasion, but the identity of the resection nuclease and factors required for the post-invasion step are largely unknown. We have developed a simple genetic assay for one-ended strand invasion that requires extensive DNA synthesis to complete recombinational repair. This assay will be used to characterize the post-invasion steps of HR using physical monitoring methods and to identify the replication factors involved. The role of BIR in gross chromosomal rearrangements will also be investigated. The second aim is directed at identification of the resection nuclease. Because formation of ssDNA at break sites is considered an essential prerequisite for Rad51 -promoted strand invasion, it is surprising that none of the RAD52 group genes encodes the resection nuclease. This suggests redundancy for this step, or that the resection nuclease is essential for viability. Several genetic screens are proposed to identify genes encoding nucleases. Finally, we have shown high frequency R/AD57-independentrecombination of repeated sequences; genetic studies are proposed to identify components of the R/\D57-independent recombination pathway.