MOLECULAR MECHANISMS OF DNA REPAIR AND RECOMBINATION IN YEAST

我们正在研究与此相关的独特的染色体代谢事件 与减数分裂和染色体修复接触DNA后 破坏剂。 许多修复DNA所必需的基因 链断裂（DSB）是成功完成 减数分裂周期 已经鉴定出两种蛋白质/酶， 在减数分裂和/或修复中起作用; Mg（2+）依赖性核酸酶（RhoNUC， 以前称为yNucR的蛋白质，以及能够进行免疫调节的蛋白质。 链交换反应（SEP）。 这两种蛋白质的活性似乎 在RAD 52基因的控制下，RAD 52基因是修复 辐射诱导的DSB和减数分裂的完成。 探讨 RAD 52的作用，我们已经创建了含有染色体居民的菌株 在控制下缺失质粒上的RAD 52和RAD 52基因 酵母GAL 1启动子。 这使我们能够严格控制 RAD 52基因 我们发现，对于DSB的修复， 在用电离辐射治疗之前需要RAD 52蛋白。 有趣的是，RhoNUC蛋白的表达并不需要免疫调节。 功能性RAD 52基因 这表明RAD 52对RhoNUCH的控制是 而不是在转录或翻译水平上。 以便识别 其他可能与RAD 52蛋白相互作用的蛋白质，我们已经开始 筛选“蛋白质相互作用”文库。 这个系统可以让我们 鉴定与RAD 52物理相互作用的其他蛋白质。 RAD 52在DNA复制过程中的可能作用也正在研究中。 研究了 利用一个独特的系统，我们可以测量 “切除”置于酵母LYS 2基因中的细菌转座子Tn 5， 我们已经在编码DNA聚合酶I或III的基因中发现了突变， 导致高水平Tn 5“切除”。 未观察到DNA增加 聚合酶II突变体。 在polIII或polIII中观察到的“切除”水平升高， 当将RAD 52的无效突变体引入到pol I菌株中时， 这些菌株。 这表明所提出的酶之间的相互作用 负责DNA滞后链合成（POLIII和POLI）， RAD52。 目前，我们正在研究这种相互作用的性质。

We are investigating the unique chromosomal metabolic events associated with meiosis and the repair of chromosomes following exposure to DNA damaging agents. Many of the genes necessary for the repair of DNA double strand breaks (DSB) are required for the successful completion of the meiotic cycle. Two proteins/enzyme have been identified that appear to play a role in meiosis and/or repair; a Mg(2+) dependent nuclease (RhoNUC, previously referred to as yNucR, and a protein that is able to carry out a strand exchange reaction (SEP). Both of these protein activities appears to under the control of the RAD52 gene, a gene required for the repair of radiation-induced DSBs and the completion of meiosis. To investigate the role of RAD52, we have created strains containing a chromosomal resident deletion of the RAD52 and a RAD52 gene on a plasmid under the control of the yeast GALl promoter. This allows us to tightly regulate the expression of the RAD52 gene. We have found that for the repair of DSBs the presence of RAD52 protein is required prior to treatment with ionizing radiation. Interestingly, the expression of the RhoNUC protein does not require a functional RAD52 gene. This suggests the control of RhoNUCH by RAD52 is not at the transcriptional or translational level. In order to identify other proteins which may interact with the RAD52 protein, we have begun screening a "protein interaction" library. This system will allow us to identify other proteins which physically interact with RAD52. The possible role of RAD52 during DNA replication is also being investigated. Using a unique system whereby we measure the frequency of "excision" of a bacterial transposon Tn5 placed into the yeast LYS2 gene, we have found the mutations in the genes encoding DNA polymerase I or III lead to high levels of Tn5 "excision". No increase is observed in DNA polymerase II mutants. The elevated levels of "excision" seen in polIII or polI strains is reduced when a null mutant of RAD52 is introduced into these strains. This suggests an interaction between the enzymes proposed to be responsible for DNA lagging strand synthesis (POLIII and POLI) and RAD52. Currently we are investigating the nature of this interaction.