S CEREVISIAE RAD54P IN HOMOLOGOUS DNA RECOMBINATION

RAD52上位性基团负责通过同源重组修复S.啤酒。 这一组至少包含9个基因：RAD50、RAD51、RAD52、RAD54、RAD55、RAD57、RAD59、MRE 11和XRS2（1，2）。 许多这些基因的突变导致对DNA损伤剂的敏感性增加，例如电离辐射或甲基甲磺酸盐，产生双链DNA断裂（3，4）。 此外，当细胞进行减数分裂时，需要同源重组（4，5），在该上位性组的许多成员中含有突变的菌株是不能存活的。 虽然在阐明T4噬菌体和E. coli中，我们目前对真核DNA重组的理解水平仍然有限。 本建议的重点是扩大我们的知识，这一过程中的S。啤酒。 目前，Rad51蛋白、Rad52蛋白和RPA的作用正由Kowalczykowski小组等进行研究。 我的目标是使用生物化学方法来确定Rad54蛋白在同源重组中的功能。 鉴于迄今为止的研究，我相信Rad54蛋白可能作为DNA染色质重塑因子或作为能够刺激同源重组的分支迁移阶段的酶发挥作用。 我计划使用噬菌体T4和E.大肠杆菌重组系统，以帮助指导初步研究。

The RAD52 epistasis group is responsible for repairing the majority of double-stranded DNA breaks via homologous recombination in S. cerevisiae. This group contains at least nine genes: RAD50, RAD51, RAD52, RAD54, RAD55, RAD57, RAD59, MRE11, and XRS2 (1,2). Mutations in many of these genes result in an increased sensitivity to DNA damaging agents, such as ionizing radiation or methylmethanesulfonate, that produce double stranded DNA breaks (3,4). In addition, strains containing mutations in many members of this epistasis group are inviable when the cells go through meiosis, where homologous recombination is required (4,5). While considerable progress has been made toward elucidating the mechanistic steps of and determining the enzymes responsible for homologous recombination in bacteriophage T4 and in E. coli, our current level of understanding of eukaryotic DNA recombination remains limited. The focus of this proposal is to expand our knowledge of this process in S. cerevisiae. Currently, the roles of Rad5l protein, Rad52 protein and RPA are being investigated by the Kowalczykowski group and others. My goal is to use biochemical methods to define the function of the Rad54 protein in homologous recombination. Given the studies to date, I believe that Rad54 protein may have a role either as a DNA chromatin remodeling factor, or as an enzyme capable of stimulating the branch migration phase of homologous recombination. I plan to examine the enzymatic characteristics of Rad54 protein using the bacteriophage T4 and E. coli recombination systems to help guide the initial studies.