MEIOTIC INTERACTIONS TO THE RECA HOMOLOGUE DMC1

减数分裂期间需要同源重组以促进同源染色体在第一次减数分裂时的准确分离。已鉴定出酵母属中减数分裂重组所需的几种蛋白质，但这些蛋白质在重组中的具体作用在很大程度上是未知的。 这项工作的长期目标是确定减数分裂重组的分子机制，以及细胞周期机制如何与其他减数分裂事件协调重组。 该项目的重点是一个减数分裂蛋白，Dmcl，这是需要减数分裂重组。 Dmc 1及其相关的Rad51与RecA蛋白具有结构和功能上的同源性。 RecA是大肠杆菌DNA损伤重组修复所需的中心蛋白。杆菌 酵母Rad51，可能还有Dmc1，共享RecAs促进两个分子之间链交换的能力。 Dmcl蛋白将被纯化和表征，以确定它与RecA共享什么活性。 重组产物的形成需要除了RecA所展示的那些之外的许多酶活性。 减数分裂重组所需的许多蛋白质形成多蛋白质复合物。 亲和层析和串联质谱将用于鉴定重组过程中与Dmcl形成复合物的蛋白质。 时间过程和突变分析将用于表征功能重组复合物的组装和拆卸的总体进展。 此外，共聚焦显微镜将被用来检查个人重组活细胞中使用绿色荧光蛋白的事件。 Rad51和Dmc 1的功能只是部分冗余。 我们提出了突变体筛选，旨在将Dmc 1的冗余功能与其独特的功能分开。然后，这些突变将被用来表征减数分裂重组的机制特征，将其与有丝分裂重组修复区分开来。 Dmc 1的功能由检查点控制机制监测，该机制确保在染色体分离发生之前完成重组。 我们最近发现，检查点控制所需的基因在突触和抑制异位重组中具有额外的功能。 这些新确定的功能和检查点控制之间的关系将被检查。

Homologous recombination is required during meiosis to promote accurate segregation of homologous chromosomes at the first meiotic division. Several proteins have been identified that re required for MEIOTIC RECOMBINATION in the YEAST Saccharomyces, but the specific role of these proteins in recombination is largely unknown. The long term goals of this work are to determine the molecular mechanism of meiotic recombination and how the cell cycle machinery coordinates recombination with other meiotic events. This project focuses on one meiotic protein, Dmcl, that is required for meiotic recombination. Dmc1 and its relative Rad51 share structural and functional homology to RecA protein. RecA is the central protein required fo recombinogenic REPAIR OF DNA DAMAGE in E. Coli. Yeast Rad51, and probably Dmc1 as well, share RecAs ability to promote strand exchange between two molecules. Dmcl protein will be purified and characterized to determine what activities it shares with RecA. The formation of recombination products requires numerous enzymatic activities in addition to those displayed by RecA. Many of the proteins required for meiotic recombination form a multi protein complex. Affinity chromatography and tandem mass spectroscopy will be used to identify proteins that form complexes with Dmcl during recombination. Time-course and mutational analyses will be used to characterize overall progress in the assembly and disassembly of functional recombination complexes. In addition, confocal microscopy will be used to examine individual recombination events in living cells using green fluorescent protein. The functions of Rad51 and Dmc1 are only partially redundant. We propose mutant screens designed to separate the redundant function of Dmc1 from its unique functions. These mutations will then be used to characterize the mechanistic features of meiotic recombination that distinguish it from mitotic recombinational repair. The function of Dmc1 is monitored by a checkpoint control mechanism that ensures recombination is complete before chromosome segregation occurs. We recently showed the genes required for checkpoint control have additional functions in synapsis and suppression of ectopic recombination. The relationship between these newly identified functions and checkpoint control will be examined.