Meiotic Interactions of the RecA Homologue Dmc1

描述（申请人提供）：同源重组在减数分裂中染色体的还原分离中起着关键作用。减数分裂重组是由DNA双链断裂（DSBs）的程序化诱导引发的，涉及有丝分裂细胞中DSBs的重组修复机制。减数分裂重组和有丝分裂重组的关键区别在于减数分裂过程必须发生在同源染色单体之间，而有丝分裂过程通常发生在姐妹染色单体之间。重组的核心步骤是同源链的侵入和交换。这一过程由重组酶催化，重组酶在结构和功能上与细菌链交换蛋白RecA相关。在大多数真核生物中存在两种reca样重组酶，包括芽殖酵母和人类；Rad51是唯一参与有丝分裂重组的reca样重组酶。Dmc1是一种减数分裂特异性重组酶，可以在没有Rad51的情况下发挥作用，但通常与Rad51合作。为了促进链入侵，重组酶必须首先在单链DNA （ssDNA）束上聚合成螺旋状细丝，这些单链DNA束形成于dsb位点。辅助因子，称为介质，允许重组酶在单链DNA结合蛋白形成细丝时取代它们。最近的证据表明，第二种辅助因子消耗能量，通过ATP水解依赖的DNA转位酶活性来促进重组酶与DNA的分离。提出的工作旨在阐明芽殖酵母的减数分裂重组。本建议的目的是：1。利用高分辨率光学显微镜观察减数分裂重组体的结构，目的是确定先前检测到的并排Rad51和Dmc1结构是否具有相对于参与重组事件的两条染色体轴的特定组织。2. 目的：探讨DNA易位Tid1/Rdh54和Rad54之间功能差异的机制。3. 确定Dmc1的辅助因子相互作用刺激重组的机制。减数分裂重组缺陷导致染色体不分离和丢失，两者都导致出生缺陷和自然流产。有丝分裂中重组修复的失败与乳腺癌和其他恶性肿瘤的病因有关。

DESCRIPTION (provided by applicant): Homologous recombination plays a critical role in reductional segregation of chromosomes in meiosis. Meiotic recombination is initiated by the programmed induction of DNA double strand breaks (DSBs) and involves a mechanism related to recombinational repair of DSBs in mitotic cells. The key difference between meiotic and mitotic recombination is that the meiotic process must occur between homologous chromatids and the mitotic process usually occurs between sister chromatids. The central step of recombination is homologous strand invasion and exchange. This process is catalyzed by recombinases that are structurally and functionally related to the bacterial strand exchange protein, RecA. There are two RecA-like recombinases in most eukaryotic organisms, including budding yeast and humans; Rad51 is the only RecA-like recombinase involved in mitotic recombination. Dmc1 is a meiosis-specific recombinase that can function in the absence of Rad51, but often cooperates with it. In order to promote strand invasion, recombinases must first polymerize into helical filaments on the tracts of single strand DNA (ssDNA) that form at sites of DSBs. Accessory factors, called mediators, allow recombinase to displace single strand DNA binding proteins as they form filaments. Recent evidence indicates that a second type of accessory factor expends energy to promote dissociation of recombinases from DNA using ATP hydrolysis dependent DNA-translocase activity. The proposed work seeks to elucidate meiotic recombination in budding yeast. The aims of this proposal are: 1. To visualize the architecture of the meiotic recombinosome using high-resolution light microscopy with the aim of determining if the previously detected side-by-side Rad51 and Dmc1 structures have a specific organization relative to the axes of the two chromosomes engaged in a recombination event. 2. To determine the mechanism underlying the functional differences between the DNA translocases Tid1/Rdh54 and Rad54. 3. To determine the mechanisms through which Dmc1's accessory factors interact to stimulate recombination. Defects in meiotic recombination cause chromosome non-disjunction and loss, both of which lead to birth defects and spontaneous abortion. Failure of recombinational repair in mitosis is implicated in the etiology of breast cancer and other malignancies.