Joint Molecule Formation During Recombination

重组过程中联合分子的形成

• 批准号: 9258438
• 负责人: NEIL HUNTER
• 金额: $ 29.9万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-05-01 至 2018-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9258438
• 关键词: Address; Biological; Biological Assay; Chemicals; Chromosome Pairing; Chromosome Segregation; Chromosomes; Complex; Complication; DNA; DNA Repair; DNA biosynthesis; Defect; Disease; Down Syndrome; Drosophila polo protein; Electron Microscopy; Enzymes; Family; Genetic Crossing Over; Genetic Recombination; Genome Stability; Goals; Hereditary Disease; Homologous Gene; Human; Infertility; Investigation; Joints; Link; Malignant Neoplasms; Mass Spectrum Analysis; Mediating; Meiosis; Meiotic Recombination; Mismatch Repair; Monitor; One-Step dentin bonding system; Organism; Pathologic; Phosphorylation; Phosphorylation Site; Phosphotransferases; Post-Translational Protein Processing; Pregnancy; Process; Proteins; Reaction; Regulation; Reproduction; Resolution; Role; Spontaneous abortion; Structure; Time; helicase; homologous recombination; in vivo; insight; joint formation; molecular phenotype; mutant; public health relevance; repaired; segregation; ubiquitin-protein ligase

DESCRIPTION (provided by applicant): SUMMARY Homologous recombination is a template-dependent DNA repair process that underpins DNA replication and genome stability, and is essential for chromosome segregation during meiosis. The fundamental recombination reaction is the formation of Joint Molecule (JM) intermediates via homologous pairing and DNA strand-exchange between a broken chromosome end and a homologous template. JMs are matured and resolved in distinct ways to produce products with or without an associated crossover. This proposal will investigate the in vivo roles of factors that regulate the formation and resolution of JMs during meiosis. The general hypothesis is that specialized JM processing factors are subject to distinct spatial and temporal regulation. Assigning in vivo functions to specific JM processing factors and understanding how they interact during recombination remain challenging issues, which are complicated by overlapping/compensatory activities and the need for specialized assays to monitor molecular phenotypes. DNA assays to monitor the chemical steps of recombination in vivo will form the cornerstone of this investigation. There are three distinct aims: AIM1. To investigate the regulation of pro-crossover factors by post-translational modification. We have identified phosphorylation sites on three meiosis-specific factors that stabilize JMs and promote crossing-over. These factors are the DNA helicase, Mer3, the MutS� complex related to DNA mismatch repair factors, and a SUMO E3 ligase, Zip3. In this, aim our goal is to understand the functional consequences of these phosphorylations and identify the responsible kinases. AIM2. To characterize factors that promote crossover-specific resolution of JMs. We have identified factors that promote biased resolution of JMs into crossovers. In this aim, we will further characterize these factors in order to: (i) identify activities and interactions required for crossng-over; (ii) demonstrate that specific factors act at the time of JM resolution; (iii) understand how polo-kinase, Cdc5, activates crossover-specific resolution. AIM3. To understand how the Smc5/6 complex regulates processing of JMs during meiosis. We have shown that the SMC-family complex, Smc5/6 regulates both the formation and resolution of JMs. We will address the idea that smc5/6 mutants form pathological JM structures that cannot be processed by resolving enzymes. We will also investigate the general theme that the temporally distinct roles of the Smc5/6 complex in JM formation and JM resolution are mediated through regulation of distinct partner proteins, with a focus on the DNA helicases Mph1 and Sgs1. Results obtained from these studies will be broadly relevant for understanding meiotic recombination in all organisms and will be especially important for discerning meiotic processes that make crossing-over and homolog disjunction incredibly accurate processes.

描述（由申请人提供）： 同源重组是一种模板依赖性DNA修复过程，其支持DNA复制和基因组稳定性，并且是减数分裂期间染色体分离所必需的。基本的重组反应是通过断裂的染色体末端和同源模板之间的同源配对和DNA链交换形成接合分子（JM）中间体。JM以不同的方式成熟和解决，以生产具有或不具有相关交叉的产品。这项建议将调查在减数分裂过程中，调节JM的形成和解决的因素在体内的作用。一般的假设是，专门的JM加工因素受到不同的空间和时间调节。研究特定JM加工因子的体内功能并了解它们在重组过程中如何相互作用仍然是具有挑战性的问题，这些问题由于重叠/补偿活性以及需要专门的测定来监测分子表型而变得复杂。监测体内重组的化学步骤的DNA测定将构成本研究的基石。有三个不同的目标：目标1。探讨翻译后修饰对前交换因子的调控作用。我们已经确定了三个减数分裂特异性因子的磷酸化位点，这些因子稳定JM并促进交换。这些因子是DNA解旋酶Mer 3、与DNA错配修复因子相关的MutS '复合物以及SUMO E3连接酶Zip 3。在这方面，我们的目标是了解这些磷酸化的功能后果，并确定负责激酶。 目标2.表征促进JM交叉特异性分离度的因素。我们已经确定了促进JM到交叉的有偏分辨率的因素。为此，我们将进一步描述这些因素，以便：（i）确定交叉所需的活动和相互作用;（ii）证明特定因素在JM解决时起作用;（iii）了解如何 polo激酶Cdc 5激活交叉特异性分辨。 AIM3.了解Smc 5/6复合体在减数分裂过程中如何调节JM的加工。我们已经表明，SMC家族复合物，Smc 5/6调节JM的形成和解决。我们将解决的想法，smc 5/6突变体形成病理JM结构，不能通过分解酶处理。我们还将调查的一般主题，时间不同的Smc 5/6复合物在JM形成和JM决议的作用是通过调节不同的合作伙伴蛋白介导的，重点是DNA解旋酶Mph 1和SGS 1。 从这些研究中获得的结果将是广泛相关的了解减数分裂重组在所有生物体，将是特别重要的识别减数分裂过程，使交换和同源分离令人难以置信的准确过程。