Roles of Checkpoint and DNA Repair Proteins in Fission Yeast Telomere Maintenance

检查点和 DNA 修复蛋白在裂殖酵母端粒维持中的作用

• 批准号: 7134092
• 负责人: Toru Nakamura
• 金额: $ 27.12万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-08-01 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7134092
• 关键词: DNA binding protein; DNA damage; DNA repair; Schizosaccharomyces pombe; cell cycle proteins; chromatin immunoprecipitation; enzyme complex; flow cytometry; fungal genetics; fungal proteins; gene deletion mutation; genetic regulation; nucleic acid structure; protein protein interaction; protein structure function; southern blotting; telomere; temperature sensitive mutant; tissue /cell culture

DESCRIPTION (provided by applicant): Proper maintenance of telomere structure is crucial for stable inheritance of the genome. Various checkpoint and DNA repair proteins, including evolutionarily highly conserved checkpoint kinases Tel1 (ATM) and Rad3 (ATR), play important roles in stable maintenance of telomeres. However, no clear mechanistic roles for various checkpoint and DNA repair proteins in telomere maintenance have been established. Major goal of our research is to understand how checkpoint and DNA repair proteins contribute to telomere maintenance. The current proposal will utilize fission yeast Schizosaccharomyces pombe as a model system. Highly conserved DNA damage responses and telomere maintenance mechanisms between fission yeast and humans should be helpful in extrapolating our findings to build testable models for human cells. Deregulation of telomere maintenance mechanisms has been found to be a key event in tumorigenesis, thus mechanistic insights on how various proteins collaborate to generate functional telomeres are first needed to devise effective methods for preventing tumorigenesis. In order to be stably maintained, telomeres must fulfill two major functions. First, telomeres must protect telomeric DNA ends from fusions and degradation. Second, telomeres must provide access to telomerase to prevent loss of telomeric DNA after DNA replication. Thus, telomeres must undergo dynamic switches from the highly protected state to the more accessible state that allows recruitment of telomerase. We hypothesize that checkpoint and DNA repair proteins are recruited to telomeres in a cell cycle-regulated manner to trigger appropriate changes in telomere structure and telomere protein composition. This model will be directly tested in Aim 1. Additionally, newly developed fission yeast reporter strains will be utilized to search for potential telomere targets of Rad3 and Tel1 kinases in Aim 2. Our preliminary studies with MRN (Mre11-Rad50-Nbs1) complex mutants have suggested that recruitment of Tel1 to telomeres does not require Tel1-MRN interaction unlike recruitment of Tel1 to other DNA breaks. Therefore, attempts will be made in Aim 3 to understand the mechanistic basis for the MRN-independent recruitment of Tel1 to telomeres.

描述(由申请人提供)：适当地维持端粒结构对于基因组的稳定遗传至关重要。各种检查点和DNA修复蛋白，包括进化上高度保守的检查点激酶Tel1(ATM)和Rad3(ATR)，在端粒的稳定维持中发挥着重要作用。然而，各种检查点和DNA修复蛋白在端粒维持中的作用机制尚不明确。我们研究的主要目标是了解检查点和DNA修复蛋白是如何促进端粒维持的。目前的提议将利用裂殖酵母庞贝裂殖酵母作为模型系统。分裂酵母和人类之间高度保守的DNA损伤反应和端粒维持机制应该有助于推断我们的发现，为人类细胞建立可测试的模型。端粒维持机制的失控已被发现是肿瘤发生中的一个关键事件，因此首先需要对各种蛋白质如何协作产生功能端粒的机械性见解来设计有效的方法来预防肿瘤的发生。为了保持稳定，端粒必须完成两个主要功能。首先，端粒必须保护端粒DNA末端免受融合和降解。其次，端粒必须提供与端粒酶的通路，以防止DNA复制后端粒DNA的丢失。因此，端粒必须经历从高度保护状态到更容易接近的状态的动态切换，才能重新招募端粒酶。我们假设检查点和DNA修复蛋白以细胞周期调节的方式招募到端粒，以触发端粒结构和端粒蛋白组成的适当变化。该模型将直接在AIM 1中进行测试。此外，新开发的裂变酵母报告菌株将被用来寻找AIM 2中Rad3和Tel1激酶的潜在端粒靶点。我们对MRN(Mre11-Rad50-Nbs1)复合突变体的初步研究表明，与Tel1与其他DNA断裂的招募不同，Tel1招募到端粒不需要Tel1-MRN相互作用。因此，在目标3中，将试图了解MRN非依赖性地将Tel1招募到端粒的机制基础。