Molecular Modulator of RPA and RAD51 in Maintaining Genome Stability

RPA 和 RAD51 维持基因组稳定性的分子调节剂

• 批准号: 10055860
• 负责人: Weihang Chai
• 金额: $ 24.51万
• 依托单位: LOYOLA UNIVERSITY CHICAGO
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-12-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10055860
• 关键词: Molecular; Modulator; RPA; RAD51; Maintaining

DNA damage constitutes a major threat to genetic integrity, and has thus been implicated in the pathogenesis of cancer. Elucidating the mechanism safeguarding genome stability is important for understanding the mechanism underlying carcinogenesis. Genome integrity is constantly threatened by endogenous and exogenous agents arising from cellular metabolic processes as well as environmental exposure, many of which impede normal DNA replication and cause replication fork stalling. Stalled forks need to be properly repaired and rescued to prevent DNA lesions and genome instabilities that contribute to tumorigenesis. To repair and rescue stalled replication, a network of proteins regulating DNA damage response, DNA repair, replication, and cell cycle checkpoints are activated in response to stalled replication in order to stabilize and restart stalled forks. However, the mechanism underlying fork repair is poorly understood. Recent studies have revealed that the balance and dynamics of RAD51 and RPA at stalled forks are crucial for fork stabilization and restart. Yet how RAD51 and RPA activities are modulated remains largely elusive. Our recent findings suggest that the high-affinity single-stranded DNA binding protein complex known as CST may be a new modulator for RPA and RAD51 at GC-rich repetitive sequences in response to replication stress. The objective of this proposal is to understand the molecular relationship between CST, RAD51 and RPA in fork rescue, with the goal to provide novel insights into how cells counteract DNA damage caused by genotoxins. We propose to integrate advanced biochemical, cell biological, cell imaging, and next-gen sequencing techniques to examine how CST may regulate RPA binding at stalled sites and maintain fork progression and stability under stress (Aim 1), define how CST may modulate RAD51 activity at GC-rich stalled sites (Aim 2), and characterize the regulatory role of a surface-exposed region of CST in regulating RAD51 and RPA activities (Aim 3). Findings from the proposed research will provide novel information on the mechanism of genome stability maintenance of rescuing stalled replication and preserving genome stability.

DNA损伤是对遗传完整性的主要威胁，因此与 癌症的发病机制。阐明维护基因组稳定性的机制对于 了解致癌的机制。基因组的完整性不断受到威胁 来自细胞代谢过程和环境的内源和外源因子 暴露，其中许多阻碍了正常的DNA复制，并导致复制分叉停滞。失速的叉子需要 得到适当的修复和挽救，以防止DNA损伤和基因组不稳定 肿瘤发生学。为了修复和挽救停滞的复制，一个调节DNA损伤的蛋白质网络 响应、DNA修复、复制和细胞周期检查点被激活以响应停滞的复制 以稳定并重新启动失速的叉子。然而，人们对叉子修复的机制知之甚少。 最近的研究表明，RAD51和RPA在失速分叉时的平衡和动态对于 叉子稳定并重新启动。然而，RAD51和RPA活性是如何调节的在很大程度上仍然难以捉摸。我们的 最近的发现表明，被称为CST的高亲和力单链DNA结合蛋白复合体可能 作为一种新的RPA和RAD51的调节剂，在富含GC的重复序列上响应复制压力。这个 本建议的目的是了解CST、RAD51和RPA在叉树中的分子关系 救援，目的是为细胞如何中和遗传毒素造成的DNA损伤提供新的见解。 我们建议将先进的生化、细胞生物学、细胞成像和下一代测序 检查CST如何调节停滞部位的RPA结合并维持分叉进程和 压力下的稳定性(目标1)，定义CST如何调节富含GC的停滞部位的RAD51活性(目标2)， 并表征了CST表面暴露区域在调节RAD51和RPA中的调节作用 活动(目标3)。这项拟议研究的结果将提供关于心力衰竭机制的新信息。 维持基因组稳定性，挽救停滞的复制，保持基因组的稳定性。