Cellular Responses to UV Light in Ataxia Telangiectasia

共济失调毛细血管扩张症的细胞对紫外线的反应

基本信息

批准号：
7216181
负责人：
Kathleen Dixon
金额：
$ 31.37万
依托单位：
UNIVERSITY OF ARIZONA
依托单位国家：
美国
项目类别：
财政年份：
1997
资助国家：
美国
起止时间：
1997-01-01 至 2009-04-30
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): We propose to investigate the functional significance of DNA-damage-induced, ATM/ATR-dependent phosphoryIation of the major cellular single-stranded DNA-binding protein, RPA. Since RPA is essential for DNA replication and DNA repair, phosphorylation-dependent alteration in RPA function has the potential of altering the cellular responses to DNA damage. Indeed, we have demonstrated that DNA damage-induced ATM-dependent phosphorylation of RPA occurs, and it changes RPA function in vitro. Specifically, RPA phosphorylation alters DNA binding activity and protein/protein interactions. We postulate that a failure to phosphorylate RPA and other proteins required for DNA replication and DNA repair in ATM mutants compromises the cellular switch from DNA replication to DNA repair that would normally protect cells from the deleterious consequences of DNA damage. We propose that these abnormal responses in A-T lead to the enhanced cell death and genetic instability associated with the disease. We propose to test the hypothesis that the failure to phosphorvlate RPA (and other DNA repair proteins) in response to DNA damage compromises the DNA repair capacity of A-T ceils. The specific aims of this project are: 1. To determine the sites of ATM- and ATR-mediated RPA phosphorylation and construct site-specific mutants to test their role in RPA function. 2. To determine the influence of RPA (and mutant RPA) phosphorylation on protein/protein interactions in vivo and in vitro. A newly-developed RPA phospho-specific antibody will greatly facilitate these studies. 3. To clarify the role of the ATM kinase activity vs. other ATM functions in DSB repair: We are concentrating on abnormal responses to DNA damage in A-T cells that may influence the DNA repair capacity of these cells and may contribute to the high cancer risk and neurodegeneration. The ultimate goal of this project is to improve our understanding of the mechanisms that cells use to maintain genomic stability. This improved understanding may ultimately lead to the development of strategies for preventing and/or treating diseases that are characterized by genomic instability and neurological defects.

描述（由申请人提供）：我们建议研究主要细胞单链 DNA 结合蛋白 RPA 的 DNA 损伤诱导的、ATM/ATR 依赖性磷酸化的功能意义。由于 RPA 对于 DNA 复制和 DNA 修复至关重要，因此 RPA 功能的磷酸化依赖性改变有可能改变细胞对 DNA 损伤的反应。事实上，我们已经证明 DNA 损伤诱导的 RPA 发生 ATM 依赖性磷酸化，并且它在体外改变了 RPA 功能。具体来说，RPA 磷酸化会改变 DNA 结合活性和蛋白质/蛋白质相互作用。我们假设，ATM 突变体中 DNA 复制和 DNA 修复所需的 RPA 和其他蛋白质磷酸化失败会损害细胞从 DNA 复制到 DNA 修复的转换，而 DNA 修复通常可以保护细胞免受 DNA 损伤的有害后果。我们认为 A-T 中的这些异常反应导致细胞死亡增加和与疾病相关的遗传不稳定性。我们建议检验以下假设：在 DNA 损伤反应中未能磷酸化 RPA（和其他 DNA 修复蛋白）会损害 A-T 细胞的 DNA 修复能力。该项目的具体目标是： 1. 确定ATM和ATR介导的RPA磷酸化位点并构建位点特异性突变体以测试其在RPA功能中的作用。 2. 确定RPA（和突变型RPA）磷酸化对体内和体外蛋白质/蛋白质相互作用的影响。新开发的 RPA 磷酸化特异性抗体将极大地促进这些研究。 3. 澄清 ATM 激酶活性与其他 ATM 功能在 DSB 修复中的作用：我们专注于 A-T 细胞对 DNA 损伤的异常反应，这可能会影响这些细胞的 DNA 修复能力，并可能导致高癌症风险和神经退行性变。该项目的最终目标是提高我们对细胞维持基因组稳定性的机制的理解。这种加深的理解可能最终导致开发预防和/或治疗以基因组不稳定和神经缺陷为特征的疾病的策略。