Repair of Oxidatively Damaged Guanines in Human

人体氧化损伤鸟嘌呤的修复

• 批准号: 6929941
• 负责人: A-Lien L Lu-Chang
• 金额: $ 27.92万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-08-14 至 2008-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6929941
• 关键词: DNA damage; DNA repair; HeLa cells; SDS polyacrylamide gel electrophoresis; Schizosaccharomyces pombe; aging; chemical carcinogen; chemical carcinogenesis; endonuclease; enzyme activity; flow cytometry; guanine; immunoprecipitation; molecular oncology; mutagens; oxidative stress; phosphorylation; proliferating cell nuclear antigen; protein protein interaction; tissue /cell culture; western blottings

DESCRIPTION (provided by applicant): Defects in DNA repair can lead to genome instability, a hallmark of cancer. Oxidative DNA damage is a major source of mutation load in living organisms and plays a role in carcinogenesis and aging. The 8-oxo-7,8-dihydrodeoxyguanine (8-oxoG) lesion is a major stable product of oxidative damage and has the most deleterious effects because it can mispair with adenine during DNA replication. Thus, repair ol A/8-oxoG and C/8-oxoG by hMYH (MutY adenine glycosylase homolog), hOGG1 (an 8-oxoG glycosylase), and hMSH2/hMSH6 dependent mismatch repair pathways provide levels of defense against oxidative stress The overall goals of this project is to understand the role of hMYH pathway in controlling carcinogenesis and its interplay with other repair pathways in response to oxidative stress such as treatments by hydrogen peroxide and ionizing radiation. The interactions of hMYH with replication proteins hPCNA and hRPA, with repai] enzymes hAPE1 and hMSH6, as well as cell cycle checkpoint proteins hHusl/hRadl/hRad9 will be investigated. (I) Alterations in the interactions of hMYH with hPCNA and hHusl under oxidative stress will be investigated. The interacting domains on hMYH and hHusl will be mapped and the significance of their functional interaction will be tested in vivo. Working models that a molecular switch of hMYH-hPCNA tc hMYH-hRad9/hRadl/hHusl occurs to induce DNA damage response and that hMYH may recruit_ hRad9/hRadl/hHusl to the lesion sites will be tested. (II) Interaction between hMYH and hMSH6 will be elucidated both in vitro and in vivo. Fission yeast S. pombe will be used as a model system to test the in viw function of hMYH mutants. (III) Co-localization of hMYH with hAPE 1, hPCNA, hHus 1, hRPA, and hMSH6 ir repair and replication loci will be examined. (IV) The phosphorylation state of hMYH after DNA damage, the kinase responsible for the hMYH phosphorylation, and the phosphorylation site(s) of hMYH will be determined. The correlation of hMYH activities with protein phosphorylation or certain protein-protein interaction(s) after DNA damage will be examined. This study will advance our understanding of the role of DNA repair in tumor susceptibility.

描述（由申请人提供）：DNA 修复缺陷可能导致基因组不稳定，这是癌症的一个标志。 DNA 氧化损伤是生物体突变负荷的主要来源，在致癌和衰老中发挥着重要作用。 8-oxo-7,8-二氢脱氧鸟嘌呤 (8-oxoG) 损伤是氧化损伤的主要稳定产物，具有最有害的影响，因为它可以在 DNA 复制过程中与腺嘌呤错配。因此，通过 hMYH（MutY 腺嘌呤糖基化酶同源物）、hOGG1（8-oxoG 糖基化酶）和 hMSH2/hMSH6 依赖性错配修复途径修复 A/8-oxoG 和 C/8-oxoG 提供了针对氧化应激的防御水平。该项目的目标是了解 hMYH 通路在控制癌发生中的作用及其与其他修复通路响应氧化反应的相互作用压力，例如过氧化氢和电离辐射治疗。将研究hMYH与复制蛋白hPCNA和hRPA、修复酶hAPE1和hMSH6以及细胞周期检查点蛋白hHus1/hRad1/hRad9的相互作用。 (I) 将研究氧化应激下 hMYH 与 hPCNA 和 hHusl 相互作用的变化。 hMYH 和 hHusl 上的相互作用结构域将被绘制，并且它们的功能相互作用的重要性将在体内进行测试。将测试hMYH-hPCNA tc hMYH-hRad9/hRad1/hHusl发生分子开关以诱导DNA损伤反应以及hMYH可能将hRad9/hRad1/hHusl募集至损伤位点的工作模型。 (II) hMYH 和 hMSH6 之间的相互作用将在体外和体内得到阐明。裂殖酵母粟酒裂殖酵母将用作模型系统来测试 hMYH 突变体的可视功能。 (III) 将检查 hMYH 与 hAPE 1、hPCNA、hHus 1、hRPA 和 hMSH6 ir 修复和复制基因座的共定位。 (IV)将确定DNA损伤后hMYH的磷酸化状态、负责hMYH磷酸化的激酶以及hMYH的磷酸化位点。将检查 DNA 损伤后 hMYH 活性与蛋白质磷酸化或某些蛋白质-蛋白质相互作用的相关性。这项研究将加深我们对 DNA 修复在肿瘤易感性中的作用的理解。