Repair of Oxidatively Damaged Guanines in Human

描述(由申请人提供)：DNA修复缺陷会导致基因组不稳定，这是癌症的一个标志。氧化DNA损伤是生物体中突变负荷的主要来源，并在癌症发生和衰老中发挥作用。8-氧代-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，其危害最大，因为它可以在DNA复制过程中与腺嘌呤错误配对。因此，通过hMYH、hOGG1和hMSH2/hMSH6依赖的错配修复途径来修复A/8-oxoG和C/8-oxoG提供了抵抗氧化应激的能力。本项目的总体目标是了解hMYH途径在控制肿瘤发生中的作用及其与其他修复途径的相互作用，以响应氧化应激，如过氧化氢和电离辐射处理。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hHusl/hRad1/hRad9的相互作用。(I)将研究hMYH与hPCNA和hHusl在氧化应激下相互作用的变化。HMYH和hHusl上的相互作用结构域将被绘制出来，并将在体内测试它们功能相互作用的意义。将测试hMYH-hPCNA TC hMYH-hRad9/hRad1/hHusl的分子开关诱导DNA损伤反应以及hMYH可能招募_hRad9/hRad1/hHus1到病变部位的工作模型。(Ii)hMYH和hMSH6之间的相互作用将在体外和体内得到阐明。裂解酵母S.pombe将作为一个模型系统来测试hMYH突变体的体内功能。(Iii)研究hMYH与Hape 1、hPCNA、hHus 1、HrpA和hMSH6 IR修复和复制基因座的共定位。(4)DNA损伤后hMYH的磷酸化状态、负责hMYH磷酸化的激酶和hMYH的磷酸化位点(S)将被确定。在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(S)的相关性将被检验。这项研究将促进我们对DNA修复在肿瘤易感性中的作用的理解。

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.