Processing Site-Specific DNA Lesions by DNA Repair/Recom

通过 DNA 修复/重组处理位点特异性 DNA 损伤

• 批准号: 6990365
• 负责人: Karen M Vasquez
• 金额: $ 16.16万
• 依托单位: UNIVERSITY OF TX MD ANDERSON CAN CTR
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-04-21 至 2009-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6990365
• 关键词: CHO cells; DNA damage; DNA repair; cell free system; crosslink; genetic recombination; mitomycin C; psoralens; triple helix; ultraviolet radiation

DNA interstrand crosslinks (ICLs) present a formidable block to DNA metabolic processes and must be repaired for cell survival. While much work has been done to define the mechanism(s) of DNA interstrand crosslink repair in bacteria and yeast, very little is known about their repair in mammalian cells. It is has been proposed that both homologous recombination (HR) and nucleotide excision repair pathways are involved in DNA crosslink repair. It is becoming increasingly evident that there is overlap among the DNA repair pathways for certain types of DNA lesions. Our working hypothesis is that a number of proteins already defined as essential in DNA repair mechanisms are also required for efficient HR, or work together with HR in DNA interstrand crosslink processing and removal. The long-term objectives of the proposed research are to elucidate the molecular mechanism(s) involved in the removal of site-specific DNA interstrand crosslinks and complex DNA lesions (psoralen plus UVA radiation and mitomycin C crosslinks directed by triplex formation) from the mammalian genome, to identify interactions among the DNA repair pathways, and to determine the role(s) of homologous recombination, nucleotide excision, and mismatch repair mechanisms in the removal and processing of ICLs and complex DNA lesions. Specifically we propose to: 1) direct sitespecific DNA crosslinks to the genome in normal and recombination-deficient Chinese hamster ovary cells to elucidate the role of recombination in the removal of interstrand crosslinks and complex DNA lesions; 2) determine the mechanism(s) of removal of site-specific DNA interstrand crosslinks and complex DNA lesions directed by triplex-forming oligonucleotides in mammalian cell-free extracts; and 3) identify interactions between proteins involved in DNA repair pathways in the removal of site-specific DNA interstrand crosslinks and complex DNA lesions.

DNA链间交联（ICLs）是DNA代谢过程的一个强大障碍，必须修复才能使细胞存活。虽然已经做了很多工作来定义细菌和酵母中DNA链间交联修复的机制，但对哺乳动物细胞中的修复知之甚少。已经提出同源重组（HR）和核苷酸切除修复途径都参与DNA交联修复。越来越明显的是，某些类型的DNA损伤的DNA修复途径之间存在重叠。我们的工作假设是，许多已被定义为DNA修复机制必需的蛋白质也是高效HR或与HR共同工作所必需的 在DNA链间交联处理和去除中。拟议研究的长远目的是阐明去除位点特异性DNA链间交联和复杂DNA损伤的分子机制（通过三链体形成指导的peptien加UVA辐射和丝裂霉素C交联），以鉴定DNA修复途径之间的相互作用，并确定同源重组，核苷酸切除，以及ICL和复杂DNA损伤的去除和处理中的错配修复机制。具体而言，我们建议：1）直接特定地点 在正常和重组缺陷的中国仓鼠卵巢细胞中DNA与基因组的交联，以阐明重组在去除链间交联和复杂DNA损伤中的作用; 2）确定在哺乳动物无细胞提取物中由三链体形成寡核苷酸指导的位点特异性DNA链间交联和复杂DNA损伤的去除机制;和3）鉴定在去除位点特异性DNA链间交联和复杂DNA损伤中参与DNA修复途径的蛋白质之间的相互作用。