Processing Site-Specific DNA Lesions by DNA Repair/Recom

DNA链间交联（ICLs）对DNA代谢过程造成了严重的阻碍，细胞存活必须对其进行修复。虽然已经做了很多工作来确定细菌和酵母中DNA链间交联修复的机制，但对它们在哺乳动物细胞中的修复知之甚少。同源重组（homologous recombination， HR）和核苷酸切除修复途径都参与了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，许多已经被定义为DNA修复机制中必不可少的蛋白质也需要有效的HR，或者与HR一起工作

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.