ISOLATION AND ANALYSIS OF HUMAN DNA REPAIR GENES

这项提议的长期目标是阐明核苷酸 真核系统中的切除修复机制。NER是一家 维持化学完整性的少量生化途径 通过一个持续的监测和修复过程来获得遗传物质。 NER的缺陷被发现是导致NER发展的原因 几种人类疾病，包括着色性干皮病(XP)、Cockayne‘s 综合征和毛发硫代营养不良。此外，NER中的缺陷加速 致癌，并可能参与衰老过程。NER途径 在真核生物中是复杂的，可能涉及多达30种不同的 完成修复过程的基因。这个项目的重点是 主要是关于NER的早期阶段，这涉及到对 损伤、损伤部位的引爆以备后续切开和 切开步骤本身。这些早期的步骤似乎涉及到大约12到 15个基因。其中相当数量的基因已经被克隆和 这些成就为更详细的生物化学奠定了基础 学习。拟议项目的具体目的是：L)调查 XPC基因在NER中的功能进一步研究和确定 先前证明的蛋白质-蛋白质的功能相关性 DNA修复因子之间的相互作用，以及3)分析最近分离的 确定它们是否与NER或NER有关的新基因 重组修复途径。这些研究将使用 生化和遗传方法的结合，包括双杂交 系统，细胞无修复试验，体外试验检测蛋白质-蛋白质 相互作用和定点突变。

The long term objective of this proposal is the elucidation of nucleotide excision repair (NER) mechanisms in eucaryotic systems. NER is one of a small number of biochemical pathways that maintain the chemical integrity of the genetic material by a constant monitoring and repair process. Defects in NER have been found to be responsible for the development of several human diseases including xeroderma pigmentosum (XP), Cockayne's syndrome, and trichothiodystrophy. In addition, defects in NER accelerate carcinogenesis and may be involved in the aging process. The NER pathway is complex in eucaryotes involving perhaps as many as thirty different genes for the complete repair process. The focus of this project is primarily on the early stages of NER which involve the recognition of damage, priming of the damaged site for subsequent incision and the incision step itself. These early steps appear to involve about twelve to fifteen genes. A significant number of these genes have been cloned and these accomplishments have set the stage for more detailed biochemical studies. The specific aims of the proposed project are to: l) investigate the function of the XPC gene in NER 2) further investigate and determine the functional relevance of previously demonstrated protein-protein interactions among DNA repair factors, and 3) analyze recently isolated novel genes to determine if they have an involvement in either NER or recombinational repair pathways. These studies will be conducted using a combination of biochemical and genetic approaches including the two-hybrid system, cell free repair assays, in vitro assays to detect protein-protein interactions, and site directed mutagenesis.