Ubiquitin-Mediated Regulation of DNA Repair

泛素介导的 DNA 修复调节

• 批准号: 7077712
• 负责人: JAMES M. FORD
• 金额: $ 28.83万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-07-07 至 2008-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7077712
• 关键词: DNA binding protein; DNA damage; DNA repair; RNA interference; biological signal transduction; cell line; covalent bond; gene expression; immunoprecipitation; posttranslational modifications; proteasome; protein binding; protein degradation; protein structure function; site directed mutagenesis; ubiquitin; ultraviolet radiation

DESCRIPTION (provided by applicant): Nucleotide excision repair (NER) is a highly conserved DNA repair pathway that functions to protect the genome from the detrimental effects of DNA lesions introduced by endogenous and environmental sources, including UV light. Defects in the NER system result in genomic instability and the development of cancers. The initial step in NER requires lesion recognition, and is performed by the heterodimeric DNA damage binding proteins XPC-HR23B and UV-DDB (composed of the p48 and p127 proteins). The DDB2 (encoding the p48 protein) and XPC genes are transcriptionally induced by the p53 tumor suppressor gene following DNA damage. In addition, both p48 and XPC are post-translationally regulated by the ubiquitin-proteasome degradation pathway. Covalent attachment of mono- and polyubiquitin chains to cellular proteins has emerged as a predominant cellular regulatory mechanism with roles in cell division, signal transduction, endocytic trafficking, and DNA repair. Thus, the DNA damage recognition step of NER is highly regulated through transcriptional and post-transcriptional mechanisms. The long-term objectives of this project are to understand how the ubiquitin-proteasome pathway regulates human NER. The hypothesis is that UV-induced ubiquitination of the NER proteins t>48 and XPC specifically regulate their degradation, protein- and DNA damage binding activities, and affect on NER. These studies will have broad applicability to understanding the role of DNA repair in health and disease, and provide important insight into the role of NER in the etiology of human cancer. The project will be approached through the following specific aims. 1. To determine if the human DNA damage recognition factors p48 and XPC are ubiquitinated and if this process is regulated by HR23B. 2. To determine whether ubiquitination of these NER proteins is required for DNA damage binding activity, in vivo. 3. To determine the functional consequences of ubiquitination on NER activity in human cells.

描述（由申请人提供）：核苷酸切除修复（NER）是一种高度保守的DNA修复途径，其作用是保护基因组免受内源性和环境来源（包括紫外线）引入的DNA损伤的有害影响。NER系统的缺陷导致基因组不稳定和癌症的发展。NER的初始步骤需要损伤识别，并由异二聚体DNA损伤结合蛋白XPC-HR 23 B和UV-DDB（由p48和p127蛋白组成）执行。DDB 2（编码p48蛋白）和XPC基因在DNA损伤后由p53肿瘤抑制基因转录诱导。此外，p48和XPC均受泛素-蛋白酶体降解途径的后调节。单链和多聚泛素链与细胞蛋白的共价连接已经成为一种主要的细胞调节机制，在细胞分裂、信号转导、内吞运输和DNA修复中发挥作用。因此，NER的DNA损伤识别步骤通过转录和转录后机制高度调节。本项目的长期目标是了解泛素-蛋白酶体途径如何调节人类NER。假设是UV诱导的NER蛋白t>48和XPC的泛素化特异性地调节它们的降解、蛋白质和DNA损伤结合活性，并影响NER。这些研究将广泛适用于理解DNA修复在健康和疾病中的作用，并为NER在人类癌症病因学中的作用提供重要见解。该项目将通过以下具体目标来实施。 1.确定人DNA损伤识别因子p48和XPC是否被泛素化，以及这一过程是否受HR 23 B的调控。 2.确定这些NER蛋白的泛素化是否是体内DNA损伤结合活性所必需的。 3.确定泛素化对人类细胞NER活性的功能影响。