Ubiquitin-Mediated Regulation of DNA Repair

泛素介导的 DNA 修复调节

• 批准号: 6918588
• 负责人: JAMES M. FORD
• 金额: $ 29.52万
• 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/6918588
• 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，其功能是保护基因组免受内源性和环境源（包括紫外线）引起的 DNA 损伤的有害影响。 NER 系统的缺陷会导致基因组不稳定和癌症的发生。 NER 的第一步需要损伤识别，由异二聚体 DNA 损伤结合蛋白 XPC-HR23B 和 UV-DDB（由 p48 和 p127 蛋白组成）执行。 DDB2（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 均受到泛素蛋白酶体降解途径的翻译后调节。单泛素链和多泛素链与细胞蛋白的共价连接已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受到高度调控。该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。假设紫外线诱导的 NER 蛋白 t>48 和 XPC 泛素化特异性调节它们的降解、蛋白质和 DNA 损伤结合活性，并对 NER 产生影响。这些研究将具有广泛的适用性，有助于理解 DNA 修复在健康和疾病中的作用，并为了解 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现。 1. 确定人类DNA损伤识别因子p48和XPC是否被泛素化，以及该过程是否受HR23B调控。 2. 确定体内 DNA 损伤结合活性是否需要这些 NER 蛋白的泛素化。 3. 确定泛素化对人类细胞 NER 活性的功能影响。