SWI/SNF chromatin remodeling in nucleotide excision repair

核苷酸切除修复中的 SWI/SNF 染色质重塑

• 批准号: 8240084
• 负责人: FENG GONG
• 金额: $ 33.88万
• 依托单位: UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-08-10 至 2015-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8240084
• 关键词: Binding; Biochemical; Biochemistry; Biological Assay; Cells; Chromatin; Chromatin Remodeling Factor; Chromatin Structure; Complement; Complex; DNA; DNA Binding; DNA Damage; DNA Double Strand Break; DNA Repair; DNA Repair Pathway; DNA lesion; Defense Mechanisms; Detection; Development; Due Process; Environment; Environmental Exposure; Eukaryotic Cell; Genetic Transcription; Genome; Genome Stability; Goals; Guanine Nucleotide Exchange Factors; Health; Hereditary Disease; Histones; Human; Human Genetics; In Vitro; Knowledge; Label; Laboratories; Link; Mammalian Cell; Metabolism; Methods; Microscopy; Molecular; Monitor; Nucleosomes; Nucleotide Excision Repair; Organism; Pathway interactions; Play; Positioning Attribute; Prevention strategy; Process; Proteins; Recruitment Activity; Repair Complex; Research; Role; Saccharomyces cerevisiae; Site; System; Techniques; Testing; Ultraviolet Rays; Yeasts; cancer therapy; chromatin immunoprecipitation; chromatin modification; chromatin remodeling; cigarette smoking; disease-causing mutation; gene function; histone modification; innovation; protein protein interaction; prototype; public health relevance; reconstitution; repaired; research study; skin cancer prevention; ultraviolet; ultraviolet damage; ultraviolet irradiation; ultraviolet lesions; yeast genetics

DESCRIPTION (provided by applicant): A wide variety of human genetic diseases are caused by mutations in genes that function in DNA repair pathways. As an important defense mechanism to maintain genome stability, DNA repair also contributes to the normal development and health of organisms. Bulky DNA lesions resulting from ultraviolet (UV) radiation, cigarette smoke and other endogenous and exogenous agents are repaired by the conserved nucleotide excision repair (NER) pathway. The assembly of DNA into chromatin in eukaryotic cells interferes with the NER process. How NER operates in the context of chromatin is largely unknown. Our previous studies in Saccharomyces cerevisiae first linked the prototype ATP-dependent chromatin remodeling complex SWI/SNF to NER. Very recent findings from our group and others have established a connection between SWI/SNF and NER in human cells. However, the process by which SWI/SNF remodels chromatin to facilitate NER remains undefined. Also unknown is the SWI/SNF recruitment mechanism in NER facilitation. The central objective of this proposal is to elucidate the mechanism by which SWI/SNF facilitates damage detection and repair in S. cerevisiae and cultured mammalian cells. Additionally, in vitro biochemical studies will be undertaken to validate cellular findings and provide detailed biochemical mechanisms on the coordination of chromatin remodeling and NER. We hypothesize that DNA damage recognition factors recruit SWI/SNF to sites of DNA damage via protein-protein interactions. We will also test an alternative mechanism that histone modifications are involved in SWI/SNF recruitment/or retention at sites of UV damage. In Aim I, we will determine the role of chromatin modifying activities required for efficient global genome NER (GG-NER) in yeast. We will investigate SWI/SNF remodeling at the Sir complex coated HML locus during NER. Aim II will discern the role of chromatin modification and remodeling during NER in mammalian cells. We will investigate the consequence of SWI/SNF inactivation on the dynamic NER process. In Aim III, we will ascertain the mechanisms of damage recognition and NER of nucleosomes in vitro. A purified system will be used to examine how a DNA lesion 'buried' in a mononucleosome is detected by DDB2 and XPC and how SWI/SNF remodels the mononucleosome to facilitate damage detection. PUBLIC HEALTH RELEVANCE: Nucleotide excision repair is the major DNA repair pathway that removes bulky DNA damage induced by ultraviolet (UV) radiation to protect genome stability and prevent skin cancers. Our proposed experiments will help understand how bulky DNA lesions are repaired in human cells in the context of chromatin and reveal molecular mechanisms important to overcome the hazardous health effects of environmental exposures. Knowledge gained from these efforts could be exploited to develop strategies for the prevention and/or treatment of cancers induced by genotoxic environmental exposures.

描述（由申请人提供）：多种人类遗传疾病是由在 DNA 修复途径中起作用的基因突变引起的。 DNA修复作为维持基因组稳定性的重要防御机制，也有助于生物体的正常发育和健康。由紫外线 (UV) 辐射、香烟烟雾和其他内源性和外源性因素引起的大量 DNA 损伤可通过保守的核苷酸切除修复 (NER) 途径进行修复。真核细胞中 DNA 组装成染色质会干扰 NER 过程。 NER 如何在染色质环境中发挥作用尚不清楚。我们之前对酿酒酵母的研究首先将原型 ATP 依赖性染色质重塑复合物 SWI/SNF 与 NER 连接起来。我们小组和其他人的最新研究结果已经在人类细胞中建立了 SWI/SNF 和 NER 之间的联系。然而，SWI/SNF 重塑染色质以促进 NER 的过程仍不清楚。同样未知的是 NER 促进中的 SWI/SNF 招募机制。该提案的中心目标是阐明 SWI/SNF 促进酿酒酵母和培养哺乳动物细胞损伤检测和修复的机制。此外，还将进行体外生化研究来验证细胞研究结果，并提供染色质重塑和 NER 协调的详细生化机制。我们假设 DNA 损伤识别因子通过蛋白质-蛋白质相互作用将 SWI/SNF 招募到 DNA 损伤位点。我们还将测试另一种机制，即组蛋白修饰参与 SWI/SNF 募集/或在 UV 损伤位点的保留。在目标 I 中，我们将确定酵母中高效全基因组 NER (GG-NER) 所需的染色质修饰活性的作用。我们将在 NER 期间研究 Sir 复合物包被的 HML 基因座的 SWI/SNF 重塑。目标 II 将了解哺乳动物细胞 NER 过程中染色质修饰和重塑的作用。我们将研究 SWI/SNF 失活对动态 NER 过程的影响。在目标III中，我们将在体外确定核小体损伤识别和NER的机制。纯化系统将用于检查 DDB2 和 XPC 如何检测“埋藏”在单核小体中的 DNA 损伤，以及 SWI/SNF 如何重塑单核小体以促进损伤检测。 公共卫生相关性： 核苷酸切除修复是主要的 DNA 修复途径，可消除紫外线 (UV) 辐射引起的大量 DNA 损伤，以保护基因组稳定性并预防皮肤癌。我们提出的实验将有助于了解在染色质背景下人类细胞中庞大的 DNA 损伤是如何修复的，并揭示对于克服环境暴露对健康的危害至关重要的分子机制。从这些努力中获得的知识可用于制定预防和/或治疗由遗传毒性环境暴露诱发的癌症的策略。