Chromatin Modulation Associated with DNA Breakage and Repair in Human Cells

与人类细胞 DNA 断裂和修复相关的染色质调节

• 批准号: 8202555
• 负责人: Michael B Kastan
• 金额: $ 10.39万
• 依托单位: ST. JUDE CHILDREN'S RESEARCH HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2011-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8202555
• 关键词: Acetylation; Address; Affect; Ataxia-Telangiectasia-Mutated protein kinase; BRCA1 Protein; BRCA1 gene; Binding; Biochemical; Cancer-Predisposing Gene; Cell Line; Cell Survival; Cells; Cellular biology; Chromatin; Complex; DNA; DNA Damage; DNA Double Strand Break; DNA Ligation; DNA Repair; Development; Disease; Double Strand Break Repair; Endonuclease I; Enzymes; Eukaryota; Event; Functional disorder; Fungal Genome; Generations; Genetic Transcription; Homing; Human; Human Genome; Kinetics; Laboratories; Lead; Ligation; Malignant Neoplasms; Mammalian Cell; Medicine; Methods; Modeling; Modification; Molecular; Molecular Probes; Movement; Mutagenesis; Nature; Nucleosomes; Outcome; Phosphorylation; Predisposition; Process; Protein Binding; Proteins; Radiation therapy; Regulation; Reporting; Role; Signal Pathway; Signal Transduction Pathway; Site; Stress; System; Techniques; Ubiquitination; base; cancer therapy; chemotherapy; chromatin protein; endonuclease; human disease; improved; insight; interest; malignant breast neoplasm; novel; novel strategies; nuclease; prevent; repaired; research study; response; restoration; tumor

DESCRIPTION (provided by applicant): Cellular responses to DNA damage and other stresses are important determinants of cell viability and mutagenesis and impact the development of a wide range of human diseases. DNA damage responses, in particular, are major determinants of both cancer development and outcomes of cancer therapies. Induction of signal transduction pathways is a critical aspect of cellular responses to these stresses and significant advances have been made in recent years elucidating the biochemical steps in such signaling pathways. Clarification of such steps enables modulation of these responses, which can enhance research studies and can lead to the generation of new medicines to prevent and treat these diseases. This application describes an enhancement of a novel technique developed in the Kastan laboratory in which DNA double strand breaks (DSBs) are introduced at defined sites in the human genome. Introduction of site-directed DNA damage permits studies of molecular events occurring at and around the DSB and direct assessment of repair (re- ligation) of DNA breaks. The enhancements of this technique permit greater temporal control of introduction of the DSBs and allow assessment of the kinetics and completeness of DSB repair. This enhanced approach will be used in the experiments proposed in this application to elucidate the dynamic changes in protein movements/modifications occurring at and around the DSB and explore factors which may determine efficiency of repair of the DNA breaks. One major focus is exploration of the molecular controls of nucleosome disruption at the DSB site and elucidating the functional importance of this nucleosomal disruption. It is well established that numerous modifications occur in chromatin-associated proteins surrounding DSBs during the damage/repair process; however, the functional role(s) of many of these modifications, especially related to DNA repair, remain to be clarified. Experiments are proposed that will use this novel system to explore the potential impact of these chromatin changes surrounding the DSBs on the ability of the cell to repair (re-ligate) DNA breaks. Further, the mechanisms and impact of the breast cancer susceptibility gene product, Brca1, on this process will be investigated. The proposed studies will provide novel insights into molecular mechanisms associated with the DNA DSB repair process in human cells. PUBLIC HEALTH RELEVANCE: DNA damage and repair are critical determinants of cancer development and responses of tumors to chemotherapy and radiation therapy. Based on a novel system developed to probe molecular events associated with DNA breakage in human cells, studies are proposed to explore factors that affect the ability of human cells to repair DNA double strand breaks. Insights gained into these mechanisms associated with cellular responses to DNA damage will improve our understanding of cancer predisposition and development and have the potential to lead to development of new approaches to enhancing tumor responses to therapy.

描述(申请人提供)：细胞对DNA损伤和其他压力的反应是细胞存活和突变的重要决定因素，并影响一系列人类疾病的发展。尤其是DNA损伤反应，是癌症发展和癌症治疗结果的主要决定因素。信号转导通路的诱导是细胞对这些胁迫反应的一个重要方面，近年来在阐明这些信号转导通路中的生化步骤方面取得了重大进展。澄清这些步骤可以调整这些反应，从而加强研究，并产生预防和治疗这些疾病的新药。这项申请描述了Kastan实验室开发的一项新技术的增强，在该技术中，DNA双链断裂(DSB)被引入到人类基因组的特定位置。引入定点DNA损伤可以研究发生在DSB及其周围的分子事件，并直接评估DNA断裂的修复(重新连接)。这项技术的增强允许更好地在时间上控制DSB的引入，并允许评估DSB修复的动力学和完整性。这种增强的方法将用于本申请中提出的实验，以阐明发生在DSB及其周围的蛋白质运动/修饰的动态变化，并探索可能决定DNA断裂修复效率的因素。一个主要的焦点是探索DSB位点核小体破坏的分子控制，并阐明这种核小体破坏的功能重要性。众所周知，在DNA损伤/修复过程中，DSB周围的染色质相关蛋白会发生许多修饰；然而，其中许多修饰的功能作用(S)仍不清楚，尤其是与DNA修复有关的修饰。建议进行实验，使用这一新系统来探索DSB周围的这些染色质变化对细胞修复(重新连接)DNA断裂的能力的潜在影响。此外，还将研究乳腺癌易感基因产物BRCA1在这一过程中的机制和影响。拟议的研究将为人类细胞中与DNA DSB修复过程相关的分子机制提供新的见解。 公共卫生相关性：DNA损伤和修复是癌症发展和肿瘤对化疗和放射治疗反应的关键决定因素。在开发了一种新的系统来探测与人类细胞DNA断裂相关的分子事件的基础上，提出了探索影响人类细胞修复DNA双链断裂能力的因素的研究。对这些与细胞对DNA损伤的反应相关的机制的洞察将提高我们对癌症易感性和发展的理解，并有可能导致开发新的方法来增强肿瘤对治疗的反应。