Mechanisms of R-loop-Associated Genome Instability

R 环相关基因组不稳定性的机制

• 批准号: 10206172
• 负责人: Karlene A Cimprich
• 金额: $ 35.52万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-07-01 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10206172
• 关键词: Address; Affect; Automobile Driving; BRCA1 gene; Biological; Cell Cycle; Cell physiology; Cells; Complex; Conflict (Psychology); Cytoplasm; DNA; DNA Damage; DNA Double Strand Break; DNA Repair; DNA biosynthesis; DNA replication fork; Data; Disease; Event; Gene Expression; Generations; Genetic Transcription; Genome; Genome Stability; Genomic Instability; Genomic approach; Genomics; Goals; Human; Hybrids; Immune response; Innate Immune Response; Location; Malignant Neoplasms; Mammalian Cell; Maps; Mediating; Molecular; Molecular Probes; Mutation; Nature; Nuclear; Nucleic Acids; Pathogenesis; Physiological Processes; Play; Poly(ADP-ribose) Polymerases; Population; Population Process; Process; Product R; Publishing; RNA; Research; Resolution; Role; SETX gene; Signal Transduction; Single-Stranded DNA; Site; Spatial Distribution; Structure; Syndrome; System; Techniques; Testing; Therapeutic; Transcription-Coupled Repair; Work; developmental disease; endonuclease; experimental study; human disease; insight; mammalian genome; nervous system disorder; novel; nuclease; nucleic acid structure; programs; protein complex; repair enzyme

PROJECT SUMMARY Replication and transcription are fundamental physiological processes, yet paradoxically they both threaten genome stability. Replication forks encounter various types of endogenous and exogenous obstacles that keep them from accurately completing DNA replication. Recent studies suggest that the deleterious effects of transcription could be a consequence of R-loops, three- stranded nucleic acid structures containing an RNA-DNA hybrid and a region of single-stranded DNA. R-loops occur throughout the genome of mammalian cells and regulate various aspects of gene expression, but their accumulation leads to DNA damage, particularly when cells are undergoing DNA replication. Increasing evidence suggests that conflicts between transcription- associated R-loops and replication protein complexes are important factors underlying genome instability, but the specific mechanisms driving this instability are currently unknown. The long- term goal of this research program is to understand how cells distinguish and resolve regulatory and deleterious R-loops, and how this is perturbed in human disease. It is hypothesized that R- loops are dynamic structures that become susceptible to processing when they accumulate, leading to the formation of DNA breaks and ultimately resulting in genome instability. The object of this application is to define how R-loops are recognized and processed in the cells throughout the cell cycle, to determine where in the genome processing occurs, and to determine how conflicts with the replication machinery contribute to R-loop processing. In the first aim, the processing of R-loops by cellular endonucleases involved in DNA repair will be explored in cells using molecular and cell biological approaches. In the second aim, the sites and products of R- loop formation and processing will be identified and mapped. These studies will take advantage of cutting-edge genomic approaches that have been developed to map the spatial distribution of R-loops and R-loop processing products throughout the genome. Finally, in the third aim, the impact of an R-loop on collisions between replication and transcription machineries will be studied in cells. These studies will take advantage of a novel system recently developed to control such collisions and R-loop formation in the context of the replication fork and recent break-mapping strategies.

项目摘要 复制和转录是基本的生理过程，但从矛盾的角度来看 两者都威胁着基因组稳定性。复制叉会遇到各种类型的内生和 外源性障碍，可防止它们准确地完成DNA复制。最近的研究 表明转录的有害影响可能是R环的结果，三 滞留的核酸结构，该结构含有RNA-DNA杂交和单链区域 脱氧核糖核酸。 R环发生在整个哺乳动物细胞的基因组中，并调节 基因表达，但它们的积累会导致DNA损伤，尤其是当细胞为 进行DNA复制。越来越多的证据表明转录之间的冲突 - 相关的R环和复制蛋白复合物是基因组的重要因素 不稳定，但是目前尚不清楚推动这种不稳定的特定机制。长期 该研究计划的术语目标是了解细胞如何区分和解决调节 和有害的R环，以及如何在人类疾病中扰动。假设R- 循环是动态结构，在积累时容易处理， 导致DNA断裂的形成，并最终导致基因组不稳定性。对象 此应用的是定义如何在整个单元格中识别和处理R-loops 细胞周期，以确定基因组加工中发生的位置，并确定如何 与复制机械的冲突有助于R环处理。在第一个目标中， 将在细胞中探索参与DNA修复的细胞内切酶处理R环 使用分子和细胞生物学方法。在第二个目标中，r-的站点和产品 循环形成和处理将被识别和映射。这些研究将利用 已经开发出的尖端基因组方法来绘制的空间分布 整个基因组中的R环和R环处理产物。最后，在第三个目标中， 将研究R环对复制和转录机械之间的碰撞的影响 在细胞中。这些研究将利用最近开发的新型系统来控制这种 在复制叉和最近的断裂映射的背景下的碰撞和R环形成 策略。