Mechanisms of programmed chromosome breakage

程序性染色体断裂的机制

• 批准号: 10552369
• 负责人: Andreas Hochwagen
• 金额: $ 51.86万
• 依托单位: NEW YORK UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-02-01 至 2028-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10552369
• 关键词: Architecture; Cell Cycle Progression; Cell physiology; Cells; Chromosome Breakage; Chromosome abnormality; Chromosomes; Congenital Abnormality; Copy Number Polymorphism; DNA; DNA Double Strand Break; DNA Repair; DNA copy number; Defect; Development; Disease; Environment; Eukaryota; Fertility; Gene Dosage; Genes; Genetic; Genetic Variation; Genome; Genomic Instability; Genomics; Germ Cells; Germ Lines; Goals; Health; Human Cell Line; Infertility; Laboratories; Length; Lesion; Life Cycle Stages; Location; Loss of Heterozygosity; Malignant Neoplasms; Measures; Meiosis; Meiotic Recombination; Mental disorders; Molecular; Molecular Biology; Organism; Process; Research; Ribosomal DNA; Risk; Role; Saccharomyces cerevisiae; Syndrome; Testing; Time; Work; Yeasts; cancer predisposition; developmental disease; experimental study; genetic variant; genome integrity; genome-wide; genomic locus; high risk; insight; neuronal cell body; novel; phosphoproteomics; programs; repaired; surveillance network

Program Summary/Abstract The overall goal of this research is to understand how organisms safely break their own DNA. DNA double- strand breaks (DSBs) are highly hazardous lesions whose improper repair can cause loss of heterozygosity and copy-number variations, leading to numerous psychiatric and developmental disorders, as well as cancers. Despite these risks, most eukaryotes introduce programmed DSBs into their genomes at one or more points in their life cycle. These breaks occur in the soma and the germ line and function to create genetic diversity, remove unwanted DNA, and support adaptation to changing environments. Cells go to great lengths to keep programmed DSBs safe. They control the location and timing of DSBs, promote correct repair-template choice, and use surveillance mechanisms to coordinate DSB formation with other cellular processes, including cell-cycle progression. Defects in any of these layers of control leave organisms with a higher risk of genome instability, and thus provide key insights into the genome instability associated with cancers and the chromosomal abnormalities that lead to birth defects and infertility. To investigate safe DSB formation, the proposed work focuses on two highly conserved instances of developmentally induced DNA breakage: (1) meiotic recombination, which involves hundreds of DSBs per meiotic germ cell, and (2) programmed copy-number changes in the ribosomal DNA (rDNA), the most highly expressed gene locus in eukaryotes. The proposed work uses genetics, molecular biology, and genomics to investigate these processes. Most of the work is conducted in the yeast Saccharomyces cerevisiae, but conservation of rDNA copy-number control is also tested in human cell lines. To investigate meiotic DSBs, research over the next 5 years will build on results of a phospho-proteomics screen to dissect the surveillance network that coordinates many meiotic processes with DSB formation. Experiments will also define the role of chromosome architecture in making DSB hotspots hot, and a genome- wide approach will be developed to measure meiotic repair-template choice across the genome. To analyze copy-number dynamics in the rDNA, research will focus on the mechanisms that drive re- expansion of critically short rDNA clusters and investigate the role of a novel DNA repair intermediate in this process. In addition, the proposed work will investigate the spreading of genetic variants among repeats of an rDNA cluster over evolutionary time scales and upon selection in the laboratory. Together, these analyses will provide fundamental insights into the dynamics of developmentally induced DSBs, open avenues for understanding how these endogenous processes contribute to genomic plasticity in health and disease.

项目概要/摘要 这项研究的总体目标是了解生物体如何安全地破坏自己的DNA。DNA双- 链断裂（DSB）是高度危险的损伤，其不适当的修复可导致杂合性丢失 和拷贝数变异，导致许多精神和发育障碍，以及癌症。 尽管有这些风险，大多数真核生物还是在基因组的一个或多个位点上引入了程序化的DSB。 生命周期。这些断裂发生在索马和生殖细胞系中，其功能是创造遗传多样性， 去除不需要的DNA，并支持适应不断变化的环境。 细胞竭尽全力保护编程的DSB的安全。他们控制着DSB的位置和时间， 促进正确的修复模板的选择，并使用监督机制，以协调DSB的形成， 其他细胞过程，包括细胞周期进程。这些控制层中的任何一层出现缺陷， 基因组不稳定性风险较高的生物，从而提供了基因组不稳定性的关键见解 与癌症和导致出生缺陷和不育的染色体异常有关。 为了研究安全的DSB形成，建议的工作集中在两个高度保守的情况下， 发育诱导的DNA断裂：（1）减数分裂重组，其中涉及数百个DSB， 减数分裂生殖细胞，和（2）核糖体DNA（rDNA）的程序性拷贝数变化，最高 在真核生物中表达的基因位点。这项拟议中的工作利用遗传学、分子生物学和基因组学， 研究这些过程。大部分工作是在酿酒酵母中进行的，但 还在人细胞系中测试了rDNA拷贝数控制的保守性。 为了研究减数分裂DSB，未来5年的研究将建立在磷酸化蛋白质组学的基础上。 筛选以剖析协调许多减数分裂过程与DSB形成的监视网络。 实验还将确定染色体结构在使DSB热点变热中的作用，以及基因组- 将开发广泛的方法来测量整个基因组的减数分裂修复模板选择。 为了分析rDNA中的拷贝数动态，研究将集中在驱动重新编码的机制上。 扩展极短的rDNA簇并研究新型DNA修复中间体在其中的作用 过程此外，拟议的工作将调查遗传变异在重复序列中的传播， rDNA簇在进化的时间尺度和在实验室中的选择。 总之，这些分析将提供基本的见解，发展诱导的动态 DSB，为理解这些内源性过程如何促进基因组可塑性开辟了途径， 健康和疾病。