Role of regulation of eukaryotic DNA replication in preserving genomic stability

真核 DNA 复制调控在保持基因组稳定性中的作用

基本信息

批准号：
8489302
负责人：
JOACHIM J LI
金额：
$ 28.2万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
财政年份：
2000
资助国家：
美国
起止时间：
2000-07-01 至 2015-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8489302
关键词：
Address Affect Aneuploidy Animal Model Biological Biological Assay Biological Models Cell Cycle Cells Cellular biology Centromere Chromosomal Breaks Chromosome Breakage Chromosomes Cis-Acting Sequence Complex Cyclin-Dependent Kinases DNA DNA Sequence Rearrangement DNA biosynthesis Data Dependency Diploidy Elements Ensure Eukaryotic Cell Event Evolution Frequencies Funding Gene Amplification Genetic Nondisjunction Genetic Variation Genome Genome Stability Genomic Instability Genomics Goals Haploidy Hereditary Disease Human Human Genetics Impairment Lead Malignant Neoplasms Maps Measures Mediating Minor Modeling Molecular Evolution Molecular Genetics Monitor Movement Mutation Pathway interactions Proteins Regulation Repetitive Sequence Replication Initiation Replication Origin Research Role Saccharomyces cerevisiae Saccharomycetales Science Side Site Source Staging Structure Techniques Technology Temperature Testing Time Yeast Model System base cancer cell cancer genetics disorder control genetic analysis genome analysis homologous recombination insight prevent promoter research study segregation tool tumorigenesis

项目摘要

DESCRIPTION (provided by applicant): A central tenet of eukaryotic cell biology is that DNA replication must be tightly controlled so that it occurs only once per cell cycle. It is presumed, but largely untested, that this control is vital for preserving genome integrity. Our long-term goal is to understand how the re-initiation of DNA replication is reliably prevented at the thousands of replication origins scattered throughout eukaryotic genomes, and to discern the effect of disrupting this control on genome stability. We study replication control in budding yeast because this model system offers an exceptional opportunity to dissect the complex, overlapping mechanisms that are required to achieve this control with such extraordinary fidelity. Additionally, the molecular genetic tools available in budding yeast allow us to apply both simple and sophisticated technologies to query the effects of disrupting replication controls. In previous funding periods, we demonstrated that cyclin-dependent kinases (CDKs) use multiple overlapping mechanisms to prevent origins from re-initiating within a single cell cycle. We have also shown that re-replication arising from loss of these controls leads to significant chromosomal breakage and lethality, providing a previously unknown justification for the importance of replication control. More recently, we provided the first evidence that re-replication is a highly efficient means to induce gene amplification (Green et al, Science, in press). Re-replication induced gene amplification (RRIGA) occurred with extraordinary efficiency (roughly 1/20 re-initiation events). This finding supports the compelling hypothesis that even minor impairment of replication control may contribute to genome instability. Ultimately, we hope to demonstrate that re-replication can drive the copy number changes observed in tumorigenesis, human genetic variation, and evolution. Here we propose to expand our understanding of how the loss of replication control leads to genomic instability, both by probing the mechanisms that underlie RRIGA as well as by further investigating the biological consequences and significance of loss of replication control. We propose to (1) define the mechanism and parameters enabling RRIGA; (2) determine how local regulatory factors modulate replication control at origins that are highly susceptible to re-initiation; (3) determine whether RRIGA participates in a model of evolutionary adaptation; and (4) establish whether re-replication can induce chromosome missegregation. These data will significantly enhance our understanding of how re-replication promotes genomic instability, as well as give insight into the biological significance of loss of replication control.

描述（由申请人提供）：真核细胞生物学的核心原则是 DNA 复制必须受到严格控制，以便每个细胞周期仅发生一次。据推测，但基本上未经测试，这种控制对于保持基因组完整性至关重要。我们的长期目标是了解如何在散布于真核基因组中的数千个复制起点处可靠地阻止 DNA 复制的重新启动，并辨别破坏这种控制对基因组稳定性的影响。我们研究芽殖酵母的复制控制，因为该模型系统提供了一个绝佳的机会来剖析以如此非凡的保真度实现这种控制所需的复杂、重叠的机制。此外，芽殖酵母中可用的分子遗传工具使我们能够应用简单和复杂的技术来查询破坏复制控制的影响。在之前的资助期间，我们证明了细胞周期蛋白依赖性激酶（CDK）使用多种重叠机制来防止起源在单个细胞周期内重新启动。我们还表明，由于失去这些控制而引起的重新复制会导致显着的染色体断裂和致死性，这为复制控制的重要性提供了以前未知的理由。最近，我们提供了第一个证据，证明重新复制是诱导基因扩增的高效手段（Green 等人，《科学》，出版中）。再复制诱导基因扩增 (RRIGA) 的发生效率极高（大约为再启动事件的 1/20）。这一发现支持了一个令人信服的假设，即即使复制控制的微小损害也可能导致基因组不稳定。最终，我们希望证明重新复制可以驱动在肿瘤发生、人类遗传变异和进化中观察到的拷贝数变化。在这里，我们建议通过探索 RRIGA 的机制以及进一步研究复制控制丧失的生物学后果和意义，扩大我们对复制控制丧失如何导致基因组不稳定的理解。我们建议 (1) 定义启用 RRIGA 的机制和参数； (2) 确定局部调控因素如何调节极易重新启动的起始点的复制控制； (3)确定RRIGA是否参与进化适应模型； (4)确定重新复制是否会导致染色体错误分离。这些数据将显着增强我们对再复制如何促进基因组不稳定性的理解，并深入了解复制控制丧失的生物学意义。