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DNA damage response and repair of a broken chromosome

DNA 损伤反应和断裂染色体的修复

基本信息

批准号：
10622121
负责人：
JAMES E HABER
金额：
$ 97.34万
依托单位：
BRANDEIS UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-06-01 至 2028-05-31
项目状态：
未结题

项目摘要

PROJECT SUMMARY The overall goal of NIGMS-funded research in my lab is to describe the molecular and cellular mechanisms by which cells sense the presence of DNA damage and how they repair chromosomal double-strand breaks (DSBs). Primarily using budding yeast as a model system, it is possible to induce site-specific DSBs with a high degree of synchrony that is not generally possible in mammalian cells, allowing “in vivo biochemistry” approaches to monitor intermediate steps in DSB repair and DNA damage signaling. We also wish to apply our understanding of DNA repair and recombination mechanisms to elucidate how the Lyme disease bacterium, Borrelia burgdorferi, is able to “change its coat” by repeated gene conversion events. The goals for the next five years of this project focus on understanding how homologous donor sequences are found and used to repair a DSB and how mismatches are tolerated and repaired during different steps of recombination. A second objective is to understand the basis of the 1000- fold increase in mutations associated with DSB repair and how microhomologies are used in repair-dependent template switching, creating complex chromosome rearrangements analogous to events recently found in human cancers. We employ similar approaches to elucidating how CRISPR/as9-mediated gene editing is accomplished using single-stranded DNA templates. A third area of concern is to understand how the DNA damage checkpoint is regulated. We wish to determine how the DNA damage response affects DSB repair and how the DNA damage checkpoint is maintained and turned off. These studies will provide new insights and guidance in defining the DSB repair and checkpoint signaling in human cells. Finally, we will continue our investigation of the mechanisms by which Borrelia initiates and mediates gene conversion events between one expressed gene and a set of adjacent pseudogenes.

项目摘要在我的实验室里，NIGMS资助的研究的总体目标是描述细胞感知DNA损伤的存在以及它们如何修复的机制染色体双链断裂（DSB）。首先使用芽殖酵母作为模型系统，可以诱导具有高度同步性的位点特异性DSB，可能在哺乳动物细胞中，允许“体内生物化学”方法监测中间体 DSB修复和DNA损伤信号的步骤。我们也希望运用我们对DNA的理解修复和重组机制，以阐明莱姆病细菌，疏螺旋体， burgdorferi能够通过重复的基因转换事件“改变它的外套”。该项目未来五年的目标集中在了解同源供体如何序列被发现并用于修复DSB以及如何容忍和修复错配在不同的重组步骤中。第二个目标是了解1000- 与DSB修复相关的突变增加了一倍，以及微同源性如何用于修复依赖的模板转换，创造复杂的染色体重排类似最近在人类癌症中发现的事件。我们采用类似的方法来阐明如何 CRISPR/as 9介导的基因编辑使用单链DNA模板完成。一第三个值得关注的领域是了解DNA损伤检查点是如何调节的。我们希望确定DNA损伤反应如何影响DSB修复以及DNA损伤如何影响DSB修复。检查点被保持并关闭。这些研究将提供新的见解和指导，在人类细胞中定义DSB修复和检查点信号传导。最后，我们将继续研究疏螺旋体启动和介导一个表达基因和一组相邻的假基因