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Mechanisms of chromatin remodeling and roadblock clearance by DNA motor proteins

DNA 运动蛋白的染色质重塑和路障清除机制

基本信息

批准号：
8829295
负责人：
ILYA J FINKELSTEIN
金额：
$ 24.9万
依托单位：
UNIVERSITY OF TEXAS AT AUSTIN
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-04-05 至 2017-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8829295
关键词：
ABCG2 gene Address Bacteria Binding Biochemical Biochemistry Biological Assay Biology Cells Chromatin Complex Crowding DNA DNA Repair DNA Repair Pathway DNA-Binding Proteins Data Set Deoxyribonucleases Disease Enzymes Escherichia coli Event Excision Fellowship Genetic Recombination Genome Stability Genomics Goals Human Individual Institution Laboratories Light Malignant Neoplasms Mediation Motor Motor Activity Mutation Nucleoproteins Nucleosomes Obstruction Outcome Phase Physics Process Property Proteins Reaction Research Research Personnel Role Rothmund-Thomson syndrome S cerevisiae DNA2 protein Staging System Technology Testing Time Work Yeasts abstracting career chromatin remodeling design helicase homologous recombination human disease in vivo nanoscale new technology nuclease prevent professor protein complex repair enzyme repaired research study single molecule skills tool translocase

项目摘要

Project Summary/Abstract Repairing broken DNA is essential for preventing mutations that can cause diseases such as cancer. Homologous recombination is an error-free DNA repair pathway that is conserved from bacteria to human. In the first step of recombination, Sgs1 and other specialized DNA motor proteins move along the broken DNA to process damaged strands for repair. Loss of Sgs1 in humans leads to devastating diseases such as Bloom, Werner and Rothmund-Thomson syndromes. The process by which Sgs1 and related DNA motors navigate on highly condensed chromatin and deal with other nucleoprotein collisions remains unresolved. Our hypothesis is that DNA motors collaborate to destabilize nucleosomes and other roadblocks by sequentially displacing and evicting the obstacles, thereby allowing other repair enzymes to gain access to the DNA. I have begun to address how DNA motors negotiate roadblocks by directly visualizing these collisions at the single molecule level. I observed that RecBCD, a prokaryotic DNA repair motor, displaces multiple types of obstacles as it moves along DNA. In the K99 phase, I will extend my single molecule assay to study the motor properties of Sgs1. During the R00 phase, I will elucidate the role of Top3/Rmi1 and Dna2 in facilitating Sgs1-dependent eukaryotic DNA repair. My second aim in the R00 phase is to determine how the Sgs1/Top3/Rmi1 complex processes chromatin. These experiments will rely on a new technology developed in the Greene laboratory, which allows us to directly visualize hundreds of individual DNA motor proteins in real time. By rapidly gathering statistically relevant datasets, we can study homologous recombination with an unprecedented level of mechanistic detail. My ultimate career goal is to achieve tenure as a professor at a research institution. The skills that I develop during the K99 phase of the fellowship will enable me to succeed as an independent investigator.

项目概要/摘要修复断裂的 DNA 对于预防可能导致癌症等疾病的突变至关重要。同源重组是一种从细菌到人类都保守的无错误 DNA 修复途径。在重组的第一步，Sgs1 和其他专门的 DNA 运动蛋白沿着断裂的 DNA 移动处理损坏的线束进行修复。人类失去 Sgs1 会导致毁灭性的疾病，例如布卢姆病、维尔纳和罗斯蒙德-汤姆森综合征。 Sgs1 和相关 DNA 马达的导航过程关于高度浓缩的染色质和处理其他核蛋白碰撞的问题仍然悬而未决。我们的假设是 DNA 马达通过顺序协作来破坏核小体和其他障碍的稳定取代并驱逐障碍，从而使其他修复酶能够接触到 DNA。我已经开始通过直接可视化这些碰撞来解决 DNA 马达如何克服障碍在单分子水平上。我观察到 RecBCD，一种原核 DNA 修复马达，取代了多种类型当它沿着 DNA 移动时会遇到障碍。在 K99 阶段，我将扩展我的单分子测定来研究 Sgs1 的运动特性。在R00阶段，我将阐明Top3/Rmi1和Dna2在促进 Sgs1 依赖性真核 DNA 修复。我在 R00 阶段的第二个目标是确定如何 Sgs1/Top3/Rmi1 复合物处理染色质。这些实验将依赖于开发的新技术在 Greene 实验室中，我们可以直接真实地观察数百个单独的 DNA 马达蛋白时间。通过快速收集统计相关的数据集，我们可以研究同源重组机械细节达到前所未有的水平。我的最终职业目标是在某大学获得终身教授职位研究机构。我在奖学金 K99 阶段培养的技能将使我取得成功作为一名独立调查员。