Mechanisms of RNA-DNA recombination

RNA-DNA 重组机制

• 批准号: 1615335
• 负责人: Francesca Storici
• 金额: $ 69万
• 依托单位: Georgia Tech Research Corporation
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-08-15 至 2020-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1615335&HistoricalAwards=false
• 关键词: Mechanisms; RNA; DNA; recombination

A break in both strands of the DNA double helix is one of the most harmful DNA lesions because if not properly repaired it leads to mutations, chromosome rearrangements or cell death. The safest way to repair a DNA double-strand break (DSB) is through a process called homologous recombination (HR). The known mechanisms of HR utilize an intact copy of the DNA sequence as a template to retrieve the information lost at the site of a DSB. However, recent results, which will be expanded in this study, indicate a new role of RNA in this process of DNA repair. Therefore, the study will provide important new biological insights to better understanding the physiological function of RNA in DNA repair, and the impact of RNA on genome maintenance and evolution. A postdoctoral fellow, a graduate student and several undergraduate students will be involved in the research. The work and findings of this proposal will be integrated in class topics and activities for many graduate and undergraduate students. Through the inclusion of a Research Experience for Teachers (RET), the project will also target students from a local, 100%-minority High School to support student interest and participation in Science, Technology, Engineering and Math (STEM) programs.To characterize the mechanism of RNA-DNA recombination, novel genetic systems will be developed to study repair of chromosomal double-strand breaks (DSBs) by homologous transcript RNA in budding yeast cells. One Objective of this project will be to identify the DNA polymerization function/s that use RNA as template in RNA-DNA recombination. Another focus will be to examine whether the process of DSB repair by transcript RNA can be visualized in cells arrested at two distinct points in the cell cycle: before and after DNA replication. It was shown that DSB repair by transcript RNA in yeast cells is initiated efficiently, but it is quickly suppressed by cellular ribonuclease H enzymes. Here, refined genetic systems to detect DSB repair by RNA will be utilized to determine the efficiency of DSB repair by transcript RNA in wild-type and ribonuclease-defective cells. Overall, the work of this project will help to better understand the conditions and mechanism in which transcript RNA is a preferred direct template for DSB repair in cells.This project is funded by the Genetic Mechanisms Program in the Division of Molecular and Cellular Biosciences.

DNA双螺旋的两条链的断裂是最有害的DNA损伤之一，因为如果没有正确修复，它会导致突变，染色体重排或细胞死亡。修复DNA双链断裂（DSB）的最安全方法是通过一个称为同源重组（HR）的过程。HR的已知机制利用DNA序列的完整拷贝作为模板来恢复在DSB位点处丢失的信息。然而，最近的研究结果表明，RNA在DNA修复过程中发挥了新的作用。因此，该研究将为更好地理解RNA在DNA修复中的生理功能以及RNA对基因组维持和进化的影响提供重要的新生物学见解。一名博士后、一名研究生和几名本科生将参与这项研究。这项提案的工作和发现将被整合到许多研究生和本科生的课堂主题和活动中。通过纳入教师研究经验（RET），该项目还将针对当地100%少数民族高中的学生，以支持学生对科学，技术，工程和数学（STEM）课程的兴趣和参与。为了表征RNA-DNA重组的机制，将开发新的遗传系统来研究芽殖酵母细胞中同源转录RNA对染色体双链断裂（DSB）的修复。本研究的目的之一是鉴定以RNA为模板的DNA聚合功能。另一个重点将是检查转录RNA的DSB修复过程是否可以在细胞周期中的两个不同点（DNA复制之前和之后）被阻止的细胞中可视化。结果表明，在酵母细胞中转录RNA的DSB修复是有效启动的，但它很快被细胞核糖核酸酶H酶抑制。在这里，将利用改进的遗传系统来检测RNA的DSB修复，以确定野生型和核糖核酸酶缺陷细胞中转录RNA的DSB修复效率。总的来说，本项目的工作将有助于更好地理解转录本RNA作为细胞中DSB修复的首选直接模板的条件和机制。