Determining the factors that impact single stranded DNA mutagenesis

确定影响单链 DNA 突变的因素

• 批准号: 10713599
• 负责人: Natalie Saini
• 金额: $ 37.75万
• 依托单位: MEDICAL UNIVERSITY OF SOUTH CAROLINA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-08-01 至 2028-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10713599
• 关键词: Acetaldehyde; Alkylating Agents; Bypass; Cell Culture Techniques; Cell Cycle; Cells; Complex; DNA Binding; DNA Damage; DNA Repair; DNA damage checkpoint; DNA glycosylase; Data; Genetic; Genomic Instability; Goals; Human; Individual; Knowledge; Laboratories; Malignant Neoplasms; Mutagenesis; Mutagens; Mutation; Outcome; Pathway interactions; Pattern; Polymerase; Positioning Attribute; Predisposition; Preventive measure; Proteins; Reporter; Research; Research Project Grants; Role; SS DNA BP; Scientific Advances and Accomplishments; Single-Stranded DNA; Specificity; Stretching; System; Work; Yeasts; bioinformatics tool; interdisciplinary approach; large datasets; next generation sequencing; prevent; programs; replication factor A; tool

Abstract Single stranded DNA (ssDNA) has been demonstrated to by extremely vulnerable to DNA damage. Cancers often carry long stretches of clustered mutations that likely arose due to damage of ssDNA. While, various studies have demonstrated that certain mutagens preferentially damage ssDNA, the mechanisms that alter mutation specificity due to damage in ssDNA and the pathways that prevent mutagenesis at ssDNA are unknown. The overarching goal of this proposal is to specifically identify the roles of DNA damage checkpoint proteins, translesion polymerases, ssDNA-specific glycosylases and ssDNA binding proteins in altering the mutation patterns obtained upon ssDNA-specific damage. My laboratory is in a unique position to advance this scientific front based on my strong track record in DNA damage and repair, assembled team of collaborators, and multidisciplinary approach. My expertise in using highly sensitive yeast reporter systems, human cell culture techniques and the use of bioinformatics tools to probe large data sets and to analyze next generation sequencing data allow us to develop our research program to understand the pathways modulating ssDNA mutagenesis in yeast and human cells. Previously, I have demonstrated that alkylating agents and acetaldehyde have an ssDNA-specific mutation signature in yeast and in cancers. These mutation signatures provide us with a highly sensitive tool to determine how changes in various DNA repair, damage bypass and damage sensing pathways alter mutagenesis by ssDNA-specific mutagens. Here, we propose to determine 1) How cell cycle dependent translesion polymerase expression alters mutation signatures in ssDNA; 2) The role of ssDNA binding Replication Protein A complex in protecting ssDNA from exogenous damage; 3) The role of DNA damage checkpoint activation in modulating the mutation signatures associated with ssDNA damage; 4) Which DNA glycosylases function on ssDNA and alter the mutation signatures due to ssDNA damage; and 5) What are the mutagenic outcomes when translesion polymerases are unable to bypass ssDNA damage. This set of research projects will address a key gap in knowledge in understanding the mechanisms that alter the hypermutability of ssDNA in cells. Our work will enable us to identify and develop better cancer preventative measures for individuals who are prone to increased genome instability and ssDNA formation in their cells.

抽象的 非常容易受到DNA损伤的影响，已经证明了单链DNA（ssDNA）。癌症 通常携带长长的聚集突变，这些突变可能由于ssDNA损坏而产生。而，各种各样 研究表明，某些诱变者优先损害ssDNA，这是改变的机制 由于ssDNA的损害和预防ssDNA诱变的途径引起的突变特异性是 未知。该提案的总体目标是专门确定DNA损伤检查点的作用 蛋白质，跨质聚合酶，ssDNA特异性糖基酶和ssDNA结合蛋白在改变 在ssDNA特异性损伤上获得的突变模式。我的实验室处于独特的位置，可以推进这一点 根据我在DNA损伤和维修方面的良好往绩，科学阵线，组装的合作者团队， 和多学科方法。我使用高敏感酵母记系统的专业知识，人类细胞 文化技术和使用生物信息学工具探测大型数据集并分析下一代 测序数据使我们能够制定我们的研究计划，以了解调节ssDNA的途径 酵母和人类细胞中的诱变。以前，我已经证明了烷基化剂和 乙醛在酵母和癌症中具有ssDNA特异性突变。这些突变特征 为我们提供高度敏感的工具，以确定各种DNA修复，损坏旁路和如何变化 损伤传感途径改变了ssDNA特异性诱变剂的诱变。在这里，我们建议确定1） 细胞循环依赖性跨性酶聚合酶表达如何改变ssDNA中的突变特征； 2）角色 ssDNA结合复制蛋白A在保护ssDNA免受外源损伤的过程中的复合物； 3） DNA损伤检查点在调节与ssDNA损伤相关的突变特征时激活； 4） 哪种DNA糖基酶在ssDNA上起作用并因ssDNA损伤而改变突变特征。 5） 当转移聚合酶无法绕过ssDNA损伤时，诱变的结果是什么。这 一组研究项目将解决了解改变机制的知识的关键差距 细胞中ssDNA的过度可显性性。我们的工作将使我们能够识别和发展更好的癌症预防效果 对于容易增加基因组不稳定性和细胞中ssDNA形成的个体的措施。