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.

摘要 单链DNA（ssDNA）已被证明非常容易受到DNA损伤。癌症 通常携带很长一段簇状突变，可能是由于ssDNA的损伤而引起的。虽然，各种 研究表明，某些诱变剂优先损伤ssDNA，改变ssDNA的机制是， 由于ssDNA中的损伤引起的突变特异性和防止ssDNA处的诱变的途径是 未知该提案的总体目标是明确确定DNA损伤检查点的作用 蛋白质、跨损伤聚合酶、ssDNA特异性糖基化酶和ssDNA结合蛋白在改变肿瘤细胞中的表达中的作用。 在ssDNA特异性损伤后获得的突变模式。我的实验室处于一个独特的位置来推进这一点 科学前沿基于我在DNA损伤和修复方面的良好记录，组建了合作者团队， 多学科的方法。我在使用高灵敏度酵母报告系统，人类细胞 培养技术和使用生物信息学工具来探测大型数据集并分析下一代 测序数据使我们能够发展我们的研究计划，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，烷基化剂和 乙醛在酵母和癌症中具有ssDNA特异性突变特征。这些变异特征 为我们提供了一个高度敏感的工具，以确定如何在各种DNA修复，损害旁路和 损伤感应途径改变ssDNA特异性诱变剂的诱变。在这里，我们建议确定1） 细胞周期依赖性跨损伤聚合酶表达如何改变ssDNA中的突变特征; 2） ssDNA结合复制蛋白A复合物在保护ssDNA免受外源性损伤中的作用; 在调节与ssDNA损伤相关的突变特征中的DNA损伤检查点激活; 4） 哪些DNA糖基化酶在ssDNA上起作用并由于ssDNA损伤而改变突变特征;以及5） 当跨损伤聚合酶不能绕过ssDNA损伤时，会产生什么样的致突变结果？这 一系列研究项目将解决在理解改变人类行为的机制方面的一个关键知识缺口。 ssDNA在细胞中的超突变性。我们的工作将使我们能够识别和开发更好的癌症预防 这些措施适用于那些容易在细胞中增加基因组不稳定性和ssDNA形成的个体。