CAREER: Uncovering Mechanisms of DNA-protein Crosslink Repair

职业：揭示 DNA-蛋白质交联修复机制

• 批准号: 2146384
• 负责人: Jaime Lopez-Mosqueda
• 金额: $ 95万
• 依托单位: South Dakota State University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-05-01 至 2023-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2146384&HistoricalAwards=false
• 关键词: CAREER; Uncovering; Mechanisms; DNA; protein

Timely and accurate duplication of DNA is essential for the growth of all eukaryotic organisms, but the replication process is subject to obstruction by DNA damage such as DNA-protein crosslinks. Sustained DNA-protein crosslinks prevent segregation and transmission of chromosomes to daughter cells; hence, removal of DNA-protein crosslinks is critically important for cell survival. The overarching goal of this project is to determine how cells choose distinct cellular pathways to bypass or repair DNA-protein crosslinks. A multidisciplinary strategy combining biochemistry, molecular biology and yeast genetics will be employed to investigate how cells mark DNA-protein crosslinks for proteolytic degradation. Additionally, the project will engage a diverse group of students living in rural areas to provide training, skills, and experiences in scientific research and promote their participation in STEM careers.The research project aims to elucidate the biochemical and molecular mechanisms underlying selective targeting and removal of DNA-protein crosslinks that physically impede DNA replication, gene transcription and chromatin remodeling. Multiple, non-redundant pathways cooperate to repair or bypass DNA-protein crosslinks. SPRTN is the founding member of mammalian proteases dedicated to DNA-protein crosslink repair and several paralogues and orthologues have since been identified in species across multiple kingdoms, underscoring the conserved nature of DNA-protein crosslink repair. Given the multitude of proteins that can be covalently trapped on DNA, how SPRTN selectively targets them for removal in a crowded landscape of chromatin-associated proteins remains unknown. Aim 1 will investigate a unifying mechanism to target DNA-protein crosslinks, irrespective of their size and chemical composition. Aim 2 will investigate how SPRTN proteolytic activity is down-regulated to prevent aberrant destruction of chromatin-associated proteins. Aim 3 is a genetic approach to identify additional factors important for DNA-protein crosslink repair in the budding yeast Saccharomyces cerevisiae. The last goal involves high school students from rural schools performing screening experiments in their own classrooms, and thus provides them with STEM career development and mentoring opportunities.This project is jointly funded by the Genetic Mechanisms program in the Molecular and Cellular Biosciences Division of the Biological Sciences Directorate and the Established Program to Stimulate Competitive Research (EPSCoR).This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

DNA的及时和准确的复制对于所有真核生物的生长是必不可少的，但是复制过程受到DNA损伤如DNA-蛋白质交联的阻碍。持续的DNA-蛋白质交联防止染色体分离和传递到子细胞;因此，去除DNA-蛋白质交联对细胞存活至关重要。该项目的首要目标是确定细胞如何选择不同的细胞途径来绕过或修复DNA-蛋白质交联。结合生物化学，分子生物学和酵母遗传学的多学科策略将被用来研究细胞如何标记DNA-蛋白质交联蛋白水解降解。此外，该项目还将吸引生活在农村地区的不同群体的学生，为他们提供科学研究方面的培训、技能和经验，并促进他们参与STEM职业。该研究项目旨在阐明选择性靶向和去除DNA-蛋白质交联的生物化学和分子机制，这些交联在物理上阻碍DNA复制、基因转录和染色质重塑。多个非冗余途径合作修复或绕过DNA-蛋白质交联。SPRTN是致力于DNA-蛋白质交联修复的哺乳动物蛋白酶的创始成员，并且此后在多个王国的物种中鉴定了几种旁系同源物和直系同源物，强调了DNA-蛋白质交联修复的保守性质。考虑到可以共价捕获在DNA上的蛋白质的数量，SPRTN如何选择性地将它们靶向去除在拥挤的染色质相关蛋白质中仍然是未知的。目的1将研究一个统一的机制，以靶向DNA-蛋白质交联，无论其大小和化学组成。目的2将研究SPRTN蛋白水解活性是如何下调，以防止异常破坏染色质相关蛋白。目的3是一种遗传学方法，以确定在芽殖酵母酿酒酵母DNA-蛋白质交联修复的重要因素。最后一个目标是让来自农村学校的高中生在自己的教室里进行筛查实验，该项目由生物科学理事会分子和细胞生物科学部的遗传机制计划和刺激竞争性研究的既定计划（EPSCoR）共同资助。该奖项反映了NSF的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。