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: 2335208
• 负责人: Jaime Lopez-Mosqueda
• 金额: $ 95万
• 依托单位: University of Texas at Austin
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-03-15 至 2027-04-30
• 项目状态: 未结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2335208&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的及时和准确复制对所有真核生物的生长至关重要，但复制过程容易受到DNA损伤的阻碍，如DNA-蛋白质交联键。持续的DNA-蛋白质交联会阻止染色体的分离和传递到子代细胞；因此，DNA-蛋白质交联物的去除对细胞的生存至关重要。这个项目的首要目标是确定细胞如何选择不同的细胞路径来绕过或修复DNA-蛋白质交联链。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-蛋白质交联物以进行蛋白质降解。此外，该项目将吸引生活在农村地区的不同群体的学生提供培训、技能和科学研究经验，并促进他们参与STEM职业生涯。该研究项目旨在阐明选择性靶向和移除DNA-蛋白质交联物潜在的生化和分子机制，这些交联物在物理上阻碍DNA复制、基因转录和染色质重塑。多条非冗余通路协同修复或绕过DNA-蛋白质交联链。SPRTN是致力于DNA-蛋白质交联修复的哺乳动物蛋白水解酶的创始成员，此后在多个王国的物种中发现了几种类似物和直系物，突显了DNA-蛋白质交联修复的保守性。鉴于大量蛋白质可以共价捕获在DNA上，SPRTN如何在拥挤的染色质相关蛋白质中选择性地将它们移除仍是未知的。目标1将研究一种以DNA-蛋白质交联物为靶标的统一机制，而不考虑其大小和化学组成。目的2将研究SPRTN蛋白水解酶活性如何下调，以防止染色质相关蛋白的异常破坏。目的3是一种遗传方法，用来确定对酿酒酵母萌芽中的DNA-蛋白质交联修复至关重要的其他因素。最后一个目标是让农村学校的高中生在自己的教室里进行筛选实验，从而为他们提供STEM职业发展和指导机会。该项目由生物科学局分子和细胞生物科学部门的遗传机制计划和既定的激励竞争研究计划(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智力优势和更广泛的影响审查标准进行评估，被认为值得支持。