CAREER: Uncovering Mechanisms of DNA-protein Crosslink Repair

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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.