Early events in double-strand break repair in local, genomic and metabolic contexts

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Project Summary / Abstract Chromosomal rearrangements have multiple impacts on human health in cancer, inherited genetic disease, and normal tissue function. While less frequent than single-nucleotide changes, structural variants are disproportionately impactful because they alter genome continuity and change many base pairs at once. Rearrangements mainly arise through DNA double-strand breaks (DSBs) because they disrupt chromosomal integrity. Over many years this project has engaged basic studies of the molecular mechanisms of DSB repair and chromosomal mutagenesis conducted mainly in yeast, specifically nonhomologous end joining (NHEJ) and how it and other repair pathways such as homologous recombination (HR) contribute to genome maintenance. We will continue to exploit yeast to explore a set of critical contextual influences that determine the flux through DSB repair toward variable outcomes, focusing on contexts for which a single-cell eukaryote is an ideal experimental system. This project will specifically investigate some of the earliest events in DSB repair that occur soon after recognition of the break and coincident with the commitment to a repair mechanism. At the smallest scale, we will exploit novel single-base resolution resection and protein occupancy assays to explore the impact of local sequence on DSB repair, in particular on specific protein functions in the initial stages of DSB resection in vivo. At the genomic scale, we will follow up our recent findings to understand how nuclear and functional properties of different DSB locations influence mutagenic outcome frequencies through DSB movement. At the cellular scale, we will expand our focus to investigate the mechanisms by which metabolic signaling in response to carbon source interacts with the DNA damage response and cell cycle, which our prior work identifies as an important and underexplored aspect of DSB repair regulation. Results will provide novel insights into how these cellular processes normally preserve the genome and how rearrangements result when they are perturbed by environmental or genetic factors.