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• W4386933953 abstract "Replication fork reversal is a fundamental process required for resolution of encounters with DNA damage. A key step in the stabilization and eventual resolution of reversed forks is formation of RAD51 nucleoprotein filaments on exposed ssDNA. To avoid genome instability, RAD51 filaments are tightly controlled by a variety of positive and negative regulators. RADX is a recently discovered negative regulator that binds tightly to ssDNA, directly interacts with RAD51, and regulates replication fork reversal and stabilization in a context-dependent manner. Here we present a structure-based investigation of RADX's mechanism of action. Mass photometry experiments showed that RADX forms multiple oligomeric states in a concentration dependent manner, with a predominance of trimers in the presence of ssDNA. The structure of RADX, which has no structurally characterized orthologs, was determined ab initio by cryo-electron microscopy (EM) from maps in the 2-3 Å range. The structure reveals the molecular basis for RADX oligomerization and binding of ssDNA binding. The binding of RADX to RAD51 filaments was imaged by negative stain EM, which showed a RADX oligomer at the end of filaments. Based on these results, we propose a model in which RADX functions by capping and restricting the growing end of RAD51 filaments.Despite the central role of RAD51 in DNA replication and repair processes, the mechanisms of action of its many modulators are poorly understood. Here we combine structural and biophysical data to determine how the negative regulator RADX functions. We show that RADX oligomerizes upon binding DNA, and caps RAD51 filaments at the ends to prevent extension. This work advances knowledge of how RAD51 filaments can be modulated to regulate replication fork reversal and maintain genomic stability." @default.
• W4386933953 created "2023-09-22" @default.
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• W4386933953 date "2023-09-20" @default.
• W4386933953 modified "2023-10-18" @default.
• W4386933953 title "Structure of RADX and mechanism for regulation of RAD51 nucleofilaments" @default.
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• W4386933953 doi "https://doi.org/10.1101/2023.09.19.558089" @default.
• W4386933953 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/37786681" @default.
• W4386933953 hasPublicationYear "2023" @default.
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