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bioRxiv · 10.1101/2025.11.17.685403

The trimeric Shu complex in C. elegans is an ATPase that remodels RAD51 filaments in the homologous recombination-associated DNA damage response.

Chu, S. S. H.·Xing, G.·Ling, H.
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Abstract

Homologous recombination (HR) is critical to error-free lesion bypass in the DNA damage response. This error-free lesion tolerance pathway is initiated by the RAD51 recombinase, which forms nucleoprotein filaments on single-stranded DNA (ssDNA) to facilitate template-directed repair using homologous sister chromatids. RAD51 filaments are tightly regulated by RAD51 mediator proteins. Among these, the Shu complexes facilitate HR-directed DNA damage tolerance and are evolutionarily conserved from yeast to humans. The Caenorhabditis elegans Shu complex is a heterotrimer consisting of three protein subunits: RFS1, RIP1, and SWS1. However, the biochemical properties of this trimeric complex remain unclear. Here, we report the biochemical characterization of the C. elegans Shu complex and interactions with DNA, ATP, and Rad51 filaments. We first revealed that the Shu trimer preferentially binds DNA with an exposed 5' end, particularly favoring a fork-shaped double-stranded DNA (dsDNA). Then, we found that the trimer binds to ATP and exhibits DNA-dependent ATPase activity. Through site-specific mutagenesis, we identified the catalytic residues in the RFS1 domain and validated the ATPase activity. Using fluorescence-based assays, we further demonstrated that the Shu trimer remodels RAD51 filaments in an ATP-hydrolysis-dependent manner and stabilizes the filaments in an ATP-binding-dependent manner. These findings provide key mechanistic insights into how the C. elegans Shu complex regulates RAD51 filaments, priming them for downstream HR-mediated DNA repair processes.

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Chu, S. S. H., Xing, G., Ling, H.. 2025-11-18. The trimeric Shu complex in C. elegans is an ATPase that remodels RAD51 filaments in the homologous recombination-associated DNA damage response.. https://doi.org/10.1101/2025.11.17.685403

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