The human Shu complex promotes RAD51 activity by modulating RPA dynamics on ssDNA

Sarah R. Hengel; Katherine G. Oppenheimer; Chelsea M. Smith; Matthew A. Schaich; Hayley L. Rein; Julieta Martino; Kristie E. Darrah; Maggie Witham; Oluchi C. Ezekwenna; Kyle R. Burton; Bennett Van Houten; Maria Spies; Kara A. Bernstein

doi:10.1038/s41467-024-51595-0

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The human Shu complex promotes RAD51 activity by modulating RPA dynamics on ssDNA

Journal article

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The human Shu complex promotes RAD51 activity by modulating RPA dynamics on ssDNA

Sarah R. Hengel, Katherine G. Oppenheimer, Chelsea M. Smith, Matthew A. Schaich, Hayley L. Rein, Julieta Martino, Kristie E. Darrah, Maggie Witham, Oluchi C. Ezekwenna, Kyle R. Burton, …

Nature communications, Vol.15(1), 7197

08/21/2024

DOI: 10.1038/s41467-024-51595-0

PMCID: PMC11339404

PMID: 39169038

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Abstract

Templated DNA repair that occurs during homologous recombination and replication stress relies on RAD51. RAD51 activity is positively regulated by BRCA2 and the RAD51 paralogs. The Shu complex is a RAD51 paralog-containing complex consisting of SWSAP1, SWS1, and SPIDR. We demonstrate that SWSAP1-SWS1 binds RAD51, maintains RAD51 filament stability, and enables strand exchange. Using single-molecule confocal fluorescence microscopy combined with optical tweezers, we show that SWSAP1-SWS1 decorates RAD51 filaments proficient for homologous recombination. We also find SWSAP1-SWS1 enhances RPA diffusion on ssDNA. Importantly, we show human sgSWSAP1 and sgSWS1 knockout cells are sensitive to pharmacological inhibition of PARP and APE1. Lastly, we identify cancer variants in SWSAP1 that alter Shu complex formation. Together, we show that SWSAP1-SWS1 stimulates RAD51-dependent high-fidelity repair and may be an important new cancer therapeutic target. The human Shu complex promotes homologous recombination by regulating RAD51. Here the authors reveal that the Shu complex proteins, SWSAP1-SWS1, decorate the RAD51 filament on ssDNA and facilitate its strand exchange reaction by stimulating RPA diffusion on ssDNA. Lastly, that SWSAP1-SWS1 knockouts are Olaparib sensitive.

Details

Title: Subtitle: The human Shu complex promotes RAD51 activity by modulating RPA dynamics on ssDNA
Creators: Sarah R. Hengel - UPMC Hillman Cancer Center
Katherine G. Oppenheimer - UPMC Hillman Cancer Center
Chelsea M. Smith - UPMC Hillman Cancer Center
Matthew A. Schaich - UPMC Hillman Cancer Center
Hayley L. Rein - UPMC Hillman Cancer Center
Julieta Martino - University of Pittsburgh School of Medicine
Kristie E. Darrah - University of Pennsylvania
Maggie Witham - Tufts University
Oluchi C. Ezekwenna - Tufts University
Kyle R. Burton - Tufts University
Bennett Van Houten - UPMC Hillman Cancer Center
Maria Spies - University of Iowa
Kara A. Bernstein - University of Pennsylvania
Resource Type: Journal article
Publication Details: Nature communications, Vol.15(1), 7197
Publisher: Nature Publishing Group UK
DOI: 10.1038/s41467-024-51595-0
PMID: 39169038
PMCID: PMC11339404
ISSN: 2041-1723
eISSN: 2041-1723
Grant note: U.S. Department of Health & Human Services | National Institutes of Health (NIH): MIRA R35GM131704, 1S10OD032158-01A1 Basser Center for BRCA: K99/R00-ES033738 NIEHS: 2P30CA047904 Hillman Postdoctoral Fellowship for Innovative Cancer Research (SRH): 133947-PF-19-132-01-DMC American Cancer Society Postdoctoral Fellowship: R35ES031638 NIH: 2P30CA047904 Hillman Postdoctoral Fellowship for Innovative Cancer Research: R35GM131704 University of Pittsburgh Health Sciences Research Fellowship: 1T32GM12274
The authors would like to thank Dr. Luke Lavis for generously sharing the Janelia Farm HaloTag dyes. The authors would like to thank Artur Karczmarczyk from David Rueda's lab for helping modulate their scripts to enable our blue and green data analysis. The authors would like to thank Dr. Shixin Liu (Rockefeller University) for providing the AlexaFluor488-RPA protein for use in these studies. K.A.B. is supported by the National Institutes of Health (R01 ES030335 and R01 ES031796), the Penn Center for Genome Integrity, and the Basser Center for BRCA. S.R.H. is supported by the NIEHS [K99/R00-ES033738], Hillman Postdoctoral Fellowship for Innovative Cancer Research (SRH) [2P30CA047904 to the UPMC Hillman Cancer Center], and the American Cancer Society Postdoctoral Fellowship (133947-PF-19-132-01-DMC). NIH [R35ES031638 to B.V.H.]; Hillman Postdoctoral Fellowship for Innovative Cancer Research (M.A.S.) [2P30CA047904 to the UPMC Hillman Cancer Center]. M. S. is supported by the National Institutes of Health MIRA R35GM131704. Lumicks C-trap supported by 1S10OD032158-01A1 (B.V.H.). The M.S. lab and Refyen TwoMP instrument were supported by the NIGMS R35GM131704 grant. C.M.S. is supported by the University of Pittsburgh Health Sciences Research Fellowship and the National Institute of General Medicine Sciences (1T32GM12274).DAS:Authors can confirm that all relevant data are included in the published article in addition to supplementary information files. Source data are provided in this paper.
Language: English
Date published: 08/21/2024
Academic Unit: Radiation Oncology; Biochemistry and Molecular Biology
Record Identifier: 9984699522002771

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