Redox regulation of RAD51 Cys319 and homologous recombination by peroxiredoxin 1
John J. Skoko,
Juxiang Cao,
David Gaboriau,
Myriam Attar,
Alparslan Asan,
Lisa Hong,
Candice E. Paulsen,
Hongqiang Ma,
Yang Liu,
Hanzhi Wu,
Trey Harkness,
Cristina M. Furdui,
Yefim Manevich,
Ciaran G. Morrison,
Erika T. Brown,
Daniel Normolle,
Maria Spies,
Michael Ashley Spies,
Kate Carroll,
Carola A. Neumann
Affiliations
John J. Skoko
Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, PA, 15261, USA; Women's Cancer Research Center, University of Pittsburgh Cancer Institute, Pittsburgh, PA, 15213, USA; Magee-Women's Research Institute, Magee-Women's Research Hospital of University of Pittsburgh Medical Center, Pittsburgh, PA, 15213, USA
Juxiang Cao
Department of Cell and Molecular Pharmacology, The Medical University of South Carolina, Charleston, SC, 29425, USA
David Gaboriau
Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland; Facility for Imaging By Light Microscopy, Imperial College London, London, SW7 2AZ, United Kingdom
Myriam Attar
Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, PA, 15261, USA; Women's Cancer Research Center, University of Pittsburgh Cancer Institute, Pittsburgh, PA, 15213, USA; Magee-Women's Research Institute, Magee-Women's Research Hospital of University of Pittsburgh Medical Center, Pittsburgh, PA, 15213, USA
Alparslan Asan
Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, PA, 15261, USA; Women's Cancer Research Center, University of Pittsburgh Cancer Institute, Pittsburgh, PA, 15213, USA; Magee-Women's Research Institute, Magee-Women's Research Hospital of University of Pittsburgh Medical Center, Pittsburgh, PA, 15213, USA
Lisa Hong
Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, PA, 15261, USA; Women's Cancer Research Center, University of Pittsburgh Cancer Institute, Pittsburgh, PA, 15213, USA; Magee-Women's Research Institute, Magee-Women's Research Hospital of University of Pittsburgh Medical Center, Pittsburgh, PA, 15213, USA
Candice E. Paulsen
Department of Chemistry, Scripps Research Institute Florida, Jupiter, FL, 33458, USA
Hongqiang Ma
Biomedical Optical Imaging Laboratory, Departments of Medicine and Bioengineering, University of Pittsburgh, Pittsburgh, PA, 15261, USA
Yang Liu
Biomedical Optical Imaging Laboratory, Departments of Medicine and Bioengineering, University of Pittsburgh, Pittsburgh, PA, 15261, USA
Hanzhi Wu
Department of Internal Medicine, Wake Forest School of Medicine, Winston-Salem, NC, 27157, USA; Center for Redox Biology and Medicine, Wake Forest School of Medicine, Winston-Salem, NC, 27157, USA
Trey Harkness
Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, PA, 15261, USA
Cristina M. Furdui
Department of Internal Medicine, Wake Forest School of Medicine, Winston-Salem, NC, 27157, USA; Center for Redox Biology and Medicine, Wake Forest School of Medicine, Winston-Salem, NC, 27157, USA
Yefim Manevich
Department of Cell and Molecular Pharmacology, The Medical University of South Carolina, Charleston, SC, 29425, USA
Ciaran G. Morrison
Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland
Erika T. Brown
Dartmouth Geisel School of Medicine, Hanover, NH, 03755, USA
Daniel Normolle
Department of Biostatistics, University of Pittsburgh, Pittsburgh, PA, 15213, USA
Maria Spies
Department of Biochemistry and Molecular Biology, University of Iowa, IA, 52242, USA
Michael Ashley Spies
Department of Biochemistry and Molecular Biology, Department of Pharmaceutical Sciences and Experimental Therapeutics, University of Iowa, IA, 52242, USA
Kate Carroll
Department of Chemistry, Scripps Research Institute Florida, Jupiter, FL, 33458, USA
Carola A. Neumann
Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, PA, 15261, USA; Women's Cancer Research Center, University of Pittsburgh Cancer Institute, Pittsburgh, PA, 15213, USA; Magee-Women's Research Institute, Magee-Women's Research Hospital of University of Pittsburgh Medical Center, Pittsburgh, PA, 15213, USA; Corresponding author. Department of Pharmacology and Chemical Biology University of Pittsburgh 204 Craft Ave, Pittsburgh, 15221, PA, USA.
RAD51 is a critical recombinase that functions in concert with auxiliary mediator proteins to direct the homologous recombination (HR) DNA repair pathway. We show that Cys319 RAD51 possesses nucleophilic characteristics and is important for irradiation-induced RAD51 foci formation and resistance to inhibitors of poly (ADP-ribose) polymerase (PARP). We have previously identified that cysteine (Cys) oxidation of proteins can be important for activity and modulated via binding to peroxiredoxin 1 (PRDX1). PRDX1 reduces peroxides and coordinates the signaling actions of protein binding partners. Loss of PRDX1 inhibits irradiation-induced RAD51 foci formation and represses HR DNA repair. PRDX1-deficient human breast cancer cells and mouse embryonic fibroblasts display disrupted RAD51 foci formation and decreased HR, resulting in increased DNA damage and sensitization of cells to irradiation. Following irradiation cells deficient in PRDX1 had increased incorporation of the sulfenylation probe DAz-2 in RAD51 Cys319, a functionally-significant, thiol that PRDX1 is critical for maintaining in a reduced state. Molecular dynamics (MD) simulations of dT-DNA bound to a non-oxidized RAD51 protein showed tight binding throughout the simulation, while dT-DNA dissociated from an oxidized Cys319 RAD51 filament. These novel data establish RAD51 Cys319 as a functionally-significant site for the redox regulation of HR and cellular responses to IR.
