RRC ID 54484
Author Inano S, Sato K, Katsuki Y, Kobayashi W, Tanaka H, Nakajima K, Nakada S, Miyoshi H, Knies K, Takaori-Kondo A, Schindler D, Ishiai M, Kurumizaka H, Takata M.
Title RFWD3-Mediated Ubiquitination Promotes Timely Removal of Both RPA and RAD51 from DNA Damage Sites to Facilitate Homologous Recombination.
Journal Mol Cell
Abstract RFWD3 is a recently identified Fanconi anemia protein FANCW whose E3 ligase activity toward RPA is essential in homologous recombination (HR) repair. However, how RPA ubiquitination promotes HR remained unknown. Here, we identified RAD51, the central HR protein, as another target of RFWD3. We show that RFWD3 polyubiquitinates both RPA and RAD51 in vitro and in vivo. Phosphorylation by ATR and ATM kinases is required for this activity in vivo. RFWD3 inhibits persistent mitomycin C (MMC)-induced RAD51 and RPA foci by promoting VCP/p97-mediated protein dynamics and subsequent degradation. Furthermore, MMC-induced chromatin loading of MCM8 and RAD54 is defective in cells with inactivated RFWD3 or expressing a ubiquitination-deficient mutant RAD51. Collectively, our data reveal a mechanism that facilitates timely removal of RPA and RAD51 from DNA damage sites, which is crucial for progression to the late-phase HR and suppression of the FA phenotype.
Volume 66(5)
Pages 622-634.e8
Published 2017-6-1
DOI 10.1016/j.molcel.2017.04.022
PII S1097-2765(17)30278-2
PMID 28575658
MeSH Adenosine Triphosphatases / genetics Adenosine Triphosphatases / metabolism Ataxia Telangiectasia Mutated Proteins / metabolism Binding Sites Cell Cycle Proteins / genetics Cell Cycle Proteins / metabolism Cell Line, Tumor Chromatin / drug effects Chromatin / enzymology* Chromatin / genetics Chromatin / radiation effects DNA / genetics DNA / metabolism* DNA Damage* Fanconi Anemia / enzymology* Fanconi Anemia / genetics Humans Minichromosome Maintenance Proteins / metabolism Mitomycin / pharmacology Mutation Phosphorylation Proteasome Endopeptidase Complex / metabolism Protein Binding Proteolysis RNA Interference Rad51 Recombinase / genetics Rad51 Recombinase / metabolism* Recombinational DNA Repair* / drug effects Recombinational DNA Repair* / radiation effects Replication Protein A / genetics Replication Protein A / metabolism* Transfection Ubiquitin-Protein Ligases / genetics Ubiquitin-Protein Ligases / metabolism* Ubiquitination* Valosin Containing Protein
IF 14.548
Times Cited 44
DNA material CSII-CMV-MCS-IRES2-Bsd (RDB04385) CSIV-TRE-RfA-UbC-KT (RDB12878) pcDNA3.1 FLAG-hRFWD3 wt (RDB17490) pcDNA3.1 FLAG-hRFWD3 C315A (RDB17491) pcDNA3.1 FLAG-hRFWD3 I639K (RDB17492) pcDNA3.1 FLAG-hRFWD3 S46/63A (RDB17493) pcDNA3.1 FLAG-hRFWD3 aa 1-278 (RDB17494) pcDNA3.1 FLAG-hRFWD3 LQP/AAA (RDB17495) CSII FLAG-hRFWD3 wt (RDB17496) CSII FLAG-hRFWD3 C315A (RDB17497) CSII FLAG-hRFWD3 I639K (RDB17498) CSII FLAG-hRFWD3 S46/63A (RDB17499) CSII FLAG-hRFWD3 C315A/I639K (RDB17500) CSII FLAG-hRFWD3 LQP/AAA (RDB17501) pENTR hRFWD3 (RDB17502)