TY - JOUR
T1 - RFWD3-Mediated Ubiquitination Promotes Timely Removal of Both RPA and RAD51 from DNA Damage Sites to Facilitate Homologous Recombination
AU - Inano, Shojiro
AU - Sato, Koichi
AU - Katsuki, Yoko
AU - Kobayashi, Wataru
AU - Tanaka, Hiroki
AU - Nakajima, Kazuhiro
AU - Nakada, Shinichiro
AU - Miyoshi, Hiroyuki
AU - Knies, Kerstin
AU - Takaori-Kondo, Akifumi
AU - Schindler, Detlev
AU - Ishiai, Masamichi
AU - Kurumizaka, Hitoshi
AU - Takata, Minoru
N1 - Funding Information:
We would like to thank Professor James Hejna (Graduate School of Biostudies, Kyoto University) for critical reading of the manuscript; Drs. Makoto Nakanishi, Yoshikazu Johmura, Maria Jasin, Kiyoshi Miyagawa, and Aziz Sancar for reagents; and Ms. Akiko Watanabe, Seiko Arai, and Fan Peng for technical or secretarial support. Our work is supported by JSPS KAKENHI grant numbers JP23114010 (to M.T.), JP26550026 (to M.T.), JP15H01738 (to M.T.), JP26281021 (to M.I.), JP25116002 (to H.K.), JP25250023 (to H.K.), and JP15H01183 (to S.N.) and grants from The Ministry of Health, Labour and Welfare (16ek0109099h0002, to M.T.). H.K. was also supported by Waseda University.
Publisher Copyright:
© 2017 Elsevier Inc.
PY - 2017/6/1
Y1 - 2017/6/1
N2 - 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.
AB - 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.
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U2 - 10.1016/j.molcel.2017.04.022
DO - 10.1016/j.molcel.2017.04.022
M3 - Article
C2 - 28575658
AN - SCOPUS:85020104262
VL - 66
SP - 622-634.e8
JO - Molecular Cell
JF - Molecular Cell
SN - 1097-2765
IS - 5
ER -