Orchestration of the DNA-Damage Response by the RNF8 Ubiquitin Ligase
Nadine K. Kolas,1*
J. Ross Chapman,2*
Shinichiro Nakada,1*
Jarkko Ylanko,1,3
Richard Chahwan,2
Frédéric D. Sweeney,1,3
Stephanie Panier,1
Megan Mendez,1
Jan Wildenhain,1
Timothy M. Thomson,4
Laurence Pelletier,1,3
Stephen P. Jackson,2
Daniel Durocher1,3
Cells respond to DNA double-strand breaks by recruiting factors such as the DNA-damage mediator protein MDC1, the p53-binding protein 1 (53BP1), and the breast cancer susceptibility protein BRCA1 to sites of damaged DNA. Here, we reveal that the ubiquitin ligase RNF8 mediates ubiquitin conjugation and 53BP1 and BRCA1 focal accumulation at sites of DNA lesions. Moreover, we establish that MDC1 recruits RNF8 through phosphodependent interactions between the RNF8 forkhead-associated domain and motifs in MDC1 that are phosphorylated by the DNA-damage activated protein kinase ataxia telangiectasia mutated (ATM). We also show that depletion of the E2 enzyme UBC13 impairs 53BP1 recruitment to sites of damage, which suggests that it cooperates with RNF8. Finally, we reveal that RNF8 promotes the G2/M DNA damage checkpoint and resistance to ionizing radiation. These results demonstrate how the DNA-damage response is orchestrated by ATM-dependent phosphorylation of MDC1 and RNF8-mediated ubiquitination.
1 Samuel Lunenfeld Research Institute, Mount Sinai Hospital, 600 University Avenue, Toronto M5G1X5, Ontario, Canada.
2 The Wellcome Trust and Cancer Research UK Gurdon Institute, and the Department of Zoology, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK.
3 Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada.
4 Department of Molecular and Cellular Biology, Instituto de Biología Molecular de Barcelona calle Jordi Girona 18-26, 08034 Barcelona, Spain.
* These authors contributed equally to this work.
To whom correspondence should be addressed. E-mail: durocher{at}mshri.on.ca (D.D.); s.jackson{at}gurdon.cam.ac.uk (S.P.J.)
