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Mol. Cell. Biol. doi:10.1128/MCB.00987-08
Copyright (c) 2008, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Non-canonical E2 variant-independent function of UBC13 in promoting checkpoint protein assembly

Department of Therapeutic Radiology, Yale University School of Medicine, New Haven, CT 06520; Department of Host Defense, Research Institute for Microbial Diseases, Osaka University, Osaka 565-0871, Japan; Department of Microbiology and Immunology, University of Saskatchewan, Saskatoon, SK, Canada S7N 5E5

^* To whom correspondence should be addressed. Email: junjie.chen{at}yale.edu.

	Abstract

The E2 ubiquitin conjugating enzyme UBC13 plays pivotal roles in diverse biological processes. Recent studies have elucidated that UBC13, in concert with the E3 ubiquitin ligase RNF8, propagates the DNA damage signal via an ubiquitylation-dependent signaling pathway. However, mechanistically how UBC13 mediates its role in promoting checkpoint protein assembly and its genetic requirement for E2 variants remains elusive. Here we provide evidence to support that the E3 ubiquitin ligase complex RNF8/UBC13 functions independently of E2 variants, and is sufficient in facilitating ubiquitin conjugations and accumulation of DNA damage mediator 53BP1 at DNA breaks. The RNF8 RING domain serves as the molecular platform to anchor UBC13 at the damaged chromatin, where localised ubiquitylation events allow sustained accumulation of checkpoint proteins. Intriguingly, we found that only a group of RING domains derived from E3 ubiquitin ligases, which have been shown to interact with UBC13, enabled UBC13-mediated FK2 and 53BP1 foci formation at DNA breaks. We propose that the RNF8 RING domain selects and loads a subset of UBC13 molecules, distinct from those that exist as heterodimers, to sites of DSBs which facilitates the amplification of DNA damage signals.