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Published March 31, 2016 | Submitted + Published
Journal Article Open

Structural and kinetic analysis of the COP9-Signalosome activation and the cullin-RING ubiquitin ligase deneddylation cycle


The COP9-Signalosome (CSN) regulates cullin–RING ubiquitin ligase (CRL) activity and assembly by cleaving Nedd8 from cullins. Free CSN is autoinhibited, and it remains unclear how it becomes activated. We combine structural and kinetic analyses to identify mechanisms that contribute to CSN activation and Nedd8 deconjugation. Both CSN and neddylated substrate undergo large conformational changes upon binding, with important roles played by the N-terminal domains of Csn2 and Csn4 and the RING domain of Rbx1 in enabling formation of a high affinity, fully active complex. The RING domain is crucial for deneddylation, and works in part through conformational changes involving insert-2 of Csn6. Nedd8 deconjugation and re-engagement of the active site zinc by the autoinhibitory Csn5 glutamate-104 diminish affinity for Cul1/Rbx1 by ~100-fold, resulting in its rapid ejection from the active site. Together, these mechanisms enable a dynamic deneddylation-disassembly cycle that promotes rapid remodeling of the cellular CRL network.

Additional Information

© 2016, Mosadeghi et al. This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited. Received October 5, 2015. Accepted March 30, 2016. Published March 31, 2016. We thank B Schulman for Skp1/Skp2 protein as well as for expression plasmids and E Morris for thin carbon for EM grids. We thank A Bernini and A Ragheb for technical assistance with protein work, P Tittmann for technical support at the ScopeM facility, members of the Ban lab for advice and Annie Moradian and Roxana Eggleston-Rangel for mass spectrometry support at PEL. We also are grateful to SO Shan, E Morris and T Stuwe for advice and A Smith, SO Shan, and D Barford for comments on the manuscript. RM was supported by a Lee-Ramo Life Sciences Fellowship, RIE was supported by an ETH Pioneer, a Marie Curie and an EMBO short-term fellowship, and AS by a Marie Curie fellowship. FS acknowledges funding from the Wellcome Trust (Grant 095951) and the German Science Foundation Collaborative Research Center (SFB) 969. The Peter laboratory is funded by an ERC advanced grant, the SNF and ETHZ, and the Aebersold laboratory is supported by ETH Zurich, SystemsX.ch and an ERC advanced grant. This work was supported in part by NIH CA164803 to RJD RJD is an Investigator of and was supported by the Howard Hughes Medical Institute. MJS and SH were supported by the Gordon and Betty Moore Foundation, through Grant GBMF775 and the Beckman Institute. JMR was supported by National Institute of General Medical Sciences award F32GM112308. Competing interests: Raymond J Deshaies, Reviewing editor, eLife. The other authors declare that no competing interests exist.

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