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The CRL4^(DCAF1) cullin-RING ubiquitin ligase is activated following a switch in oligomerization state

Mohamed, Weaam I. and Schenk, Andreas D. and Kempf, Georg and Cavadini, Simone and Basters, Anja and Potenza, Alessandro and Abdul Rahman, Wassim and Rabl, Julius and Reichermeier, Kurt and Thomä, Nicolas H. (2021) The CRL4^(DCAF1) cullin-RING ubiquitin ligase is activated following a switch in oligomerization state. EMBO Journal, 40 (22). Art. No. e108008. ISSN 0261-4189. doi:10.15252/embj.2021108008. https://resolver.caltech.edu/CaltechAUTHORS:20211021-201507835

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Abstract

The cullin-4-based RING-type (CRL4) family of E3 ubiquitin ligases functions together with dedicated substrate receptors. Out of the ˜29 CRL4 substrate receptors reported, the DDB1- and CUL4-associated factor 1 (DCAF1) is essential for cellular survival and growth, and its deregulation has been implicated in tumorigenesis. We carried out biochemical and structural studies to examine the structure and mechanism of the CRL4^(DCAF1) ligase. In the 8.4 Å cryo-EM map of CRL4^(DCAF1), four CUL4-RBX1-DDB1-DCAF1 protomers are organized into two dimeric sub-assemblies. In this arrangement, the WD40 domain of DCAF1 mediates binding with the cullin C-terminal domain (CTD) and the RBX1 subunit of a neighboring CRL4^(DCAF1) protomer. This renders RBX1, the catalytic subunit of the ligase, inaccessible to the E2 ubiquitin-conjugating enzymes. Upon CRL4^(DCAF1) activation by neddylation, the interaction between the cullin CTD and the neighboring DCAF1 protomer is broken, and the complex assumes an active dimeric conformation. Accordingly, a tetramerization-deficient CRL4^(DCAF1) mutant has higher ubiquitin ligase activity compared to the wild-type. This study identifies a novel mechanism by which unneddylated and substrate-free CUL4 ligases can be maintained in an inactive state.


Item Type:Article
Related URLs:
URLURL TypeDescription
https://doi.org/10.15252/embj.2021108008DOIArticle
https://www.ebi.ac.uk/emdb/entry/EMD-12964Related ItemData
https://www.ebi.ac.uk/emdb/entry/EMD-12965Related ItemData
https://www.rcsb.org/structure/7OKQRelated ItemData
ORCID:
AuthorORCID
Mohamed, Weaam I.0000-0002-9389-8686
Schenk, Andreas D.0000-0001-9214-2505
Kempf, Georg0000-0002-0228-6730
Cavadini, Simone0000-0003-2777-7584
Basters, Anja0000-0003-1064-3555
Rabl, Julius0000-0001-6375-852X
Reichermeier, Kurt0000-0003-0613-6905
Thomä, Nicolas H.0000-0003-2685-906X
Alternate Title:The CRL4DCAF1 cullin-RING ubiquitin ligase is activated following a switch in oligomerization state
Additional Information:© 2021 The Authors. Published under the terms of the CC BY NC ND 4.0 license. September 2021; Accepted 10 September 2021; Published online 30 September 2021. We thank Felix Freuler from Novartis Institute of Biomedical Research (NIBR) in Basel for providing the DCAF1 plasmid. Members of the Thomä laboratory for critical discussions. N.H.T was supported by the European Research Council (ERC) under the European Union’s Horizon 2020 Research and Innovation program grant agreement (grant no. 666068), the Novartis Research Foundation, and by the SNF (31003A_179541; 310030_201206). W.I.M. was supported by a PhD fellowship of the Boehringer Ingelheim Fonds from August 2013 to September 2015. We are grateful for Matthias Peter at ETH-Zürich for providing laboratory access to W.I.M. for the completion of this study. We thank C-CINA for providing time on the Polara microscope for imaging the CRL4^(DCAF1)-CSN sample. Data availability: The cryo-EM maps of the CRL4DCAF1 and CRL4DCAF1-CSN complexes are deposited in the Electron Microscopy databank under the accession codes EMD-12964 (https://www.ebi.ac.uk/emdb/entry/EMD-12964) and EMD-12965 (https://www.ebi.ac.uk/emdb/entry/EMD-12965), respectively. The model coordinates for the CRL4DCAF1 structure are deposited in the Protein Data Bank under the accession code 7OKQ (https://www.rcsb.org/structure/7OKQ). Table EV1 summaries cryo-EM data collection, refinement, and validation statistics. Table EV2 includes the validation statistics for re-refined template models. Author contributions: WIM prepared the specimens for EM data collection. WIM, SC, and ADS. collected EM data. WIM performed EM data processing, with help from SC and ADS. Model building and refinement was carried out by GK. WIM performed the activity assays with input from AP and WAR. WIM performed the biochemical assays. JR and KR contributed in data discussion. NHT, WIM, and AB wrote the manuscript with input from all authors. The authors declare that they have no conflict of interest.
Funders:
Funding AgencyGrant Number
European Research Council (ERC)666068
Novartis Research FoundationUNSPECIFIED
Swiss National Science Foundation (SNSF)31003A_179541
Swiss National Science Foundation (SNSF)310030_201206
Boehringer Ingelheim FondsUNSPECIFIED
Subject Keywords:CRL4/DCAF1; E3 ligases; Oligomerization; Ubiquitin; VprBP
Issue or Number:22
DOI:10.15252/embj.2021108008
Record Number:CaltechAUTHORS:20211021-201507835
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20211021-201507835
Official Citation:The CRL4DCAF1 cullin-RING ubiquitin ligase is activated following a switch in oligomerization state. Weaam I Mohamed, Andreas D Schenk, Georg Kempf, Simone Cavadini, Anja Basters, Alessandro Potenza, Wassim Abdul Rahman, Julius Rabl, Kurt Reichermeier, Nicolas H Thomä. The EMBO Journal (2021) 40: e108008; DOI: https://doi.org/10.15252/embj.2021108008
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:111584
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:26 Oct 2021 18:28
Last Modified:15 Nov 2021 22:57

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