title: Ubiquitylation-dependent oligomerization regulates activity of Nedd4 ligases.
abstract: Ubiquitylation controls protein function and degradation. Therefore, ubiquitin ligases need to be tightly controlled. We discovered an evolutionarily conserved allosteric restraint mechanism for Nedd4 ligases and demonstrated its function with diverse substrates: the yeast soluble proteins Rpn10 and Rvs167, and the human receptor tyrosine kinase FGFR1 and cardiac IKSpotassium channel. We found that a potential trimerization interface is structurally blocked by the HECT domain α1-helix, which further undergoes ubiquitylation on a conserved lysine residue. Genetic, bioinformatics, biochemical and biophysical data show that attraction between this α1-conjugated ubiquitin and the HECT ubiquitin-binding patch pulls the α1-helix out of the interface, thereby promoting trimerization. Strikingly, trimerization renders the ligase inactive. Arginine substitution of the ubiquitylated lysine impairs this inactivation mechanism and results in unrestrained FGFR1 ubiquitylation in cells. Similarly, electrophysiological data and TIRF microscopy show that NEDD4 unrestrained mutant constitutively downregulates the IKSchannel, thus confirming the functional importance of E3-ligase autoinhibition.
authors.affiliation: Department of Biochemistry and Molecular Biology, the George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel.
authors.forename: Ilan
authors.initials: I
authors.lastname: Attali
authors.affiliation: Department of Physiology & Pharmacology, Sackler Tel Aviv University, Tel Aviv, Israel.
authors.forename: William Sam
authors.initials: WS
authors.lastname: Tobelaim
authors.affiliation: Cell Biology Program, The Hospital for Sick Children and Biochemistry Department, University of Toronto, Toronto, ON, Canada.
authors.forename: Avinash
authors.initials: A
authors.lastname: Persaud
authors.affiliation: Tumor Initiation & Maintenance Program and NCI Cancer Centre Proteomics Facility, Sanford Burnham Prebys Medical Discovery Institute, La Jolla, CA, USA.
authors.forename: Khatereh
authors.initials: K
authors.lastname: Motamedchaboki
authors.affiliation: Department of Molecular Microbiology and Biotechnology, the George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel.
authors.forename: Kobi J
authors.initials: KJ
authors.lastname: Simpson-Lavy
authors.affiliation: Department of Biochemistry and Molecular Biology, Institute for Medical Research Israel-Canada, Hebrew University-Hadassah Medical School, Jerusalem, Israel.
authors.forename: Bayan
authors.initials: B
authors.lastname: Mashahreh
authors.affiliation: Department of Biochemistry and Molecular Biology, the George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel.
authors.forename: Olga
authors.initials: O
authors.lastname: Levin-Kravets
authors.affiliation: Department of Biochemistry and Molecular Biology, the George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel.
authors.forename: Tal
authors.initials: T
authors.lastname: Keren-Kaplan
authors.affiliation: Department of Biochemistry and Molecular Biology, the George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel.
authors.forename: Inbar
authors.initials: I
authors.lastname: Pilzer
authors.affiliation: Department of Molecular Microbiology and Biotechnology, the George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel.
authors.forename: Martin
authors.initials: M
authors.lastname: Kupiec
authors.affiliation: Department of Biochemistry and Molecular Biology, Institute for Medical Research Israel-Canada, Hebrew University-Hadassah Medical School, Jerusalem, Israel.
authors.forename: Reuven
authors.initials: R
authors.lastname: Wiener
authors.affiliation: Tumor Initiation & Maintenance Program and NCI Cancer Centre Proteomics Facility, Sanford Burnham Prebys Medical Discovery Institute, La Jolla, CA, USA.|School of Pharmaceutical Sciences, Xiamen University, Xiamen, China.
authors.forename: Dieter A
authors.initials: DA
authors.lastname: Wolf
authors.affiliation: Cell Biology Program, The Hospital for Sick Children and Biochemistry Department, University of Toronto, Toronto, ON, Canada.
authors.forename: Daniela
authors.initials: D
authors.lastname: Rotin
authors.affiliation: Department of Biochemistry and Molecular Biology, the George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel prag@post.tau.ac.il.|Sagol School of Neuroscience, Tel Aviv University, Tel Aviv, Israel.
authors.forename: Gali
authors.identifier: 0000-0003-4703-7161
authors.initials: G
authors.lastname: Prag
mesh_terms: D056827:Endosomal Sorting Complexes Required for Transport / Q000378:metabolism*; D006801:Humans; D008840:Microfilament Proteins / Q000737:chemistry / Q000378:metabolism*; D000075702:Nedd4 Ubiquitin Protein Ligases; D024642:Potassium Channels, Voltage-Gated / Q000737:chemistry / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex / Q000737:chemistry / Q000378:metabolism*; D055503:Protein Multimerization; D051496:Receptor, Fibroblast Growth Factor, Type 1 / Q000737:chemistry / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*; D054875:Ubiquitination
chemical_list: D056827:Endosomal Sorting Complexes Required for Transport; D008840:Microfilament Proteins; D024642:Potassium Channels, Voltage-Gated; C122019:RPN10 protein, S cerevisiae; C082282:RVS167 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C059983:potassium channel protein I(sk); D000075702:Nedd4 Ubiquitin Protein Ligases; C000618483:Nedd4 protein, human; D044767:Ubiquitin-Protein Ligases; C496348:FGFR1 protein, human; D051496:Receptor, Fibroblast Growth Factor, Type 1; D046988:Proteasome Endopeptidase Complex
keywords: Nedd4; Rsp5; inactivation; oligomerization; ubiquitylation