title: E3 ligases determine ubiquitination site and conjugate type by enforcing specificity on E2 enzymes.
abstract: Ubiquitin-conjugating enzymes (E2s) have a dominant role in determining which of the seven lysine residues of ubiquitin is used for polyubiquitination. Here we show that tethering of a substrate to an E2 enzyme in the absence of an E3 ubiquitin ligase is sufficient to promote its ubiquitination, whereas the type of the ubiquitin conjugates and the identity of the target lysine on the substrate are promiscuous. In contrast, when an E3 enzyme is introduced, a clear decision between mono- and polyubiquitination is made, and the conjugation type as well as the identity of the target lysine residue on the substrate becomes highly specific. These features of the E3 can be further regulated by auxiliary factors as exemplified by MDMX (Murine Double Minute X). In fact, we show that this interactor reconfigures MDM2-dependent ubiquitination of p53. Based on several model systems, we propose that although interaction with an E2 is sufficient to promote substrate ubiquitination the E3 molds the reaction into a specific, physiologically relevant protein modification.
authors.affiliation: Department of Biological Regulation, The Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Yael
authors.initials: Y
authors.lastname: David
authors.affiliation: The Henry Wellcome Building for Molecular Physiology, Nuffield Department of Clinical Medicine, University of Oxford, Roosevelt Drive, Oxford OX3 7BN, United Kingdom.
authors.forename: Nicola
authors.initials: N
authors.lastname: Ternette
authors.affiliation: The Henry Wellcome Building for Molecular Physiology, Nuffield Department of Clinical Medicine, University of Oxford, Roosevelt Drive, Oxford OX3 7BN, United Kingdom.
authors.forename: Mariola J
authors.initials: MJ
authors.lastname: Edelmann
authors.affiliation: Smoler Proteomics Center, Technion-Israel Institute of Technology, Haifa 31096, Israel.
authors.forename: Tamar
authors.initials: T
authors.lastname: Ziv
authors.affiliation: Department of Biological Regulation, The Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Batya
authors.initials: B
authors.lastname: Gayer
authors.affiliation: Faculty of Biochemistry, The Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Rotem
authors.initials: R
authors.lastname: Sertchook
authors.affiliation: Department of Biological Regulation, The Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Yakir
authors.initials: Y
authors.lastname: Dadon
authors.affiliation: The Henry Wellcome Building for Molecular Physiology, Nuffield Department of Clinical Medicine, University of Oxford, Roosevelt Drive, Oxford OX3 7BN, United Kingdom.
authors.forename: Benedikt M
authors.initials: BM
authors.lastname: Kessler
authors.affiliation: Department of Biological Regulation, The Weizmann Institute of Science, Rehovot 76100, Israel. Electronic address: ami.navon@weizmann.ac.il.
authors.forename: Ami
authors.initials: A
authors.lastname: Navon
mesh_terms: D018797:Cell Cycle Proteins; D002853:Chromatography, Liquid / Q000379:methods; D016158:Genes, p53; D006801:Humans; D008239:Lysine / Q000737:chemistry; D009687:Nuclear Proteins / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051736:Proto-Oncogene Proteins c-mdm2 / Q000737:chemistry / Q000378:metabolism; D053719:Tandem Mass Spectrometry / Q000379:methods; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D025801:Ubiquitin / Q000737:chemistry*; D044763:Ubiquitin-Conjugating Enzymes / Q000737:chemistry*; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry*
chemical_list: D018797:Cell Cycle Proteins; C482775:MDM4 protein, human; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; D025801:Ubiquitin; D044763:Ubiquitin-Conjugating Enzymes; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2; D044767:Ubiquitin-Protein Ligases; D008239:Lysine