title: A high-coverage shRNA screen identifies TMEM129 as an E3 ligase involved in ER-associated protein degradation.
abstract: Misfolded ER proteins are retrotranslocated into the cytosol for degradation via the ubiquitin-proteasome system. The human cytomegalovirus protein US11 exploits this ER-associated protein degradation (ERAD) pathway to downregulate HLA class I molecules in virus-infected cells, thereby evading elimination by cytotoxic T-lymphocytes. US11-mediated degradation of HLA class I has been instrumental in the identification of key components of mammalian ERAD, including Derlin-1, p97, VIMP and SEL1L. Despite this, the process governing retrotranslocation of the substrate is still poorly understood. Here using a high-coverage genome-wide shRNA library, we identify the uncharacterized protein TMEM129 and the ubiquitin-conjugating E2 enzyme UBE2J2 to be essential for US11-mediated HLA class I downregulation. TMEM129 is an unconventional C4C4-type RING finger E3 ubiquitin ligase that resides within a complex containing various other ERAD components, including Derlin-1, Derlin-2, VIMP and p97, indicating that TMEM129 is an integral part of the ER-resident dislocation complex mediating US11-induced HLA class I degradation.
authors.affiliation: Medical Microbiology, University Medical Center Utrecht, 3584CX Utrecht, The Netherlands.
authors.forename: Michael L
authors.initials: ML
authors.lastname: van de Weijer
authors.affiliation: 1] Department of Cellular and Molecular Pharmacology, California Institute for Quantitative Biomedical Research, Howard Hughes Medical Institute, University of California, San Francisco, California 94158, USA [2].
authors.forename: Michael C
authors.initials: MC
authors.lastname: Bassik
authors.affiliation: Medical Microbiology, University Medical Center Utrecht, 3584CX Utrecht, The Netherlands.
authors.forename: Rutger D
authors.initials: RD
authors.lastname: Luteijn
authors.affiliation: Medical Microbiology, University Medical Center Utrecht, 3584CX Utrecht, The Netherlands.
authors.forename: Cornelia M
authors.initials: CM
authors.lastname: Voorburg
authors.affiliation: Medical Microbiology, University Medical Center Utrecht, 3584CX Utrecht, The Netherlands.
authors.forename: Mirjam A M
authors.initials: MA
authors.lastname: Lohuis
authors.affiliation: Helmholtz Zentrum München, German Research Center for Environmental Health, Institute of Molecular Immunology, 81377 Munich, Germany.
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Kremmer
authors.affiliation: Department of Molecular Cell Biology, Leiden University Medical Center, 2333ZC Leiden, The Netherlands.
authors.forename: Rob C
authors.initials: RC
authors.lastname: Hoeben
authors.affiliation: 1] Genomics Solution Unit, Agilent Technologies Inc., Santa Clara, California 95051, USA [2].
authors.forename: Emily M
authors.initials: EM
authors.lastname: LeProust
authors.affiliation: 1] Genomics Solution Unit, Agilent Technologies Inc., Santa Clara, California 95051, USA [2].
authors.forename: Siyuan
authors.initials: S
authors.lastname: Chen
authors.affiliation: Medical Microbiology, University Medical Center Utrecht, 3584CX Utrecht, The Netherlands.
authors.forename: Hanneke
authors.initials: H
authors.lastname: Hoelen
authors.affiliation: 1] Medical Microbiology, University Medical Center Utrecht, 3584CX Utrecht, The Netherlands [2] Department of Molecular Cell Biology, Leiden University Medical Center, 2333ZC Leiden, The Netherlands.
authors.forename: Maaike E
authors.initials: ME
authors.lastname: Ressing
authors.affiliation: 1] Department of Cellular and Molecular Pharmacology, California Institute for Quantitative Biomedical Research, Howard Hughes Medical Institute, University of California, San Francisco, California 94158, USA [2] Department of Microbiology and Immunology, University of California, San Francisco, California 94143, USA [3].
authors.forename: Weronika
authors.initials: W
authors.lastname: Patena
authors.affiliation: Department of Cellular and Molecular Pharmacology, California Institute for Quantitative Biomedical Research, Howard Hughes Medical Institute, University of California, San Francisco, California 94158, USA.
authors.forename: Jonathan S
authors.initials: JS
authors.lastname: Weissman
authors.affiliation: Department of Microbiology and Immunology, University of California, San Francisco, California 94143, USA.
authors.forename: Michael T
authors.initials: MT
authors.lastname: McManus
authors.affiliation: 1] Medical Microbiology, University Medical Center Utrecht, 3584CX Utrecht, The Netherlands [2].
authors.forename: Emmanuel J H J
authors.initials: EJ
authors.lastname: Wiertz
authors.affiliation: 1] Medical Microbiology, University Medical Center Utrecht, 3584CX Utrecht, The Netherlands [2].
authors.forename: Robert Jan
authors.initials: RJ
authors.lastname: Lebbink
mesh_terms: D000251:Adenosine Triphosphatases / Q000235:genetics; D045744:Cell Line, Tumor; D064112:Clustered Regularly Interspaced Short Palindromic Repeats / Q000235:genetics; D003587:Cytomegalovirus / Q000235:genetics; D003586:Cytomegalovirus Infections; D015536:Down-Regulation; D004721:Endoplasmic Reticulum / Q000473:pathology; D060746:Endoplasmic Reticulum-Associated Degradation; D057809:HEK293 Cells; D015395:Histocompatibility Antigens Class I / Q000096:biosynthesis*; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics; D009687:Nuclear Proteins / Q000235:genetics; D017510:Protein Folding; D011506:Proteins / Q000235:genetics; D034622:RNA Interference; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins / Q000235:genetics*; D051140:Selenoproteins / Q000235:genetics; D020298:U937 Cells; D044763:Ubiquitin-Conjugating Enzymes / Q000235:genetics*; D044767:Ubiquitin-Protein Ligases / Q000235:genetics*; D014764:Viral Proteins / Q000235:genetics*
chemical_list: C487709:DERL1 protein, human; D015395:Histocompatibility Antigens Class I; D008565:Membrane Proteins; D009687:Nuclear Proteins; D011506:Proteins; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins; C117384:SEL1L protein, human; C487710:SELENOS protein, human; D051140:Selenoproteins; C074546:US11 protein, herpesvirus; D014764:Viral Proteins; C489966:UBE2J2 protein, human; D044763:Ubiquitin-Conjugating Enzymes; C000591994:TMEM129 protein, human; D044767:Ubiquitin-Protein Ligases; D000251:Adenosine Triphosphatases; C404942:p97 ATPase