title: Proteomic analysis of the human KEOPS complex identifies C14ORF142 as a core subunit homologous to yeast Gon7.
abstract: The KEOPS/EKC complex is a tRNA modification complex involved in the biosynthesis of N6-threonylcarbamoyladenosine (t6A), a universally conserved tRNA modification found on ANN-codon recognizing tRNAs. In archaea and eukaryotes, KEOPS is composed of OSGEP/Kae1, PRPK/Bud32, TPRKB/Cgi121 and LAGE3/Pcc1. In fungi, KEOPS contains an additional subunit, Gon7, whose orthologs outside of fungi, if existent, remain unidentified. In addition to displaying defective t6A biosynthesis, Saccharomyces cerevisiae strains harboring KEOPS mutations are compromised for telomere homeostasis, growth and transcriptional co-activation. To identify a Gon7 ortholog in multicellular eukaryotes as well as to uncover KEOPS-interacting proteins that may link t6A biosynthesis to the diverse set of KEOPS mutant phenotypes, we conducted a proteomic analysis of human KEOPS. This work identified 152 protein interactors, one of which, C14ORF142, interacted strongly with all four KEOPS subunits, suggesting that it may be a core component of human KEOPS. Further characterization of C14ORF142 revealed that it shared a number of biophysical and biochemical features with fungal Gon7, suggesting that C14ORF142 is the human ortholog of Gon7. In addition, our proteomic analysis identified specific interactors for different KEOPS subcomplexes, hinting that individual KEOPS subunits may have additional functions outside of t6A biosynthesis.
authors.affiliation: Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON M5G 1X5, Canada.|Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 3E1, Canada.
authors.forename: Leo C K
authors.initials: LC
authors.lastname: Wan
authors.affiliation: Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON M5G 1X5, Canada.
authors.forename: Pierre
authors.initials: P
authors.lastname: Maisonneuve
authors.affiliation: Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON M5G 1X5, Canada.
authors.forename: Rachel K
authors.initials: RK
authors.lastname: Szilard
authors.affiliation: Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON M5G 1X5, Canada.
authors.forename: Jean-Philippe
authors.initials: JP
authors.lastname: Lambert
authors.affiliation: Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON M5G 1X5, Canada.|Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 3E1, Canada.
authors.forename: Timothy F
authors.initials: TF
authors.lastname: Ng
authors.affiliation: Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON M5G 1X5, Canada.|Department of Biochemistry, University of Toronto, Toronto, ON M5S 3E1,Canada.
authors.forename: Noah
authors.initials: N
authors.lastname: Manczyk
authors.affiliation: Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON M5G 1X5, Canada.|School of Chemical Biology and Biotechnology, Shenzhen Graduate School of Peking University, Shenzen, 518055, China.
authors.forename: Hao
authors.initials: H
authors.lastname: Huang
authors.affiliation: School of Chemical Biology and Biotechnology, Shenzhen Graduate School of Peking University, Shenzen, 518055, China.
authors.forename: Rob
authors.initials: R
authors.lastname: Laister
authors.affiliation: Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 3E1, Canada.|Terrence Donnelly Centre for Cellular & Biomolecular Research, University of Toronto, ON, M5S 3E1, Canada.
authors.forename: Amy A
authors.initials: AA
authors.lastname: Caudy
authors.affiliation: Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON M5G 1X5, Canada.|Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 3E1, Canada.
authors.forename: Anne-Claude
authors.initials: AC
authors.lastname: Gingras
authors.affiliation: Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON M5G 1X5, Canada durocher@lunenfeld.ca.|Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 3E1, Canada.
authors.forename: Daniel
authors.initials: D
authors.lastname: Durocher
authors.affiliation: Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON M5G 1X5, Canada sicheri@lunenfeld.ca.|Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 3E1, Canada.|Department of Medical Oncology and Hematology, Princess Margaret Cancer Centre, Toronto, ON M5G 2M9, Canada.
authors.forename: Frank
authors.initials: F
authors.lastname: Sicheri
mesh_terms: D000595:Amino Acid Sequence; D002460:Cell Line; D006801:Humans; D064267:Intrinsically Disordered Proteins / Q000378:metabolism; D046912:Multiprotein Complexes / Q000737:chemistry; D016366:Open Reading Frames; D025941:Protein Interaction Mapping; D060066:Protein Interaction Maps; D021122:Protein Subunits; D040901:Proteomics / Q000379:methods; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*
chemical_list: D064267:Intrinsically Disordered Proteins; C509947:KEOPS complex, S cerevisiae; D046912:Multiprotein Complexes; D021122:Protein Subunits; D029701:Saccharomyces cerevisiae Proteins