title: Functional Proteomics Identifies Acinus L as a Direct Insulin- and Amino Acid-Dependent Mammalian Target of Rapamycin Complex 1 (mTORC1) Substrate.
abstract: The serine/threonine kinase mammalian target of rapamycin (mTOR) governs growth, metabolism, and aging in response to insulin and amino acids (aa), and is often activated in metabolic disorders and cancer. Much is known about the regulatory signaling network that encompasses mTOR, but surprisingly few direct mTOR substrates have been established to date. To tackle this gap in our knowledge, we took advantage of a combined quantitative phosphoproteomic and interactomic strategy. We analyzed the insulin- and aa-responsive phosphoproteome upon inhibition of the mTOR complex 1 (mTORC1) component raptor, and investigated in parallel the interactome of endogenous mTOR. By overlaying these two datasets, we identified acinus L as a potential novel mTORC1 target. We confirmed acinus L as a direct mTORC1 substrate by co-immunoprecipitation and MS-enhanced kinase assays. Our study delineates a triple proteomics strategy of combined phosphoproteomics, interactomics, and MS-enhanced kinase assays for the de novo-identification of mTOR network components, and provides a rich source of potential novel mTOR interactors and targets for future investigation.
authors.affiliation: From the ‡Faculty of Biology, Institute of Biology III, University of Freiburg, 79104 Freiburg, Germany; §Faculty of Biology, Institute of Biology II, University of Freiburg, 79104 Freiburg, Germany; ¶Spemann Graduate School of Biology and Medicine (SGBM), University of Freiburg;
authors.forename: Jennifer Jasmin
authors.initials: JJ
authors.lastname: Schwarz
authors.affiliation: §Faculty of Biology, Institute of Biology II, University of Freiburg, 79104 Freiburg, Germany;
authors.forename: Heike
authors.initials: H
authors.lastname: Wiese
authors.affiliation: From the ‡Faculty of Biology, Institute of Biology III, University of Freiburg, 79104 Freiburg, Germany; §Faculty of Biology, Institute of Biology II, University of Freiburg, 79104 Freiburg, Germany; ‖Department of Dermatology, Medical Center, and Centre for Biological Systems Analysis (ZBSA), University of Freiburg, 79104 Freiburg, Germany;
authors.forename: Regine Charlotte
authors.initials: RC
authors.lastname: Tölle
authors.affiliation: ‖Department of Dermatology, Medical Center, and Centre for Biological Systems Analysis (ZBSA), University of Freiburg, 79104 Freiburg, Germany; **BIOSS Centre for Biological Signaling Studies, University of Freiburg, 79104 Freiburg, Germany;
authors.forename: Mostafa
authors.initials: M
authors.lastname: Zarei
authors.affiliation: ¶Spemann Graduate School of Biology and Medicine (SGBM), University of Freiburg; ‖Department of Dermatology, Medical Center, and Centre for Biological Systems Analysis (ZBSA), University of Freiburg, 79104 Freiburg, Germany; **BIOSS Centre for Biological Signaling Studies, University of Freiburg, 79104 Freiburg, Germany;
authors.forename: Jörn
authors.initials: J
authors.lastname: Dengjel
authors.affiliation: §Faculty of Biology, Institute of Biology II, University of Freiburg, 79104 Freiburg, Germany; ¶Spemann Graduate School of Biology and Medicine (SGBM), University of Freiburg; **BIOSS Centre for Biological Signaling Studies, University of Freiburg, 79104 Freiburg, Germany; k.thedieck@umcg.nl kathrin.thedieck@uni-oldenburg.de bettina.warscheid@biologie.uni-freiburg.de.
authors.forename: Bettina
authors.initials: B
authors.lastname: Warscheid
authors.affiliation: From the ‡Faculty of Biology, Institute of Biology III, University of Freiburg, 79104 Freiburg, Germany; **BIOSS Centre for Biological Signaling Studies, University of Freiburg, 79104 Freiburg, Germany; ‡‡Department of Pediatrics, University of Groningen, University Medical Center Groningen (UMCG), 9713 AV Groningen, The Netherlands; §§Department for Neurosciences, Faculty VI - School of Medicine and Health Sciences, Carl von Ossietzky University Oldenburg, 26129 Oldenburg, Germany k.thedieck@umcg.nl kathrin.thedieck@uni-oldenburg.de bettina.warscheid@biologie.uni-freiburg.de.
authors.forename: Kathrin
authors.initials: K
authors.lastname: Thedieck
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D006367:HeLa Cells; D006801:Humans; D007328:Insulin / Q000378:metabolism*; D000076222:Mechanistic Target of Rapamycin Complex 1; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes / Q000378:metabolism*; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D025941:Protein Interaction Mapping; D020543:Proteome / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D000076223:Regulatory-Associated Protein of mTOR; D013379:Substrate Specificity; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*
chemical_list: C121310:ACIN1 protein, human; D048868:Adaptor Proteins, Signal Transducing; D000596:Amino Acids; D007328:Insulin; D046912:Multiprotein Complexes; D009687:Nuclear Proteins; D010750:Phosphoproteins; D020543:Proteome; C463659:RPTOR protein, human; D000076223:Regulatory-Associated Protein of mTOR; D000076222:Mechanistic Target of Rapamycin Complex 1; D058570:TOR Serine-Threonine Kinases