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Mol Cell Proteomics. 2015 Aug;14(8):2042-55. doi: 10.1074/mcp.M114.045807. Epub 2015 Apr 23.

Functional Proteomics Identifies Acinus L as a Direct Insulin- and Amino Acid-Dependent Mammalian Target of Rapamycin Complex 1 (mTORC1) Substrate.

Author information

1
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;
2
§Faculty of Biology, Institute of Biology II, University of Freiburg, 79104 Freiburg, Germany;
3
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;
4
‖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;
5
¶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;
6
§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.
7
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.

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.

PMID:
25907765
PMCID:
PMC4528236
DOI:
10.1074/mcp.M114.045807
[Indexed for MEDLINE]
Free PMC Article

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