Functional Proteomics Identifies Acinus L as a Direct Insulin- and Amino Acid-Dependent Mammalian Target of Rapamycin
Jennifer Jasmin Schwarz1, Heike Wiese2, Regine Charlotte Tölle3
1From 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;
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.
Insights
Researchers identified acinus L as a direct substrate of the mammalian target of rapamycin complex 1 (mTORC1) using a novel triple proteomics approach. This finding advances our understanding of mTORC1 signaling in growth and metabolism.
Area of Science:
- Cellular Biology
- Biochemistry
- Proteomics
Background:
- The mammalian target of rapamycin (mTOR) pathway is crucial for regulating cell growth, metabolism, and aging.
- mTOR signaling is frequently dysregulated in metabolic disorders and cancer.
- Despite extensive knowledge of mTOR's regulatory network, direct substrates remain largely uncharacterized.
Purpose of the Study:
- To identify novel direct substrates of mTOR complex 1 (mTORC1).
- To develop and apply a comprehensive proteomics strategy for discovering mTOR network components.
Main Methods:
- Quantitative phosphoproteomics analysis of insulin- and amino acid-responsive phosphoproteome after raptor inhibition.
- Interactomic analysis of endogenous mTOR.
- Integration of phosphoproteomic and interactomic data.
- Validation using co-immunoprecipitation and MS-enhanced kinase assays.
Main Results:
- Identification of acinus L as a potential novel mTORC1 target through integrated phosphoproteomic and interactomic analyses.
- Confirmation of acinus L as a direct substrate of mTORC1.
- Demonstration of a triple proteomics strategy for novel mTOR target discovery.
Conclusions:
- Acinus L is a newly identified direct substrate of mTORC1.
- The developed triple proteomics strategy is effective for de novo identification of mTOR network components.
- This study provides a valuable resource for future research into mTOR signaling and its targets.
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