Continuous sensing of nutrients and growth factors by the mTORC1-TFEB axis
- University of California, Davis, United States
Abstract
mTORC1 senses nutrients and growth factors and phosphorylates downstream targets, including the transcription factor TFEB, to coordinate metabolic supply and demand. These functions position mTORC1 as a central controller of cellular homeostasis, but the behavior of this system in individual cells has not been well characterized. Here, we provide measurements necessary to refine quantitative models for mTORC1 as a metabolic controller. We developed a series of fluorescent protein-TFEB fusions and a multiplexed immunofluorescence approach to investigate how combinations of stimuli jointly regulate mTORC1 signaling at the single-cell level. Live imaging of individual MCF10A cells confirmed that mTORC1-TFEB signaling responds continuously to individual, sequential, or simultaneous treatment with amino acids and the growth factor insulin. Under physiologically relevant concentrations of amino acids, we observe correlated fluctuations in TFEB, AMPK, and AKT signaling that indicate continuous activity adjustments to nutrient availability. Using partial least squares regression modeling, we show that these continuous gradations are connected to protein synthesis rate via a distributed network of mTORC1 effectors, providing quantitative support for the qualitative model of mTORC1 as a homeostatic controller and clarifying its functional behavior within individual cells.
Data availability
The following files contain the per-cell fluorescence intensity data, extracted from microscope image data, that were used to generate each figure in the paper:Sparta2023_Figure1_SourceData1.xlsSparta2023_Figure1_SourceData2.xlsSparta2023_Figure2_SourceData1.xlsxSparta2023_Figure3_SourceData1.xlsSparta2023_Figure4_SourceData1.xlsxSparta2023_Figure4_SourceData2.xlsxSparta2023_Figure4_SourceData3.xlsxSparta2023_Figure4_SourceData4.xlsxSparta2023_Figure4_SourceData5.xlsxSparta2023_Figure4_SourceData6.xlsxSparta2023_Figure5_SourceData1.xlsxSparta2023_Figure5_SourceData2.xlsxSparta2023_Figure5_SourceData3.xlsxSparta2023_Figure5_SourceData4.xlsxSparta2023_Figure6_SourceData1.xlsx
Article and author information
Author details
Funding
National Institute of General Medical Sciences (R35GM139621)
- John G Albeck
National Institute of General Medical Sciences (R01GM115650)
- John G Albeck
National Science Foundation (2136040)
- John G Albeck
National Heart, Lung, and Blood Institute (T32HL007013)
- Nicholaus DeCuzzi
National Institute of General Medical Sciences (F31GM120937)
- Breanne Sparta
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2023, Sparta et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Continuous sensing of nutrients and growth factors by the mTORC1-TFEB axis
eLife 12:e74903.
https://doi.org/10.7554/eLife.74903
Further reading
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- Cell Biology
Niemann–Pick disease type C (NPC) is a devastating lysosomal storage disease characterized by abnormal cholesterol accumulation in lysosomes. Currently, there is no treatment for NPC. Transcription factor EB (TFEB), a member of the microphthalmia transcription factors (MiTF), has emerged as a master regulator of lysosomal function and promoted the clearance of substrates stored in cells. However, it is not known whether TFEB plays a role in cholesterol clearance in NPC disease. Here, we show that transgenic overexpression of TFEB, but not TFE3 (another member of MiTF family) facilitates cholesterol clearance in various NPC1 cell models. Pharmacological activation of TFEB by sulforaphane (SFN), a previously identified natural small-molecule TFEB agonist by us, can dramatically ameliorate cholesterol accumulation in human and mouse NPC1 cell models. In NPC1 cells, SFN induces TFEB nuclear translocation via a ROS-Ca2+-calcineurin-dependent but MTOR-independent pathway and upregulates the expression of TFEB-downstream genes, promoting lysosomal exocytosis and biogenesis. While genetic inhibition of TFEB abolishes the cholesterol clearance and exocytosis effect by SFN. In the NPC1 mouse model, SFN dephosphorylates/activates TFEB in the brain and exhibits potent efficacy of rescuing the loss of Purkinje cells and body weight. Hence, pharmacological upregulating lysosome machinery via targeting TFEB represents a promising approach to treat NPC and related lysosomal storage diseases, and provides the possibility of TFEB agonists, that is, SFN as potential NPC therapeutic candidates.
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- Cell Biology
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