Abstract
The activation of the mechanistic target of rapamycin complex 1 (mTORC1), a master regulator of protein synthesis, by anabolic stimuli (such as muscle contraction or essential amino acids) involves its translocation to the cell periphery. Leucine is generally considered the most anabolic of amino acids for its ability to independently modulate muscle protein synthesis. However, it is currently unknown if free leucine impacts region-specific mTORC1-mediated phosphorylation events and protein–protein interactions. In this clinical trial (NCT03952884; registered May 16, 2019), we used immunofluorescence methods to investigate the role of dietary leucine on the postprandial regulation of mTORC1 and ribosomal protein S6 (RPS6), an important downstream readout of mTORC1 activity. Eight young, healthy, recreationally active males (n = 8; 23 ± 3 yrs) ingested 2 g of leucine with vastus lateralis biopsies collected at baseline, 30, 60, and 180 min postprandial. Leucine promoted mTOR translocation to the periphery (~ 18–29%; p ≤ 0.012) and enhanced mTOR localization with the lysosome (~ 16%; both p = 0.049) at 30 and 60 min post-feeding. p-RPS6Ser240/244 staining intensity, a readout of mTORC1 activity, was significantly elevated at all postprandial timepoints in both the total fiber (~ 14–30%; p ≤ 0.032) and peripheral regions (~ 16–33%; p ≤ 0.014). Additionally, total and peripheral p-RPS6Ser240/244 staining intensity at 60 min was positively correlated (r = 0.74, p = 0.036; r = 0.80, p = 0.016, respectively) with rates of myofibrillar protein synthesis over 180 min. The ability of leucine to activate mTORC1 in peripheral regions favors an enhanced rate of MPS, as this is the intracellular space thought to be replete with the cellular machinery that facilitates this anabolic process.
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Data availability statement
The datasets generated during and/or analysed during the current study are available from the corresponding author on reasonable request.
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Acknowledgements
We thank all participants for their engagement in completing this study.
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NH was supported by a MITACS Accelerate Postdoctoral Fellowship (IT15730). The study was supported in part by Ingenious Ingredients, L.P., Lewisville, TX, United States to NAB and Natural Sciences and Engineering Research Council of Canada Discovery Award (RGPIN-2015–04251) to DRM.
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KJMP, NAB, RJ, MP, NH, and DRM designed the experiment; MNHH, MJL, SAS, KJMP, SP, NAB, and NH collected and analyzed the data; MNHH, MJL, and DRM drafted the manuscript. All authors revised and approved the final manuscript.
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RJ and MP are inventors of US Patent No. 11026691B2 and pending patent applications for the use of dileucine and have not been involved in the data collection or analysis. All other authors report no conflict of interest.
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Holowaty, M.N.H., Lees, M.J., Abou Sawan, S. et al. Leucine ingestion promotes mTOR translocation to the periphery and enhances total and peripheral RPS6 phosphorylation in human skeletal muscle. Amino Acids 55, 253–261 (2023). https://doi.org/10.1007/s00726-022-03221-w
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DOI: https://doi.org/10.1007/s00726-022-03221-w