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Regulation of mTORC1 by the Rag GTPases is necessary for neonatal autophagy and survival

Abstract

The mechanistic target of rapamycin complex 1 (mTORC1) pathway regulates organismal growth in response to many environmental cues, including nutrients and growth factors1. Cell-based studies showed that mTORC1 senses amino acids through the RagA–D family of GTPases2,3 (also known as RRAGA, B, C and D), but their importance in mammalian physiology is unknown. Here we generate knock-in mice that express a constitutively active form of RagA (RagAGTP) from its endogenous promoter. RagAGTP/GTP mice develop normally, but fail to survive postnatal day 1. When delivered by Caesarean section, fasted RagAGTP/GTP neonates die almost twice as rapidly as wild-type littermates. Within an hour of birth, wild-type neonates strongly inhibit mTORC1, which coincides with profound hypoglycaemia and a decrease in plasma amino-acid concentrations. In contrast, mTORC1 inhibition does not occur in RagAGTP/GTP neonates, despite identical reductions in blood nutrient amounts. With prolonged fasting, wild-type neonates recover their plasma glucose concentrations, but RagAGTP/GTP mice remain hypoglycaemic until death, despite using glycogen at a faster rate. The glucose homeostasis defect correlates with the inability of fasted RagAGTP/GTP neonates to trigger autophagy and produce amino acids for de novo glucose production. Because profound hypoglycaemia does not inhibit mTORC1 in RagAGTP/GTP neonates, we considered the possibility that the Rag pathway signals glucose as well as amino-acid sufficiency to mTORC1. Indeed, mTORC1 is resistant to glucose deprivation in RagAGTP/GTP fibroblasts, and glucose, like amino acids, controls its recruitment to the lysosomal surface, the site of mTORC1 activation. Thus, the Rag GTPases signal glucose and amino-acid concentrations to mTORC1, and have an unexpectedly key role in neonates in autophagy induction and thus nutrient homeostasis and viability.

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Figure 1: Characterization of RagA GTP/GTP mice.
Figure 2: Profound glucose homeostasis defect in RagA GTP/GTP mice.
Figure 3: Impaired autophagy in RagA GTP/GTP neonates.
Figure 4: The Rag GTPases mediate inhibition of mTORC1 by glucose deprivation.

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Acknowledgements

We thank members of the Sabatini laboratory for suggestions, and A. Hutchins for technical assistance. We thank R. Shaw for providing the AMPK-DKO MEFs, D. Ron for the GCN2-KO MEFs and M. Barbacid for the transcriptional STOP cassette. This work was supported by grants from the National Institutes of Health (R01 CA129105, R01 CA103866 and R37 AI047389) and awards from the American Federation for Aging, Starr Foundation, Koch Institute Frontier Research Program, and the Ellison Medical Foundation to D.M.S., fellowships from the Human Frontiers Science Program to A.E., and the Jane Coffin Childs Memorial Fund for Medical Research and the LAM Foundation to R.Z. D.M.S. in an investigator of Howard Hughes Medical Institute.

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A.E. and D.M.S. conceived the project. A.E. designed and performed most experiments with input from D.M.S, and assistance from S.C., R.L.W. and O.K. R.Z. performed experiments and participated in discussion of the results. I.G., H.S. and D.D.S. performed electron microscopy experiments and interpretations. D.D.S. helped with discussion and interpretation of results. A.E. wrote and D.M.S. edited the manuscript.

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Correspondence to David M. Sabatini.

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The authors declare no competing financial interests.

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Efeyan, A., Zoncu, R., Chang, S. et al. Regulation of mTORC1 by the Rag GTPases is necessary for neonatal autophagy and survival. Nature 493, 679–683 (2013). https://doi.org/10.1038/nature11745

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