Signal transduction in human cells by metabolites derived from methionine and glucose

Thesis: Ph. D., Massachusetts Institute of Technology, Department of Biology, February, 2021Cataloged from the official PDF of thesis. "February 2021."Includes bibliographical references.Organisms sense nutrients to match physiological responses to their environment. The mechanistic Target of Rapamycin complex I (mTORC1) pathway integrates information from a wide range of nutrient inputs to appropriately control cell growth. Nutrients like amino acids and glucose are required for mTORC1 to localize to the lysosome, its site of activation. The Rag-GTPases dictate mTORC1 localization and are regulated by GATOR1 and GATOR2 in response to nutrients, but how the GATOR-Rag axis senses nutrients is not completely understood. We found a novel protein, which we named SAMTOR, that interacts with GATOR1, and promotes GATOR1 activity and mTORC1 inhibition. Moreover, we show that SAMTOR is an S-adenosylmethionine binding protein, and that SAM binding disrupts the SAMTOR-GATOR1 interaction. SAM is a derivative of methionine, and we show that methionine starvation promotes SAMTOR action on GATOR1 following a decrease in SAM levels.The consequence of this series of events is to inhibit mTORC1. In contrast, when SAM levels are high, it disrupts the interaction between SAMTOR and GATOR1, maintaining mTORC1 activity high. We concluded that SAMTOR is a SAM sensor in the mTORC1 pathway. Glucose is required for full activity and lysosomal localization of mTORC1. However, the mechanism of glucose sensing and the identity of the metabolite derived from glucose that is sensed by the mTORC1 pathway remain unknown. To identify the metabolite that signals glucose to mTORC1, we used metabolically engineered human cells lacking the canonical energy sensor AMPK to identify glucose-derived metabolites required to activate mTORC1 independent of energetic stress. We show that mTORC1 senses a metabolite downstream of the aldolase and upstream of the glyceraldehyde 3-phosphate dehydrogenase steps of glycolysis and pinpoint dihydroxyacetone phosphate (DHAP) as the key molecule.We also show that an intact GATOR-Rag axis is required for glucose sensing upstream of mTORC1. Altogether, we identified two metabolites that regulate the mTORC1 pathway, one-SAM-derived from methionine and the other-DHAP-from glucose. Greater mechanistic insights into both SAM- and DHAP-sensing will reveal how these two signals, along with that of other amino acids and nutrients, are integrated by the GATOR-Rag signaling axis.by Jose M. Orozco.Ph. D.Ph.D. Massachusetts Institute of Technology, Department of Biolog

SAMTOR is an S-adenosylmethionine sensor for the mTORC1 pathway

mTOR complex 1 (mTORC1) regulates cell growth and metabolism in response to multiple environmental cues. Nutrients signal via the Rag guanosine triphosphatases (GTPases) to promote the localization of mTORC1 to the lysosomal surface, its site of activation. We identified SAMTOR, a previously uncharacterized protein, which inhibits mTORC1 signaling by interacting with GATOR1, the GTPase activating protein (GAP) for RagA/B. We found that the methyl donor S-adenosylmethionine (SAM) disrupts the SAMTOR-GATOR1 complex by binding directly to SAMTOR with a dissociation constant of approximately 7 μM. In cells, methionine starvation reduces SAM levels below this dissociation constant and promotes the association of SAMTOR with GATOR1, thereby inhibiting mTORC1 signaling in a SAMTOR-dependent fashion. Methionine-induced activation of mTORC1 requires the SAM binding capacity of SAMTOR. Thus, SAMTOR is a SAM sensor that links methionine and one-carbon metabolism to mTORC1 signaling.National Institutes of Health (U.S.) (Grant R01 CA103866)National Institutes of Health (U.S.) (Grant R37 AI47389)United States. Department of Defense (Grant W81XWH-07-0448)National Institutes of Health (U.S.) (Fellowship T32 GM007753)National Institutes of Health (U.S.) (Grant T32 GM007753)National Institutes of Health (U.S.) (Grant F30 CA210373)National Institutes of Health (U.S.) (Grant F31 GM121093-01A1)National Science Foundation (U.S.) (Grant 2016197106)Massachusetts Institute of Technology. Paul Gray UROP Fund (Grant 3143900