title: Dynamics of mTORC1 activation in response to amino acids.
abstract: Amino acids are essential activators of mTORC1 via a complex containing RAG GTPases, RAGULATOR and the vacuolar ATPase. Sensing of amino acids causes translocation of mTORC1 to lysosomes, an obligate step for activation. To examine the spatial and temporal dynamics of this translocation, we used live imaging of the mTORC1 component RAPTOR and a cell permeant fluorescent analogue of di-leucine methyl ester. Translocation to lysosomes is a transient event, occurring within 2 min of aa addition and peaking within 5 min. It is temporally coupled with fluorescent leucine appearance in lysosomes and is sustained in comparison to aa stimulation. Sestrin2 and the vacuolar ATPase are negative and positive regulators of mTORC1 activity in our experimental system. Of note, phosphorylation of canonical mTORC1 targets is delayed compared to lysosomal translocation suggesting a dynamic and transient passage of mTORC1 from the lysosomal surface before targetting its substrates elsewhere.
authors.affiliation: Signalling Programme, Babraham Institute, Cambridge, United Kingdom.
authors.forename: Maria
authors.initials: M
authors.lastname: Manifava
authors.affiliation: Signalling Programme, Babraham Institute, Cambridge, United Kingdom.
authors.forename: Matthew
authors.initials: M
authors.lastname: Smith
authors.affiliation: Signalling Programme, Babraham Institute, Cambridge, United Kingdom.
authors.forename: Sergio
authors.initials: S
authors.lastname: Rotondo
authors.affiliation: Signalling Programme, Babraham Institute, Cambridge, United Kingdom.
authors.forename: Simon
authors.identifier: 0000-0001-9185-4922
authors.initials: S
authors.lastname: Walker
authors.affiliation: Signalling Programme, Babraham Institute, Cambridge, United Kingdom.
authors.forename: Izabella
authors.initials: I
authors.lastname: Niewczas
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States.
authors.forename: Roberto
authors.initials: R
authors.lastname: Zoncu
authors.affiliation: Signalling Programme, Babraham Institute, Cambridge, United Kingdom.
authors.forename: Jonathan
authors.initials: J
authors.lastname: Clark
authors.affiliation: Signalling Programme, Babraham Institute, Cambridge, United Kingdom.
authors.forename: Nicholas T
authors.identifier: 0000-0001-9397-2914
authors.initials: NT
authors.lastname: Ktistakis
mesh_terms: D000596:Amino Acids / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D000069416:Intravital Microscopy; D008247:Lysosomes / Q000378:metabolism*; D000076222:Mechanistic Target of Rapamycin Complex 1 / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism; D021381:Protein Transport; D000076223:Regulatory-Associated Protein of mTOR / Q000032:analysis; D062211:Spatio-Temporal Analysis; D025262:Vacuolar Proton-Translocating ATPases / Q000378:metabolism; D000084622:Sestrins
chemical_list: D000596:Amino Acids; D000076222:Mechanistic Target of Rapamycin Complex 1; D009687:Nuclear Proteins; D000076223:Regulatory-Associated Protein of mTOR; D025262:Vacuolar Proton-Translocating ATPases; C465398:SESN2 protein, human; D000084622:Sestrins
keywords: amino acids; biochemistry; cell biology; human; mtor; signaling