GSK3-mediated raptor phosphorylation supports amino-acid-dependent mTORC1-directed signalling.
The mammalian or mechanistic target of rapamycin (mTOR) complex 1 (mTORC1) is a ubiquitously expressed multimeric protein kinase complex that integrates nutrient and growth factor signals for the co-ordinated regulation of cellular metabolism and cell growth. Herein, we demonstrate that suppressing the cellular activity of glycogen synthase kinase-3 (GSK3), by use of pharmacological inhibitors or shRNA-mediated gene silencing, results in substantial reduction in amino acid (AA)-regulated mTORC1-directed signalling, as assessed by phosphorylation of multiple downstream mTORC1 targets. We show that GSK3 regulates mTORC1 activity through its ability to phosphorylate the mTOR-associated scaffold protein raptor (regulatory-associated protein of mTOR) on Ser(859). We further demonstrate that either GSK3 inhibition or expression of a S859A mutated raptor leads to reduced interaction between mTOR and raptor and under these circumstances, irrespective of AA availability, there is a consequential loss in phosphorylation of mTOR substrates, such as p70S6K1 (ribosomal S6 kinase 1) and uncoordinated-51-like kinase (ULK1), which results in increased autophagic flux and reduced cellular proliferation.
Division of Cell Signalling & Immunology, James Black Centre, College of Life Sciences, University of Dundee, Dundee DD1 5EH, U.K.
Clare
C
Stretton
Division of Cell Signalling & Immunology, James Black Centre, College of Life Sciences, University of Dundee, Dundee DD1 5EH, U.K.
Thorsten M
TM
Hoffmann
MRC Protein Phosphorylation Unit, James Black Centre, College of Life Sciences, University of Dundee, Dundee DD1 5EH, U.K.
Michael J
MJ
Munson
Division of Cell Signalling & Immunology, James Black Centre, College of Life Sciences, University of Dundee, Dundee DD1 5EH, U.K.
Alan
A
Prescott
Division of Cell Signalling & Immunology, James Black Centre, College of Life Sciences, University of Dundee, Dundee DD1 5EH, U.K.
Peter M
PM
Taylor
MRC Protein Phosphorylation Unit, James Black Centre, College of Life Sciences, University of Dundee, Dundee DD1 5EH, U.K.
Ian G
IG
Ganley
Division of Cell Signalling & Immunology, James Black Centre, College of Life Sciences, University of Dundee, Dundee DD1 5EH, U.K. h.s.hundal@dundee.ac.uk.
Harinder S
HS
Hundal
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism*; D000818:Animals; D001343:Autophagy; D002460:Cell Line; D049109:Cell Proliferation; D020868:Gene Silencing; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008247:Lysosomes / Q000378:metabolism; D000076222:Mechanistic Target of Rapamycin Complex 1; D051379:Mice; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes / Q000378:metabolism*; D009154:Mutation; D010766:Phosphorylation; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D000076223:Regulatory-Associated Protein of mTOR; D012694:Serine / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; D000596:Amino Acids; D046912:Multiprotein Complexes; D034741:RNA, Small Interfering; C463659:RPTOR protein, human; D000076223:Regulatory-Associated Protein of mTOR; D012694:Serine; D000076222:Mechanistic Target of Rapamycin Complex 1; D058570:TOR Serine-Threonine Kinases; D038362:Glycogen Synthase Kinase 3
L-type (leucine) amino acid transporter 1 (LAT1); amino acid; autophagy; growth; insulin; leucine; p70S6K1; proliferation; sodium-coupled neutral amino acid transporter 2 (SNAT2); transcription factor EB (TFEB); uncoordinated-51-like kinase (ULK1)