title: Insulin activation of vacuolar protein sorting 34 mediates localized phosphatidylinositol 3-phosphate production at lamellipodia and activation of mTOR/S6K1.
abstract: The class III phosphatidylinositol 3-kinase, VPS34, phosphorylates the D3 hydroxyl of inositol generating phosphatidylinositol 3-phosphate (ptdins(3)p). Initial studies suggested that ptdins(3)p solely functioned as a component of vesicular and endosomal membranes and that VPS34 did not function in signal transduction. However, VPS34 has recently been shown to be required for insulin-mediated activation of S6 kinase 1 (S6K1). Whether VPS34 activity is directly regulated by insulin is unclear. It is also not known whether VPS34 activity can be spatially restricted in response to extracellular stimuli. Data presented here demonstrate that in response to insulin, VPS34 is activated and translocated to lamellipodia where it produces ptdins(3)p. The localized production of ptdins(3)p is dependent on Src phosphorylation of VPS34. In cells expressing VPS34 with mutations at Y231 or Y310, which are Src-phosphorylation sites, insulin-stimulated VPS34 translocation to the plasma membrane and lamellipodia formation are blocked. mTOR also colocalizes with VPS34 and ptdins(3)p at lamellipodia following insulin-stimulation. In cells expressing the VPS34-Y231F mutant, which blocks lamellipodia formation, mTOR localization at the plasma membrane and insulin-mediated S6K1 activation are reduced. This suggests that mTOR localization at lamellipodia is important for full activation of S6K1 induced by insulin. These data demonstrate that insulin can spatially regulate VPS34 activity through Src-mediated tyrosine phosphorylation and that this membrane localized activity contributes to lamellipodia formation and activation of mTOR/S6K1signaling.
authors.affiliation: Division of Monoclonal Antibodies, Office of Biotechnology Products, Office of Pharmaceutical Science, Center for Drug Evaluation and Research, U.S. Food and Drug Administration (FDA), Bethesda, MD, USA.
authors.forename: Dianne S
authors.initials: DS
authors.lastname: Hirsch
authors.affiliation: Division of Monoclonal Antibodies, Office of Biotechnology Products, Office of Pharmaceutical Science, Center for Drug Evaluation and Research, U.S. Food and Drug Administration (FDA), Bethesda, MD, USA.
authors.forename: Yi
authors.initials: Y
authors.lastname: Shen
authors.affiliation: Division of Monoclonal Antibodies, Office of Biotechnology Products, Office of Pharmaceutical Science, Center for Drug Evaluation and Research, U.S. Food and Drug Administration (FDA), Bethesda, MD, USA.
authors.forename: Milos
authors.initials: M
authors.lastname: Dokmanovic
authors.affiliation: Division of Monoclonal Antibodies, Office of Biotechnology Products, Office of Pharmaceutical Science, Center for Drug Evaluation and Research, U.S. Food and Drug Administration (FDA), Bethesda, MD, USA.
authors.forename: Joyce
authors.initials: J
authors.lastname: Yu
authors.affiliation: Division of Monoclonal Antibodies, Office of Biotechnology Products, Office of Pharmaceutical Science, Center for Drug Evaluation and Research, U.S. Food and Drug Administration (FDA), Bethesda, MD, USA.
authors.forename: Nishant
authors.initials: N
authors.lastname: Mohan
authors.affiliation: Division of Monoclonal Antibodies, Office of Biotechnology Products, Office of Pharmaceutical Science, Center for Drug Evaluation and Research, U.S. Food and Drug Administration (FDA), Bethesda, MD, USA; Interagency Oncology Task Force (IOTF) Fellowship: Program 4 - Cancer Prevention Fellow, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA.
authors.forename: Mohammed Khair
authors.initials: MK
authors.lastname: Elzarrad
authors.affiliation: Division of Monoclonal Antibodies, Office of Biotechnology Products, Office of Pharmaceutical Science, Center for Drug Evaluation and Research, U.S. Food and Drug Administration (FDA), Bethesda, MD, USA. Electronic address: wen.wu@fda.hhs.gov.
authors.forename: Wen Jin
authors.initials: WJ
authors.lastname: Wu
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000201:enzymology; D058546:Class III Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism*; D004789:Enzyme Activation; D007328:Insulin / Q000502:physiology*; D051379:Mice; D020125:Mutation, Missense; D041681:NIH 3T3 Cells; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism*; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D021381:Protein Transport; D011554:Pseudopodia / Q000201:enzymology*; D038762:Ribosomal Protein S6 Kinases, 70-kDa / Q000378:metabolism*; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000378:metabolism; D019061:src-Family Kinases / Q000378:metabolism
chemical_list: D007328:Insulin; D018129:Phosphatidylinositol Phosphates; C107953:RAC1 protein, human; C055525:phosphatidylinositol 3-phosphate; C546842:MTOR protein, human; D058546:Class III Phosphatidylinositol 3-Kinases; D019061:src-Family Kinases; D038762:Ribosomal Protein S6 Kinases, 70-kDa; D058570:TOR Serine-Threonine Kinases; C448901:ribosomal protein S6 kinase, 70kD, polypeptide 1; D020830:rac1 GTP-Binding Protein
keywords: Class III phosphatidylinositol 3-kinase/vacuolar protein sorting 34 (VPS34); Insulin; S6 kinase 1 (S6K1); Src; mTOR; ptdins(3)p