title: Insulin-like growth factor-I stimulates Shc-dependent phosphatidylinositol 3-kinase activation via Grb2-associated p85 in vascular smooth muscle cells.
abstract: Insulin-like growth factor-I (IGF-I) stimulates vascular smooth muscle cell proliferation and migration by activating both MAPK and phosphatidylinositol 3-kinase (PI3K). Vascular smooth muscle cells (VSMCs) maintained in 25 mm glucose sustain MAPK activation via increased Shc phosphorylation and Grb2 association resulting in an enhanced mitogenic response compared with cells grown in 5 mm glucose. PI3K plays a major role in IGF-I-stimulated VSMC migration, and hyperglycemia augments this response. In contrast to MAPK activation the role of Shc in modulating PI3K in response to IGF-I has not been determined. In this study we show that impaired Shc association with Grb2 results in decreased Grb2-p85 association, SHPS-1-p85 recruitment, and PI3K activation in response to IGF-I. Exposure of VSMCs to cell-permeable peptides, which contained polyproline sequences from p85 proposed to mediate Grb2 association, resulted in inhibition of Grb2-p85 binding and AKT phosphorylation. Transfected cells that expressed p85 mutant that had specific prolines mutated to alanines resulted in less Grb2-p85 association, and a Grb2 mutant (W36A/W193A) that attenuated p85 binding showed decreased association of p85 with SHPS-1, PI3K activation, AKT phosphorylation, cell proliferation, and migration in response to IGF-I. Cellular exposure to 25 mm glucose, which is required for Shc phosphorylation in response to IGF-I, resulted in enhanced Grb2 binding to p85, activation of PI3K activity, and increased AKT phosphorylation as compared with cells exposed to 5 mm glucose. We conclude that in VSMCs exposed to hyperglycemia, IGF-I stimulation of Shc facilitates the transfer of Grb2 to p85 resulting in enhanced PI3K activation and AKT phosphorylation leading to enhanced cell proliferation and migration.
authors.affiliation: Department of Medicine, University of North Carolina School of Medicine, Chapel Hill, North Carolina 27599, USA.
authors.forename: Yashwanth
authors.initials: Y
authors.lastname: Radhakrishnan
authors.forename: Laura A
authors.initials: LA
authors.lastname: Maile
authors.forename: Yan
authors.initials: Y
authors.lastname: Ling
authors.forename: Lee M
authors.initials: LM
authors.lastname: Graves
authors.forename: David R
authors.initials: DR
authors.lastname: Clemmons
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000818:Animals; D002465:Cell Movement; D049109:Cell Proliferation; D004730:Endothelium, Vascular / Q000378:metabolism*; D004789:Enzyme Activation; D051380:GRB2 Adaptor Protein / Q000378:metabolism*; D005947:Glucose / Q000378:metabolism; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000378:metabolism*; D020935:MAP Kinase Signaling System; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D011392:Proline / Q000737:chemistry; D038762:Ribosomal Protein S6 Kinases, 70-kDa / Q000378:metabolism*; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D013552:Swine
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; C528024:SHC1 protein, human; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D007334:Insulin-Like Growth Factor I; D011392:Proline; D019869:Phosphatidylinositol 3-Kinases; D038762:Ribosomal Protein S6 Kinases, 70-kDa; C448901:ribosomal protein S6 kinase, 70kD, polypeptide 1; D005947:Glucose