Insulin-like growth factor (IGF) binding protein-3 potentiation of IGF action is mediated through the phosphatidylinositol-3-kinase pathway and is associated with alteration in protein kinase B/AKT sensitivity.
Cell-association and processing of insulin-like growth factor binding protein-3 (IGFBP-3) by cultured bovine fibroblasts results in markedly enhanced type I IGF receptor signaling at a step distal to ligand binding. The purpose of the present study was to determine the intracellular mediators of IGFBP-3's potentiating effect. Preincubation of cultured bovine fibroblasts with 50 nM IGFBP-3 had no effect alone, but enhanced by 3- to 4-fold IGF-I-stimulated 3H-aminoisobutryric acid (AIB) uptake. IGFBP-3-induced potentiation was specifically prevented if an inhibitor of phosphatidylinositol 3 (PI3)-kinase activation (LY294002), but not an inhibitor of mitogen-activated protein kinase activation (PD98059), was present during the preincubation period. IGFBP-3 did not directly activate the downstream effector of PI3-kinase, protein kinase B (PKB)/Akt. However, the sensitivity of PKB/Akt to activation by IGF-I was increased by 2- to 4-fold with IGFBP-3 pretreatment. This increased sensitivity was accompanied by altered mobility of PKB/Akt on SDS-polyacrylamide gels, suggestive of a diminished phosphorylation state. Consistent with this, okadaic acid, a potent serine/threonine phosphatase inhibitor, was able to block the potentiation effect of IGFBP-3 and prevent the altered mobility of the PKB/Akt molecule in response to IGFBP-3 treatment. PKB/Akt immunoprecipitated from IGFBP-3-pretreated cells was no longer recognized by an antibody specific for phosphorylated threonine followed by proline. These data indicate that IGFBP-3 modulates type I IGF receptor signaling through an effect on PI-3-kinase pathway substrates and suggest a novel mechanism of dephosphorylation whereby PKB/Akt is transformed into a more sensitive substrate of type I IGF receptor signaling.
Conover
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CA
Endocrine Research Unit, Mayo Clinic and Mayo Foundation, Rochester, Minnesota 55905, USA. conover.cheryl@mayo.edu
Bale
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LK
Durham
S K
SK
Powell
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DR
D000621:Aminoisobutyric Acids / Q000378:metabolism; D000818:Animals; D015152:Blotting, Northern; D002417:Cattle; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts; D005419:Flavonoids / Q000494:pharmacology; D006801:Humans; D018972:Insulin-Like Growth Factor Binding Protein 3 / Q000494:pharmacology*; D009025:Morpholines / Q000494:pharmacology; D019319:Okadaic Acid / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D011233:Precipitin Tests; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D011994:Recombinant Proteins; D015398:Signal Transduction / Q000187:drug effects*; D013002:Somatomedins / Q000494:pharmacology*
D000621:Aminoisobutyric Acids; D002867:Chromones; D004791:Enzyme Inhibitors; D005419:Flavonoids; D018972:Insulin-Like Growth Factor Binding Protein 3; D009025:Morpholines; D019319:Okadaic Acid; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D011994:Recombinant Proteins; D013002:Somatomedins; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C494918:AKT1 protein, human; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one