Specific role for p85/p110beta in GTP-binding-protein-mediated activation of Akt.
We prepared CHO (Chinese hamster ovary) cells expressing both IR (insulin receptor) and A1R (A1 adenosine receptor). Treatment of the cells with insulin or PIA [N6-(2-phenylisopropyl)adenosine], a specific A(1)R agonist increased Akt activity in the cells in a PI3K- (phosphoinositide 3-kinase) dependent manner. Transfection of p110beta into the cells augmented the action of PIA with little effect on insulin. Introduction of a pH1 vector producing shRNA (short hairpin RNA) that targets p110beta abolished PIA-induced Akt activation. By contrast, an shRNA probe targeting p110alpha did not impair the effects of PIA. The effect of PIA in p110alpha-deficient cells was attenuated effectively by both Deltap85 and betaARK-CT (beta-adrenergic receptor kinase-C-terminal peptide). A Deltap85-derived protein possessing point mutations in its two SH2 domains did not impair PIA action. These results suggest that tyrosine-phosphorylated proteins and Gbetagamma (betagamma subunits of GTP-binding protein) are necessary for the specific function of p110beta in intact cells. The p110beta-middle (middle part of p110beta) may play an important role in signal reception from GPCRs (GTP-binding-protein-coupled receptor), because transfection of the middle part impaired PIA sensitivity.
Kubo
Hiroshi
H
Division of Molecular Medical Science, Graduate School of Biomedical Sciences, Hiroshima University, Hiroshima 734-8551, Japan.
Hazeki
Kaoru
K
Takasuga
Shunsuke
S
Hazeki
Osamu
O
D000818:Animals; D016466:CHO Cells; D002417:Cattle; D006224:Cricetinae; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000378:metabolism*; D005786:Gene Expression Regulation; D006801:Humans; D007328:Insulin; D051379:Mice; D019869:Phosphatidylinositol 3-Kinases / Q000737:chemistry* / Q000378:metabolism*; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D043682:Receptor, Adenosine A1 / Q000378:metabolism*; D011972:Receptor, Insulin / Q000378:metabolism
D007328:Insulin; D018129:Phosphatidylinositol Phosphates; D043682:Receptor, Adenosine A1; C060974:phosphatidylinositol 3,4,5-triphosphate; D019869:Phosphatidylinositol 3-Kinases; D011972:Receptor, Insulin; D051057:Proto-Oncogene Proteins c-akt; D019204:GTP-Binding Proteins