title: Dual regulation of Akt/protein kinase B by heterotrimeric G protein subunits.
abstract: While positive regulation of c-Akt (also known as protein kinase B) by receptor tyrosine kinases is well documented, compounds acting through G protein-coupled receptors can also activate Akt and its downstream targets. We therefore explored the role of G protein subunits in the regulation of Akt in cultured mammalian cells. In HEK-293 and COS-7 cells transiently transfected with beta(2)-adrenergic or m2 muscarinic receptors, respectively, treatment with agonist-induced phosphorylation of Akt at serine 473 as evidenced by phosphoserine-specific immunoblots. This effect was blocked by the phosphatidylinositol-3-OH kinase inhibitor LY294002 and wild-type Galpha(i1), and was not duplicated by co-transfection of the constitutively active Galpha(s)-Q227L or Galpha(i)-Q204L mutant. Co-transfection of Gbeta(1), Gbeta(2) but not Gbeta(5) together with Ggamma(2) activated the kinase when assayed in vitro following immunoprecipitation of the epitope-tagged enzyme. In contrast, constitutively activated G protein subunits representing the four Galpha subfamilies were found unable to activate Akt in either cell line. The latter results are in disagreement with a report by Murga et al. (Murga, C., Laguinge, L., Wetzker, R., Cuadrado, A., and Gutkind, J. S. (1998) J. Biol. Chem. 273, 19080-19085) that described activation of Akt in response to mutationally activated Galpha(q) and Galpha(i) transfection in COS cells. To the contrary, in our experiments Galpha(q)-Q209L inhibited Akt activation resulting from betagamma or mutationally activated H-Ras co-transfection in these cells. In HEK-293 cells Galpha(q)-Q209L transfection inhibited insulin-like growth factor-1 activation of epitope-tagged Akt. In m1 muscarinic receptor transfected HEK-293 cells, carbachol inhibited insulin-like growth factor-1 stimulated phosphorylation at Ser(473) of endogenous Akt in an atropine-reversible fashion. We conclude that G proteins can regulate Akt by two distinct and potentially opposing mechanisms: activation by Gbetagamma heterodimers in a phosphatidylinositol-3-OH kinase-dependent fashion, and inhibition mediated by Galpha(q). This work identifies Akt as a novel point of convergence between disparate signaling pathways.
authors.affiliation: Metabolic Diseases Branch, NIDDK, National Institutes of Health, Bethesda, Maryland 20802-1752, USA.
authors.forename: R K
authors.initials: RK
authors.lastname: Bommakanti
authors.forename: S
authors.initials: S
authors.lastname: Vinayak
authors.forename: W F
authors.initials: WF
authors.lastname: Simonds
mesh_terms: D000818:Animals; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D002867:Chromones / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D020962:Heterotrimeric GTP-Binding Proteins / Q000378:metabolism*; D006801:Humans; D007545:Isoproterenol / Q000494:pharmacology; D009025:Morpholines / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases; D021122:Protein Subunits; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D018343:Receptors, Adrenergic, beta-2 / Q000187:drug effects / Q000502:physiology; D011976:Receptors, Muscarinic / Q000187:drug effects / Q000502:physiology; D011994:Recombinant Proteins / Q000378:metabolism; D013379:Substrate Specificity; D014162:Transfection
chemical_list: D002867:Chromones; D004791:Enzyme Inhibitors; D009025:Morpholines; D021122:Protein Subunits; D011518:Proto-Oncogene Proteins; D018343:Receptors, Adrenergic, beta-2; D011976:Receptors, Muscarinic; D011994:Recombinant Proteins; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D011505:Protein-Tyrosine Kinases; C494918:AKT1 protein, human; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D020962:Heterotrimeric GTP-Binding Proteins; D007545:Isoproterenol