G Alpha-q/11 Protein Plays a Key Role in Insulin-Induced Glucose Transport in 3T3-L1 Adipocytes

Abstract

We evaluated the role of the G alpha-q (Gαq) subunit of heterotrimeric G proteins in the insulin signaling pathway leading to GLUT4 translocation. We inhibited endogenous Gαq function by single cell microinjection of anti-Gαq/11 antibody or RGS2 protein (a GAP protein for Gαq), followed by immunostaining to assess GLUT4 translocation in 3T3-L1 adipocytes. Gαq/11 antibody and RGS2 inhibited insulin-induced GLUT4 translocation by 60 or 75%, respectively, indicating that activated Gαq is important for insulin-induced glucose transport. We then assessed the effect of overexpressing wild-type Gαq (WT-Gαq) or a constitutively active Gαq mutant (Q209L-Gαq) by using an adenovirus expression vector. In the basal state, Q209L-Gαq expression stimulated 2-deoxy-d-glucose uptake and GLUT4 translocation to 70% of the maximal insulin effect. This effect of Q209L-Gαq was inhibited by wortmannin, suggesting that it is phosphatidylinositol 3-kinase (PI3-kinase) dependent. We further show that Q209L-Gαq stimulates PI3-kinase activity in p110α and p110γ immunoprecipitates by 3- and 8-fold, respectively, whereas insulin stimulates this activity mostly in p110α by 10-fold. Nevertheless, only microinjection of anti-p110α (and not p110γ) antibody inhibited both insulin- and Q209L-Gαq-induced GLUT4 translocation, suggesting that the metabolic effects induced by Q209L-Gαq are dependent on the p110α subunit of PI3-kinase. In summary, (i) Gαq appears to play a necessary role in insulin-stimulated glucose transport, (ii) Gαq action in the insulin signaling pathway is upstream of and dependent upon PI3-kinase, and (iii) Gαq can transmit signals from the insulin receptor to the p110α subunit of PI3-kinase, which leads to GLUT4 translocation.

ACKNOWLEDGMENTS

This work was supported in part by NIH grant DK-33651 and the V.A. Medical Research Service. Takeshi Imamura is supported through an ADA Mentor Board Fellowship Award. Peter Vollenweider was supported by a grant from the Schweizerische Stiftung fur Medizinisch-Biologische Stipendien, and Martin Clodi was supported by grants J01287-Med and J1584-Med from the Ewin Schrödinger Stipendium by the Austrian Fonds zur Förderany der Wissenschaftlichen Forschung.

We thank John R. Hepler (Washington University, St. Louis, Mo.) for providing the RGS2 expression vector, David W. Rose (University of California, San Diego, La Jolla, Calif.) for technical assistance with microinjection, and Elizabeth Hansen and Augustus P. Lestick for editorial assistance.