Activation and translocation of Rho (and ADP ribosylation factor) by insulin in rat adipocytes. Apparent involvement of phosphatidylinositol 3-kinase.
Insulin reportedly (Standaert, M. L., Avignon, A., Yamada, K., Bandyopadhyay, G., and Farese, R. V. (1996) Biochem. J. 313, 1039-1046) activates phospholipase D (PLD)-dependent hydrolysis of phosphatidylcholine (PC) in plasma membranes of rat adipocytes by a mechanism that may involve wortmannin-sensitive phosphatidylinositol (PI) 3-kinase. Because Rho and ADP ribosylation factor (ARF) activate PC-PLD, we questioned whether these small G-proteins are regulated by insulin and PI 3-kinase. We found that insulin provoked a rapid translocation of both Rho and ARF to the plasma membrane and increased GTP loading of Rho. Wortmannin and LY294002 inhibited Rho translocation in intact adipocytes, and the polyphosphoinositide, PI 4,5-(PO4)2, stimulated Rho translocation in adipocyte homogenates. On the other hand, wortmannin did not block GTP loading of Rho. Guanosine 5'-3-O-(thio)triphosphate stimulated both Rho and ARF translocation and activated PC-PLD in homogenates. C3 transferase, which inhibits and depletes Rho, inhibited PC-PLD activation by insulin in intact adipocytes. C3 transferase also inhibited insulin stimulation of [3H]2-deoxyglucose uptake. Our findings suggest that: (a) insulin translocates Rho by a PI 3-kinase-dependent mechanism, but another factor is responsible for GTP loading of Rho; (b) both Rho and ARF may contribute to PC-PLD activation during insulin action; and (c) Rho may be required for insulin stimulation of glucose transport.
Karnam
P
P
J. A. Haley Veterans' Hospital Research Service and the Departments of Internal Medicine and Biochemistry/Molecular Biology, University of South Florida, Tampa, Florida 33612, USA.
Standaert
M L
ML
Galloway
L
L
Farese
R V
RV
D020727:ADP-Ribosylation Factors; D017667:Adipocytes / Q000201:enzymology*; D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D002451:Cell Compartmentation; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism; D007328:Insulin / Q000378:metabolism* / Q000502:physiology*; D009025:Morpholines / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism*; D051381:Rats; D000077191:Wortmannin; D020741:rho GTP-Binding Proteins
D000730:Androstadienes; D002867:Chromones; D007328:Insulin; D009025:Morpholines; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D006160:Guanosine Triphosphate; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D019204:GTP-Binding Proteins; D020727:ADP-Ribosylation Factors; D020741:rho GTP-Binding Proteins; D000077191:Wortmannin