Functional interactions of phosphatidylinositol 3-kinase with GTPase-activating protein in 3T3-L1 adipocytes.
The role of phosphatidylinositol (PI) 3-kinase in specific aspects of insulin signaling was explored in 3T3-L1 adipocytes. Inhibition of PI 3-kinase activity by LY294002 or wortmannin significantly enhanced basal and insulin-stimulated GTPase-activating protein (GAP) activity in 3T3-L1 adipocytes. Furthermore, removal of the inhibitory influence of PI 3-kinase on GAP resulted in dose-dependent decreases in the ability of insulin to stimulate p21ras. This effect was specific to adipocytes, as inhibition of PI 3-kinase did not influence GAP in either 3T3-L1 fibroblasts, Rat-1 fibroblasts, or CHO cells. Immunodepletion of either of the two subunits of the PI 3-kinase (p85 or p110) yielded similar activation of GAP, suggesting that catalytic activity of p110 plays an important role in controlling GAP activity in 3T3-L1 adipocytes. Inhibition of PI 3-kinase activity in 3T3-L1 adipocytes resulted in abrogation of insulin-stimulated glucose uptake and thymidine incorporation. In contrast, effects of insulin on glycogen synthase and mitogen-activated protein kinase activity were inhibited only at higher concentrations of LY294002. It appears that in adipocytes, P1 3-kinase prevents activation of GAP. Inhibition of PI 3-kinase activity or immunodepletion of either one of its subunits results in activation of GAP and decreases in GTP loading of p21ras.
DePaolo
D
D
Medical Research Service, Veterans Affairs Medical Center, Denver, Colorado 80220, USA.
Reusch
J E
JE
Carel
K
K
Bhuripanyo
P
P
Leitner
J W
JW
Draznin
B
B
D016475:3T3 Cells; D017667:Adipocytes / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D016466:CHO Cells; D002867:Chromones / Q000494:pharmacology; D006224:Cricetinae; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts; D020690:GTPase-Activating Proteins; D005947:Glucose / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D007328:Insulin / Q000494:pharmacology; D051379:Mice; D008934:Mitogens / Q000494:pharmacology; D009025:Morpholines / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism*; D016283:Proto-Oncogene Proteins p21(ras) / Q000378:metabolism*; D051381:Rats; D020703:ras GTPase-Activating Proteins
D002867:Chromones; D004791:Enzyme Inhibitors; D020690:GTPase-Activating Proteins; D007328:Insulin; D008934:Mitogens; D009025:Morpholines; D011506:Proteins; D020703:ras GTPase-Activating Proteins; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); C501009:HRAS protein, human; D016283:Proto-Oncogene Proteins p21(ras); D005947:Glucose