Activation of protein kinase C (alpha, beta, and zeta) by insulin in 3T3/L1 cells. Transfection studies suggest a role for PKC-zeta in glucose transport.
We presently studied (a) insulin effects on protein kinase C (PKC) and (b) effects of transfection-induced, stable expression of PKC isoforms on glucose transport in 3T3/L1 cells. In both fibroblasts and adipocytes, insulin provoked increases in membrane PKC enzyme activity and membrane levels of PKC-alpha and PKC-beta. However, insulin-induced increases in PKC enzyme activity were apparent in both non-down-regulated adipocytes and adipocytes that were down-regulated by overnight treatment with 5 microM phorbol ester, which largely depletes PKC-alpha, PKC-beta, and PKC-epsilon, but not PKC-zeta. Moreover, insulin provoked increases in the enzyme activity of immunoprecipitable PKC-zeta. In transfection studies, stable overexpression of wild-type or constitutively active forms of PKC-alpha, PKC-beta1, and PKC-beta2 failed to influence basal or insulin-stimulated glucose transport (2-deoxyglucose uptake) in fibroblasts and adipocytes, despite inhibiting insulin effects on glycogen synthesis. In contrast, stable overexpression of wild-type PKC-zeta increased, and a dominant-negative mutant form of PKC-zeta decreased, basal and insulin-stimulated glucose transport in fibroblasts and adipocytes. These findings suggested that: (a) insulin activates PKC-zeta, as well as PKC-alpha and beta; and (b) PKC-zeta is required for, and may contribute to, insulin effects on glucose transport in 3T3/L1 cells.
Bandyopadhyay
G
G
J. A. Haley Veterans Hospital Research Service, and Departments of Internal Medicine and Biochemistry, University of South Florida, Tampa, Florida 33612, USA.
Standaert
M L
ML
Zhao
L
L
Yu
B
B
Avignon
A
A
Galloway
L
L
Karnam
P
P
Moscat
J
J
Farese
R V
RV
D016475:3T3 Cells; D000818:Animals; D002454:Cell Differentiation; D003847:Deoxyglucose / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D007328:Insulin / Q000494:pharmacology*; D007527:Isoenzymes / Q000378:metabolism*; D051379:Mice; D008970:Molecular Weight; D009004:Monosaccharide Transport Proteins / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014162:Transfection; D000099221:Protein Kinase C zeta
D003847:Deoxyglucose; D007328:Insulin; D007527:Isoenzymes; D009004:Monosaccharide Transport Proteins; D011493:Protein Kinase C; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha; D013755:Tetradecanoylphorbol Acetate; D000099221:Protein Kinase C zeta; C496630:Prkca protein, mouse