Insulin-stimulated tyrosine phosphorylation of caveolin is specific for the differentiated adipocyte phenotype in 3T3-L1 cells.
Previous work from this laboratory has shown that insulin stimulates the tyrosine phosphorylation of caveolin in 3T3-L1 adipocytes (Mastick, C. C., Brady, M. J., and Saltiel, A. R. (1995) J. Cell Biol. 129, 1523-1531). This phosphorylation is specific for insulin and involves the activation of a tyrosine kinase downstream of the insulin receptor. We report here that tyrosine phosphorylation of caveolin is detected only in fully differentiated adipocytes, not in fibroblasts (preadipocytes), despite the fact that both cell types express caveolin-1 and active insulin receptor. Caveolin copurifies with caveolin tyrosine kinase activity in both preadipocytes and adipocytes. Accumulating evidence indicates that this kinase is the Src family kinase Fyn. Fyn is expressed in the preadipocytes and the adipocytes and is colocalized with caveolin in low density Triton-insoluble complexes in both cell types. In adipocytes, overexpression of wild type Fyn leads to increased basal phosphorylation of caveolin and hyperphosphorylation of caveolin in response to insulin. In vitro kinase assays suggest that Fyn may be activated in response to insulin through the binding of a tyrosine-phosphorylated insulin receptor substrate protein. Previous work suggested that this protein may be c-Cbl (Ribon, V., and Saltiel, A. R. (1997) Biochem. J. 324, 839-846). In 3T3-L1 adipocytes, Cbl binds to Fyn in an insulin-dependent manner, and Cbl phosphorylation is adipocyte-specific. Here we show that phosphorylated Cbl is translocated into caveolin-enriched Triton-insoluble complexes after insulin stimulation. This may lead to the cell type-specific, compartmentalized activation of Fyn and the specific phosphorylation of proteins in the caveolae in response to insulin in adipocytes.
Mastick
C C
CC
Department of Cell Biology, Parke-Davis Pharmaceutical Research Division, Warner-Lambert Co., Ann Arbor, Michigan 48105, USA.
Saltiel
A R
AR
D016475:3T3 Cells; D017667:Adipocytes / Q000737:chemistry / Q000378:metabolism*; D000818:Animals; D051242:Caveolin 1; D022461:Caveolins; D007328:Insulin / Q000494:pharmacology*; D008565:Membrane Proteins / Q000032:analysis / Q000378:metabolism*; D051379:Mice; D010641:Phenotype; D010766:Phosphorylation; D011518:Proto-Oncogene Proteins / Q000032:analysis; D051076:Proto-Oncogene Proteins c-fyn; D013234:Stem Cells / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism*
C495541:Cav1 protein, mouse; D051242:Caveolin 1; D022461:Caveolins; D007328:Insulin; D008565:Membrane Proteins; D011518:Proto-Oncogene Proteins; D014443:Tyrosine; C494960:Fyn protein, mouse; D051076:Proto-Oncogene Proteins c-fyn