VANADATE ACTIVATES MEMBRANOUS NONRECEPTOR PROTEIN-TYROSINE KINASE IN RAT ADIPOCYTES

The insulin-like effects of vanadate are independent of the insulin re ceptor and insulin receptor substrate 1 (IRS-1) phosphorylation. A cyt osolic protein tyrosine kinase (CytPTK), sensitive to inhibition by na nomolar concentrations of staurosporine (concentration at which 50% in hibition occurs [IC50], 1-2 nmol/l), has been implicated in some (i.e. , glucose oxidation, lipogenesis) but not all (i.e., hexose uptake, in hibition of lipolysis) of the insulin-like effects of vanadate. We rep ort here the existence of another nonreceptor protein tyrosine kinase in rat adipocytes, located exclusively in the plasma membranes (MembPT K), which we suggest is associated with hexose uptake and the antilipo lytic activity of vanadate. MembPTK is a nonglycoprotein with an estim ated molecular weight of 55-60 kDa. In a cell-free experiment, vanadat e activates MembPTK seven-to ninefold (median effective dose, 17 +/- 2 mu mol/l). Vanadate-activated MembPTK is inhibited by staurosporine ( IC50, 60 +/- 5 nmol/l). In intact adipocytes, staurosporine antagonize d vanadate-induced hexose uptake (IC50, 6.0 +/- 0.3 mu mol/l) and sign ificantly reversed the antilipolytic effect of vanadate (IC50, 5.0 +/- 0.4 mu mol/l). After vanadate treatment, a phosphorylated P55 protein is immunoprecipitated by antibodies to both phosphotyrosine and phosp hatidylinositol (PI) 3-kinase. In conclusion, rat adipocytes contain a n additional vanadate-activatable nonreceptor membranous protein tyros ine kinase that may participate in the effects of vanadate not carried out by CytPTK. We also suggest that after treatment with vanadate, Me mbPTK is activated by autophosphorylation and interacts with PI 3-kina se. This may explain how vanadate activates PI 3-kinase without involv ing receptor activation and IRS-1 phosphorylation.