Role of cytosolic tyrosine kinase in mediating insulin-like actions of vanadate in rat adipocytes.
In previous studies we have identified a cytosolic protein tyrosine kinase (CytPTK) in rat adipocytes that is largely activated in vanadate-pretreated cells (Shisheva, A., and Shechter, Y. (1992) FEBS Lett. 300, 93-96). We report here that staurosporine and its analog K-252a are highly potent (ID50 = 3 and 100 nM, respectively) in inhibiting CytPTK activity of crude cell extract or partially purified enzyme preparations. Staurosporine and K-252a were less effective by more than 2 and 1 orders of magnitude, respectively, in inhibiting insulin receptor-catalyzed PolyGlu4Tyr phosphorylation in cell-free experiments. Preincubation of rat adipocytes with either staurosporine or K-252a selectively blocked the action of vanadate in activating glucose incorporation into lipids and its oxidation. Thus, staurosporine inhibited vanadate-stimulated lipogenesis and glucose oxidation (via glycolysis and the pentose phosphate pathway) in a concentration-dependent manner with ID50 of 75 and 300 nM, respectively. Insulin-stimulated bioeffects were not inhibited at this low range of staurosporine concentration. Staurosporine had no effect on vanadate-stimulated hexose uptake or on vanadate's antilipolytic action. Using staurosporine, we probed those insulinomimetic agents which facilitate their biological activity via the insulin receptor kinase (insulin, wheat germ agglutinin, concanavalin A, and pervanadate) or via CytPTK (vanadate and to a certain degree Mn2+ and Zn2+). These results suggest that (a) vanadate facilitates its insulin-like actions on glucose utilization via the cytosolic tyrosine kinase and (b) this enzyme does not participate in vanadate effects in stimulating hexose uptake and in inhibiting lipolysis. These findings explain further vanadate's post-insulin receptor actions and raise possible application in the management of glucose metabolism in insulin-independent fashion in pathological conditions.
Shisheva
A
A
Department of Hormone Research, Weizmann Institute of Science, Rehovot, Israel.
Shechter
Y
Y
D000273:Adipose Tissue / Q000166:cytology / Q000201:enzymology*; D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D002478:Cells, Cultured; D003600:Cytosol / Q000201:enzymology*; D007328:Insulin / Q000494:pharmacology; D008055:Lipids / Q000096:biosynthesis; D008297:Male; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D011972:Receptor, Insulin / Q000378:metabolism; D019311:Staurosporine; D013552:Swine; D014638:Vanadates / Q000494:pharmacology*
D000470:Alkaloids; D007328:Insulin; D008055:Lipids; D014638:Vanadates; D011505:Protein-Tyrosine Kinases; D011972:Receptor, Insulin; D019311:Staurosporine