ISOPROTERENOL INHIBITS INSULIN-STIMULATED TYROSINE PHOSPHORYLATION OFTHE INSULIN-RECEPTOR WITHOUT INCREASING ITS SERINE THREONINE PHOSPHORYLATION

The effect of a beta-adrenergic agonist (isoproterenol) on the tyrosin e kinase activity of the insulin receptor was studied in intact adipoc ytes. Isoproterenol treatment rapidly (5 min) inhibited the insulin-in duced autophosphorylation of the insulin receptor on tyrosine residues in intact adipocytes. The effect of insulin on the phosphorylation of cellular proteins on tyrosine residues was also inhibited by isoprote renol. In order to understand the mechanism responsible for this inhib ition, two-dimensional phosphopeptide mapping of the insulin receptor was performed. The pattern of phosphorylation of the insulin receptor in freshly isolated adipocytes showed marked differences from that pre viously observed in cultured cells overexpressing insulin receptors. T hese differences include a larger proportion of receptors being phosph orylated on the three tyrosines from the kinase domain and no apparent phosphorylation of the two tyrosines close to the C-terminus after in sulin stimulation. Isoproterenol markedly inhibited the effect of insu lin on the phosphorylation of the three tyrosines from the kinase doma in. However, this inhibition was not associated with an increase in th e phosphorylation of serine/threonine peptides. Thus, this direct anal ysis of insulin receptor phosphorylation sites in intact adipocytes do es no support the idea that beta-adrenegic agents inhibit the tyrosine kinase activity of the receptor through a serine/threonine phosphoryl ation-dependent mechanism.