Insulin-stimulated Protein Tyrosine Phosphorylation in Intact Cells Evaluated by Giant Two-dimensional Gel Electrophoresis

摘要: We have studied the insulin-stimulated phosphorylation of proteins in NIH 3T3 cells expressing high numbers human insulin receptors (HIR 3.5 cells) using technique giant two-dimensional gel electrophoresis. In serum-deprived cells, stimulated more than 25 proteins; all but two these were also phosphorylated response to 15% (v/v) fetal bovine serum, which additional thought be direct substrates for protein kinase C. pretreated specifically at least 26 predominantly cytosolic proteins, only one was observed insulin-treated not exposed phenylarsine oxide. Serum without effect with oxide-pretreated phosphoamino acid analysis 10 most highly labeled phosphoproteins showed that or exclusively on tyrosine residues. The several could vitro by addition a detergent extract presence Mn2+ and ATP. general, phosphorylations oxide rapid those its absence. Finally, variety other growth factors mitogens did stimulate any Thus, use this inhibitor apparently unmasked number novel insulin-specific ordinarily undetectable. suggest some may receptor play significant roles action.