Rapid stimulation by insulin of a serine/threonine kinase in 3T3-L1 adipocytes that phosphorylates microtubule-associated protein 2 in vitro.
Insulin treatment (Kact, 5 X 10(-9) M) of serum-starved 3T3-L1 adipocytes stimulates a soluble serine/threonine kinase that catalyzes phosphorylation of microtubule-associated protein 2 (MAP-2) in vitro. Maximal activation of MAP-2 kinase activity by 80 nM insulin was observed after 10 min of hormonal stimulation, prior to maximal stimulation of S6 kinase activity (20 min). The insulin-stimulatable MAP-2 kinase activity is not adsorbed to phosphocellulose, whereas the principal S6 kinase activity is retained and elutes at approximately 0.5 M NaCl. The insulin-stimulatable MAP-2 kinase is less stable during incubation at 30 degrees C than S6 kinase activity. Inclusion of phosphatase inhibitors decreases the rate at which the stimulated MAP-2 kinase activity is lost from extract supernatants incubated at 30 degrees C. p-Nitrophenyl phosphate is more effective than DL-phosphotyrosine, whereas DL-phosphoserine is without effect at the concentration used (40 mM). The difference in MAP-2 kinase activity in extract supernatants from control and insulin-treated cells is also preserved after rapid chromatography on Sephadex G-25. These results show that a soluble serine/threonine kinase is rapidly activated by insulin, possibly by phosphorylation of either the kinase itself or an interacting modulator.
D000273:Adipose Tissue / Q000201:enzymology*; D000596:Amino Acids / Q000032:analysis; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D007328:Insulin / Q000494:pharmacology*; D008869:Microtubule-Associated Proteins / Q000032:analysis / Q000378:metabolism*; D009596:Nitrophenols / Q000494:pharmacology; D009943:Organophosphorus Compounds / Q000494:pharmacology; D010766:Phosphorylation; D011494:Protein Kinases / Q000032:analysis*; D017346:Protein Serine-Threonine Kinases
D000596:Amino Acids; D007328:Insulin; D008869:Microtubule-Associated Proteins; D009596:Nitrophenols; D009943:Organophosphorus Compounds; C008644:nitrophenylphosphate; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases