Rapid stimulation by insulin of a serine/threonine kinase in 3T3-L1 adipocytes that phosphorylates microtubule-associated protein 2 in vitro.

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Abstract
Insulin treatment (Kact, 5 .times. 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 eluates at .simeq. 5 M NaCl. The insulin-stimulatable MAP-2 kinase is less stable during incubation ats 30.degree.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.degree.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.