Insulin activates glycogen synthase by a novel PI 3-kinase/p70s6k dependent pathway in 3T3-L1 adipocytes.

Abstract

Insulin causes the activation of a tyrosine kinase activity in the intracellular domain of its receptor, the major substrate of this kinase being the multifunctional docking protein E l . Phosphorylation of specific tyrosine residues on IRSl allows this protein to interact with and activate a number of downstream signalling molecules including phosphoinositide 3-kinase (PI 3-kinase), SHPTP2 and the Grb-2/Sos/ras system (and hence the MAP kinase cascade) 111. Insulin also causes the activation of ribosomal protein S6-kinase (p70dk) by unknown mechanisms [2]. However it is not clear how insulin stimulation of these signalling pathways exerts control over metabolic pathways. We have used rapamyan, which specifically blocks growth factor activation of p70dk, and the specific PI 3kinase inhibitor wortmannin, to investigate the involvement of PI 3-kinase and p706k in insulin stimulation of glucose transport and glycogen synthesis in 3T3-Ll adipocytes. We find that wortmannin inhibits PI 3-kinase activity in anti IRS-1 and anti-PI 3-kinase immunoprecipitates from 3T3-Ll adipocytes in a dose dependent manner (ICm of a 10 nM), with inhibtion being complete at 100 nM (data not shown). Wortmannin inhibited insulin stimulated glucose transport in a similar dose dependent manner in the 3T3-Ll adipocytes (ICm = 15nM) (Fig. 1). The potency of these inhibitory effects of wortmannin agrees with previous reports [3,41. Insulin stimulated glycogen synthase activity 2-3 fold in these cells. Wortmannin inhibited insulin stimulated activation of glycogen synthase with a similar dose dependency to its inhibition of insulin stimulated