Compartmentalization and insulin-induced translocations of insulin receptor substrates, phosphatidylinositol 3-kinase, and protein kinase B in rat liver

Physiological doses of insulin in rats resulted in a rapid redistribution o f key signaling proteins between subcellular compartments in rat liver. In plasma membranes (PM) and microsomes, insulin induced a rapid decrease in i nsulin receptor substrate-1/2 (IRS1/2) within 30 sec and an increase in the se proteins in endosomes (EN) and cytosol. The level of p85 in PM increased 2.3-foId at 30 sec alter insulin stimulation followed by a decrease at 2 m in. In this interval, 60-85% and 10-20% of p85 in PM was associated with IR S1 and IRS2, respectively. Thus, in PM, IRS1/2 accounts for almost all of t he protein involved in phosphatidylinositol 3-kinase activation. In ENs ins ulin induced a maximal increase of 40% in p85 recruitment. As in PM, almost all p85 was associated with IRS1/2. The greater level of p85 recruitment t o PM was associated with a higher level of insulin-induced recruitment of A kt1 to this compartment (4.0-fold in PM us. 2.4-fold in EN). There was a cl ose correlation between Akt1 activity and Akt1 phosphorylation at Thr(308) and Ser(473) in PM and cytosol. However, in ENs the level of Akt1 activity per unit of phosphorylated Akt1 was significantly greater than in PM, indic ating that in addition to phosphorylation, another factor(s) modulates Akt1 activation by insulin in rat liver. Our results demonstrate that activatio n of the insulin receptor kinase and modulation of key components of the in sulin signaling cascade occur at the cell surface and within the endosomal system. These data provide further support for the role of the endocytic pr ocess in cell signaling.