Insulin receptor substrate-2 (IRS-2) can mediate the action of insulin to stimulate translocation of GLUT4 to the cell surface in rat adipose cells

Insulin receptor substrates-1 and -2 (IRS-1 and -2) are important substrates of the insulin receptor tyrosine kinase, Previous studies have focused upon the role of IRS-1 in mediating the actions of insulin, In the present study, we demonstrate that IRS-2 can mediate translocation of the insulin responsive glucose transporter GLUT4 in a physiologically relevant target cell for insulin action, Co-immunoprecipitation experiments performed on cell lysates derived from freshly isolated rat adipose cells incubated in the presence or absence of insulin indicated that twice as much phosphatidylinositol 3-kinase was associated with endogenous IRS-1 as with IRS-2 after insulin stimulation, When rat adipose cells in primary culture were transfected with expression vectors for IRS-1 or IRS-2, we observed 40-fold over-expression of human IRS-1 or murine IRS-2. In addition, anti-phosphotyrosine immunoblotting experiments confirmed that the recombinant substrates were phosphorylated in response to insulin stimulation, To examine the role of IRS-2 in insulin-stimulated translocation of GLUT4, we studied the effects of overexpression of IRS-1 and -2 on translocation of a co-transfected epitope-tagged GLUT4 (GLUT4-HA). Overexpression of IRS-1 or IRS-2 in adipose cells resulted in a significant increase in the basal level of cell surface GLUT4 (in the absence of insulin). Interestingly, at maximally effective concentrations of insulin (60 nM), the level of cell surface GLUT4 in cells overexpressing IRS-1 or -2 significantly exceeded the maximal recruitment observed in the control cells (160 and 135% of control, respectively; p < 0.003), Our data directly demonstrate that IRS-2, like IRS-1, is capable of participating in insulin signal transduction pathways leading to the recruitment of GLUT4, Thus, IRS-2 may provide an alternative pathway for critical metabolic actions of insulin.