FIG. 2.

PKC activity in L6 cells transfected with IRS-1 rib. (Top) L6hIR2 cells were transfected with either IRS1-Rib or c-Rib, solubilized, separated by SDS-PAGE, and blotted with IRS-1 or IRS-2 antibodies as indicated (left). WB, Western blot. Alternatively (right), the cells were exposed to 100 nM insulin for 5 min, and the cell lysates were precipitated with IRS-1 or IRS-2 antibodies, subjected to SDS-PAGE, and Western blotted with phosphotyrosine antibodies (PTYR). Blots were revealed by ECL as described in Materials and Methods. The autoradiographs shown are representative of six (left) and four (right) independent experiments. IP, immunoprecipitation. (Bottom) PKC activity was assayed in lysates from L6hIR cells either in the absence or upon transfection of c-rib or IRS1-Rib as described in the legend to Fig. 1. Bars represent the means ± standard deviations of triplicate determinations in four independent experiments.