Figure 1.

HGF activates Akt in the absence of IRS2. A, Coimmunoprecipitation of PI3K and IRS2 in INS-1 cells after stimulation with 25 ng/mL HGF or 0.1 μM IGF-1 or left untreated. INS-1 cells were serum depleted for 6 hours and then were incubated with HGF or IGF-1 for 10 minutes. PI3K was immunoprecipitated as indicated in the Materials and Methods, and immunoblotting (IB) was performed against IRS2 and PI3K. Densitometric analysis was performed using ImageJ and quantitation appears in the bottom graph. Results are means ± SEM of 4 experiments. *, P < .05 vs. untreated (Unt) or HGF-treated cells. Notice the increase in IRS2/PI3K coimmunoprecipitation with IGF-1 but not with HGF. B, Western blot analysis of protein extracts from WT and IRS2 KO islets treated with 25 ng/mL HGF for 10 minutes or left untreated. Islets were serum depleted for 6 hours and then were treated with HGF. Immunoblots were performed against phospho-Ser473-AKT and actin as a housekeeping protein to control for loading. Densitometry of 3 different blots was performed using ImageJ, and ratios of the phospho-AKT against actin are represented on the bottom graph. Results are means ± SEM. *, P < .05 vs control (C) untreated.