Figure 2
Knockdown of G i α proteins down regulates Akt and ERK activation, and reintroduction of G i α proteins restores their activation by EGF , bFGF and IGF - 1. (A-C) WT MEFs transfected with siRNA duplexes specific for Gi1α and Gi3α were treated with (A) EGF (100 ng/ml), (B) bFGF (25 ng/ml), or (C) IGF-1 (20 ng/ml) for the indicated time points. p-AKT(S473), p-ERK(T202/Y204), p-S6(S235/236), β-actin, Gi1α (Gi1) and Gi3α (Gi3) were detected by IB. The experiments were repeated two to three times and similar results were obtained. (D-F) WT MEFs were transfected with empty vector and Gi1/3 DKO MEFs were transfected with WT Gi1α, WT Gi3α or empty expression vectors (5 μg). Cells were treated with (D) EGF (100 ng/ml), (E) bFGF (25 ng/ml) or (F) IGF-1 (20 ng/ml) for 5, 15 min, or left untreated. p-ERK(T202/Y204), p-AKT(S473), p-S6(S235/236), or β-actin, Gi1α and Gi3α were detected by IB. The experiments were repeated two to three times and similar results were obtained.