Figure 1. Effects of RGS protein expression and Rho kinase inhibition on p70S6K, Akt phosphorylation, and ASM [³H]-thymidine incorporation stimulated by EGF and thrombin. ASM cells infected with GFP or GRK2NT-GFP (A, C, D, F) or GFP or p115RhoGEFRGS-GFP (B, C, D, F) were growth-arrested then stimulated with 10 ng/ml of EGF (E), 1 U/ml of thrombin (T), or both (E+T) for 30 min or 4 h. In (E) naive ASM cells were pretreated with Rho kinase inhibitor Y27623 (10µM) for 30 min prior to stimulation. Levels of phospho-p70S6K (T389), phospho-Akt (S473) and total Akt in cell lysates were assessed by immunoblotting. Representative blots (A, B, and E) and graphs depicting mean ± SE values of quantified phosphoprotein bands (C and D, n =5–7) are presented. Raw values for the 30 min EGF-stimulated condition were set to a value of 1.0 and all other values normalized accordingly. F) [³H]-thymidine incorporation stimulated by EGF, thrombin, or both in GFP-, GRK2NT-GFP-, or p115RhoGEFRGS-GFP-expressing lines (mean ±SE values, n =7 paired observations for GFP vs. GRK2NT-GFP, and n =4 paired observations for GFP vs. p115 RhoGEFRGS-GFP). *P <0.05, GFP vs. GRK2NT-GFP or p115 RhoGEFRGS-GFP group value for the indicated stimulatory condition.