FIGURE 5.

ERK1/2 mediates thrombin-induced phosphorylation of CREB-Ser¹³³. A, cultured VSM cells were pretreated with the MEK inhibitor U0126 (5 μm) or vehicle control (dimethyl sulfoxide, DMSO) for 30 min prior to stimulation with thrombin (50 nm) or ionomycin (Iono; 0. 5 μm) for the indicated times. Cell lysates were resolved by SDS-PAGE and immunoblotted for phospho-ERK1/2, phospho-Ser¹³³-CREB (pSer133), total CREB, and GAPDH. B, P-ERK ECL signals, and C, CREB-Ser(P)¹³³ signals were quantified with a FujiLAS 4000. Data were normalized for loading using GAPDH signals and expressed as fold-values compared with unstimulated. Values shown are mean ± S.E., n = 3 and were analyzed by one-way ANOVA. *, p < 0.05; **, p < 0.01; ***, p < 0.001, by Bonferroni post hoc test.