MRE responds to MEK1,2-dependent MEKK-1 stimulation in a copy number—dependent manner. Isolated MRE region was inserted into pGL3 promoter to generate MRE/Luc, (MRE)2/Luc, and (MRE)4/Luc. These constructs were transfected into VSMCs with pCDNA3.1 blank vector (hatched bars) or MEKK-1* (solid bars). Transfected cells were treated with 50 μM PD98059 (PD), 5 μM SB203580 (SB), or 20 μM SP600125 (SP) 6 hours after the start of transfection. Cells were lysed 1 day after the start of transfection, and firefly luciferase activities were normalized to Renilla luciferase activities of cotransfected pRL-TK. Results show normalized luciferase activity (mean ± SEM) from at least 3 separate experiments. Fold increases induced by MEKK-1* compared with blank vector, pCDNA3.1, are shown. **Statistical significance with P < .001.
