Figure 3

RAMP-1 promoter/luciferase activity in RAMP-1 positive and negative cell lines. RAMP-1 promoter-deletion constructs were generated by PCR and ligated into the luciferase expression vector pGL3-basic. The amount of 5' flanking region relative to the start site of translation (ATG = 0) is indicated for each construct. Constructs were transiently transfected into RAMP-1 positive expressing cells (C2C12 and NIH3T3) and RAMP-1 negative cells (HEK293). Forty-eight hours after transfection, cells were harvested and assayed for luciferase activity. The co-transfection control plasmid pRL-TK was employed for all transfections. Relative luciferase activities were determined by comparing luciferase activity of cells transfected with RAMP-1/luciferase deletion constructs to cells transfected with the promoterless pGL3-basic plasmid. Background levels of luciferase = 1. The data are an average of three independent sets of transfections each giving similar results.