Fig. 2. Up-regulation of transcription from the murine VDR promoter induced by ZEB. In A, transcriptional activity of the VDR promoter fused to the luciferase gene in the pGL3Basic vector was assayed by cotransfection of this reporter gene (0.2 µg/well) with increasing amounts of ZEB expression vector (0.125, 0.25, and 0.5 µg) in COS 7 cells. B, requirement of CACCTG boxes in the VDR promoter region for the activation of the VDR promoter by ZEB in COS 7 cells. The activity of the wild-type VDR promoter reporter gene (0.2 µg/well) was measured in the presence of 0.5 µg of ZEB expression vector per well (ZEB) or the presence of 0.5 µg of control vector per well (CTRL) and compared with the activities of mutated VDR reporter genes with only one mutated E-box (Z1 or Z2) or with mutations in both E-boxes (Z1,2) in the presence of 0.5 µg of ZEB expression vector per well. Luciferase activities were normalized against the activities of the control vector pRL-TK. The average of three to seven independent transfections each with triplicate samples and SDs are shown. In some cases, the SDs were very low and do not show up as observable error bars.