FIG. 9.

E1 and E3 E boxes are important for ngn3-E47-mediated activation of the BETA2 promoter. (A) Mutational analyses of the three proximal E boxes in the BETA2 promoter in the presence of ngn3 and E47. All constructs (except the control) derived from a TATA-luciferase reporter driven by a 155-bp BETA2 promoter fragment containing three E boxes in either a wild type (WT-3E; open box) or mutated (hatched box) form. (B) Illustration of mutated E box sequences, in each of which the sequence CANNTG was replaced by TANNCT. Each construct was cotransfected into HeLa cells with both ngn3 and E47 expression vectors. The activation of each construct by ngn3-E47 is represented as fold activation (±standard deviation of the mean). (C) Synergistic activation of individual E box reporters by ngn3 and E47. Three copies of each E box (E1, E2, and E3) were cloned into pGL3-TATA vector. The resultant reporters (300 ng) were then cotransfected into HeLa cells with ngn3 and/or E47 expression vectors (100 ng). The transfections were repeated three times. Top four rows, E1 results; middle four rows, E3 results; bottom four rows, E2 results.