Fig. 5.
Luciferase reporter assays. (A) Alignment of a region of the human and mouse Myc promoter sequences containing a recognition site for Cbf1 binding (site A). The start sites of promoters P1 and P2 are indicated. Segments of the plasmid constructs used as reporters are shown (for definition of binding sites, see Fig. 4A). In the diagrams, the putative Cbf1-binding sites (closed ovals) are intact, whereas, in the other constructs, one or more sites have been mutagenized (open ovals), as indicated. Constructs in which shorter segments of the mouse and human Myc promoters were used in association with a minimal Junb gene promoter (hatched rectangle) are also shown. (B) Results of reporter assays. 293T cells were cotransfected with Renilla luciferase plasmid and an effector plasmid expressing Cbf1-VP16 or a control plasmid (empty pcDNA3 vector; mock) in combination with one of the indicated reporter plasmids shown in A containing either intact or mutated Cbf1-binding sites. After normalization to Renilla luciferase activity, firefly luciferase activity relative to that of the control plasmid was calculated for each of the reporters. These relative values (mean ± SEM), measured in at least three independent experiments, are represented by the bars in the bar chart.