Figure 2

5' deletion of the Jab1 promoter analysis reveals increased activity in breast cancer cells compared with normal cells. (a) Identification of the minimal promoter region of Jab1 using progressive deletion of the 5' region in luciferase reporter constructs. The plasmids were cotransfected with Renilla luciferase plasmid pRL-null (transfection control) into MCF7 cells, and luciferase activity was assayed. Values represent mean ± standard error of triplicate experiments. Luciferase assays shown in subsequent figures were performed similarly. (b) Jab1 promoter 5' deletion constructs were transfected into MCF7 and MCF-10A cells and subjected to luciferase reporter assays. Promoter activity was higher in MCF7 epithelial breast cancer cells than in non-tumorigenic epithelial MCF-10A cells. Deletion of the region -472 to -344 resulted in a loss of promoter activity in both cell lines. (c) A panel of breast cancer cells and normal mammary cells were transfected with the -472 Jab1-Luc construct. Significance is noted as * P < 0.05 and ** P < 0.01 and was determined by t-test. (d) Jab1 protein levels for the cell lines in (c) were detected by western blotting, β-actin is shown as a control. Jab1, c-Jun activation domain-binding protein-1.