Figure 2
p53-independent activation of p21Waf1/Cip1 promoter activity by CPZ. (A) C6 cells were treated with 20 µM CPZ for the indicated time periods. Cellular extracts were subjected to Western blot analysis using the indicated antibodies. Each blot represents three individual experiments. (B) p53-dependent transcriptional activity assay. C6 cells were transfected with 0.5 µg of 5Xp53-Luc cis-reporting plasmid containing five repeats of the p53 binding site, along with 50 ng of the pRL-null vector. After 48 h, the cells were irradiated with UV (40 J/m2) or CPZ (20 µM) for an additional 8 h. (C) C6 cells were transfected with 0.5 µg of p21-Luc (-2400/+1 or -150/+38) plasmid. After 48 h, the cells were either left untreated or were treated with 20 µM CPZ for 8 h, and then analyzed for luciferase activity. The firefly luciferase activity was normalized to the Renilla activity. Error bars represent the mean ± SD of three independent experiments performed in triplicate.
