Figure 5. Overexpression of p300/CBP overcomes p53-mediated repression of Rad51 core promoter activity.

H1299 cells were cotransfected with a plasmid expressing luciferase under the control of the Rad51core promoter, in the presence or absence of plasmids encoding wild-type p53 or p300. Increasing amounts of the p300 expression were used (as indicated); the total amount of amount of transfected plasmid DNA in each reaction was maintained at a constant 4 µg by adding the necessary amount of irrelevant pCMV-GFPflag plasmid DNA. (A) 48 hours post transfection, cells were collected, lysed and transcriptional activity measured by Luciferase assay. Results shown represent the mean of independent experimental triplicates; error bars represent the standard deviation of the data. (B) Western blot analysis was performed to confirm expression of p53 and p300. 10 µg of total protein from each sample was separated on a 7.5% polyacrylamide gel, transferred to a nitrocellulose membrane, and probed for each respective protein using antibodies specific for the indicated proteins (p300, p53 and β-tubulin as a loading control).