FIGURE 3.

Ski suppresses the levels of acetylated p53 and reduces the DNA binding activity of p53. A, Ski expression reduces the amount of p53 on the promoters of p53 target genes. MCF7 cells and MCF7 cells stably expressing Ski were treated with or without 5 nm ActD for 8 h and then analyzed by ChIP to determine the amount of p53 that was bound to the indicated promoters. B, knockdown of endogenous Ski augments p53 occupancy on the promoters of p53 target genes. MCF7 cells were transfected with a control siRNA or Ski siRNA for 48 h and then treated with 5 nm ActD for 4 h. ChIP was performed using an anti-p53 antibody, and quantitative real-time PCR was performed for the indicated promoters. C, Ski suppresses the levels of acetylated p53 after DNA damage. MCF7 cells and MCF7 cells stably expressing Ski were treated with 2 nm ActD for the indicated periods. The levels of acetylated p53 were determined by immunoprecipitation with an anti-acetylated p53 antibody (IP: α-Acp53) and then Western blotting with an anti-p53 antibody. D, knockdown of endogenous Ski increases p53 acetylation after DNA damage. MCF7 cells were transfected with a control siRNA or Ski siRNA. Forty-eight hours after transfection, the cells were treated with 2 nm ActD for the indicated time periods. The levels of acetylated p53 were determined as described in C.