Fig. 7. Cdk5 induces acetylation of p53 and its cooperation with p300. (A) SH-SY5Y cells were treated with 5 µM mitomycin C or 2 mM SNP for the indicated times. The amounts of acetyl-p53 () and GAPDH were determined by immunoblotting. (B) SH-SY5Y cells were transfected with Cdk5-targeting or scrambled control siRNA, and then treated with 5 µM mitomycin C or 2 mM SNP for 9 hours in the presence of 20 µM MG132. Total proteins were subjected to western blotting using the indicated antibodies. (C) SH-SY5Y cells were transiently transfected with vectors encoding Flag-p53-WT or Flag-p53-SDM (a mutant p53 substituted with alanine at Ser15, Ser33 and Ser46), and then treated with 5 µM mitomycin C or 2 mM SNP for 9 hours. Total proteins were subjected to western blotting using the indicated antibodies. (D) SH-SY5Y cells were transfected with vectors encoding Flag-p53-WT or Flag-p53-SDM and then treated with 5 µM mitomycin C or vehicle for 6 hours. Cell lysates were immunoprecipitated with antibodies against Flag-p53, and membranes were immunoblotted with anti-p300 and/or anti-Flag, with 5% of the cell lysates used as input controls.