Fig. 1. Cdk5 regulates expression of the p53 tumor suppressor protein in response to genotoxic and oxidative stress. (A) SH-SY5Y cells were treated with 5 µM mitomycin C or 2 mM SNP for the indicated times, and cell lysates were subjected to immunoblotting with the indicated antibodies. (B) SH-SY5Y cells were transfected with Cdk5 targeting or scrambled control siRNA, treated with 5 µM mitomycin C or 2 mM SNP for 9 hours, and then subjected to immunoblotting with the indicated antibodies. (C) SH-SY5Y cells were pretreated with 10 µM roscovitine or Cdk2/5 inhibitor for 30 minutes and then incubated with 5 µM mitomycin C or 2 mM SNP for 9 hours. Cell lysates were subjected to immunoblot analysis with the indicated antibodies. (D) SH-SY5Y cells were treated with 5 µM mitomycin C for 6 hours and then incubated with 60 µg/ml cycloheximide for indicated times. Cdk2/5 inhibitor was added 30 minutes before the incubation with cycloheximide. (i) The amounts of p53 and GAPDH were determined by immunoblotting. (ii) Protein levels were quantified. (E) SH-SY5Y cells were treated as described in C for the indicated times, total RNA was isolated and quantitative real-time RT-PCR was performed. The results are from three independent experiments and are presented as the mean ± s.d. Each experiment was performed in triplicate.