Fig. 4

Stabilization of p53 by inhibition of MDM2 expression.

(A) Inhibition of MDM2 expression results in an increase of p53 level. Cells were treated with 200 nM of AS2 or control oligonucleotide K for 20 hr. p53 protein levels were detected by Western blot with antibody DO-1. Identical amounts of total protein were loaded on each lane. The double band in MCF-7 is due to a p53 polymorphic allele. (B) Determination of p53 half-life. SJSA cells were treated with 200 nM AS2 for 20 hr and the rate of p53 degradation was determined by a pulse-chase experiment. The half-life of p53 was ∼0.5 hr in untreated SJSA cells and >4 hr in AS2 treated cells as determined by densitometric analysis. The total CPM used for untreated samples was 2-fold higher than for AS2-treated samples, thus the p53 band in 0 hr untreated sample is darker and does not indicate a different rate of synthesis.