Post-transcriptional inactivation of p53 in immortalized murine embryo fibroblast cells

The steady-state levels of p53 mRNA and protein were barely detectable by N orthern and Western blot analysis in spontaneously immortalized (10)3 and ( 10)7 murine embryo fibroblast (MEF) cells, But when cells were treated with cycloheximide (CHX) or emetine, expression levels were restored to those o bserved in primary and immortal (10)10 MEF cells. However, levels of p53 mR NA were not changed in primary or (10)10 MEF cells by CHX treatment. De nov o p53 mRNA synthetic rates were similar in primary, (10)10, (10)3, and (10) 7 MEF cells treated with or without CHX. Treatment with actinomycin D (ActD ) showed that p53 mRNA in primary and (10)10 MEF cells had a relatively lon g half-life of 22 h, compared to less than 2 h for (10)3 and (10)7 MEF cell s. Pulse-chase analysis of p53 mRNA turnover using CHX and ActD showed that the rapid destabilization of p53 mRNA in (10)3 and (10)7 MEF cells could b e regulated at the transcriptional and translational levels. In addition, t he destabilization of p53 mRNA appeared to occur in the nucleus for (10)3 a nd (10)7 cells, but not for primary and (10)10 MEF cells. Taken together, t he present study demonstrates: that inactivation of the p53 gene occurs at the posttranscriptional level by rapid destabilization of its mRNA in the n ucleus of spontaneously immortalized (10)3 and (10)7 MEF cells.