Fig. 5. Analysis of PLK1 expression in response to p53 signaling. The levels of PLK1 protein and mRNA were measured using western and northern blots, respectively. To assess loading, northern and western blots were stripped and reprobed for the expression of gapdh (northern) or actin (western). (A) Expression of plk1 and p21/waf1 mRNAs in response to the overexpression of p53. TR9-7 cells containing a tetracycline-regulated p53 were incubated in the presence (p53 off) or absence (p53 on) of tetracycline. (B) The effects of p53 induced by DNA damage or high levels of p14ARF. NARF2 cells containing IPTG-inducible p14ARF were incubated in the presence of IPTG for 24 hours or in the presence of adriamycin (345 nM) for 24 hours without adding IPTG. (C) The role of p53 and p21/WAF1 in the downregulation of the PLK1 protein in response to DNA damage. HT1080 cells with (GSE) and without (WT) the dominant-negative p53 fragment GSE56 were analyzed, as well as HCT116 cells in which p21/waf1 was deleted and the corresponding parental cells. The cells were exposed to adriamycin (345 nM), and the total protein was extracted and analyzed by western blot.