FIG. 2.

p53 regulates the expression of igf2 and c-jun in hAFS cells. (A) hAFS cells were transfected in duplicates with a siRNA targeted against p53 (hAFSp53sir) or with control siRNA (hAFSCsir). Seventy-two hours after transfection, cells were harvested. From one of the duplicates, RNA was prepared, transcribed into cDNA, and analyzed by qRT-PCR. Abundance of specific cDNAs was normalized by determining the abundance of the housekeeping gene β-actin. Relative abundance of the specific RNA in the cells with control siRNA was set to one. The graph shows mean values and standard deviations of four independent experiments (**P < 0.02; ***P < 0.005). The second duplicate of the cells was used to monitor p53 abundance by western blotting. (B) hAFS cells (with a female karyotype) were transfected with a plasmid encoding p53 (hAFSp53) or with vector DNA (hAFSVector). The relative abundance of igf2 mRNA was monitored by qRT-PCR as described in the legend to part A. The graph shows mean values and experimental variation of two independent experiments. A second duplicate was used to monitor p53 abundance by western blotting. (C) H1299 cells were transfected with a plasmid encoding wild-type p53 (H1299wtp53) or with vector DNA (H1299Vector). The relative amount of igf2 mRNA was monitored by qRT-PCR as described in the legend to part A. The graph shows mean values and experimental variation of two independent experiments. A second duplicate was used to monitor p53 abundance by western blotting. igf2, insulin-like growth factor 2 gene; qRT-PCR, quantitative real time-polymerase chain reaction.