Figure 4.

E2F1 activates hPTTG1 transcription. E2F1 activates hPTTG1 transcriptional activity in H1299 (A) and Hela cells (C) but not in MCF7 (D) and HCT116 cells (E). Cells (each well of a 24-well plate) were cotransfected with 200 ng pGL_3-Basic or either of hPTTG1 promoter −2642/−1 or −1272/−1, together with 800 ng empty vector pcDNA_3.1 (hatched bar), 400 ng E2F1 and 400 ng pcDNA_3.1 (dotted bar), 400 ng E2F1 and 400 ng DP1 (black bar), or 400 ng E2F1 and 400 ng E2F1-DN (white bar). B, Dose dependency of E2F1-induced hPTTG1 transcriptional activity in H1299 cells. H1299 cells were cotransfected with 200 ng hPTTG1 promoter −2642/−1 (dotted bar) or −1272/−1 (black bar), together with increasing amounts of E2F1 and DP1 (each from 100–500 ng). Empty vector pcDNA_3.1 was used to adjust equivalent amounts of DNA for transfection. Results are plotted as arbitrary units with controls (transfected with empty vector pcDNA_3.1) set as 1. F, Deleted p53 in HCT116 cells abolished p53 suppression of E2F1-induced hPTTG1 elevation. Relative activities were normalized by empty vector pGL₃-Basic as background. Five nanograms of pRL-TK plasmid were cotransfected to normalize transfection efficiency. Assays were performed 24 h after transfection. Transfections were performed in triplicate for each experiment and repeated four times.