Figure 7.

Modulation of E2F1 transcriptional activity by EPC1. (A) Regulation of E2F1 transcriptional activity by EPC1. SK-Mel-29 cells were transfected with 0.5 μg Bcl-2-luc (left) or BIRC5-luc reporter plasmids (right) together with expression vectors for E2F1 (0.5 μg) in the absence or presence of EPC1 expression vector (1, 2, 3 μg). E2F1 and EPC1 expression was confirmed by Western blot. Actin was used as loading control. (B) SK-Mel-29 cells were cotransfected with the p27-luc reporter plasmid (0.5 μg) and expression plasmids for E2F1 (0.5 μg) in the absence or presence of EPC1 expression vectors (3 μg). E2F1, EPC1 and actin expression was confirmed by Western blot. (C and D) H1299 cells were transfected with Bcl-2-luc (C) or p27-luc reporter plasmids (D) together with expression vectors for Flag-tagged full-length E2F1 or Flag-tagged E2F1 mutant in the presence or absence of EPC1 expression vector. Luciferase assays were performed after 24 h using pcDNA plasmid as control that was set to 1. Data are presented as the mean with standard deviation of technical replicates (duplicates or triplicates) in an experiment representative of several independent others. (E) Schematic representation of full-length and truncated E2F1 proteins. Western blot shows expression of Flag-tagged full-length E2F1 or Flag-tagged E2F1 mutant and EPC1 in H1299 cells. Actin was used as loading control. Statistical significance was determined by two-sided Student's t-test. *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001.