FIG.4.
Overexpression of ACTR selectively induces the expression of E2F1-responsive genes, but not ER target genes. (A) The expression of cell cycle regulatory proteins in T-47D cells was analyzed by Western blotting. Cells were treated with ICI and infected with adeno-ACTR (labeled with an A) or adeno-GFP (labeled with a G) as described in the legend for Fig. 3A, and at different hours after infection cells were harvested for Western blotting. Cells were refed every 2 days with fresh medium containing 10−8 M ICI. T-47D cells treated with 10−7 M E2 for 48 h or continuously treated with ICI were included for comparison. (B, panels a to g) Quantitative RT-PCR analysis of E2F-responsive gene expression in T-47D cells that were treated and/or infected with adeno-ACTR and adeno-GFP as described for panel A. Details of the procedures and calculation of difference are described in Materials and Methods. (h) RT-PCR analysis (28 cycles at a Tm of 55°C) of E2F1 target genes that did not show any significant changes of expression due to elevated levels of ACTR. E2F1 is shown for comparison.