FIG. 4.
E1A overcomes G1 arrest induced in p21+/+ or p21+/− cells after doxorubicin treatment. p21+/+ and p21+/− cells were transfected in parallel with pCMV-E1A12S or the control plasmid pCMV, and immediately after, doxorubicin was added to the cultures. At 36 h posttransfection, the cells were collected and stained with propidium iodide and anti-E1A for DNA content and E1A expression, respectively. The cell cycle distribution of these cells was assessed by flow cytometry, as described in Materials and Methods. The gates were established with cells containing empty vector (pCMV), thereby allowing the analysis of only E1A-expressing cells. Each plot represents the analysis of 20,000 gated events. The data, from at least three different experiments, are presented in histograms which show the percentage of cells with G1 (2 N), S, G2/M (4N), and >4N DNA content, respectively. Note the presence of a sizeable population of doxorubicin-treated cells with greater-than-4N DNA content in the p21+/− cells transfected with E1A. The error bars indicate standard deviations.