Figure 6
Effects of stable silencing of endogenous Egr-1 by siRNA on CPZ-induced p21Waf1/Cip1 expression. (A) Knockdown of endogenous Egr-1 expression was verified in stable transfectants of C6 cells expressing Egr-1 siRNA (C6/siEgr1), as compared with control cells (C6/vec). (B) A cell proliferation assay was performed in C6/vec and C6/siEgr1 cells as in Figure 1B. The data shown represent the mean ± SD for one experiment performed in triplicate. Similar results were obtained from two other independent experiments. (C) C6/vec and C6/siEgr1 cells were left untreated or treated with 20 µM CPZ for the indicated time periods. Cells were collected and subjected to Western blot analysis for p21Waf1/Cip1. The same blot was re-probed with anti-ERK2 antibody as an internal control. Each blot represents at least three individual experiments. (D) Exponentially growing C6/vec and C6/siEgr-1 cells were treated with 20 µM CPZ. After 24 h, the cells were harvested and the DNA content was analyzed using FACS analysis. The x-axis represents DNA content. The y-axis represents the number of cells. The percentages of the cell population at each phase of the cell cycle are indicated in each histogram. Similar results were obtained from two other independent experiments.
