Figure 1
Regnase-2 inhibits proliferation, and stalls the cell cycle at the G1 phase. (a) U87-MG cells were treated with dox (1 μg/mL) or left untreated and counted 24, 48, and 72 h after the addition of dox. The levels of Reg-2 and Reg-2 D/A were examined using western blotting. The graph shows results from three independent experiments + / − SD. (b) U87-luc, Reg-2, or Reg-2D/A cells were labeled using CellTrace Far Red reagent and divided into two dishes. One dish was induced with doxycycline (1 μg/ml), the other was left untreated. After 48 h of induction, the fluorescence was measured by flow cytometry. The graphs show histogram overlays of noninduced (gray) and induced (white) cells. Representative results of three independent experiments are shown. (c) U87-MG cells were plated on 6-well plates (100 cells/well), and treated with dox (1 μg/ml) daily until visible colonies were formed (10 days). Then the colonies were fixed and stained. Graph presents the ratio of the surface covered by colonies formed by dox-induced to non-induced cells normalized to control cells (luc) (mean value of three independent experiments ± SD) (d) U87-luc/Reg-2/Reg-2D/A cells were treated with dox (1 µg/mL) for 48 h, and cell cycle analysis was performed. The graphs show a histogram of propidium iodide fluorescence in particular cell lines. Representative results of three independent experiments are shown. The graph and table show the percentages of cells in each cell cycle phase. The data were analyzed using two-way ANOVA and Bonferroni's post-hoc test (***p < 0.001; *p < 0.05).
