Fig. 5. Cell-cycle analysis of Ku80-knockdown cells. (A) Asynchronous cultures of control, scr-siRNA-transfected and Ku80-siRNA-transfected HeLa cells were harvested after 96 hours of Ku80 silencing and stained with propidium iodide for monitoring of their cell-cycle distribution. Representative flow cytometric analysis is shown (top). (Bottom) The table shows the means (n=3) of the quantification of the percentage of cells present in each phase of the cell cycle. In brackets, 1 s.d. of three individual experiments is shown. (B) Western blot analysis of the cyclin A, cyclin D1 and cyclin E protein levels in whole cell extracts prepared from untreated, control, scr-siRNA-transfected and Ku80-siRNA-transfected HeLa cells. (C) Quantification of the relative optical densities of cyclin A, cyclin D1 and cyclin E. Values are expressed as the average of three experiments ± 1 s.d. (D) Schematic presentation of the cyclin/Cdk cycle and the various checkpoints. Silencing of Ku80 blocks the cells at the G1-S transition point, as evidenced by high levels of cyclin E (dark-orange arrow).