Fig. 5
CDC6 knockdown sensitizes CNE2-R cells to X-ray radiation. CNE2-R cells were transfected with CDC6 siRNA, and cells transfected with or without nontarget siRNA were used as controls. a Equal numbers of cells were exposed to 0, 2, 4, 6, and 8 Gy X-ray radiation. The living cells formed cell colonies after 3 weeks, and the radiobiological parameters D0, Dq, N, and sensitizing enhancement ratio (SER) were calculated using a single-hit multitarget model of cell survival. b The cell colonies were stained by crystal violet when the cells were treated with 6 Gy radiation or/and CDC6 siRNA. c CDC6 was depleted, CDC6 protein levels were assessed by western blot, and apoptotic cells were analyzed by Annexin V-FITC/PI apoptosis detection assay. d The cell cycle was analyzed. e The subpopulation at sub-G1 represents the apoptotic cells. f The cell apoptosis-associated proteins were assessed by western blot. g CDC6 knockdown enhanced the premature senescence of CNE2 cells. The senescent cells were inoculated with β-galactosidase staining solution at PH 6, and positive cells were counted and compared (100×). h The cell senescence-associated proteins were assessed by western blot. i The comparison of morphology of CNE2 and CNE2-R cells. j EMT-associated proteins including E-cadherin and Vimentin were assessed by western blot. k, l Cell migration abilities were assessed by scratch wound healing assay. m, n Cell invasion abilities were assessed by transwell assay. *P < 0.05, ** P < 0.01, *** P < 0.001
