Fig. 4: Effect of KCNN1 knockdown on ES cell proliferation.

A KCNN1 expression in A-673 cell line stably transduced with doxycycline-inducible shRNA control cells (shRNA CT) or 3 shRNA cells targeting KCNN1 (shRNA 511, shRNA 908 and shRNA 576) after 72 h of treatment with doxycycline (0.5 µg/mL). Bars indicate mean ± SD of relative and normalized KCNN1 mRNA expression obtained after RT-qPCR (N = 5 for shRNA CT and shRNA 908, N = 4 for shRNA 511 and N = 3 for shRNA 576). B Clonogenicity assays of A-673 cell line stably transduced with shRNA control (shRNA CT) or three shRNA targeting KCNN1 cells (shRNA 511, shRNA 908 and shRNA 576) after 72 h of doxycycline treatment (0.5 µg/mL), with the colonies numbers observed and the relative surface area of clones. Bars indicate mean ± SD of counted colonies or the clones surface area observed on the left panel relative to the untreated cells (N = 6 for shRNA CT, N = 3 for shRNA 511 and shRNA 908 and N = 7 for shRNA 908). C Cell cycle assays on shRNA 908 cells treated (shRNA 908 +Doxycycline) or not (shRNA 908 −Doxycycline) with doxycycline 0, 6, 8 or 10 h after thymidine double block. Dark blue: G0/G1 phase, Light blue: S phase, Purple: G2/M phase. D Percentage of cell distribution 6, 8, and 10 h after thymidine double block (N = 5 for 6 and 10 h after thymidine double block; N = 6 for 8 h after thymidine double block, Wilcoxon test). E Western blots of p21 and Aurora A and their quantification normalized to β-tubuline 10 h after the thymidine double block (N = 3). *p < 0.05; **p < 0.01 (A, B, D, E: Mann–Whitney test).