Fig. 2.

hGwl knockdown promotes heterogeneity of phenotypes of varying severity in human cells. (A) HeLa cells stably expressing EB3-GFP (green) and H2B-CherryFP (red) were transfected with 50 nM of Scramble (Control) or hGwl siRNA for 24 h. After transfection, cells were synchronized with thymidine block. Movies were started 5 h after release and images were taken every 15 min. Notations indicate, time of the frame in minutes (white), lagging chromosomes (yellow arrows), metaphase plate orientation (dotted white lines), and cell death (yellow “d”). (Scale bars, 5 μm.) (B) Mitotic HeLa cells were treated with scramble or hGwl siRNA and stained with anti-Aurora B or anti-BubR1 (red) antibodies and with DAPI (DNA, blue). Shown are the deconvoluted maximum projections from 0.3-μm Z- sections. (Scale bars, 5 μm.) (C) Cells from A were separated according to their mitotic phenotype and plotted against mitotic length. Mitotic entry was determined by analyzing the first signs of DNA condensation cross-reinforced with cell rounding. Mitotic exit was scored based on chromosome segregation at anaphase or DNA decondensation. Shown are box-plots with 5 to 95% confidence intervals (black dots indicate outliers). Two-tailed unpaired Student t tests were performed to determine statistical relevance; significant P values are shown.