Fig. 1

COASY knockdown induced multinucleation and prolonged mitosis. a, b COASY knockdown by siRNA in MDA-MB231 triggered cobble-stone morphological change (a) and multinucleation (b). MDA-MB-231 cells transfected with control or COASY siRNA for 72 h were stained with DAPI (nuclei) and Alexa Fluor 488 phalloidin (F-actin). Scale bars, 50 mm (a), 20 mm (b). c The percentage of multi-nucleated A549 cells after transfection with control or two independent COASY siRNAs for 72 h. For each sample, more than 150 cells were examined by immunofluorescence microscopy. d Live cell time-lapse imaging showed that COASY knockdown extended mitosis and induced cytokinesis failure. A549 cells expressing histone 2B (H2B)-mCherry (nucleus marker) were transfected with control or COASY siRNA for 24 h before live cell imaging. Scale bars, 10 mm. e COASY knockdown increased the time in mitosis as determined by the time of nuclear envelope breakdown (NEBD) to anaphase onset (in min). For each sample, more than 35 cells were examined by live cell imaging. f COASY knockdown increased percentage of cytokinesis failure during mitosis. For each sample, more than 35 cells were examined by live cell imaging. g Western blots show that COASY knockdown extend the time of mitosis by using cyclin B1 expression as a mitotic marker. A549 cells were synchronized by thymidine-nocodazole block and released in fresh media. The samples were then harvested every 20 min and analyzed by western blots. Bars show standard error of the mean. **p < 0.01, ***p < 0.001, two-tailed Student’s t-test, n = 3 independent repeats