FIGURE 8.
Inhibition of Cdk1 during mitotic arrest in HT29 cells reverses effects on Mcl-1 and Bcl-xL in concert with decreased mitotic death. A, HT29 cells were synchronized by single thymidine block for 16 h, released for 4.5 h, untreated or treated with 0.1 μm Taxol (Tax) for 8 h as indicated, and then co-treated with vehicle or 10 μm RO3306 (RO) or 10 μm aminopurvalanol A (PA) for an additional 16 h, as indicated. Cells were harvested and subjected to immunoblotting with MPM-2 antibody or for the proteins indicated. B, HT29 cells were synchronized, untreated or treated with 0.1 μm Taxol for 8 h, and then co-treated with vehicle or 10 μm RO3306 or 10 μm aminopurvalanol A for an additional 16 h (24 h after Taxol), and cell death was analyzed by phycoerythrin-annexin V staining, as described under “Experimental Procedures.” *** = p < 0.001 (Student's t test). C, HT29 cells were synchronized, treated with 0.1 μm Taxol for 8 h, then co-treated with 10 μm RO3306 for an additional 40 h, and monitored by time-lapse microscopy. Fate profiles of 50 cells are shown on the right and compared with fate profiles for synchronized HT29 cells treated with 0.1 μm Taxol 4.5 h after release and monitored for 72 h (Fig. 1A). D, graphical representation of data from panel C, showing proportions of cells undergoing different fates in response to Taxol alone versus Taxol plus RO3306 co-treatment, which was highly significantly different (p < 0.0001, Fisher's exact test).