HSP90 Inhibitors, Geldanamycin and Radicicol, Enhance Fisetin-Induced Cytotoxicity via Induction of Apoptosis in Human Colonic Cancer Cells
Figure 5
Decreased p53 and its stability, and increased ubiquitination in fisetin (FIS)+geldanamycin (GA)- or FIS+radicicol (RAD)-treated COLO205 cells. (a) Reduction of the p53 protein level and induction of ubiquitin-tagged proteins (Ub) in FIS+GA- or FIS+RAD-treated COLO205 cells. Cells were treated with FIS (60 μM), GA (2 μM), RAD (5 μM), or their combinations for 4 h, and expressions of p53 and Ub-tagged proteins were detected by Western blotting. Intensities of (Ub)n and the p53 protein were quantified, and the ratio of p53/α-TUB and (Ub)n/α-TUB in the respective control group was described as 1. (b) GA and RAD decreased p53 protein stability in FIS-treated COLO205 cells. Cells were treated with cycloheximide (CHX; 1 μg/mL) for 1 h, followed by the addition of FIS (FIS/CHX), FIS+GA (FIS+GA/CHX), FIS+RAD (FIS+RAD/CHX), GA (GA/CHX), or RAD (RAD/CHX) for different times (0, 0.5, 1, 2, 4, and 8 h). At the various time points, expressions of p53 and the α-tubulin protein were detected by western blotting using specific antibodies. Intensities of (Ub)n and the p53 protein were quantified, and the ratio of p53/α-TUB in the respective control group was described as 1. Data were repeated at least three times, and similar results were obtained.
