Figure 2

Bortezomib downregulated CIP2A, p-Akt and induced cleavage of caspase 3 in a dose- and time-dependent manner. A, Dose-dependent analysis of CIP2A, p-Akt and caspase-3. Cells were exposed to bortezomib at the indicated doses for 36 hours. Cell lysates were prepared and assayed for these molecules by western blotting. Data are representative of three independent experiments. CF, cleaved form (activated form). Note that PARP cleavage, instead of caspase-3, was used for assessment of apoptosis in MCF-7 cells since previous studies have shown that MCF-7 cells do not express caspase-3 due to the functional deletion of the CASP-3 gene [34–36]. Lower right, relative expression levels of CIP2A, p-Akt and Akt among all the cell lines tested are also shown. B, Ratio of CIP2A to actin levels from the cell lysates in (A). Immunoblots were scanned by a UVP BioSpectrum AC image system and quantitated using VisionWork LS software. Columns, mean (n = 3); bars, SD. C, Time-dependent analysis of CIP2A, p-Akt and caspase-3. Cells were exposed to bortezomib at 500 nM for the indicated time points. Data are representative of three independent experiments. CIP2A, cancerous inhibitor of PP2A; PARP, poly ADP-ribose polymerase; SD, standard deviation.