Effect of abolishing Hsp90β expression by RNAi on the antiapoptotic activity of Bcl-2. (A) After transduction with RNAi (96 hours), RNA was extracted from BMMCs, and mRNA levels of Hsp90α and Hsp90β were tested by PCR using specific primers, as described in “Materials and methods.” Results of 1 of 3 representative experiments are shown. (B) After transduction with RNAi (96 hours), whole-cell proteins from BMMCs were extracted, and the level of Hsp90 was tested by Western blotting using monoclonal antibody against Hsp90, as described in “Materials and methods.” Results of 1 of 3 representative experiments are shown. (C) After transduction (96 hours), the activity of caspases 3 and 7 was tested in RBL cells using a fluorometric assay, as described in “Materials and methods.” Data are represented as mean ± SE (n = 3). Results of 1 of 3 representative experiments are shown. (D) Effect of RNAi on BMMCs discriminated by annexin V–FITC and PI double stain. Seventy-two hours after lentiviral transduction, BMMCs were double stained with annexin V–FITC and PI and were analyzed by FACS, as described in “Materials and methods.” Representative dot plots of annexin V–FITC and PI staining are shown. The bottom left quadrant in each dot plot contains the vital (double-negative) population. The bottom right quadrant in each dot plot contains the early apoptotic (annexin V⁺, PI^-) population.