Figure 3
Effect of nutlin-3 treatment on Bax relocalization and activation in EBV (−) and EBV (+) cells. (a) Cells were treated with 10 μM nutlin-3 or the solvent dimethyl sulfoxide for 24 h. Cytosolic and mitochondrial fractions were analyzed by western blotting with an anti-Bax pAb. Vinculin and Bcl-2 were used as cytosolic and mitochondrial markers, respectively. Fold-change values versus the untreated control (0 h), after normalization with respect to vinculin or Bcl-2 protein levels, are shown under the blots. Results are representative of three independent experiments. (b) BL2 EBV (−) cells and BL2/B95, LY47 and Remb1 EBV (+) cells were treated with 10 μM nutlin-3 or the solvent dimethyl sulfoxide for 24 h. Cells were fixed in 0.25% paraformaldehyde and labeled with a conformation-specific 6A7 Bax antibody, which recognizes only the active conformation of the protein and Alexa 488-conjugated goat anti-mouse IgG. Cells were then analyzed with a FACSCalibur flow cytometer
