Figure 3

Increased apoptosis sensitivity to death ligands by silencing Siah2/POSH. a: Phase contrast micrographs of PPC-1 cells transfected with siRNA specific for Siah2 or a non-targeting control and treated +/- TRAIL (100 ng/mL) for 10 hours. b: Quantification of cell viability in PPC-1 cells treated as described (A) for 24 hours using an MTT assay. c: Apoptosis analysis. PPC-1 cells were transfected with siRNA against Siah2 or non-targeting control for 72 hours. Cells were left untreated or treated with 100 ng/mL TRAIL for 16 hours and subjected to annexin V staining. Cells in lower left quadrant are viable, annexin V negative; lower right, viable annexin V positive; upper right, dead annexin V positive. The percentage of viable, annexin V positive cells is depicted in lower right quadrant. d: Quantification of viable annexin V positive cells in three independent experiments. e: Siah2 or POSH was silenced in PC-3 cells and either treated with TRAIL for 16 hours (right) or transfected with Fas ligand or a LacZ (control) expression construct for 24 hours (left). Annexin V staining was performed and the percentage of viable, annexin V positive cells quantified. All statistical analysis was carried out using a Student's t-test. **P < 0.01, ***P < 0.001.