Figure 7.

 IP₃R-3 silencing abrogates cell death regulation induced by VDAC1. (a) HeLa cells were transfected for 72 h with control or different shRNAs against IP₃R-1 or IP₃R-3. Cells were harvested, total protein was extracted and subjected to western blotting analysis with specific antibodies anti-IP₃R-1/3 and anti-β-Tubulin as loading control. shRNA-IP₃R-1#58 and IP₃R-3#67 were selected for subsequent experiments. (b, c and d) Cells were co-transfected with one construct containing both the shRNAs for control (b, shScrambled), IP₃R-1 (c, shIP₃R-1), or IP₃R-3 (d, shIP₃R-3) and a fluorescent marker (GFP), together with the siRNA of interest. The graph bar shows the change in percentage of fluorescent cells before treatment with 100 μM H₂O₂ for 2 h (b, Control −7.5±2.5% siRNA-hVDAC1 27.4±1.5% siRNA-hVDAC2 −45.9±3.5% c, Control −6.5±2.1% siRNA-hVDAC1 +30.9±3.9% siRNA-hVDAC2 −45.2±2.3% d, Control −4±5.3% siRNA-hVDAC1 +24.3±5.1% siRNA-hVDAC2 −50.1±6.8%)