FIGURE 1.

Reconstitution of tBID-triggered BAKΔC permeabilizing function in MOM-like liposomes. A, rat liver mitochondria were incubated with the indicated recombinant apoptotic proteins for 20 min followed by enzyme-linked immunosorbent assay-based analysis of cytochrome c release. tBID, BIMLΔC, p53, histone 1.2 (H1.2), and gliotoxin (Gliot.) were added at 200 nm. The double asterisk denotes significant differences between experiments, as revealed by Student's t test (p < 0.01). All data points represent mean values and S.E. of three independent experiments (except p53, n = 2). B, ANTS/DPX-loaded MOM-like LUVs (100 μm) were treated with the indicated recombinant apoptotic proteins for 20 min followed by determination of the extents of vesicular contents release by spectrofluorimetry. tBID, BIMLΔC, p53, histone 1.2, and gliotoxin were added at 200 nm. Data represent the mean values and S.E. from 3–6 independent experiments. C, shown is the extent of ANTS/DPX release from MOM-like LUVs (100 μm) elicited by BAKΔC/BAKΔCG126S (200 nm) incubated for 20 min with increasing concentrations of indicated forms of tBID for 20 min. Data represent the mean values and S.E. of three independent experiments (except 3000 nm, n = 2). Experimental data were fitted with a sigmoidal dose-response nonlinear regression model. D, shown is concentration dependence of the inhibitory effect elicited by BCL-2ΔC, BCL-X_LΔC, and MCL-1ΔC on the vesicular ANTS/DPX release induced by 200 nm BAKΔC plus 100 nm tBID. LUVs (100 μm) were first treated with antiapoptotic proteins for 5 min followed by incubation of the mixture with proapoptotic proteins for 20 min, and finally, extents of vesicular contents release were determined. When indicated, experimental data were fitted with a sigmoidal dose-response nonlinear regression model. Data represent mean values and S.E. from at least two independent experiments.