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. 2005 Nov;25(21):9369–9382. doi: 10.1128/MCB.25.21.9369-9382.2005

FIG. 2.

FIG. 2.

Knockdown of Bif-1 inhibits Bax translocation to mitochondria, cytochrome c release, caspase 3 activation, and PARP cleavage. (A) HeLa cells transfected with siGFP or siBif-1 were pretreated with 75 μM z-VAD-fmk for 30 min before incubation with 20 μM ActD for 4 h. The cells were stained with anti-cytochrome c (anti-cyt c) monoclonal antibody (red) plus anti-Bax polyclonal antiserum (green), and cells with released cytochrome c (indicated by red arrows) and mitochondrion-translocated Bax (indicated by green arrowheads) were counted (n > 500) under fluorescence microscopy and shown in bar graphs. (B) The siRNA-transfected HeLa cells were exposed to either DMSO as a control, 1 μM ADR for 12 h, or 0.5 μM STS for 6 h, and the caspase 3-like activity was measured by using the caspase 3 fluorometric assay kit (Sigma) according to the manufacturer's protocol. (C) HeLa cells with siGFP or siBif-1 were treated with 20 μM ActD for the indicated periods and subjected to SDS-PAGE/immunoblot analysis with anti-PARP antibody.