Fig. 5: BBM induces upregulation of BNIP3 and the interaction of SNAP29 and BNIP3.

MCF-7 cells were treated with various concentrations of BBM for 24 h. a The expression of BNIP3 was determined by western blot. b The mRNA levels of BNIP3 were determined by qRT-PCR in three independent experiments (mean ± SD, *P < 0.05, **P < 0.01 compared with control). c MCF-7 cells were treated with BBM (5 μM) or Rapa (0.25 μM) for 24 h. After immunostaining with BNIP3 (green), the colocalization of MitoTracker (Deep Red FM, red) and BNIP3 was examined by confocal microscopy. Scale bars: 10 μm. d MCF-7 cells were treated with BBM (5 µM) or Rapa (0.25 µM) for 24 h, whole-cell lysate was prepared and subjected to immunoprecipitation using anti-BNIP3, and the associated SNAP29 and VAMP8 were determined using immunoblotting. e Confocal microscopy images of MCF-7 cells immunostained for BNIP3 (green) and SNAP29 (red) after treatment with BBM (5 μM) or Rapa (0.25 µM) for 24 h. The colocalization puncta was indicated by arrowheads. Scale bars: 10 μm. f Confocal microscopy images of MCF-7 cells immunostained for BNIP3 (green) and VAMP8 (red) after treatment with BBM (5 μM) or Rapa (0.25 µM) for 24 h. Scale bars:10 μm