Figure 6.
Proteasome inhibitors block the decrease of Mcl-1, the translocation of Bcl-xL, and other apoptotic events after UV treatment. (A) Mitochondria and S100 were fractionated from HeLa cells treated with UV for different amounts of time and pretreated for 1 h with DMSO (lanes 1-4), 10 μM MG132 (lanes 5-8), or 10 μM epoxomicin (lanes 9-12). The levels of Mcl-1, Bcl-xL, Bax, and cytochrome c oxidase were measured in the mitochondria. The levels of Mcl-1, Bcl-xL, cytochrome c, and cleaved caspase-3 were measured in the cytosol. (B) Six 6-well dishes of HeLa cells were transfected with Luciferase or Mcl-1 siRNA as described in Materials and Methods. For each sample, two dishes were pretreated (30 min) 24 h later with DMSO (lanes 1,2,4,5) or 10 μM MG132 (lanes 3,6). Cells were either left untreated (lanes 1,4) or UV-treated (lanes 2,3,5,6) and harvested 4 h later. A small aliquot of the cell pellet was used to make whole cell lysate in 0.5% CHAPS for the caspase assay. The remaining cells were fractionated for mitochondria and S100. The levels of Mcl-1, Bcl-xL, Bax, and cytochrome c oxidase were measured in the mitochondria. The levels of Mcl-1, Bcl-xL, Bax, and cytochrome c were measured in the cytosol.