FIG 4.

CHOP is not essential for activation of cytoprotective responses and maintaining cellular viability. (A and B) Transcript levels of CHOP and Hsp60 were determined by qRT-PCR of cells transfected with 20 nM siRNA against CHOP or 20 nM control siRNA against luciferase, followed by transfection with either GFP or Endo G-GFP. (C) Endogenous levels of FOXO3A, lipidated LC3B, SirT3, and MnSOD were evaluated by Western blotting of the crude extract of cells treated as described above for panel A. (D) Transcript levels of endogenous FOXO3A, SirT3, and MnSOD were measured by qRT-PCR of cells treated with 20 nM siRNA against CHOP, followed by transfection with either GFP or Endo G-GFP. (E) Mitochondrial superoxide anion levels were determined by FACS analysis of cells transfected as described above for panel A and stained with MitoSox Red. (F) Viability of MDA-MB231 cells, transfected as described above for panel A, was determined by trypan blue staining at 48 h.