Figure 6.

Detection of S-nitrosylated GAPDH, ERp72 and ERp5 and evaluation of ER stress. HepG2 cells were treated with thapsigargin (5 μM) for 12 h, and lysates prepared from the treated cells were analyzed by SNO-RAC assay, in which ascorbate-dependent capture of proteins by resin indicates the presence of S-nitrosylated Cys residues. Elutants were subjected to immunoblot analysis using anti-GAPDH (A), anti-ERp72 (B) and anti-ERp5 antibodies (C). (D) Lysates prepared from LPS-treated liver tissues of wild-type and GSNOR^−/− mice were subjected to immunoblot analysis using anti-Xbp-1 antibodies. Anti-β-actin antibody was used as a control to ensure that equal amount protein was loaded in each lane. (E) Summary of proteins whose expression levels were evaluated by immunoblot analysis. Proteins enclosed within the solid line are members of the PDI family, whereas proteins enclosed within the dotted lines are members of the heat shock protein (HSP) family. *S-nitrosylated protein.