Fig. 3.

Effects of N₃-AZA on GAPDH and GSTP1 protein levels, GAPDH localization and their enzymatic activity. (A) Lysates prepared from N₃-AZA (or 0.02% DMSO) treated normoxic and hypoxic FaDu cells were processed for western blotting. N₃-AZA treatment did not alter GAPDH and GSTP1 protein levels regardless of O₂ conditions. Representative immunoblots and quantitation [mean ± S.E.M.] from three independent experiments are displayed. (B) Cells treated with N₃-AZA (or 0.02% DMSO) under normoxia and hypoxia were processed for immunocytochemistry to monitor GAPDH localization; no change in cellular localization of GAPDH was observed in response to N₃-AZA treatment. The micrographs are representative of at least three independent experiments; scale bar = 20 μm. (C and D) FaDu cells treated with N₃-AZA (or 0.02% DMSO) under normoxia and hypoxia were processed for GAPDH activity assay (C) or GST activity assay (D); the enzymatic activities of GAPDH and GST were significantly reduced only in N₃-AZA treated hypoxic cells. Data represent mean ± S.E.M. from three independent experiments.