Figure 7

GSH attenuates the repression and nitrosylation of FOXA2 by PCN. (A-B) GSH treatment overcomes PCN-mediated inhibition of FOXA2 expression. NCI-H292 cells were pretreated with various concentrations of GSH for 60 min before exposure to PCN (6.25 μg/ml). Nuclear proteins were harvested for Western blot using anti-FOXA2 (A) and quantified with densitometry (B). The same membrane was stripped and probed with antibody nuclear protein Histone H3 for loading control. (C-D) GSH reduces nitrosylation of FOXA2 by PCN. NCI-H292 cells were pretreated with PBS (negative control) or with GSH for 60 min before exposing to PCN. FOXA2 was immunoprecipitated from nuclear extracts, and equal amounts were loaded for western blot analyses using anti-FOXA2 and anti-nitrotyrosine antibodies, respectively (C). FOXA2 nitrosylation was quantified by densitometry (D). The experiments were independently performed three times with similar results. Western blots from one typical experiment are shown. Densitometry data represent the mean ± SD from all three experiments. *p < 0.05 when comparing GSH-pretreated cells against cells exposed only to PCN.