Fig. 3.

Infection of HepG2 cells with siRNA adenoviruses against NCOA6 (A) leads to a decrease in FXR-mediated transactivation of BSEP promoter while NCOA6 overexpression (B) using a plasmid encoded cDNA results in a dose-dependent increase in promoter activity. A: GFP-containing adenoviruses encoding siRNAs to fragments of NCOA6 along with a scrambled control siRNA were prepared and infection of HepG2 cells were performed as described by us earlier (10). Infection was monitored by GFP fluorescence that was 70–80%. Farnesoid x receptor (FXR) transactivation of BSEP promoter was assayed by luciferase activity. NC III and NC IV siRNA significantly (P < 0.05, compared scrambled siRNA adenovirus treatment) downregulated promoter activity while NC II did not. A representative Western blot of the protein levels is shown. B: overexpression of NCOA6 was achieved by transfection with increasing amounts of full-length cDNA plasmid and FXR transactivation of BSEP promoter was monitored by luciferase activity. Significant (P < 0.05, compared with activity in cells without NCOA6 plasmid cotransfection) stimulation of promoter activity was seen with 1 and 2 μg of NCOA6 cDNA.