Higher expression of HBZ partially suppressed activation of TGF-β signaling via AP-1. (A) HepG2 cells were cotransfected with 3TP-Lux (0.5 μg), phRL-TK (2 ng), pCDNA3-c-Fos (0, 0.1 μg), FLAG-Smad3 (0, 0.1 μg), and pcDNA3.1-mycHis-sHBZ (0, 20, 200, and 4000 ng). At 24 hours after transfection, the cells were treated with or without TGF-β (10 ng/mL). After 24 hours, the cells were harvested, and luciferase activity was determined. Expression of sHBZ, Smad3, and c-Fos was detected by Western blot (middle panel). CBB staining was shown as the loading control (bottom panel). (B) HepG2 cells were cotransfected with 9 × CAGA-Luc (0.5 μg), phRL-TK (2 ng), and pME18Sneo-sHBZ (0, 2, 5, 10, 20, 50, 100, and 200 ng). Luciferase activity was measured 24 hours after 10 ng/mL TGF-β stimulation. (C) sHBZ inhibited AP-1 signaling via its bZIP domain. Jurkat cells were cotransfected with AP-1-Luc (1 μg), phRL-TK (10 ng), pCG-Tax (1 μg), and pME18Sneo-sHBZ mutants (1 μg). After 48 hours, luciferase activity was measured.