A natural mutation of the hepatitis B virus X gene affects cell cycle progression and apoptosis in Huh7 cells

Abstract

Hepatocellular carcinoma (HCC), one of the most prevalent cancers, and HCC-associated diseases are caused by the hepatitis B virus (HBV). Viral sequences often contain mutations in the basal core promoter (BCP), which overlaps with the open reading frame encoding HBV X protein (HBx). HBx protein plays a very important role in the development of HCC. Influence of naturally-occurring mutation of HBx (BCP1) on cell cycle progression was investigated in the Huh7 HCC line. BCP1 mutation was cloned from clinical samples into recombinant green fluorescent protein-fusion expression plasmid constructs that were then transfected into Huh7 cells for either transient expression studies or isolation of stable transfectants, along with wild-type HBx and empty vector controls. The mutant protein was more effective than wild-type in reducing expression of cyclindependent kinase (CDK) inhibitor p27kip1, promoting cell cycle progression, inhibiting production of cleaved poly ADP ribose polymerase (PARP) (a marker for apoptosis), and down-regulating the multifunctional cytokine transforming growth factor-β (TGF-β). Since TGF-controls the cell cycle and apoptosis, the effect of BCP1 mutation on the enhanced activity of HBx to promote cell cycle progression and to inhibit apoptosis in Huh7 cells may result from down-regulation of TGF-β.