Abstract
Hepatitis B virus X protein C-terminal 127 amino acid truncation is often found expressed in hepatocellular carcinoma (HCC) tissue samples. The present in vitro study tried to determine the role of this truncation mutant in the hepatitis B–related liver diseases such as fibrosis, cirrhosis, HCC, and metastasis. HBx gene and its 127 amino acid truncation mutant were cloned in mammalian expression vectors and transfected in human hepatoma cell line. Changes in cell growth/proliferation, cell cycle phase distribution, expression of cell cycle regulatory genes, mitochondrial depolarization, and intracellular reactive oxygen species (ROS) level were analyzed. Green fluorescent protein (GFP)–tagged version of HBx and the truncation mutant were also created and the effects of truncation on HBx intracellular expression pattern and localization were studied. Effect of time lapse on protein expression pattern was also analyzed. The truncation mutant of HBx is more efficient in inducing cell proliferation, and causes more ROS production and less mitochondrial depolarization as compared with wild type (wt) HBx. In addition, gene expression is altered in favor of carcinogenesis in the presence of the truncation mutant. Furthermore, mitochondrial perinuclear aggregation is achieved earlier in the presence of the truncation mutant. Therefore, HBx C-terminal 127 amino acid truncation might be playing important roles in the development of hepatitis B–related liver diseases by inducing cell proliferation, altering gene expression, altering mitochondrial potential, inducing mitochondrial clustering and oxidative stress, and changing HBx expression pattern.
Highlights
350 to 400 million people are chronically infected with hepatitis B virus (HBV) worldwide (Kao et al, 2000)
No amplification was observed from Complementary DNA (cDNA) from control cells transfected with emptypcDNA3 and empty-pEGFP-C3, and amplification was observed for cDNA derived from HepG2.2.15 (Figure 1B)
HBV X protein (HBx) is reported to be a crucial viral agent actively involved in liver cirrhosis and carcinogenesis in chronic hepatitis B patients (Feitelson et al, 2005)
Summary
350 to 400 million people are chronically infected with hepatitis B virus (HBV) worldwide (Kao et al, 2000). It is estimated that around 80 to 90% of chromosomal DNA isolated from hepatitis B–related HCC patients have integrated HBV fragments (Tu et al, 2001; Wang et al, 2004) Some of these random mutations and integrations cause creation of stop codons, which result in expression of truncated versions of HBV proteins, HBx at its c terminus (Tu et al, 2001; Wang et al, 2004; Caligiuri et al, 2016). The effect of HBx 127 on cell growth/proliferation, gene expression, intracellular reactive oxygen species (ROS) level, mitochondrial potential, and HBx gene expression were studied
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