[Hepatitis B virus X protein inhibits hepatoma cell growth in vitro through p14(ARF)-dependent and p14(ARF)-independent pathways].
OBJECTIVE
To explore the effects of hepatitis B virus X protein (HBx) on hepatoma cell growth through p14(ARF)-dependent and p14(ARF)-independent pathways.


METHODS
HBx and p14(ARF) were transfected either separately or in combination into HepG2 cells containing wt-p53 but not expressing p14(ARF). The cells were divided into 4 groups, namely pcDNA3 (control), pcDNA3HBx, pcDNA3p14(ARF), and pcDNA3HBx + pcDNA3p14(ARF) groups. Flow cytometry was used to examine the apoptosis rates and cell cycle progression of HepG2 cells in different groups. The expression of p14(ARF), MDM2, p53, and p21(WAF1) proteins were investigated by detecting the activity of p21(WAF1) promoter-luciferase and using Western blotting.


RESULTS
The apoptosis rates of HepG2 cells in pcDNA3HBx and pcDNA3p14(ARF) groups were significantly higher than that in the control group (14.11%, 13.72% vs 10.66%). Compared with the control group, pcDNA3HBx and pcDNA3p14(ARF) groups also showed significantly higher cell percentages arrested at G(0)/G(1) phase (63.62%, 61.75% vs 57.42%), luciferase activity of p21 promoter (1.25-/+0.05, 1.09-/+0.06 vs 0.77-/+0.03) and expressions of p53 and p21(WAF1). The cell apoptosis rate, percentage of cells in G(0)/G(1) phase and expression level of p14(ARF) were even higher in pcDNA3HBx+pcDNA3p14(ARF) group (18.61%, 66.74%, and 3.53-/+0.43, respectively) than in either p14(ARF) or HBx group.


CONCLUSIONS
HBx induces p53 expression through p14(ARF)-dependent and independent pathways to activate p21(WAF1) promoter, leading to G(0)/G(1) arrest and apoptosis of HepG2 cells.
Yu
Dang-Hui
DH
Department of Pathology, Changhai Hospital, Second Military Medical University, Shanghai, China. alexfishing@yahoo.com.cn
Lin
Jing
J
Qu
Jian-Hui
JH
Zhu
Zhi
Z
Li
Fang-Mei
FM
Ni
Can-Rong
CR
Zhu
Ming-Hua
MH
D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000473:pathology / Q000821:virology; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics; D006801:Humans; D008113:Liver Neoplasms / Q000235:genetics / Q000473:pathology* / Q000821:virology; D011401:Promoter Regions, Genetic; D015534:Trans-Activators / Q000235:genetics*; D014162:Transfection; D025803:Tumor Suppressor Protein p14ARF / Q000235:genetics*; D016159:Tumor Suppressor Protein p53 / Q000235:genetics; D054334:Viral Regulatory and Accessory Proteins
C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; C495901:TP53 protein, human; D015534:Trans-Activators; D025803:Tumor Suppressor Protein p14ARF; D016159:Tumor Suppressor Protein p53; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein