Identification of differentially expressed genes in human hepatoblastoma cell line (HepG2) and HBV-X transfected hepatoblastoma cell line (HepG2-4x).

Hepatitis B virus-X protein (HBV-X) is known to be an important factor in the formation of hepatocellular carcinoma by acting as a transcriptional activator on viral or cellular genes. To identify differentially expressed genes between the human hepatoblastoma cell line HepG2 and HBV-X gene transfected hepatoblastoma cell line HepG2-4X, we used a differential display polymerase chain reaction technique. The technique produced numerous up-regulated and down-regulated bands, each representing a partial cDNA fragment. We isolated 23 different kinds of cDNA fragments that showed marked differences in two cell lines. The fragments were used as templates for DNA sequencing analysis and as probes for Northern blot analysis. This analysis revealed that eight cDNA clones were differentially expressed in each cell line but fifteen cDNA clones were not. Among the 8 clones, 3 clones showed sequence similarities with human mitochondrial ATPase subunit 6 (mtATPase 6) and the human amidophosphoribosyl transferase (ATase) precursor, whereas 5 other clones were human novel protein encoding genes. Two genes having similarity with known genes were repressed by HBV-X. These results reflect that complex alterations of the expression of enzymes concerning the energy-generating system in mitochondria and metabolite synthesis are closely associated with the HBV-X function during the formation of hepatocellular carcinoma. These newly obtained genes will be useful for analyzing HBV-X functions. We are in the process of further characterizing these genes.