Differential expression of cellular genes in hepatitis C virus core expressing and parental cell lines

• Main Authors: Bor-I Kuo, 郭柏毅
• Other Authors: Ming-Fu Chang
• Format: Others
• Language: zh-TW
• Published: 2001
• Online Access: http://ndltd.ncl.edu.tw/handle/39243628368718438836

碩士 === 國立臺灣大學 === 生化學研究所 === 89 === Abstract Hepatitis C virus ( HCV ) is the major etiologic agent of post-transfusionally transmitted non-A, non-B hepatitis, and is highly associated with the development of hepatocellular carcinoma. The primary function of the HCV core protein is the formation of the viral nucleocapsid, but its various biochemical properties suggest that it may also play important roles in the regulation of cellular genes as well as the viral pathogenesis. For patients chronically infected with HCV, only a minority of them benefit from existing antiviral therapies, thus making highly desirable the development of a preventive, and possibly a therapeutic vaccine. One of the specific aims of this study is to induce immune response and generate HCV core-specific antibodies by DNA vaccination. Balb/c female mice were injected with a plasmid encoding an HCV core-Cr1 fusion protein into skeletal muscle. After 3 months of multiple boost immunization, the antibody titer was analyzed. Although expression of the HCV core-Cr1 protein was detected in transfected HepG2, none of the antisera of immunized mice recognize the HCV core protein specifically. The other specific aim of this study is to examine whether the expression of cellular genes are regulated by HCV core protein. Differential expression of cellular proteins among HCV core-expressing cell lines, HeLa tet-off(6), (13), (14), (16) and their parental HeLa tet-off cell line was analyzed by two dimensional electrophoresis assay. Six proteins including superoxide dismutase ( SOD ) were identified to be consistently regulated by HCV core protein in the stable cell lines. These results implied that HCV infection may disturb TNFa pathway and produce reactive oxygen species, then trigger cellular defense mechnisms and upregulate SOD expression. In addition, differential expression of cellular tyrosine kinases among HCV core-expressing cell lines and the parental HeLa tet-off cell line was analyzed by kinase display assay. Five restriction enzyme digestion patterns showed enhanced signals for all of the HCV core-expressing cell lines as compared to those of the parental cell line. From the sizes of the digested DNA fragments, these expression profiles were recognized as as the tyrosine kinase AXL/UFO/ARK. The result suggested that AXL/UFO/ARK is upregulated by HCV core protein. In addition, five differentially displayed bands were identified to be downregulated in three of the four HCV core-expressing cell lines. Nevertheless, these expression profiles could not be recognized as known tyrosine kinases to date. Additional experiments are needed to confirm the results from two-dimentional electrophoresis and kinase display analysis. The differentially expressed cellular proteins identified in these studies should be further characterized.