| Summary: | 碩士 === 國立東華大學 === 生物技術研究所 === 96 === Japanese encephalitis virus (JEV) is a member of the genus flavivirus in the family flaviviridae. The genome is positive, single-strand RNA of 10,976 nucleotides in length. Viral nonstructural protein 5 (NS5) is an RNA-dependent RNA polymerase and plays a major role in JEV replication. In this study, host metabolic enzyme glyceraldehydes 3-phosphate dehydrogenase (GAPDH) was found to colocalize with NS5 using confocal laser-scanning microscopy. GAPDH fluorescence was visible in the nucleus and the cytoplasm of uninfected cells, while only visible in the cytoplasm of infected cells at 36 hours postinfection. Direct binding of GAPDH to NS5 could not be detected in JEV-infected BHK-21 cell lysate by immunoprecipitation followed by Western blot analysis, suggesting that colocalization of these two proteins may not involve direct protein-protein interaction. In addition, GAPDH binds to 3’ stemloop of JEV genome using electrophoretic mobility shift assay. In infected cells, the amount of GAPDH remained relative constant in cytosol, while increased at 12 to 24 hours postinfection and decreased at 36 hours postinfection in nucleus fraction, suggesting that GAPDH redistribution in nucleus may play a role in antivirus or antiapoptosis.
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