Identification and characterization of Molecular Chaperone,GRP78, in Japanese Encephalitis Virus-infected cells

• Main Authors: Yi Ping Wu, 吳依蘋
• Other Authors: R. Y. L. Wang
• Format: Others
• Published: 2011
• Online Access: http://ndltd.ncl.edu.tw/handle/58345651233714802564

碩士 === 長庚大學 === 生物醫學研究所 === 99 === Japanese Encephalitis virus (JEV) is a mosquito-borne virus, which is the main cause of encephalitis diseases in Asia. Previous studies showed that some host proteins involved in positive-strand RNA viruses life cycle, included Dengue virus (DENV), Hepatitis C virus (HCV) as well as Enterovirus (EV). In this study, the proteomic analysis of the secretory proteins from JEV infected Baby hamster kidney 21 (BHK21) cells identified several host proteins：elongation factor 2 (EF2), heat shock protein 90 (HSP90), glucose regulation protein 78 (GRP78), heat shock protein 70 (Hsp70) and phosphopantothenate-cysteine ligase。. Here we focus on the role of GRP78 in JEV life cycle. The GRP78 was an important endoplasmic reticulum (ER) chaperon in unfolding protein response (UPR) that will bind with unfolding protein and regulate the ER stress. The interactions between JE viral structural and GRP78 were identified by sucrose density gradient assay, immunofluerencent assay and co-immunoprecipitation assay. Western blot analysis revealed that GPR78 was associated with viral particles, which were purified from sucrose density gradient fractions. The western blot result also showed the JE viral RNA replication was reduced when GRP78 expression level was silenced by GRP78 siRNA. The extra-cellular viral RNA was reduced in the GRP78 silenced BHK21 cells compared to GRP78 wild type cells. The antibody neutralization assay also identified that GRP78 associated with JEV particle so as to facilitate JEV entry. Altogether, our data suggested that GRP78 was associated with JE viral structural proteins and facilitate the JE viral proteins replication in host cells. The host protein GRP78 released with JEV particle from host cells, and help JEV entry.