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Screening compounds against HCV based on MAVS/IFN-β pathway in a replicon model

Author(s): Qiu-Xia Fu, Li-Cui Wang, Shuai-Zheng Jia, Bo Gao, Yong Zhou, Juan Du, Ying-Li Wang, Xiao-Hui Wang, Jian-Chun Peng, Lin-Sheng Zhan

Journal: World Journal of Gastroenterology
ISSN 1007-9327

Volume: 16;
Issue: 44;
Start page: 5582;
Date: 2010;
Original page

Keywords: Mitochondrial antiviral signaling protein | Hepatitis C virus | Interferon-β | Drug screening

AIM: To develop a sensitive assay for screening compounds against hepatitis C virus (HCV).METHODS: The proteolytic cleavage of NS3/4A on enhanced yellow fluorescent protein (eYFP)-mitochondrial antiviral signaling protein (MAVS) was examined by reporter enzyme secreted placental alkaline phosphatase (SEAP), which enabled us to perform ongoing monitoring of anti-HCV drugs through repeated chemiluminescence. Subcellular localization of eYFP-MAVS was assessed by fluorescence microscopy. Cellular localization and protein levels were examined by Western blotting.RESULTS: HCV NS3/4A protease cleaved eYFP-MAVS from mitochondria to block the activation of interferon (IFN)-β promoter, thus resulting in downregulation of SEAP activity. The decrease in SEAP activity was proportional to the dose of active NS3/4A protease. Also this reporter assay was used to detect anti-HCV activity of IFN-α and cyclosporine A.CONCLUSION: Our data show that this reporter system is a sensitive and quantitative reporter of anti-HCV inhibitors. This system will constitute a new tool to allow the efficient screening of HCV inhibitors.
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