Proteomic analysis for Type I interferon antagonism of Japanese encephalitis virus NS5 protein |
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Authors: | Chien‐Chen Lai Kai‐Zen Lu Man‐Tzu Chiu Tsung‐Han Hsieh Lei Wan Cheng‐Wen Lin |
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Affiliation: | 1. Department of Medical Genetics and Medical Research, China Medical University Hospital, , Taichung, Taiwan;2. Institute of Molecular Biology, National Chung Hsing University, , Taichung, Taiwan;3. Department of Medical Laboratory Science and Biotechnology, China Medical University, , Taichung, Taiwan;4. Department of Biotechnology, College of Health Science, Asia University, Wufeng, , Taichung, Taiwan |
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Abstract: | Japanese encephalitis virus (JEV) nonstructural protein 5 (NS5) exhibits a Type I interferon (IFN) antagonistic function. This study characterizes Type I IFN antagonism mechanism of NS5 protein, using proteomic approach. In human neuroblastoma cells, NS5 expression would suppress IFNβ‐induced responses, for example, expression of IFN‐stimulated genes PKR and OAS as well as STAT1 nuclear translocation and phosphorylation. Proteomic analysis showed JEV NS5 downregulating calreticulin, while upregulating cyclophilin A, HSP 60 and stress‐induced‐phosphoprotein 1. Gene silence of calreticulin raised intracellular Ca2+ levels while inhibiting nuclear translocalization of STAT1 and NFAT‐1 in response to IFNβ, thus, indicating calreticulin downregulation linked with Type I IFN antagonism of JEV NS5 via activation of Ca2+/calicineurin. Calcineurin inhibitor cyclosporin A attenuated NS5‐mediated inhibition of IFNβ‐induced responses, for example, IFN‐sensitive response element driven luciferase, STAT1‐dependent PKR mRNA expression, as well as phosphorylation and nuclear translocation of STAT1. Transfection with calcineurin (vs. control) siRNA enhanced nuclear translocalization of STAT1 and upregulated PKR expression in NS5‐expressing cells in response to IFNβ. Results prove Ca2+, calreticulin, and calcineurin involvement in STAT1‐mediated signaling as well as a key role of JEV NS5 in Type I IFN antagonism. This study offers insights into the molecular mechanism of Type I interferon antagonism by JEV NS5. |
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Keywords: | Calreticulin Interferon Japanese encephalitis virus Microbiology Non‐structural protein 5 STAT1 |
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