Dataset Information

NS5-V372A and NS5-H386Y variations are responsible for differences in interferon ?/? induction and co-contribute to the replication advantage of Japanese encephalitis virus genotype I over genotype III in ducklings.

ABSTRACT: Japanese encephalitis virus (JEV) genotype I (GI) replicates more efficiently than genotype III (GIII) in birds, and this difference is considered to be one of the reasons for the JEV genotype shift. In this study, we utilized duck embryo fibroblasts and domestic ducklings as in vitro and in vivo models of a JEV amplifying avian host to identify the viral determinants of the differing replication efficiency between the GI and GIII strains in birds. GI strains induced significantly lower levels of interferon (IFN)-? and ? production than GIII strains, an effect orrelated with the enhanced replication efficiency of GI strains over GIII strains. By using a series of chimeric viruses with exchange of viral structural and non-structural (NS) proteins, we identified NS5 as the viral determinant of the differences in IFN-? and ? induction and replication efficiency between the GI and III strains. NS5 inhibited IFN-? and ? production induced by poly(I:C) stimulation and harbored 11 amino acid variations, of which the NS5-V372A and NS5-H386Y variations were identified to co-contribute to the differences in IFN-? and ? induction and replication efficiency between the strains. The NS5-V372A and NS5-H386Y variations resulted in alterations in the number of hydrogen bonds formed with neighboring residues, which were associated with the different ability of the GI and GIII strains to inhibit IFN-? and ? production. Our findings indicated that the NS5-V372A and NS5-H386Y variations enabled GI strains to inhibit IFN-? and ? production more efficiently than GIII strains for antagonism of the IFN-I mediated antiviral response, thereby leading to the replication and host adaption advantages of GI strains over GIII strains in birds. These findings provide new insight into the molecular basis of the JEV genotype shift.

SUBMITTER: Li C

PROVIDER: S-EPMC7494076 | biostudies-literature | 2020 Sep

REPOSITORIES: biostudies-literature

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NS5-V372A and NS5-H386Y variations are responsible for differences in interferon α/β induction and co-contribute to the replication advantage of Japanese encephalitis virus genotype I over genotype III in ducklings.

Li Chenxi C Di Di D Huang Hui H Wang Xin X Xia Qiqi Q Ma Xiaochun X Liu Ke K Li Beibei B Shao Donghua D Qiu Yafeng Y Li Zongjie Z Wei Jianchao J Ma Zhiyong Z

PLoS pathogens 20200903 9

Japanese encephalitis virus (JEV) genotype I (GI) replicates more efficiently than genotype III (GIII) in birds, and this difference is considered to be one of the reasons for the JEV genotype shift. In this study, we utilized duck embryo fibroblasts and domestic ducklings as in vitro and in vivo models of a JEV amplifying avian host to identify the viral determinants of the differing replication efficiency between the GI and GIII strains in birds. GI strains induced significantly lower levels o ...[more]

PMID: 32881988

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Project description:Japanese encephalitis virus (JEV) genotype dominance has shifted to genotype I (GI) from genotype III (GIII) in China as demonstrated by molecular epidemiological surveillance. In this study, we performed a serological survey in JEV-non-vaccinated pigs to confirm JEV genotype shift at the sero-epidemiological level. The average ratio of GI/GIII infection was 1.87, suggesting co-circulation of GI and GIII infections with GI infection being more prevalent in pigs in China. To gain an insight into the reasons for this JEV genotype shift, the replication kinetics of seven recently-isolated JEV isolates including three GI strains and four GIII strains were compared in mosquito C6/36 cells, chicken fibroblast cells (DF-1) and porcine iliac artery endothelial cells (PIEC). We observed that GI strains replicated more efficiently than GIII strains in DF-1 and PIEC cells, particularly in DF-1 cells with titers reaching 22.9-225.3 fold higher than GIII strains. This shows an enhanced replication efficiency of GI viruses in avian cells. To examine this enhanced replication efficiency in vivo, young domestic ducklings were used as the animal model and inoculated with GI and GIII strains at day 2 post-hatching. We observed that GI-inoculated ducklings developed higher viremia titers and displayed a comparatively longer viremic duration than GIII-inoculated ducklings. These results conform to the hypothesis of an enhanced replication efficiency for GI viruses in birds. There are 36 amino acid differences between GI and GIII viruses, some of which may be responsible for the enhanced replication efficiency of GI viruses in birds. Based on these findings, we speculated that the enhanced replication of GI viruses in birds would have resulted in higher exposure and therefore infection in mosquitoes, which could result in an increased transmission efficiency of GI viruses in the birds-mosquitoes-birds enzootic transmission cycle, thereby contributing to JEV genotype shift.

Project description:Japanese Encephalitis Virus (JEV) NS2B-NS3 is a protein complex composed of NS3 proteases and a NS2B cofactor. The N-terminal protease domain (180 residues) of NS3 (NS3(pro)) interacts directly with a central 40-amino acid hydrophilic domain of NS2B (NS2B(H)) to form an active serine protease. In this study, the recombinant NS2B(H)-NS3(pro) proteases were prepared in E. coli and used to compare the enzymatic activity between genotype I (GI) and III (GIII) NS2B-NS3 proteases. The GI NS2B(H)-NS3(pro) was able to cleave the sites at internal C, NS2A/NS2B, NS2B/NS3 and NS3/NS4A junctions that were identical to the sites proteolytically processed by GIII NS2B(H)-NS3(pro). Analysis of the enzymatic activity of recombinant NS2B(H)-NS3(pro) proteases using a model of fluorogenic peptide substrate revealed that the proteolytical processing activity of GIII NS2B(H)-NS3(pro) was significantly higher than that of GI NS2B(H)-NS3(pro). There were eight amino acid variations between GI and GIII NS2B(H)-NS3(pro), which may be responsible for the difference in enzymatic activities between GI and GIII proteases. Therefore, recombinant mutants were generated by exchanging NS2B(H) and NS3(pro) domains between GI and GIII NS2B(H)-NS3(pro) and subjected to protease activity analysis. Substitution of NS2B(H) significantly altered the protease activities, as compared to the parental NS2B(H)-NS3(pro), suggesting that NS2B(H) played an essential role in regulation of NS3(pro) protease activity. To further identify the amino acids responsible for the difference in protease activities, multiple substitution mutants including the individual and combined mutations at the variant residue 55 and 65 of NS2B(H) were generated and subjected to protease activity analysis. Replacement of NS2B-55 and NS2B-65 of GI to GIII significantly increased the enzymatic activity of GI NS2B(H)-NS3(pro) protease, whereas mutation of NS2B-55 and NS2B-65 of GIII to GI remarkably reduced the enzymatic activity of GIII NS2B(H)-NS3(pro) protease. Overall, these data demonstrated that NS2B-55 and NS2B-65 variations in hydrophilic domain of NS2B co-contributed to the difference in NS2B(H)-NS3(pro) protease activities between GI and GIII. These observations gain an insight into the role of NS2B in regulation of NS3 protease activities, which is useful for understanding the replication of JEV GI and GIII viruses.

Dataset Information

NS5-V372A and NS5-H386Y variations are responsible for differences in interferon ?/? induction and co-contribute to the replication advantage of Japanese encephalitis virus genotype I over genotype III in ducklings.

Publications

NS5-V372A and NS5-H386Y variations are responsible for differences in interferon α/β induction and co-contribute to the replication advantage of Japanese encephalitis virus genotype I over genotype III in ducklings.

Similar Datasets

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