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J Virol. 2015 Apr 22. pii: JVI.00857-15. [Epub ahead of print]
FACS-based analysis reveals an asymmetric induction of interferon stimulated genes in response to seasonal influenza a virus.
von Recum-Knepper J1, Sadewasser A1, Weinheimer VK1, Wolff T2.
Author information
Abstract
Influenza A virus (IAV) infection provokes an antiviral response involving the expression of type I and III interferons (IFN) and IFN-stimulated genes (ISGs) in infected cell cultures. However, the spatio-temporal dynamics of the IFN reaction are incompletely understood as previous studies investigated mainly the population responses of virus infected cultures, although substantial cell-to-cell variability has been documented. We devised a fluorescence activated cell sorting-based assay to simultaneously quantify expression of viral antigen and ISGs such as ISG15, MxA and IFIT1 in IAV-infected cell cultures on the level of single cells. This approach revealed that seasonal IAV triggers an unexpected asymmetric response as the major cell populations expressed either viral antigen or ISG, but rarely both. Further investigations identified a role of the viral NS1 protein in blocking ISG expression in infected cells, which surprisingly did not reduce paracrine IFN signaling to non-infected cells. Interestingly, viral ISG control was impaired in cultures infected with avian-origin IAV including H7N9 virus from Eastern China. This phenotype was traced back to polymorphic NS1 amino acids known to be important for stable binding of the polyadenylation factor CPSF30 and concomitant suppression of host cell gene expression. Most significantly, mutation of two amino acids within the CPSF30 attachment site of NS1 from seasonal IAV diminished the strict control of ISG expression in infected cells and substantially attenuated virus replication. In conclusion, our approach revealed an asymmetric, NS1-dependent ISG induction in cultures infected with seasonal IAV, which appears to be essential for efficient virus propagation.
IMPORTANCE:
Interferons are expressed by infected cells in response to IAV infection and play important roles in the antiviral immune response by inducing hundreds of interferon-stimulated genes (ISGs). Unlike many previous studies, we investigated the ISG response on the level of single cells enabling novel insights into this virus-host interaction. Hence, cell cultures infected with seasonal IAV displayed an asymmetric ISG induction that was confined almost exclusively to non-infected cells. In comparison, ISG expression was observed in larger cell populations infected with avian-origin IAV suggesting a more resolute antiviral response towards these strains. Strict control of ISG expression by seasonal IAV was explained by the binding of the viral NS1 protein to the polyadenylation factor CPSF30, which reduces host cell gene expression. Mutational disruption of CPSF30 binding within NS1 concomitantly attenuated ISG control and replication of seasonal IAV illustrating the importance of maintaining an asymmetric ISG response for efficient virus propagation.
Copyright ? 2015, American Society for Microbiology. All Rights Reserved.
PMID: 25903337 [PubMed - as supplied by publisher]
FACS-based analysis reveals an asymmetric induction of interferon stimulated genes in response to seasonal influenza a virus.
von Recum-Knepper J1, Sadewasser A1, Weinheimer VK1, Wolff T2.
Author information
Abstract
Influenza A virus (IAV) infection provokes an antiviral response involving the expression of type I and III interferons (IFN) and IFN-stimulated genes (ISGs) in infected cell cultures. However, the spatio-temporal dynamics of the IFN reaction are incompletely understood as previous studies investigated mainly the population responses of virus infected cultures, although substantial cell-to-cell variability has been documented. We devised a fluorescence activated cell sorting-based assay to simultaneously quantify expression of viral antigen and ISGs such as ISG15, MxA and IFIT1 in IAV-infected cell cultures on the level of single cells. This approach revealed that seasonal IAV triggers an unexpected asymmetric response as the major cell populations expressed either viral antigen or ISG, but rarely both. Further investigations identified a role of the viral NS1 protein in blocking ISG expression in infected cells, which surprisingly did not reduce paracrine IFN signaling to non-infected cells. Interestingly, viral ISG control was impaired in cultures infected with avian-origin IAV including H7N9 virus from Eastern China. This phenotype was traced back to polymorphic NS1 amino acids known to be important for stable binding of the polyadenylation factor CPSF30 and concomitant suppression of host cell gene expression. Most significantly, mutation of two amino acids within the CPSF30 attachment site of NS1 from seasonal IAV diminished the strict control of ISG expression in infected cells and substantially attenuated virus replication. In conclusion, our approach revealed an asymmetric, NS1-dependent ISG induction in cultures infected with seasonal IAV, which appears to be essential for efficient virus propagation.
IMPORTANCE:
Interferons are expressed by infected cells in response to IAV infection and play important roles in the antiviral immune response by inducing hundreds of interferon-stimulated genes (ISGs). Unlike many previous studies, we investigated the ISG response on the level of single cells enabling novel insights into this virus-host interaction. Hence, cell cultures infected with seasonal IAV displayed an asymmetric ISG induction that was confined almost exclusively to non-infected cells. In comparison, ISG expression was observed in larger cell populations infected with avian-origin IAV suggesting a more resolute antiviral response towards these strains. Strict control of ISG expression by seasonal IAV was explained by the binding of the viral NS1 protein to the polyadenylation factor CPSF30, which reduces host cell gene expression. Mutational disruption of CPSF30 binding within NS1 concomitantly attenuated ISG control and replication of seasonal IAV illustrating the importance of maintaining an asymmetric ISG response for efficient virus propagation.
Copyright ? 2015, American Society for Microbiology. All Rights Reserved.
PMID: 25903337 [PubMed - as supplied by publisher]