Protein ISGylation modulates the JAK-STAT signaling pathway

ISG15 is one of the most strongly induced genes upon viral infection, type I interferon (IFN) stimulation, and lipopolysaccharide (LPS) stimulation. Here we report that mice lacking UBP43, a protease that removes ISG15 from ISGylated proteins, are hypersensitive to type I IFN. Most importantly, in UBP43-deficient cells, IFN-beta induces a prolonged Stat1 tyrosine phosphorylation, DNA binding, and IFN-mediated gene activation. Furthermore, restoration of ISG15 conjugation in protein ISGylation-defective K562 cells increases IFN-stimulated promoter activity. These findings identify UBP43 as a novel negative regulator of IFN signaling and suggest the involvement of protein ISGylation in the regulation of the JAK-STAT pathway.     

CXCL10/CXCR3-mediated responses promote immunity to respiratory syncytial virus infection by augmenting dendritic cell and CD8(+) T cell efficacy

The induction of inflammatory cytokines during respiratory viral infections contributes to both disease pathogenesis and resolution. The present studies investigated the role of the chemokine CXCL10 and its specific receptor, CXCR3, in the host response to pulmonary respiratory syncytial virus (RSV) infection. Antibody-mediated neutralization of CXCL10 resulted in a significant increase in disease pathogenesis, including airway hyperresponsiveness (AHR), mucus gene expression, and impaired viral clearance. When the pulmonary cytokine levels were examined, only type I IFN and IL-12p70 were significantly reduced. These latter observations were reflected in reduced dendritic cell (DC) numbers and DC maturation in the lungs of RSV-infected mice treated with anti-CXCL10. Neutralization of the only known receptor for CXCL10, CXCR3, resulted in similar increases in pathogenic responses. When bone marrow-derived DC were incubated with CXCL10 and RSV, an up-regulation of type I IFN was observed. In addition, T lymphocytes were also examined and a significant decrease in the number of RSV M2 peptide-specific CD8(+) T cells was identified. These findings highlight a previously unappreciated role for the CXCL10:CXCR3 signaling axis in RSV-infected animals by recruiting virus-specific T cells into the lung and promoting viral clearance.     

Respiratory syncytial virus activates epidermal growth factor receptor to suppress interferon regulatory factor 1-dependent interferon-lambda and antiviral defense in airway epithelium

Respiratory syncytial virus (RSV) persists as a significant human pathogen that continues to contribute to morbidity and mortality. In children, RSV is the leading cause of lower respiratory tract infections, and in adults RSV causes pneumonia and contributes to exacerbations of chronic lung diseases. RSV induces airway epithelial inflammation by activation of the epidermal growth factor receptor (EGFR), a tyrosine kinase receptor. Recently, EGFR inhibition was shown to decrease RSV infection, but the mechanism(s) for this effect are not known. Interferon (IFN) signaling is critical for innate antiviral responses, and recent experiments have implicated IFN-λ (lambda), a type III IFN, as the most significant IFN for mucosal antiviral immune responses to RSV infection. However, a role for RSV-induced EGFR activation to suppress airway epithelial antiviral immunity has not been explored. Here, we show that RSV-induced EGFR activation suppresses IFN regulatory factor (IRF) 1-induced IFN-λ production and increased viral infection, and we implicate RSV F protein to mediate this effect. EGFR inhibition, during viral infection, augmented IRF1, IFN-λ, and decreased RSV titers. These results suggest a mechanism for EGFR inhibition to suppress RSV by activation of endogenous epithelial antiviral defenses, which may be a potential target for novel therapeutics.     

Type I interferon regulates respiratory virus infected dendritic cell maturation and cytokine production

Activation of dendritic cells (DCs) by viruses is critical for both innate and adaptive immune responses. In this report, we investigated the role of type I interferon (IFN) in the activation of DCs by respiratory syncytial virus (RSV). Using DCs from type I IFNR-/- mice, these studies indicate that maturation, including upregulation of co-stimulatory molecules and optimal cytokine production, by RSV infection was dependent on type I IFN receptor signaling. Subsequently, studies using DCs from wild type mice demonstrate that continued production of type I IFN during later stages of DC maturation could alter their activation profiles. IFN-alpha and IFN-beta were upregulated in DCs grown from bone marrow of wild type mice after infection with RSV. In order to determine their function in competent DCs, blocking antibodies were used to specifically inhibit IFN-alpha/beta . The data demonstrate that production of IFN-beta, but not IFN-alpha, in RSV-infected wild type DCs promotes chemokine production and toll-like receptor (TLR) expression, while limiting IL-12 production. The inhibition of IL-12p70 by IFN-beta correlated with suppressed IL-12p40 expression levels. Furthermore, the addition of recombinant IFN-beta potently inhibited IL-12p40 expression in mature DC subsets during RSV infection, while only the highest dose of IFN-alpha had any inhibitory effect. Together, our studies provide insight into the complex regulation of DC maturation and IL-12 production co-ordinated by type I interferons in RSV-infected dendritic cells, and demonstrate that type I IFN has specific roles depending upon the stage of DC maturation.     

Respiratory syncytial virus strain A2 is resistant to the antiviral effects of type I interferons and human MxA.

Respiratory syncytial virus (RSV) belongs to Paramyxoviridae family of enveloped negative-strand RNA viruses and causes severe bronchiolitis and pneumonia in children younger than 2 years of age. As members of Paramyxoviridae family, RSV and parainfluenza type 3 (PIV3) have similar modes of infection and replication. A variety of negative-strand RNA virus infections, including that of PIV3, are inhibited by human MxA protein, a type I interferon (IFN)-inducible GTPase. We tested whether the MxA protein, induced either by type I human IFNs or by stable transfection of human MxA gene in human (U-87) or simian (Vero) cells, confers resistance to these cells against infection by RSV strain A2. RSV infection was resistant to antiviral effects induced by 0-10,000 U/ml type I IFNs (IFN-alpha or -beta) in both human lung epithelial, A549, and fibroblast, MRC-5 cells. RSV virus yield was reduced only by 10- to 20-fold, and viral protein synthesis was not significantly affected under conditions of IFN treatment where PIV3 yield was reduced by 1000- to 10,000-fold. Human or simian cell lines constitutively expressing MxA were protected against infection by PIV3 but not by RSV. Our results indicate that RSV A2 is resistant to the antiviral effects of MxA, even though RSV and PIV3 have similar replication strategies. In IFN-treated coinfected cultures, IFN-resistant RSV A2 did not prevent the IFN-mediated inhibition of PIV3 multiplication. Hence the resistance of RSV A2 to type I IFNs does not appear to be due to soluble factors released into the medium or a disruption in the cellular antiviral machinery brought about by RSV A2 infection.     

Role for herpes simplex virus 1 ICP27 in the inhibition of type I interferon signaling

Host cells respond to viral infection by many mechanisms, including the production of type I interferons which act in a paracrine and autocrine manner to induce the expression of antiviral interferon-stimulated genes (ISGs). Viruses have evolved means to inhibit interferon signaling to avoid induction of the innate immune response. Herpes simplex virus 1 (HSV-1) has several mechanisms to inhibit type I interferon production, the activities of ISGs, and the interferon signaling pathway itself. We report that the inhibition of the Jak/STAT pathway by HSV-1 requires viral gene expression and that viral immediate-early protein ICP27 plays a role in downregulating STAT-1 phosphorylation and in preventing the accumulation of STAT-1 in the nucleus. We also show that expression of ICP27 by transfection causes an inhibition of IFN-induced STAT-1 nuclear accumulation. Therefore, ICP27 is necessary and sufficient for at least some of the effects of HSV infection on STAT-1.