Interleukin-6 induces the (2'-5') oligoadenylate synthetase gene in M1 cells through an effect on the interferon-responsive enhancer

Interleukin-6 (IL-6) activates (2'-5') A synthetase (2'-5' AS) gene expression in differentiating myeloleukemic M1 cells. Antibodies to type I interferon (IFN) inhibit 2'-5' AS induction but not differentiation. Analysis of the mechanism of 2'-5' AS induction shows that it does not result from increased IFN formation, but from a synergism between IL-6 and endogenously secreted IFN. IL-6 can activate expression of a CAT construct fused to the interferon response sequence (IRS) of the 2'-5' AS gene. In extracts of IL-6-treated M1 cells, changes in protein binding to IRS DNA can be demonstrated. One of the effects of IL-6 on M1 cells is, therefore, to induce DNA binding factors, some of which act on the same enhancer sequence as IFNs, resulting in a synergistic gene activation. M1 variants resistant to differentiation by IL-6 have lost the ability to induce the 2'-5' AS gene.     

Mutational analysis of regulation of MHC and anti-viral genes.

CTL-mediated selection for loss of expression of Mta by H-2-heterozygous SV40-transformed mouse fibroblasts (line 24SV) produced an unusual phenotypic class of maternally transmitted Ag negative mutants defective in both MHC expression and in anti-viral activity. Severely reduced surface expression of class I MHC Ag from multiple loci of both haplotypes correlated with low levels of MHC H chain and beta 2-microglobulin mRNA. Inasmuch as IFN can up-regulate class I expression and some fibroblasts elaborate autocrine IFN-beta, we examined whether IFN could restore wild-type expression of class I MHC Ag. However, IFN could not restore wild-type expression. Moreover, the fold-increases in class I Ag and mRNA expression were significantly reduced in mutant cells compared to wild-type cells. These results suggested that the mutants might have generalized defects in IFN response. Inasmuch as the induction of an anti-viral state is a hallmark of IFN responses, we exposed cells to IFN-alpha, -beta, or -gamma and challenged with virus. 24SV cells, exposed to any of the three IFNs, were completely protected from destruction by vesicular stomatitis, mengovirus or respiratory syncytial viruses. In contrast, MHC and anti-viral defective mutants could not be protected from virus-induced lysis by any IFN. Somatic cell hybridization analyses indicated that both basal MHC and IFN-inducible phenotypes were recessive to wild-type, and that a trans-acting regulatory factor required for basal MHC expression is defectively expressed in the mutants. Such a factor may integrate the organismal response to virus infection, encompassing both immune and nonimmune anti-viral responses.     

Influenza virus evades innate and adaptive immunity via the NS1 protein

Both antibodies and T cells contribute to immunity against influenza virus infection. However, the generation of strong Th1 immunity is crucial for viral clearance. Interestingly, we found that human dendritic cells (DCs) infected with influenza A virus have lower allospecific Th1-cell stimulatory abilities than DCs activated by other stimuli, such as lipopolysaccharide and Newcastle disease virus infection. This weak stimulatory activity correlates with a suboptimal maturation of the DCs following infection with influenza A virus. We next investigated whether the influenza A virus NS1 protein could be responsible for the low levels of DC maturation after influenza virus infection. The NS1 protein is an important virulence factor associated with the suppression of innate immunity via the inhibition of type I interferon (IFN) production in infected cells. Using recombinant influenza and Newcastle disease viruses, with or without the NS1 gene from influenza virus, we found that the induction of a genetic program underlying DC maturation, migration, and T-cell stimulatory activity is specifically suppressed by the expression of the NS1 protein. Among the genes affected by NS1 are those coding for macrophage inflammatory protein 1beta, interleukin-12 p35 (IL-12 p35), IL-23 p19, RANTES, IL-8, IFN-alpha/beta, and CCR7. These results indicate that the influenza A virus NS1 protein is a bifunctional viral immunosuppressor which inhibits innate immunity by preventing type I IFN release and inhibits adaptive immunity by attenuating human DC maturation and the capacity of DCs to induce T-cell responses. Our observations also support the potential use of NS1 mutant influenza viruses as live attenuated influenza virus vaccines.     

Rotavirus NSP1 inhibits expression of type I interferon by antagonizing the function of interferon regulatory factors IRF3, IRF5, and IRF7

Secretion of interferon (IFN) by virus-infected cells is essential for activating autocrine and paracrine pathways that promote cellular transition to an antiviral state. In most mammalian cells, IFN production is initiated by the activation of constitutively expressed IFN regulatory factor 3, IRF3, which in turn leads to the induction of IRF7, the "master regulator" of IFN type I synthesis (alpha/beta IFN). Previous studies established that rotavirus NSP1 antagonizes IFN signaling by inducing IRF3 degradation. In the present study, we have determined that, in comparison to wild-type rotaviruses, rotaviruses encoding defective NSP1 grow to lower titers in some cell lines and that this poor growth phenotype is due to their failure to suppress IFN expression. Furthermore, we provide evidence that rotaviruses encoding wild-type NSP1 subvert IFN signaling by inducing the degradation of not only IRF3, but also IRF7, with both events occurring through proteasome-dependent processes that proceed with similar efficiencies. The capacity of NSP1 to induce IRF7 degradation may allow rotavirus to move across the gut barrier by enabling the virus to replicate in specialized trafficking cells (dendritic cells and macrophages) that constitutively express IRF7. Along with IRF3 and IRF7, NSP1 was found to induce the degradation of IRF5, a factor that upregulates IFN expression and that is involved in triggering apoptosis during viral infection. Our analysis suggests that NSP1 mediates the degradation of IRF3, IRF5, and IRF7 by recognizing a common element of IRF proteins, thereby allowing NSP1 to act as a broad-spectrum antagonist of IRF function.     

Dengue Virus Serotype 2 Blocks Extracellular Signal-Regulated Kinase and Nuclear Factor-κB Activation to Downregulate Cytokine Production

Background

Dengue virus (DENV) infection is the most common mosquito-borne viral disease threatening human health around the world. Type I interferon (IFN) and cytokine production are crucial in the innate immune system. We previously reported that DENV serotype 2 (DENV-2) induced low levels of interferon regulatory factor 3 and NF-κB activation, thus leading to reduced production of IFN-β in the early phase of infection. Here, we determined whether DENV infection not only hampers type I IFN activation but also cytokine production triggered by Toll-like receptor (TLR) signaling.

Methodology/Principal Findings

We used quantitative RT-PCR and found that only low levels of IFN-β and inflammatory cytokines such as interleukin 10 (IL-10), IL-12 and tumor necrosis factor α (TNFα) mRNA were detected in DENV-2–infected bone-marrow–derived dendritic cells. Furthermore, DENV-2 infection repressed cytokine production triggered by TLR signaling. To elucidate the molecular mechanisms underlying this suppression event, we measured NF-κB activation by p65 nuclear translocation and luciferase reporter assay and found that NF-κB activation triggered by TLR ligands was blocked by DENV-2 infection. As well, extracellular signal-regulated kinase (ERK) activity was suppressed by DENV-2 infection.

Conclusions/Significance

To downregulate the host innate immunity, DENV-2 by itself is a weak inducer of type I IFN and cytokines, furthermore DENV-2 can also block the TLR-triggered ERK–NF-κB activation and cytokine production.     

Ebola virus glycoprotein toxicity is mediated by a dynamin-dependent protein-trafficking pathway

Ebola virus infection causes a highly lethal hemorrhagic fever syndrome associated with profound immunosuppression through its ability to induce widespread inflammation and cellular damage. Though GP, the viral envelope glycoprotein, mediates many of these effects, the molecular events that underlie Ebola virus cytopathicity are poorly understood. Here, we define a cellular mechanism responsible for Ebola virus GP cytotoxicity. GP selectively decreased the expression of cell surface molecules that are essential for cell adhesion and immune function. GP dramatically reduced levels of alphaVbeta3 without affecting the levels of alpha2beta1 or cadherin, leading to cell detachment and death. This effect was inhibited in vitro and in vivo by brefeldin A and was dependent on dynamin, the GTPase. GP also decreased cell surface expression of major histocompatibility complex class I molecules, which alters recognition by immune cells, and this effect was also dependent on the mucin domain previously implicated in GP cytotoxicity. By altering the trafficking of select cellular proteins, Ebola virus GP inflicts cell damage and may facilitate immune escape by the virus.     

Infection and activation of monocytes by Marburg and Ebola viruses

In this study we investigated the effects of Marburg virus and Ebola virus (species Zaire and Reston) infections on freshly isolated suspended monocytes in comparison to adherent macrophages under culture conditions. Our data showed that monocytes are permissive for both filoviruses. As is the case in macrophages, infection resulted in the activation of monocytes which was largely independent of virus replication. The activation was triggered similarly by Marburg and Ebola viruses, species Zaire and Reston, as indicated by the release of the proinflammatory cytokines interleukin-1beta (IL-1beta), tumor necrosis factor alpha, and IL-6 as well as the chemokines IL-8 and gro-alpha. Our data suggest that infected monocytes may play an important role in the spread of filoviruses and in the pathogenesis of filoviral hemorrhagic disease.     

SOCS-1 inhibits expression of the antiviral proteins 2',5'-OAS and MxA induced by the novel interferon-lambdas IL-28A and IL-29

Recently, we have shown that SOCS-1/3 overexpression in hepatic cells abrogates signaling of type I interferons (IFN) which may contribute to the frequently observed IFN resistance of hepatitis C virus (HCV). IFN-lambdas (IL-28A/B and IL-29), a novel group of IFNs, also efficiently inhibit HCV replication in vitro with potentially less hematopoietic side effects than IFN-alpha because of limited receptor expression in hematopoietic cells. To further evaluate the potential of IFN-lambdas in chronic viral hepatitis, we examined the influence of SOCS protein expression on IFN-lambda signaling. First, we show that hepatic cell lines express the IFN-lambda receptor complex consisting of IFN-lambdaR1 (IL-28R1) and IL-10R2. Whereas in mock-transfected HepG2 cells, IL-28A and IL-29 induced STAT1 and STAT3 phosphorylation, overexpression of SOCS-1 completely abrogated IL-28A and IL-29-induced STAT1/3 phosphorylation. Similarly, IL-28A and IL-29 induced mRNA expression of the antiviral proteins 2',5'-OAS and MxA was abolished by overexpression of SOCS-1. In conclusion, we assume that despite antiviral properties of IFN-lambdas, their efficacy as antiviral agents may have similar limitations as IFN-alpha due to inhibition by SOCS proteins.     

Expression kinetics of interferon and interferon-induced genes in Atlantic salmon (Salmo salar) following infection with infectious pancreatic necrosis virus and infectious salmon anaemia virus

Infectious pancreatic necrosis virus (IPNV) and infectious salmon anaemia virus (ISAV) are economically important pathogens of the salmonid aquaculture industry. Atlantic salmon were challenged by intraperitoneal injection (i.p.) with either virus followed by time-course sampling. Cohabiting fish in the IPNV challenge were also sampled. Kidney tissue was analysed using a TaqMan real-time PCR assay to measure the expression of a range of host immune genes in relation to the endogenous control, elongation factor 1 alpha (ELF). Host genes measured included Mx, type I and type II interferon (IFN), gammaIFN induced protein (gammaIP), interleukin-1 beta (IL-1beta) and tumour necrosis factor alpha (TNF-alpha). Viral levels were also measured. In i.p. injected fish, both viruses greatly induced expression of Mx, gammaIP, type I and type II IFN by day 6 post-infection, however only ISAV caused substantial mortality. Some differences between the expression kinetics produced by both viruses were noted. Infection with ISAV increased IL-1beta expression following day 6, but no effect was seen in fish infected with IPNV. Neither virus induced TNF-alpha expression. This study confirms the presence of both type I and type II IFN responses and their induced genes in Atlantic salmon upon infection with an orthomyxovirus and a birnavirus.     

Newcastle disease virus (NDV)-based assay demonstrates interferon-antagonist activity for the NDV V protein and the Nipah virus V,W, and C proteins

We have generated a recombinant Newcastle disease virus (NDV) that expresses the green fluorescence protein (GFP) in infected chicken embryo fibroblasts (CEFs). This virus is interferon (IFN) sensitive, and pretreatment of cells with chicken alpha/beta IFN (IFN-alpha/beta) completely blocks viral GFP expression. Prior transfection of plasmid DNA induces an IFN response in CEFs and blocks NDV-GFP replication. However, transfection of known inhibitors of the IFN-alpha/beta system, including the influenza A virus NS1 protein and the Ebola virus VP35 protein, restores NDV-GFP replication. We therefore conclude that the NDV-GFP virus could be used to screen proteins expressed from plasmids for the ability to counteract the host cell IFN response. Using this system, we show that expression of the NDV V protein or the Nipah virus V, W, or C proteins rescues NDV-GFP replication in the face of the transfection-induced IFN response. The V and W proteins of Nipah virus, a highly lethal pathogen in humans, also block activation of an IFN-inducible promoter in primate cells. Interestingly, the amino-terminal region of the Nipah virus V protein, which is identical to the amino terminus of Nipah virus W, is sufficient to exert the IFN-antagonist activity. In contrast, the anti-IFN activity of the NDV V protein appears to be located in the carboxy-terminal region of the protein, a region implicated in the IFN-antagonist activity exhibited by the V proteins of mumps virus and human parainfluenza virus type 2.