Arenavirus Nucleoprotein Targets Interferon Regulatory Factor-Activating Kinase IKK epsilon

Institute of Microbiology University Hospital Center and University of Lausanne, Lausanne, Switzerland.
Journal of Virology (Impact Factor: 4.65). 04/2012; 86(15):7728-38. DOI: 10.1128/JVI.00187-12
Source: PubMed

ABSTRACT Arenaviruses perturb innate antiviral defense by blocking induction of type I interferon (IFN) production. Accordingly, the arenavirus nucleoprotein (NP) was shown to block activation and nuclear translocation of interferon regulatory factor 3 (IRF3) in response to virus infection. Here, we sought to identify cellular factors involved in innate antiviral signaling targeted by arenavirus NP. Consistent with previous studies, infection with the prototypic arenavirus lymphocytic choriomeningitis virus (LCMV) prevented phosphorylation of IRF3 in response to infection with Sendai virus, a strong inducer of the retinoic acid-inducible gene I (RIG-I)/mitochondrial antiviral signaling (MAVS) pathway of innate antiviral signaling. Using a combination of coimmunoprecipitation and confocal microscopy, we found that LCMV NP associates with the IκB kinase (IKK)-related kinase IKKε but that, rather unexpectedly, LCMV NP did not bind to the closely related TANK-binding kinase 1 (TBK-1). The NP-IKKε interaction was highly conserved among arenaviruses from different clades. In LCMV-infected cells, IKKε colocalized with NP but not with MAVS located on the outer membrane of mitochondria. LCMV NP bound the kinase domain (KD) of IKKε (IKBKE) and blocked its autocatalytic activity and its ability to phosphorylate IRF3, without undergoing phosphorylation. Together, our data identify IKKε as a novel target of arenavirus NP. Engagement of NP seems to sequester IKKε in an inactive complex. Considering the important functions of IKKε in innate antiviral immunity and other cellular processes, the NP-IKKε interaction likely plays a crucial role in arenavirus-host interaction.

Download full-text


Available from: Luis Martinez-Sobrido, Aug 12, 2014
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Lassa virus infection elicits distinctive changes in host gene expression and metabolism. We focus on changes in host gene expression that may be biomarkers that discriminate individual pathogens or may help to provide a prognosis for disease. In addition to assessing mRNA changes, functional studies are also needed to discriminate causes of disease from mechanisms of host resistance. Host responses that drive pathogenesis are likely to be targets for prevention or therapy. Host responses to Lassa or its related arenaviruses have been monitored in cell culture, in animal models of hemorrhagic fever, in Lassa-infected nonhuman primates and, to a limited extent, in infected human beings. Here, we describe results from those studies and discuss potential targets for reducing virus replication and mitigating disease.
    Future Virology 03/2015; 10(3):233-256. DOI:10.2217/fvl.15.1 · 1.00 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Arenaviruses have a significant impact in public health and pose a credible biodefense threat, but the development of safe and effective arenavirus vaccines has remained elusive and currently no Food and Drug Administration (FDA)-licensed arenavirus vaccines are available. Here, we have explored the use of a codon deoptimization (CD)-based approach as a novel strategy to develop live-attenuated arenavirus vaccines. We recoded the nucleoprotein (NP) of the prototypic arenavirus lymphocytic choriomeningitis virus (LCMV) with the least frequently used codons in mammalian cells, which caused lower LCMV NP expression levels in transfected cells that correlated with decreased NP activity in cell-based functional assays. We used reverse genetic approaches to rescue a battery of recombinant (r)LCM viruses encoding CD NPs (rLCMV/NPCD) that showed attenuated growth kinetics in vitro. Moreover, experiments using the well-characterized mouse model of LCMV infection revealed that rLCMV/NPCD1 and rLCMV/NPCD2 were highly attenuated in vivo but, upon a single immunization, conferred complete protection against a subsequent lethal challenge with wild-type (WT) recombinant LCMV (rLCMV/WT). Both rLCMV/NPCD1 and rLCMV/NPCD2 were genetically and phenotypically stable during serial passages in FDA vaccine-approved Vero cells. These results provide proof of concept of the safety, efficacy and stability of a CD-based approach for developing live-attenuated vaccine candidates against human pathogenic arenaviruses. Several arenaviruses cause severe hemorrhagic fever in humans and pose a credible bioterrorism threat. Currently, no FDA-licensed vaccines are available to combat arenavirus infections, whereas anti-arenaviral therapy is limited to the off-label use of ribavirin that is only partially effective and associated with side effects. Here we describe the generation of recombinant versions of the prototypic arenavirus LCMV encoding codon deoptimized viral nucleoproteins (rLCMV/NPCD). We identified rLCMV/NPCD1 and rLCMV/NPCD2 as being highly attenuated in vivo but able to confer protection against a subsequent lethal challenge with wild-type LCMV. These viruses displayed an attenuated phenotype during serial amplification passages in cultured cells. Our findings support the use of this approach for the development of safe, stable, and protective live-attenuated arenavirus vaccines. Copyright © 2015, American Society for Microbiology. All Rights Reserved.
    Journal of Virology 01/2015; 89(7). DOI:10.1128/JVI.03401-14 · 4.65 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Many persistent viral infections are characterized by a hypofunctional T cell response and the upregulation of negative immune regulators. These events occur days after the initiation of infection. However, the very early host-virus interactions that determine the establishment of viral persistence remain poorly uncharacterized. Here we show that to establish persistence, LCMV must counteract an innate anti-viral immune response within eight hours after infection. While the virus triggers cytoplasmic RNA sensing pathways soon after infection, LCMV counteracts this pathway through a rapid increase in viral titers leading to a dysfunctional immune response characterized by a high cytokine and chemokine expression profile. This altered immune environment allows for viral replication in the splenic white pulp as well as infection of immune cells essential to an effective anti-viral immune response. Our findings illustrate how early events during infection critically dictate the characteristics of the immune response to infection and facilitate either virus control and clearance or persistence.
    PLoS Pathogens 01/2015; 11(1):e1004588. DOI:10.1371/journal.ppat.1004588 · 8.06 Impact Factor