The antiviral state has shaped the CpG composition of the vertebrate interferome to avoid self-targeting.

Antiviral defenses can sense viral RNAs and mediate their destruction. This presents a challenge for host cells since they must destroy viral RNAs while sparing the host mRNAs that encode antiviral effectors. Here, we show that highly upregulated interferon-stimulated genes (ISGs), which encode antiviral proteins, have distinctive nucleotide compositions. We propose that self-targeting by antiviral effectors has selected for ISG transcripts that occupy a less self-targeted sequence space. Following interferon (IFN) stimulation, the CpG-targeting antiviral effector zinc-finger antiviral protein (ZAP) reduces the mRNA abundance of multiple host transcripts, providing a mechanistic explanation for the repression of many (but not all) interferon-repressed genes (IRGs). Notably, IRGs tend to be relatively CpG rich. In contrast, highly upregulated ISGs tend to be strongly CpG suppressed. Thus, ZAP is an example of an effector that has not only selected compositional biases in viral genomes but also appears to have notably shaped the composition of host transcripts in the vertebrate interferome.

Identification of TBK1 complexes required for the phosphorylation of IRF3 and the production of interferon ß.

The double-stranded RNA mimetic poly(I:C) and lipopolysaccharide (LPS) activate Toll-like receptors 3 (TLR3) and TLR4, respectively, triggering the activation of TANK (TRAF family member-associated NF-κB activator)-binding kinase 1 (TBK1) complexes, the phosphorylation of interferon regulatory factor 3 (IRF3) and transcription of the interferon ß (IFNß) gene. Here, we demonstrate that the TANK-TBK1 and optineurin (OPTN)-TBK1 complexes control this pathway. The poly(I:C)- or LPS-stimulated phosphorylation of IRF3 at Ser396 and production of IFNß were greatly reduced in bone marrow-derived macrophages (BMDMs) from TANK knockout (KO) mice crossed to knockin mice expressing the ubiquitin-binding-defective OPTN[D477N] mutant. In contrast, IRF3 phosphorylation and IFNß production were not reduced significantly in BMDM from OPTN[D477N] knockin mice and only reduced partially in TANK KO BMDM. The TLR3/TLR4-dependent phosphorylation of IRF3 and IFNß gene transcription were not decreased in macrophages from OPTN[D477N] crossed to mice deficient in IκB kinase ε, a TANK-binding kinase related to TBK1. In contrast with the OPTN-TBK1 complex, TBK1 associated with OPTN[D477N] did not undergo phosphorylation at Ser172 in response to poly(I:C) or LPS, indicating that the interaction of ubiquitin chains with OPTN is required to activate OPTN-TBK1 in BMDM. The phosphorylation of IRF3 and IFNß production induced by Sendai virus infection were unimpaired in BMDM from TANK KO × OPTN[D477N] mice, suggesting that other/additional TBK1 complexes control the RIG-I-like receptor-dependent production of IFNß. Finally, we present evidence that, in human HACAT cells, the poly(I:C)-dependent phosphorylation of TBK1 at Ser172 involves a novel TBK1-activating kinase(s).

Inhibition of interferon induction and action by the nairovirus Nairobi sheep disease virus/Ganjam virus.

The Nairoviruses are an important group of tick-borne viruses that includes pathogens of man (Crimean Congo hemorrhagic fever virus) and livestock animals (Dugbe virus, Nairobi sheep disease virus (NSDV)). NSDV is found in large parts of East Africa and the Indian subcontinent (where it is known as Ganjam virus). We have investigated the ability of NSDV to antagonise the induction and actions of interferon. Both pathogenic and apathogenic isolates could actively inhibit the induction of type 1 interferon, and also blocked the signalling pathways of both type 1 and type 2 interferons. Using transient expression of viral proteins or sections of viral proteins, these activities all mapped to the ovarian tumour-like protease domain (OTU) found in the viral RNA polymerase. Virus infection, or expression of this OTU domain in transfected cells, led to a great reduction in the incorporation of ubiquitin or ISG15 protein into host cell proteins. Point mutations in the OTU that inhibited the protease activity also prevented it from antagonising interferon induction and action. Interestingly, a mutation at a peripheral site, which had little apparent effect on the ability of the OTU to inhibit ubiquitination and ISG15ylation, removed the ability of the OTU to block the induction of type 1 and the action of type 2 interferons, but had a lesser effect on the ability to block type 1 interferon action, suggesting that targets other than ubiquitin and ISG15 may be involved in the actions of the viral OTU.