Antagonism of the interferon-induced OAS-RNase L pathway by murine coronavirus ns2 protein is required for virus replication and liver pathology.
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ABSTRACT:
SUBMITTER: Zhao L
PROVIDER: S-EPMC3377938 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:The OAS/RNase L system was one of the first characterized interferon effector pathways. It relies on the synthesis, by oligoadenylate synthetases (OAS), of short oligonucleotides that act as second messengers to activate the latent cellular RNase L. Viruses have developed diverse strategies to escape its antiviral effects. This underscores the importance of the OAS/RNase L pathway in antiviral defenses. Viral proteins such as the NS1 protein of Influenza virus A act upstream of the pathway while other viral proteins such as Theiler's virus L* protein act downstream. The diversity of escape strategies used by viruses likely stems from their relative susceptibility to OAS/RNase L and other antiviral pathways, which may depend on their host and cellular tropism.
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Project description:Viral 2',5'-phosphodiesterases (2',5'-PDEs) help disparate RNA viruses evade the antiviral activity of interferon (IFN) by degrading 2',5'-oligoadenylate (2-5A) activators of RNase L. A kinase anchoring proteins (AKAPs) bind the regulatory subunits of protein kinase A (PKA) to localize and organize cyclic AMP (cAMP) signaling during diverse physiological processes. Among more than 43 AKAP isoforms, AKAP7 appears to be unique in its homology to viral 2',5'-PDEs. Here we show that mouse AKAP7 rapidly degrades 2-5A with kinetics similar to that of murine coronavirus (mouse hepatitis virus [MHV]) strain A59 ns2 and human rotavirus strain WA VP3 proteins. To determine whether AKAP7 could substitute for a viral 2',5'-PDE, we inserted AKAP7 cDNA into an MHV genome with an inactivated ns2 gene. The AKAP7 PDE domain or N-terminally truncated AKAP7 (both lacking a nuclear localization motif), but not full-length AKAP7 or a mutant, AKAP7(H185R), PDE domain restored the infectivity of ns2 mutant MHV in bone marrow macrophages and in livers of infected mice. Interestingly, the AKAP7 PDE domain and N-terminally deleted AKAP7 were present in the cytoplasm (the site of MHV replication), whereas full-length AKAP7 was observed only in nuclei. We suggest the possibility that viral acquisition of the host AKAP7 PDE domain might have occurred during evolution, allowing diverse RNA viruses to antagonize the RNase L pathway. Importance: Early virus-host interactions determine whether an infection is established, highlighting the need to understand fundamental mechanisms regulating viral pathogenesis. Recently, our laboratories reported a novel mode of regulation of the IFN antiviral response. We showed that the coronavirus MHV accessory protein ns2 antagonizes the type I IFN response, promoting viral replication and hepatitis. ns2 confers virulence by cleaving 2',5'-oligoadenylate (2-5A) activators of RNase L in macrophages. We also reported that the rotavirus VP3 C-terminal domain (VP3-CTD) cleaves 2-5A and that it may rescue ns2 mutant MHV. Here we report that a cellular protein, AKAP7, has an analogous 2',5'-phosphodiesterase (2',5'-PDE) domain that is able to restore the growth of chimeric MHV expressing inactive ns2. The proviral effect requires cytoplasmic localization of the AKAP7 PDE domain. We speculate that AKAP7 is the ancestral precursor of viral proteins, such as ns2 and VP3, that degrade 2-5A to evade the antiviral activity of RNase L.
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Project description:Phosphodiesterases (PDEs) encoded by viruses are putatively acquired by horizontal transfer of cellular PDE ancestor genes. Viral PDEs inhibit the OAS-RNase L antiviral pathway, a key effector component of the innate immune response. Although the function of these proteins is well-characterized, the origins of these gene acquisitions are less clear. Phylogenetic analysis revealed at least five independent PDE acquisition events by ancestral viruses. We found evidence that PDE-encoding genes were horizontally transferred between coronaviruses belonging to different genera. Three clades of viruses within Nidovirales: merbecoviruses (MERS-CoV), embecoviruses (HCoV-OC43), and toroviruses encode independently acquired PDEs, and a clade of rodent alphacoronaviruses acquired an embecovirus PDE via recent horizontal transfer. Among rotaviruses, the PDE of rotavirus A was acquired independently from rotavirus B and G PDEs, which share a common ancestor. Conserved motif analysis suggests a link between all viral PDEs and a similar ancestor among the mammalian AKAP7 proteins despite low levels of sequence conservation. Additionally, we used ancestral sequence reconstruction and structural modeling to reveal that sequence and structural divergence are not well-correlated among these proteins. Specifically, merbecovirus PDEs are as structurally divergent from the ancestral protein and the solved structure of human AKAP7 PDE as they are from each other. In contrast, comparisons of rotavirus B and G PDEs reveal virtually unchanged structures despite evidence for loss of function in one, suggesting impactful changes that lie outside conserved catalytic sites. These findings highlight the complex and volatile evolutionary history of viral PDEs and provide a framework to facilitate future studies.
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Project description:Fibroblast growth factors (FGFs) play key roles in the pathogenesis of different human diseases, but the cross-talk between FGFs and other cytokines remains largely unexplored. We identified an unexpected antagonistic effect of FGFs on the interferon (IFN) signaling pathway. Genetic or pharmacological inhibition of FGF receptor signaling in keratinocytes promoted the expression of interferon-stimulated genes (ISG) and proteins in vitro and in vivo. Conversely, FGF7 or FGF10 treatment of keratinocytes suppressed ISG expression under homeostatic conditions and in response to IFN or poly(I:C) treatment. FGF-mediated ISG suppression was independent of IFN receptors, occurred at the transcriptional level and required FGF receptor kinase and proteasomal activity. It is not restricted to keratinocytes and functionally relevant, since FGFs promoted the replication of herpes simplex virus I (HSV-1), lymphocytic choriomeningitis virus and Zika virus. Most importantly, inhibition of FGFR signaling blocked HSV-1 replication in cultured human keratinocytes and in mice. These results suggest the use of FGFR kinase inhibitors for the treatment of viral infections.
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Project description:The 2'-5'-oligoadenylate synthetase (OAS)/RNase L pathway is an innate immune system that protects hosts against pathogenic viruses and bacteria through cleavage of exogenous single-stranded RNA; however, this system's selective targeting mechanism remains unclear. Here, we identified an mRNA quality control factor Dom34 as a novel restriction factor for a positive-sense single-stranded RNA virus. Downregulation of Dom34 and RNase L increases viral replication, as well as half-life of the viral RNA. Dom34 directly binds RNase L to form a surveillance complex to recognize and eliminate the exogenous RNA in a manner dependent on translation. Interestingly, the feature detected by the surveillance complex is not the specific sequence of the viral RNA but the 'exogenous nature' of the RNA. We propose the following model for the selective targeting of exogenous RNA; OAS3 activated by the exogenous RNA releases 2'-5'-oligoadenylates (2-5A), which in turn converts latent RNase L to an active dimer. This accelerates formation of the Dom34-RNase L surveillance complex, and its selective localization to the ribosome on the exogenous RNA, thereby promoting degradation of the RNA. Our findings reveal that the selective targeting of exogenous RNA in antiviral defense occurs via a mechanism similar to that in the degradation of aberrant transcripts in RNA quality control.
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Project description:The identification of a novel β coronavirus, nCoV, as the causative agent of severe respiratory illness in humans originating in Saudi Arabia, Qatar and Jordan has raised concerns about the possibility of a coronavirus pandemic similar to that of SARS-CoV. As a definitive treatment regimen has never been thoroughly evaluated for coronavirus infections, there is an urgent need to rapidly identify potential therapeutics to address future cases of nCoV. To determine an intervention strategy, the effect of interferon-α2b and ribavirin on nCoV isolate hCoV-EMC/2012 replication in Vero and LLC-MK2 cells was evaluated. hCoV-EMC/2012 was sensitive to both interferon-α2b and ribavirin alone in Vero and LLC-MK2 cells, but only at relatively high concentrations; however, when combined, lower concentrations of interferon-α2b and ribavirin achieved comparable endpoints. Thus, a combination of interferon-α2b and ribavirin, which are already commonly used in the clinic, may be useful for patient management in the event of future nCoV infections.
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Project description:Deregulated expression of oncogenes or transcription factors such as specificity protein 1 (Sp1) is observed in many human cancers and plays a role in tumor maintenance. Paradoxically in untransformed cells, Sp1 overexpression induces late apoptosis but the early intrinsic response is poorly characterized. In the present work, we studied increased Sp1 level consequences in untransformed cells and showed that it turns on an early innate immune transcriptome. Sp1 overexpression does not activate known cellular stress pathways such as DNA damage response or endoplasmic reticulum stress, but induces the activation of the OAS-RNase L pathway and the generation of small self-RNAs, leading to the upregulation of genes of the antiviral RIG-I pathway at the transcriptional and translational levels. Finally, Sp1-induced intrinsic innate immune response leads to the production of the chemokine CXCL4 and to the recruitment of inflammatory cells in vitro and in vivo. Altogether our results showed that increased Sp1 level in untransformed cells constitutes a novel danger signal sensed by the OAS-RNase L axis leading to the activation of the RIG-I pathway. These results suggested that the OAS-RNase L-RIG-I pathway may be activated in sterile condition in absence of pathogen.
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Project description:Drugs that reverse epigenetic silencing, such as the DNA methyltransferase inhibitor (DNMTi) 5-azacytidine (AZA), have profound effects on transcription and tumor cell survival. AZA is an approved drug for myelodysplastic syndromes and acute myeloid leukemia, and is under investigation for different solid malignant tumors. AZA treatment generates self, double-stranded RNA (dsRNA), transcribed from hypomethylated repetitive elements. Self dsRNA accumulation in DNMTi-treated cells leads to type I IFN production and IFN-stimulated gene expression. Here we report that cell death in response to AZA treatment occurs through the 2',5'-oligoadenylate synthetase (OAS)-RNase L pathway. OASs are IFN-induced enzymes that synthesize the RNase L activator 2-5A in response to dsRNA. Cells deficient in RNase L or OAS1 to 3 are highly resistant to AZA, as are wild-type cells treated with a small-molecule inhibitor of RNase L. A small-molecule inhibitor of c-Jun NH2-terminal kinases (JNKs) also antagonizes RNase L-dependent cell death in response to AZA, consistent with a role for JNK in RNase L-induced apoptosis. In contrast, the rates of AZA-induced and RNase L-dependent cell death were increased by transfection of 2-5A, by deficiencies in ADAR1 (which edits and destabilizes dsRNA), PDE12 or AKAP7 (which degrade 2-5A), or by ionizing radiation (which induces IFN-dependent signaling). Finally, OAS1 expression correlates with AZA sensitivity in the NCI-60 set of tumor cell lines, suggesting that the level of OAS1 can be a biomarker for predicting AZA sensitivity of tumor cells. These studies may eventually lead to pharmacologic strategies for regulating the antitumor activity and toxicity of AZA and related drugs.
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