Severe Acute Respiratory Syndrome Coronavirus Evades Antiviral Signaling: Role of nsp1 and Rational Design of an Attenuated Strain

Abstract

The severe acute respiratory syndrome (SARS) epidemic was caused by the spread of a previously unrecognized infectious agent, the SARS-associated coronavirus (SARS-CoV). Here we show that SARS-CoV could inhibit both virus-and interferon (IFN)-dependent signaling, two key steps of the antiviral response. We mapped a strong inhibitory activity to SARS-CoV nonstructural protein 1 (nsp1) and show that expression of nsp1 significantly inhibited the activation of all three virus-dependent signaling pathways. We show that expression of nsp1 significantly inhibited IFN-dependent signaling by decreasing the phosphorylation levels of STAT1 while having little effect on those of STAT2, JAK1, and TYK2. We engineered an attenuated mutant of nsp1 in SARS-CoV through reverse genetics, and the resulting mutant virus was viable and replicated as efficiently as wild-type virus in cells with a defective IFN response. However, mutant virus replication was strongly attenuated in cells with an intact IFN response. Thus, nsp1 is likely a virulence factor that contributes to pathogenicity by favoring SARS-CoV replication.Vertebrates have evolved sophisticated immunity mechanisms to manage infections by pathogens. The first line of defense is the innate immune response, which is initiated by the rapid recognition of pathogen-associated molecular patterns. In most cell types, viral RNA is sensed by RNA helicases (18) and triggers three signaling pathways, leading to the coordinated activation of the transcription factors ATF2/c-Jun, IRF3/IRF7, and NF-B (25). In addition, double-stranded RNA and other viral determinants are recognized by Toll-like receptors expressed by specialized cells involved in the immune response. Signaling downstream of the Toll-like receptors involves various adaptor molecules and ultimately results in the activation of the same or a closely related set of transcription factors to those activated in the generic response (reviewed in reference 18).Virus-activated transcription factors synergize to induce the production of a number of cytokines, including members of the type I interferon (IFN) family (which in humans comprises 14 IFN-␣ genes, 1 IFN-gene, and 1 IFN-␤ gene) and the type III IFN family (three human IFN-genes); inflammatory cytokines, such as interleukin-1 (IL-1), IL-6, IL-12, tumor necrosis factor alpha (TNF-␣), and TNF-␤; and chemokines, such as RANTES and IL-8. Together, these cytokines signal the occurrence of the infection and orchestrate the innate immune response directed against the invading virus (2, 33, 37).Binding of type I or type III IFNs to their cognate receptors triggers a second wave of signaling, where phosphorylation and activation of the receptor-associated JAK1 and TYK2 kinases result in the phosphorylation and activation of the transcription factors STAT1 and STAT2. Activated STAT1/STAT2 forms a complex with IRF9, termed ISGF3, which activates the transcription of a set of IFN-inducible genes by binding to their IFN-stimulated response elements (ISREs) (1,33,37).The functional cons...

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“…This probably illustrates the need for a potent inhibition of IFN signaling for the life cycle of this group of viruses. Similar multiple mechanisms have also been described for other viruses such as paramyxoviruses and coronaviruses (16,29,40,54). The knowledge gained by characterizing the various mechanisms through which viruses evade IFN paves the way for the targeted attenuation of pathogenic viruses, which may be used as potential live vaccines.…”

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