NSs protein of Sandfly fever Sicilian phlebovirus counteracts interferon induction by masking the DNA-binding domain of interferon regulatory factor 3.

Sandfly fever Sicilian virus (SFSV) is one of the most widespread and frequent members of the genus Phlebovirus (order Bunyavirales , family Phenuiviridae ) infecting humans. Being transmitted by Phlebotomus sandflies, SFSV causes a self-limiting acute, often incapacitating febrile disease ("sandfly fever", "pappataci fever" or "dog disease") that is known at least since the beginning of the 20th century. We show that, similar to other pathogenic phleboviruses, SFSV suppresses the induction of the antiviral type I interferon (IFN) system in an NSs-dependent manner. SFSV NSs interfered with the TBK1-IRF3 branch of the RIG-I signalling pathway, but not with NF-κB activation. Consistently, we identified interferon regulatory factor 3 (IRF3) as host interactor of SFSV NSs. In contrast to IRF3, neither the IFN master regulator IRF7, nor the related transcription factors IRF2, IRF5, or IRF9 were bound by SFSV NSs. In spite of this specificity for IRF3, NSs inhibited neither its phosphorylation, dimerization, nor nuclear accumulation, and the interaction was independent of the IRF3 activation or multimerization state. In further studies we identified the DNA-binding domain of IRF3 (amino acids 1 to 113) as sufficient for NSs binding, and found that SFSV NSs prevented the association of activated IRF3 with the IFN-β promoter. Thus, unlike highly virulent phleboviruses which either destroy antiviral host factors or sequester whole signalling chains into inactive aggregates, SFSV modulates type I IFN induction by directly masking the DNA-binding domain of IRF3. Significance Phleboviruses are receiving increased attention due to the constant discovery of new species, and the ongoing spread of long-known members of the genus. Outbreaks of Sandfly fever were reported in the nineteenth century, in World War I, and in World War II. Nowadays, SFSV is recognized as being one of the most widespread phleboviruses, exhibiting high seroprevalence rates in humans and domestic animals and causing a self-limiting but incapacitating disease predominantly in immunologically naïve troops and travellers. We show how the non-structural protein NSs of SFSV counteracts the upregulation of the antiviral interferon (IFN) system. SFSV NSs specifically inhibits promoter binding by the IFN transcription factor IRF3, a molecular strategy which is unique among phleboviruses and, to our knowledge, human pathogenic RNA viruses in general. This IRF3-specific and stoichiometric mechanism, much distinct from the ones exhibited by the highly virulent phleboviruses, correlates with the intermediate level of pathogenicity of SFSV.