Spring Viremia of Carp Virus N Protein Suppresses Fish IFNφ1 Production by Targeting the Mitochondrial Antiviral Signaling Protein

Abstract

For a virus to replicate efficiently, it must try and inhibit host IFN expression because IFN is an important host defense at early stages after viral infection. For aquatic viruses, the mechanisms used to escape the hosts IFN system are still unclear. In this study, we show that the N protein of spring viremia of carp virus (SVCV) inhibits zebrafish IFNφ1 production by degrading the mitochondrial antiviral signaling protein (MAVS). First, the upregulation of IFNφ1 promoter activity stimulated by polyinosinic:polycytidylic acid, retinoic acid–inducible gene I (RIG-I) or MAVS was suppressed by the SVCV infection. However, the upregulation by the downstream factor of the RIG-I–like receptor signaling pathway, TANK-binding kinase 1, was not affected. Notably, at the protein level, MAVS decreased remarkably when cells were infected with SVCV. Second, consistent with the result of the SVCV infection, overexpression of the N protein of SVCV blocked the IFNφ1 transcription activated by MAVS and downregulated MAVS expression at the protein level but not at the mRNA level. Further analysis demonstrated that the N protein targeted MAVS for K48-linked ubiquitination, which promoted the degradation of MAVS. These data indicated that fish MAVS could be degraded by the N protein of SVCV through the ubiquitin-proteasome pathway. To our knowledge, this is the first article of a fish RIG-I–like receptor pathway interfered by an aquatic virus in an ubiquitin-proteasome manner, suggesting that immune evasion of a virus also exists in lower vertebrates.

Zebrafish foxo3b Negatively Regulates Antiviral Response through Suppressing the Transactivity of irf3 and irf7

“…Zebrafish IFNw1 and IFNw3 promoter luciferase reporters and the EPC IFN promoter luciferase reporter could be induced by poly(I:C), whereas zebrafish IFNw2 and IFNw4 promoter luciferase reporters could not be induced by poly(I:C) (36). These inductions are validated in Fig.…”

“…It has been reported that similar to that of mammalian species, the fish RLR signaling cascade can activate IFN expression (36). To determine whether zebrafish foxo3b also inhibits IFN expression through its effect on RLR signaling pathway, we examined the activity of ISRE-luc, IFNw1-luc, IFNw3-luc, and EPC-IFN-luc stimulated by overexpression of cascaded factors in the RLR signaling pathway, including the zebrafish N-terminal domain of rig-1 (rig-1-Nter), mavs, tbk1, irf3, or irf7 together with empty Myc vector or Myc-tagged foxo3b.…”

“…The plasmids encoding rig-1-Nter and tbk1 in the pcDNA3.1(+) vector (Invitrogen) and the plasmids containing Dr-IFNw1-luc, Dr-IFNw2-luc, Dr-IFNw3-luc, Dr-IFNw4-luc, and EPC-IFN-luc in the pGL3-Basic vector (Promega) were constructed as described previously (36,37). The ISRE luciferase reporter construct (ISRE-luc) containing five ISRE motifs in a series and the pFR-luc reporter construct were purchased from Stratagene.…”

“…luc) of zebrafish and the EPC IFN promoter luciferase reporter have previously been used for detecting poly(I:C) stimulation in EPC cells (36). Zebrafish IFNw1 and IFNw3 promoter luciferase reporters and the EPC IFN promoter luciferase reporter could be induced by poly(I:C), whereas zebrafish IFNw2 and IFNw4 promoter luciferase reporters could not be induced by poly(I:C) (36).…”

Fish IRF6 is a positive regulator of IFN expression and involved in both of the MyD88 and TBK1 pathways

“…To generate the EGFP-IRF6 expression plasmid, the cDNA fragment encoding IRF6 was cloned into pEGFP-N3 vector (BD Clontech). The plasmids containing IFN41pro-Luc (NM_207640.1), IFN43pro-Luc (NM_001111083.1), and ISRE-Luc in pGL3-Basic luciferase reporter vectors (Promega) were constructed as described previously [28]. All constructs were confirmed by DNA sequencing.…”