The VP3 Factor from Viruses of Birnaviridae Family Suppresses RNA Silencing by Binding Both Long and Small RNA Duplexes

Abstract

RNA silencing is directly involved in antiviral defense in a wide variety of eukaryotic organisms, including plants, fungi, invertebrates, and presumably vertebrate animals. The study of RNA silencing-mediated antiviral defences in vertebrates is hampered by the overlap with other antiviral mechanisms; thus, heterologous systems are often used to study the interplay between RNA silencing and vertebrate-infecting viruses. In this report we show that the VP3 protein of the avian birnavirus Infectious bursal disease virus (IBDV) displays, in addition to its capacity to bind long double-stranded RNA, the ability to interact with double-stranded small RNA molecules. We also demonstrate that IBDV VP3 prevents the silencing mediated degradation of a reporter mRNA, and that this silencing suppression activity depends on its RNA binding ability. Furthermore, we find that the anti-silencing activity of IBDV VP3 is shared with the homologous proteins expressed by both insect- and fish-infecting birnaviruses. Finally, we show that IBDV VP3 can functionally replace the well-characterized HCPro silencing suppressor of Plum pox virus, a potyvirus that is unable to infect plants in the absence of an active silencing suppressor. Altogether, our results support the idea that VP3 protects the viral genome from host sentinels, including those of the RNA silencing machinery.

Exacerbated Apoptosis of Cells Infected with Infectious Bursal Disease Virus upon Exposure to Interferon Alpha

“…Moreover, the recognition of naked IBDV dsRNA by the RNA cytoplasmic sensor MDA5 in DF-1 cells was found to activate chIFN-␤ expression (59), but it was also described that VP3 interferes with this recognition (40). This is consistent with the notion that the binding of VP3 to IBDV dsRNA shields the virus genome from host RNA sensors, thus preventing PKR-mediated apoptosis (34,71). Regarding this, it is interesting to note that a stoichiometric analysis of IBDV structural components showed that infectious IBDV particles harbor four dsRNA genome segments associated (in the form of RNPs) with ca.…”

Mosquito andDrosophilaentomobirnaviruses suppress dsRNA- and siRNA-induced RNAi

“…Even a dsRNA-binding protein from E. coli , a species that is not targeted by an RNAi response, is able to suppress RNAi under overexpression conditions (53). It has been shown previously that the IBDV, IPNV and DXV VP3 proteins have the capacity to suppress RNAi (54). However, these studies were done in a heterologous system and the in vivo relevance in a relevant host remained unclear.…”

Inhibition of Antiviral Innate Immunity by Birnavirus VP3 Protein via Blockage of Viral Double-Stranded RNA Binding to the Host Cytoplasmic RNA Detector MDA5

“…VP3 was previously reported to bind IBDV genomic dsRNA to protect the viral genome from the host RNA silencing machinery and inhibit PKR-mediated apoptosis (18,21). To investigate whether viral proteins suppressed IFN-␤ expression by binding to naked IBDV genomic dsRNA, we identified the viral proteins binding to dsRNA and examined IFN-␤ expression under these conditions.…”

“…5D, ectopically expressed VP3 protein from IPNV and DXV also bound dsRNA. The basic amino acid motif 99 K 102 R 105 K 106 K in the ␣1 helix of VP3 has been shown to be responsible for binding small duplex RNA (18,21), so we speculated that this region may be involved in the inhibition of chIFN-␤ expression. Figure 5E indicates that the VP3 patch 1 mutant, in which the basic amino acids 99 K, 102 R, 105 K, and 106 K are replaced by acidic amino acid D, had almost no dsRNA binding activity, and a further 97 R-to-D substitution had no additional effect on dsRNA binding activity.…”