Myxoma Virus Protein M029 Is a Dual Function Immunomodulator that Inhibits PKR and Also Conscripts RHA/DHX9 to Promote Expanded Host Tropism and Viral Replication

The vaccinia virus (VACV) E3 protein has been shown to be important for blocking activation of the cellular innate immune system and allowing viral replication to occur unhindered. Mutation or deletion of E3L severely affects viral host range and pathogenesis. While the monkeypox virus (MPXV) genome encodes a homologue of the VACV E3 protein, encoded by the F3L gene, the MPXV gene is predicted to encode a protein with a truncation of 37 N-terminal amino acids. VACV with a genome encoding a similarly truncated E3L protein (VACV-E3L⌬37N) has been shown to be attenuated in mouse models, and infection with VACV-E3L⌬37N has been shown to lead to activation of the host antiviral protein kinase R pathway. In this report, we present data demonstrating that, despite containing a truncated E3 homologue, MPXV phenotypically resembles a wild-type (wt) VACV rather than VACV-E3L⌬37N. Thus, MPXV appears to contain a gene or genes that can suppress the phenotypes associated with an N-terminal truncation in E3. The suppression maps to sequences outside F3L, suggesting that the suppression is extragenic in nature. Thus, MPXV appears to have evolved mechanisms to minimize the effects of partial inactivation of its E3 homologue. IMPORTANCEPoxviruses have evolved to have many mechanisms to evade host antiviral innate immunity; these mechanisms may allow these viruses to cause disease. Within the family of poxviruses, variola virus (which causes smallpox) is the most pathogenic, while monkeypox virus is intermediate in pathogenicity between vaccinia virus and variola virus. Understanding the mechanisms of monkeypox virus innate immune evasion will help us to understand the evolution of poxvirus innate immune evasion capabilities, providing a better understanding of how poxviruses cause disease.

Section: Discussionmentioning

confidence: 99%

Evasion of the Innate Immune Type I Interferon System by Monkeypox Virus

Arndt

Cotsmire

Trainor

et al. 2015

“…However, some sh20 was found in the nucleus, even though this form lacks this sequence, suggesting that OV20.0 has an additional mechanism for nuclear localization. This is reminiscent of myxoma virus M029, a functional ortholog of OV20.0 and E3, which like sh20 is missing around 80 amino acids from the N terminus (50). All poxviruses replicate and transcribe their genomes exclusively in the cytoplasm, which is then the main site of viral dsRNA production.…”

Section: Discussionmentioning

confidence: 99%

Functional Analysis of the Short Isoform of Orf Virus Protein OV20.0

Tseng

Lin

Cheng

et al. 2015

Orf virus (ORFV) OV20.0L is an ortholog of vaccinia virus (VACV) gene E3L. The function of VACV E3 protein as a virulence factor is well studied, but OV20.0 has received less attention. Here we show that like VACV E3L, OV20.0L encodes two proteins, a full-length protein and a shorter form (sh20). The shorter sh20 is an N-terminally truncated OV20.0 isoform generated when a downstream AUG codon is used for initiating translation. These isoforms differed in cellular localization, with full-length OV20.0 and sh20 found throughout the cell and predominantly in the cytoplasm, respectively. Nonetheless, both OV20.0 isoforms were able to bind double-stranded RNA (dsRNA)-activated protein kinase (PKR) and dsRNA. Moreover, both isoforms strongly inhibited PKR activation as shown by decreased phosphorylation of the translation initiation factor eIF2␣ subunit and protection of Sindbis virus infection against the activity of interferon (IFN). In spite of this apparent conservation of function in vitro, a recombinant ORFV that was able to express only the sh20 isoform was attenuated in a mouse model. IMPORTANCEThe OV20.0 protein of orf virus (ORFV) has two isoforms and contributes to virulence, but the roles of the two forms are not known. This study shows that the shorter isoform (sh20) arises due to use of a downstream initiation codon and is amino-terminally truncated. The sh20 form also differs in expression kinetics and cellular localization from full-length OV20.0. Similar to the full-length isoform, sh20 is able to bind dsRNA and PKR, inactivate PKR, and thus act as an antagonist of the interferon response in vitro. In vivo, however, wild-type OV20.0 could not be replaced with sh20 alone without a loss of virulence, suggesting that the functions of the isoforms are not simply redundant. Orf virus (ORFV), a member of the Parapoxvirus genus and the Poxviridae family, is the causative agent of contagious ecthyma in sheep, goats, and other ruminants. The disease is characterized by the development of pustular lesions around the nostrils and mouth with a high incidence rate and a low mortality rate in healthy adult animals. In contrast, infection in immunosuppressed animals or in lambs may be fatal (1). ORFV is also of concern as a source of zoonotic infection because it can cause cutaneous lesions in humans in contact with infected animals. Persistent infection with ORFV can be observed in goats and sheep, and while the severity of lesions is reduced compared with that seen in primary infection, this persistence suggests that the virus is able to evade host immunity (2-4). In line with this observation, ORFV has been shown to encode several proteins that modulate the host response to infection. These include viral homologues of ovine cytokines, such as vascular endothelial growth factor, interleukin-10 (IL-10), and a granulocyte-macrophage colony-stimulating factor (GM-CSF)-inhibiting protein, as well as an apoptosis inhibitor (5-7). ORFV also antagonizes interferon (IFN) signaling, and this is done by the product...

“…Interestingly, this mutated sequence has reverted to the original sequence twice on independent branches of the tree and in viruses isolated from widely separated geographic regions. M029 has been shown to function similarly to VACV E3, binding double-stranded RNA, inhibiting protein kinase R (PKR) activation, and inhibiting IFN-␤, tumor necrosis factor alpha (TNF-␣), and interleukin-6 (IL-6) expression (45); it is a critical virulence factor in rabbit infections but also has a second function of binding RNA helicase A (RHA; DHX9), which promotes virus replication in some cell lines, so it has been described as a dual-function virulence and host range factor (46). The amino acid sequence from the related leporipoxvirus RFV is conserved from amino acid 1 to 8 and is identical to that from MYXV from amino acid 20 to 55 but is poorly conserved between residues 9 and 19; this region is also divergent in the Californian MSW strain of MYXV (47).…”

Section: Pathways To Attenuation and Virulence In Australian Isolatesmentioning

confidence: 99%

Genome Scale Evolution of Myxoma Virus Reveals Host-Pathogen Adaptation and Rapid Geographic Spread

Kerr

Rogers

Fitch

et al. 2013

The evolutionary interplay between myxoma virus (MYXV) and the European rabbit (Oryctolagus cuniculus) following release of the virus in Australia in 1950 as a biological control is a classic example of host-pathogen coevolution. We present a detailed genomic and phylogeographic analysis of 30 strains of MYXV, including the Australian progenitor strain Standard Laboratory Strain (SLS), 24 Australian viruses isolated from 1951 to 1999, and three isolates from the early radiation in Britain from 1954 and 1955. We show that in Australia MYXV has spread rapidly on a spatial scale, with multiple lineages cocirculating within individual localities, and that both highly virulent and attenuated viruses were still present in the field through the 1990s. In addition, the detection of closely related virus lineages at sites 1,000 km apart suggests that MYXV moves freely in geographic space, with mosquitoes, fleas, and rabbit migration all providing means of transport. Strikingly, despite multiple introductions, all modern viruses appear to be ultimately derived from the original introductions of SLS. The rapidity of MYXV evolution was also apparent at the genomic scale, with gene duplications documented in a number of viruses. Duplication of potential virulence genes may be important in increasing the expression of virulence proteins and provides the basis for the evolution of novel functions. Mutations leading to loss of open reading frames were surprisingly frequent and in some cases may explain attenuation, but no common mutations that correlated with virulence or attenuation were identified.