Modulation of the Myxoma Virus Plaque Phenotype by Vaccinia Virus Protein F11

Abstract

Vaccinia virus (VACV) produces large plaques consisting of a rapidly expanding ring of infected cells surrounding a lytic core, whereas myxoma virus (MYXV) produces small plaques that resemble a focus of transformed cells. This is odd, because bioinformatics suggests that MYXV carries homologs of nearly all of the genes regulating Orthopoxvirus attachment, entry, and exit. So why does MYXV produce foci? One notable difference is that MYXV-infected cells produce few of the actin microfilaments that promote VACV exit and spread. This suggested that although MYXV carries homologs of the required genes (A33R, A34R, A36R, and B5R), they are dysfunctional. To test this, we produced MYXV recombinants expressing these genes, but we could not enhance actin projectile formation even in cells expressing all four VACV proteins. Another notable difference between these viruses is that MYXV lacks a homolog of the F11L gene. F11 inhibits the RhoA-mDia signaling that maintains the integrity of the cortical actin layer. We constructed an MYXV strain encoding F11L and observed that, unlike wild-type MYXV, the recombinant virus disrupted actin stress fibers and produced plaques up to 4-fold larger than those of controls, and these plaques expanded ϳ6-fold faster. These viruses also grew to higher titers in multistep growth conditions, produced higher levels of actin projectiles, and promoted infected cell movement, although neither process was to the extent of that observed in VACV-infected cells. Thus, one reason for why MYXV produces small plaques is that it cannot spread via actin filaments, although the reason for this deficiency remains obscure. A second reason is that leporipoxviruses lack vaccinia's capacity to disrupt cortical actin. P oxviruses produce two kinds of plaques in culture. The first kind is formed rapidly by viruses like vaccinia virus (VACV), and typically they comprise a ring of infected cells surrounding a large central clearing or lytic zone. The second type of poxvirus plaque is smaller, grows slower, and consists of a clump of virusinfected cells. These plaques look more like a cluster of transformed cells and are sometimes called foci. Such plaques are produced by tumorigenic poxviruses like the leporipoxviruses myxoma virus (MYXV) and Shope fibroma virus. Although it is well established that the appearance of poxvirus plaques is determined by the genetics of both the host and the virus, why MYXV plaques look so different from VACV plaques, even when plated on the same cell type, is not well understood.The process of plaque formation depends (in part) upon how well viruses can engage the host machinery to spread efficiently from cell to cell, processes that are best understood for VACV (reviewed in references 44, 51, and 58). VACV characteristically produces several different forms of infectious virus. Mature viruses (MV) are bounded by just a single lipid bilayer. MV comprise the most abundant infectious form and are probably released by cell lysis. However, some MV migrate away from viral factories, w...

The role of signalling and the cytoskeleton during Vaccinia Virus egress

“…Lastly, by stimulating infected cell migration and the formation of long neurite-like projections, F11 enhances the spread of infection by increasing viral contact with more non-infected cells (Cordeiro et al, 2009;Valderrama et al, 2006). The ability of F11 to promote viral spread has also been elegantly illustrated using Myxoma virus, which lacks a related homologue (Irwin and Evans, 2012). Irwin et al found that a recombinant Myxoma virus expressing F11 is more effective in its cell-to-cell spread, as plaques expand ∼6 fold faster and are four times larger than controls after 4 days (Irwin and Evans, 2012).…”

“…The ability of F11 to promote viral spread has also been elegantly illustrated using Myxoma virus, which lacks a related homologue (Irwin and Evans, 2012). Irwin et al found that a recombinant Myxoma virus expressing F11 is more effective in its cell-to-cell spread, as plaques expand ∼6 fold faster and are four times larger than controls after 4 days (Irwin and Evans, 2012). The expression of F11 also improves the oncolytic efficacy of Myxoma by increasing viral spread in tumours and prolonging the survival of mice (Irwin et al, 2013).…”

A Whole-Genome RNA Interference Screen for Human Cell Factors Affecting Myxoma Virus Replication

“…However, it is notable that knocking down LIMK2, which would be expected to destabilize the actin cytoskeleton (99), and DIAPH1, which would disrupt adherens junctions (100), also promotes MYXV growth. We recently showed that MYXV has difficulties in disrupting the cortical actin layer during exit, and this can be partially ameliorated by incorporating the VACV F11L gene (101). This provides some rationale in support for the later structural hypothesis, although the two biological effects are not mutually exclusive.…”

Oncolytic Poxviruses

“…However, in a murine model bearing transplanted human gliomas on both sides of the brain, single intratumoral injection of MYXV could eliminate the tumor mass only in the hemisphere in which MYXV was administered, indicating that the virus did not efficiently spread to the tumors in the contralateral hemisphere (102). The VV F11 protein promotes virus exit from an infected cell by inhibiting Rho signaling, which maintains the integrity of the cortical actin layer (112). Irwin & Evans (112) showed that a recombinant MYXV expressing the VV F11L gene (MYXV-F11L) replicates more efficiently than wild-type MYXV in monkey or rabbit cell lines.…”

“…The VV F11 protein promotes virus exit from an infected cell by inhibiting Rho signaling, which maintains the integrity of the cortical actin layer (112). Irwin & Evans (112) showed that a recombinant MYXV expressing the VV F11L gene (MYXV-F11L) replicates more efficiently than wild-type MYXV in monkey or rabbit cell lines. In a variety of human cancerous cell lines, MYXV-F11L replicates more efficiently than control viruses (113).…”