Deletion analysis of two tandemly arranged virulence genes in myxoma virus, M11L and myxoma growth factor

Abstract

Myxoma virus (MYX) is a leporipoxvirus of rabbits that induces a lethal syndrome characterized by disseminated tumorlike lesions, generalized immunosuppression, and secondary gram-negative bacterial infection. A MYX deletion mutant (vMYX-GF-AM11L) was constructed to remove the entire myxoma growth factor (MGF) coding sequence and that for the C-terminal five amino acids of the partially overlapping upstream gene, M1lL. Unexpectedly, this deletion completely abrogates the capacity of MYX to cause the characteristic disease symptoms of myxomatosis. Upon inoculation of rabbits with vMYX-GF-AM1lL, recipient animals developed only a benign, localized nodule reminiscent of a Shope fibroma virus-induced tumor in which a single primary lesion appeared at the site of injection and then completely regressed within 14 days, leaving the animals resistant to challenge with wild-type MYX. No evidence of the purulent

Mitochondria and viruses

“…Myxoma virus (MXV) is a rabbit leporipoxvirus that induces a lethal syndrome characterized by disseminated tumor-like lesions and generalized immunosuppression. M11L, a protein in the mitochondrial outer membrane, has been shown to inhibit apoptosis by binding to the MMP-inducible, pore-forming proteins Bax and Bak (Opgenorth et al, 1992;Wang et al, 2004). Structural analysis has revealed that M11L is a Bcl-2 homolog that binds to Bax and Bak with affinities similar to those observed for cellular Bcl-2 and Bcl-xL (Douglas et al, 2007).…”

Punctuated evolution of myxoma virus: rapid and disjunct evolution of a recent viral lineage in Australia

“…These non-synonymous substitutions include an amino acid change in the M010L gene (A66V). M010 is a homologue of cellular epidermal growth factor/transforming growth factor alpha and is critical for virulence (36, 37). Interestingly, this is the only amino acid substitution in M010 we have seen in Australian isolates, although a valine is also found at this position in the Californian MYXV strain MSW and rabbit fibroma virus.…”

M11l

“…Recombinant viruses used in this study have been described previously. These include vMyxlac, a control myxoma virus that produces M11L, vMyxM11L Ϫ which fails to produce M11L owing to a targeted gene disruption (23), and vMyxM11L R , a revertant virus in which the gene disruption has been repaired (20). A vaccinia virus construct that overexpresses M11L, VVM11L (22), and a control vaccinia virus, VV601, were also employed.…”

“…The adherent cells were detached from the plastic support using warm 1 ϫ SSC (150 mM sodium chloride, 15 mM sodium citrate, pH 7.5) and infected with appropriate viruses at an MOI of 10. Typically, infection rates exceeded 70% as verified by staining with X-gal (23). The infected cells were cultured in medium containing 20% FBS for 12 h, fixed in 2% paraformaldehyde/PBS, and apoptotic cells were detected using the TUNEL reaction.…”

“…Despite its reduced virulence, however, the lesions produced by the M11L knockout virus were unusual, and histological analysis revealed signs of vigorous inflammatory activity. The knockout virus was also shown to be impaired in its ability to replicate in primary rabbit splenocytes (23). The attributes of M11L suggested a model in which this protein could act as a virulence factor by preventing apoptosis of leukocytes during host infection, thus compromising the effectiveness of cellular protective mechanisms designed to limit viral propagation.…”