RESUMO
Interleukin-18 (IL-18) is a critical proinflammatory cytokine whose extracellular bioactivity is regulated by a cellular IL-18 binding protein (IL-18BP). Many poxviruses have acquired variants of this IL-18BP gene, some of which have been shown to act as viral virulence factors. Yaba monkey tumor virus (YMTV) encodes a related family member, 14L, which is similar to the orthopoxvirus IL-18BPs. YMTV 14L was expressed from a baculovirus system and tested for its ability to bind and inhibit IL-18. We found that YMTV 14L bound both human IL-18 (hIL-18) and murine IL-18 with high affinity, at 4.1 nM and 6.5 nM, respectively. YMTV 14L was able to fully sequester hIL-18 but could only partially inhibit the biological activity of hIL-18 as measured by gamma interferon secretion from KG-1 cells. Additionally, 17 hIL-18 point mutants were tested by surface plasmon resonance for their ability to bind to YMTV 14L. Two clusters of hIL-18 surface residues were found to be important for the hIL-18-YMTV 14L interaction, in contrast to results for the Variola virus IL-18BP, which has been shown to primarily interact with a single cluster of three amino acids. The altered binding specificity of YMTV 14L most likely represents an adaptation resulting in increased fitness of the virus and affirms the plasticity of poxviral inhibitor domains that target cytokines like IL-18.
Assuntos
Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Interleucina-18/antagonistas & inibidores , Proteínas Virais/fisiologia , Vírus do Tumor do Macaco de Yaba/imunologia , Linhagem Celular , Humanos , Interferon gama/biossíntese , Mutagênese Sítio-Dirigida , Mutação Puntual , Ligação Proteica , Mapeamento de Interação de Proteínas , Ressonância de Plasmônio de SuperfícieRESUMO
Myxoma virus (MV) is a highly lethal, rabbit-specific poxvirus that induces a disease called myxomatosis in European rabbits. In an effort to understand the function of predicted immunomodulatory genes we have deleted various viral genes from MV and tested the ability of these knockout viruses to induce lethal myxomatosis. MV encodes a unique 15 kD cytoplasmic protein (M130R) that is expressed late (12h post infection) during infection. M130R is a non-essential gene for MV replication in rabbit, monkey or human cell lines. Construction of a targeted gene knockout virus (vMyx130KO) and infection of susceptible rabbits demonstrate that the M130R knockout virus is attenuated and that loss of M130R expression allows the rabbit host immune system to effectively respond to and control the lethal effects of MV. M130R expression is a bona fide poxviral virulence factor necessary for full and lethal development of myxomatosis.
Assuntos
Myxoma virus/patogenicidade , Proteínas Virais/fisiologia , Fatores de Virulência/fisiologia , Sequência de Aminoácidos , Animais , Linhagem Celular , Deleção de Genes , Técnicas de Inativação de Genes , Ordem dos Genes , Haplorrinos , Humanos , Masculino , Dados de Sequência Molecular , Myxoma virus/genética , Filogenia , Coelhos , Alinhamento de Sequência , Índice de Gravidade de Doença , Proteínas Virais/genética , Virulência , Fatores de Virulência/genéticaRESUMO
The authors have recently demonstrated that wild-type myxoma virus (MV) tagged with gfp (vMyxgfp) can generate a tumor-specific infection that productively infects and clears human tumor-derived xenografts when injected intratumorally into human gliomas transplanted into immunodeficient mice (Lun et al, 2005). To expand the understanding of MV tropism in cancer cells from a specific tissue lineage, the authors have screened a series of human glioma cells (U87, U118, U251, U343, U373) for myxoma virus replication and oncolysis. To assess the viral tropism determinants for these infections, the authors have screened myxoma virus knockout constructs that have been deleted for specific host range genes (M-T2, M-T4, M-T5, M11L, and M063), as well as a control MV gene knockout construct with no known host range function (vMyx135KO) but is highly attenuated in rabbits. The authors report wide variation in the ability of various vMyx-hrKOs to replicate and spread in the human glioma cells as measured by early and late viral gene expression. This differential ability to support vMyx-hrKO productive viral replication is consistent with levels of endogenous activated Akt in the various gliomas. The authors have identified one vMyx-hrKO virus (vMyx63KO) and one nonhost range knockout construct (vMyx135KO) that appear to replicate in the gliomas even more efficiently than the wild-type virus and that reduce the viability of the infected gliomas. These knockout viruses also inhibit the proliferation of gliomas in a manner similar to the wild-type virus. Together these data, as well as the fact that these knockout viruses are attenuated in their natural hosts, may represent environmentally safer candidate oncolytic viruses for usage in human trials.