RESUMO
OBJECTIVE: To determine the contribution of MX dynamin, oligoadenylate synthetase (OAS), and double-stranded RNA-dependent protein kinase R (PKR) to the antiviral effects of type 1 interferons (IFNs) against bovine parainfluenza-3 virus (PI-3V) infection of Vero cells. SAMPLE POPULATION: Vero cell cultures. PROCEDURES: PI-3V yield was first compared between control and transfected type 1 IFNs-incompetent Vero cells expressing recombinant OAS or MX proteins. Afterwards, phosphorylation of eukaryotic initiation factor 2 alpha (eIF2alpha) was used to scale the degree of PKR activation upon infection of Vero cells by PI-3V. RESULTS: Overexpression of OAS did not result in significantly decreased viral replication. Phosphorylated eIF2alpha forms, the hallmark of PKR activation, were not increased in IFNalpha-primed infected Vero cells. Although human MXA contributed to partial blockade of replication of bovine PI-3V, the antiviral effect was not as strong as that of IFNalpha. CONCLUSIONS AND CLINICAL RELEVANCE: The powerful anti-Paramyxovirus activity of type 1 IFNs is mediated by noncanonic pathways.
Assuntos
2',5'-Oligoadenilato Sintetase/metabolismo , Doenças dos Bovinos/virologia , Dinaminas/metabolismo , Interferon-alfa/farmacologia , Vírus da Parainfluenza 3 Bovina/efeitos dos fármacos , Infecções por Respirovirus/veterinária , eIF-2 Quinase/metabolismo , Animais , Bovinos , Doenças dos Bovinos/imunologia , Chlorocebus aethiops , Testes Imunológicos de Citotoxicidade/veterinária , Citometria de Fluxo/veterinária , Técnica Indireta de Fluorescência para Anticorpo/veterinária , Formazans/química , Proteínas de Ligação ao GTP/biossíntese , Proteínas de Ligação ao GTP/genética , Proteínas de Ligação ao GTP/imunologia , Immunoblotting/veterinária , Interferon beta/farmacologia , Proteínas de Resistência a Myxovirus , Vírus da Parainfluenza 3 Bovina/enzimologia , Vírus da Parainfluenza 3 Bovina/imunologia , Infecções por Respirovirus/imunologia , Infecções por Respirovirus/virologia , Sais de Tetrazólio/química , Transfecção/veterinária , Células Vero , Replicação Viral/efeitos dos fármacosRESUMO
Bovine parainfluenza virus type 3 (bPIV3) is under development as a live virus vaccine vector. The RNA genome of a recombinant bPIV3 harbored four nucleotide changes, one of which resulted in a mutation of the viral polymerase (A. A. Haller et al., 2000, J. Virol. 74, 11626-11635). The contribution of this conservative amino acid substitution (I1103V) in the polymerase to the temperature-sensitive and attenuation phenotypes of r-bPIV3 was investigated by creating a new virus, r-bPIV3(I), that expressed the wild-type polymerase. r-bPIV3(I) was not temperature-sensitive for growth in vitro and the replication of r-bPIV3(I) was no longer restricted in hamsters. The effect of the amino acid substitution in the polymerase was also studied in a chimeric bovine/human PIV3, a virus that displayed temperature-sensitive and attenuated phenotypes (A. A. Haller et al., 2000, J. Virol. 74, 11626-11635). It was not clear whether these defects were due to the impaired polymerase or the replacement of the bPIV3 surface glycoproteins with those of hPIV3. The results showed that the altered polymerase was indeed responsible for the temperature-sensitive phenotype of bovine/human PIV3 but did not appear to play a role in the attenuation phenotype.
