RESUMEN
Cats were experimentally infected with cell culture-adapted feline foamy virus (FFV, spumaretrovirinae) isolate FUV. FFV was consistently recovered from peripheral blood leukocytes and throat samples of FFV-infected cats starting 2 to 3 weeks postinfection (p. i.), indicative of the establishment of persistent FFV infections. Viral persistence was established, even despite neutralizing antibodies that appeared early after infection. The humoral immune response toward FFV was quantitatively and qualitatively analyzed over time. FFV Gag-specific antibodies were first detected 2 weeks p. i. and increased further; reactivities to the other structural and nonstructural FFV proteins appeared slightly delayed. Reactivities against FFV Pol and Gag proteins were detectable by immunoblotting and radioimmunoprecipitation, whereas the latter techniques had to be employed for the unambiguous detection of FFV Env-, Bet-, and Bel 1-specific antibodies.
Asunto(s)
Gatos/virología , Infecciones por Retroviridae/inmunología , Infecciones por Retroviridae/virología , Spumavirus/inmunología , Spumavirus/fisiología , Replicación Viral , Animales , Anticuerpos Antivirales/inmunología , Western Blotting , Gatos/inmunología , Línea Celular , Células Cultivadas , Cinética , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/virología , Boca/virología , Pruebas de Neutralización , Ensayo de Radioinmunoprecipitación , Infecciones por Retroviridae/veterinaria , Spumavirus/química , Proteínas Virales/análisis , Proteínas Virales/biosíntesisRESUMEN
Full-length genomes of the feline foamy virus (FFV or FeFV) isolate FUV were constructed. DNA clone pFeFV-7 stably directed the expression of infectious FFV progeny virus indistinguishable from wild-type, uncloned FFV isolate FUV. The env and bel 1 genes of pFeFV-7 were substituted for by corresponding sequences of the FFV serotype 951 since previous studies implicated a defined part of FFV Env protein as responsible for serotype-specific differences in serum neutralization (I. G. Winkler, R. M. Flügel, M. Löchelt, and R. L. P. Flower, 1998. Virology 247: 144-151). Recombinant virus derived from chimeric plasmid pFeFV-7/951 containing the hybrid env gene and the parental clone pFeFV-7 were used for neutralization studies. By means of a rapid titration assay for FFV infectivity, we show that progeny virus derived from plasmid pFeFV-7 was neutralized by FUV- but not by 951-specific antisera, whereas pFeFV-7/951-derived chimeric virus was neutralized by 951-specific antisera only. Both recombinant proviruses will be useful for repeated delivery of foreign genes for therapeutic gene applications into cats.
Asunto(s)
Anticuerpos Antivirales/inmunología , Genes env , Genoma Viral , Spumavirus/genética , Proteínas del Envoltorio Viral/inmunología , Secuencia de Aminoácidos , Animales , Gatos , Proteínas de Unión al ADN/genética , Datos de Secuencia Molecular , Pruebas de Neutralización , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/inmunología , Infecciones por Retroviridae/inmunología , Infecciones por Retroviridae/virología , Proteínas de los Retroviridae/genética , Análisis de Secuencia de ADN , Spumavirus/inmunología , Transactivadores/genética , Proteínas del Envoltorio Viral/genéticaRESUMEN
Previous results have indicated that incorporation of surface glycoprotein into retroviral particles is not a specific process and that many heterologous viral and cellular glycoproteins can be incorporated as long as they do not have long cytoplasmic C-terminal regions which were presumed to be sterically inhibitory. In this study, this concept has been directly examined by analyzing the incorporation of the wild-type human epidermal growth factor receptor (Wt-EGFR) and of a C-terminally truncated mutant of Wt-EGFR (Tr-EGFR) into human immunodeficiency virus (HIV)-like particles. Incorporation was directly analyzed at the protein level and by immunogold labelling of enriched HIV-like particles. In agreement with the above concept, Tr-EGFR, with only 7 C-terminal amino acids (aa), was efficiently incorporated into HIV-like particles. Incorporation of the Wt-EGFR species, with 542 C-terminal cytoplasmic aa, was reduced by a factor of about 5 in comparison to that of the Tr-EGFR species. However, the Wt-EGFR species was still very significantly present in the HIV-like particles. A series of control experiments verified that this represents genuine incorporation of Wt-EGFR into the membrane of HIV-like particles. These observations allow further speculation as to the processes governing glycoprotein incorporation into retroviral particles and indicate that the internal virus structure of HIV (in particular the matrix layer [MA]) can accommodate much larger heterologous cytoplasmic domains in incorporated glycoproteins than previously assumed.