RESUMO
To identify T-helper (Th)-cell epitopes, we analyzed 25 synthetic peptides, which included most of the 495-amino-acid sequence of the envelope (E)-glycoprotein of dengue 2 virus. The peptides were analyzed in three mouse strains, BALB/c (H-2d), C57BL/6 (H-2b), and outbred NIH-Swiss, for their ability to elicit antibody or prime the Th-cell compartment following two inoculations in Freund's incomplete adjuvant. Sixteen peptides were able to elicit an antipeptide antibody response in one or more mouse strain. Eleven antipeptide serum pools were able to bind to virus in ELISA. Fifteen peptides primed one or more haplotype for an in vitro antipeptide Th-cell response as measured by blastogenesis. Th-cell activation was generally confirmed by measurable in vitro production of interleukin (IL)-2/IL-4. Nine peptides that were positive for in vitro blastogenesis, 1-2, 35, 4-6, 79, 142, 208, 06, 16, and 17, elicited virus-reactive Th-cells in vitro in H-2d mice. Two of these peptides (4-6 and 17) were able to prime virus-reactive Th-cells in H-2b mice. Nine peptides primed outbred mice in vitro for an antiviral antibody response significantly greater than that seen in animals primed with an irrelevant peptide. These results correlate with, and expand on, our previous observations based on a smaller set of synthetic peptides derived from the E-glycoprotein of Murray Valley encephalitis virus and suggest that synthetic peptides can function as E-glycoprotein Th-cell epitopes. The similarity of results between two distantly related flaviviruses suggests that E-glycoprotein Th-cell epitopes are consistent in location and activity.
Assuntos
Antígenos Virais/imunologia , Vírus da Dengue/imunologia , Epitopos/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Proteínas do Envelope Viral/imunologia , Animais , Anticorpos Antivirais/biossíntese , Anticorpos Antivirais/sangue , Antígenos Virais/sangue , Glicoproteínas/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Fragmentos de Peptídeos/imunologia , Proteínas do Envelope Viral/sangueRESUMO
Stable neutralization (N) escape variants of Venezuelan equine encephalitis (VEE) virus were selected by anti-E2 glycoprotein monoclonal antibodies (MAbs) that neutralize viral infectivity, block viral hemagglutination, and passively protect mice. The nucleotide sequence of the E1, E2, and E3 genes of four variants revealed a clustering of single mutations in a domain spanning E2-182 to E2-207. The conformation of this short linear sequence affects antigenicity in the N domain because reduction and alkylation of virus disrupted binding of some E2 neutralizing MAbs. Serologic evidence for interaction of E2 epitopes also was obtained. Mutations in the N domain of VEE virus did not alter the kinetics of binding to Vero cells. They did, in some cases, produce attenuation of virulence in mice.