Identification of Western equine encephalitis virus structural proteins that confer protection after DNA vaccination

Abstract

DNA vaccines encoding different portions of the structural proteins of western equine encephalitis virus were tested for the efficacy of their protection in a 100% lethal mouse model of the virus. The 6K-E1 structural protein encoded by the DNA vaccine conferred complete protection against challenge with the homologous strain and limited protection against challenge with a heterologous strain.

FIG. 1.

Schematic illustration of the synthesis of structural proteins of the alphaviruses and DNA plasmids constructed in this study.

FIG. 2.

Expression of E1 and E2 proteins from DNA vaccines. Proteins from 293S cells transfected with the DNA vaccine were separated by electrophoresis through 12.5% polyacrylamide gels, transferred to nitrocellulose membranes, probed with an anti-E1 (A) or an anti-E2 (B) monoclonal antibody, and incubated with goat anti-mouse IgG conjugated with horseradish peroxidase (Sigma-Aldrich, St. Louis, MO). The E1 and E2 proteins were visualized by use of an Amersham ECL Plus Western blotting detection kit (GE Healthcare, Buckinghamshire, United Kingdom). Lane 1, molecular mass standard (Invitrogen); lane 2, mock-transfected cells (negative control); lane 3, pVAX-transfected cells (vector control); lane 4, pVHX-6-transfected cells; lane 5, pE3-E2-transfected cells; lane 6, pE3-E2-6K-E1-transfected cells; and lane 7, p6K-E1-transfected cells.

FIG. 3.

Kaplan-Meier survival curves for the mice (eight mice per group) vaccinated with pE3-E2-6K-E1, p6K-E1, or pE3-E2 and challenged with a lethal dose of the homologous strain of WEEV. The vaccinated mice were challenged intranasally with strain 71V-1658 and monitored for 14 days. The log rank test was used to assess the differences in the survival rates of the challenged mice. *, P < 0.01 for pE3-E2-6K-E1 versus pVAX; **, P < 0.01 for p6K-E1 versus pVAX; ***, P > 0.05 for pE3-E2 versus pVAX.

FIG. 4.

Kaplan-Meier survival curves for the mice (eight mice per group) vaccinated with pE3-E2-6K-E1, p6K-E1, or pE3-E2 and challenged with a lethal dose of a heterologous strain of WEEV. The vaccinated mice were challenged intranasally with low-virulence strain CBA87 or high-virulence strain Fleming and were monitored for 14 days. The log rank test was used to assess the differences in the survival rates of the challenged mice. *, P < 0.01 for pVHX-6 versus pVAX; **, P < 0.01 for pE3-E2-6K-E1 versus pVAX; ***, P < 0.01 for p6K-E1 versus pVAX; #, P < 0.01 for pVHX-6 versus pVAX; ##, P < 0.01 for pE3-E2-6K-E1 versus pVAX.