Abstract
A genetic vaccine for West Nile virus (WN) has been synthesized with the WN premembrane-envelope (WN preM-E) gene sequences encoded as a chimera with the transmembrane and carboxyl terminal domains of the lysosome-associated membrane protein (LAMP). The LAMP sequences are used to direct the antigen protein to the major histocompatibility class II (MHC II) vesicular compartment of transfected professional antigen-presenting cells (APCs). Vaccine constructs encoding the native WN preM-E and WN preM-E/LAMP chimera were synthesized in pVAX1 and pITR plasmid backbones. Extracts of human fibroblast 293 and monkey kidney COS-7 cells transfected with the WN preM-E/LAMP chimera constructs contained much greater amounts of E than did the cells transfected with constructs encoding the native WN preM-E. This difference in the concentration of native E and the E/LAMP chimera in transfected cells is attributed to the secretion of native E. The amount of preM protein in cell extracts, in contrast to the E protein, and the levels of DNA and RNA transcripts, did not differ between WN preM-E- and WN preM-E/LAMP-transfected cells. Additionally, confocal and immunoelectron microscopic analyses of transfected B cells showed localization of the WN preM-E/LAMP chimera in vesicular compartments containing endogenous LAMP, MHC II, and H2-M, whereas native viral preM-E lacking the LAMP sequences was distributed within the cellular vesicular network with little LAMP or MHC II association. Mice immunized with a DNA construct expressing the WN preM-E/LAMP antigen induced significant antibody and long-term neutralization titers in contrast to the minimal and short-lived neutralization titer of mice vaccinated with a plasmid expressing the untargeted antigen. These results underscore the utility of LAMP targeting of the WN envelope to the MHC II compartments in the design of a genetic WN vaccine.
Original language | English (US) |
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Pages (from-to) | 66-77 |
Number of pages | 12 |
Journal | Virology |
Volume | 332 |
Issue number | 1 |
DOIs | |
State | Published - Feb 5 2005 |
Externally published | Yes |
Keywords
- Electron microscopy
- H-2M
- LAMP
- MHC II
- MIICs
- Neutralizing antibody
- West Nile virus DNA vaccine
ASJC Scopus subject areas
- Virology