Functional mapping of protective domains and epitopes in the rotavirus VP6protein

Citation
Ahc. Choi et al., Functional mapping of protective domains and epitopes in the rotavirus VP6protein, J VIROLOGY, 74(24), 2000, pp. 11574-11580
Citations number
33
Categorie Soggetti
Microbiology
Journal title
JOURNAL OF VIROLOGY
ISSN journal
0022538X → ACNP
Volume
74
Issue
24
Year of publication
2000
Pages
11574 - 11580
Database
ISI
SICI code
0022-538X(200012)74:24<11574:FMOPDA>2.0.ZU;2-9
Abstract
The purpose of this study was to determine which regions of the VP6 protein of the murine rotavirus strain EDIM are able to elicit protection against rotavirus shedding in the adult mouse model following intranasal (i.n.) imm unization with fragments of VP6 and a subsequent oral EDIM challenge. In th e initial experiment, the first (fragment AB), middle (BC), or last (CD) pa rt of VP6 that was genetically fused to maltose-binding protein (MBP) and e xpressed in Escherichia coli was examined. Mice (BALB/c) immunized with two 9-mug doses of each of the chimeras and 10 mug of the mucosal adjuvant LT( R192G) were found to be protected against EDIM shedding (80, 92, and nearly 100% reduction, respectively; P less than or equal to 0.01) following chal lenge. Because CD produced almost complete protection, we prepared four E. coli-expressed, MBP-fused chimeras containing overlapping fragments of the CD region (i.e., CD1, CD2, CD3, and CD4) whose lengths ranged from 61 to 67 amino acid residues. Following i.n. immunization, CD1, CD2, and CD4 induce d significant (P less than or equal to 0.004) protection (88, 84, and 92% r eduction, respectively). In addition, 11 peptides (18 to 30 residues) of th e CD region with between 0 and 13 overlapping amino acids were synthesized. Two 50-mug doses of each peptide with LT(R192G) were administered i.n. to BALB/c mice. Five peptides were found to elicit significant (P less than or equal to 0.02) protection. Moreover, a 14-amino-acid region within peptide 6 containing a putative CD4(+) T-cell epitope was found to confer nearly c omplete protection, suggesting a protective role for CD4(+) T cells. Mice t hat were protected by fragments BC and CD1 and four of the five protective synthetic peptides did not develop measurable rotavirus antibodies in serum or stool, implying that protection induced by these domains was not depend ent on antibody. Together, these observations suggest that multiple regions of VP6 can stimulate protection, a region of VP6 as small as 14 amino acid s containing a CD4(+) T-cell epitope can stimulate nearly complete protecti on, and protection mediated by a subset of epitopes in the VP6 protein does not require antibodies in BALB/c mice.