|Norimine, Junzo - WASHINGTON STATE UNIV.|
|Mc Elwain, Terry - WASHINGTON STATE UNIV.|
|Florin-Christensen, Monica - WASHINGTON STATE UNIV.|
|Brown, Wendy - WASHINGTON STATE UNIV.|
Submitted to: Infection and Immunity
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: January 2, 2002
Publication Date: N/A
Interpretive Summary: In this work we describe the mapping of T-cell epitopes in the vaccine candidate rhoptry associated protein-1 (RAP-1) of B. bovis. The amino terminal domain of RAP-1 contains the conserved motifs tipical of the babesial rap-1 family. The paper demonstrates that immunodominant T -cell helper epitopes are all located in the amino terminal domain of RAP-1, thus supporting the inclusion of this region in vaccine constructs. In addition, peptides representing these immunodominant T-cell helper epitopes could be used for the development of an epitope-based vaccine against B. bovis.
Technical Abstract: Babesia bovis rhoptry-associated protein 1 (RAP-1), which confers partial protection against B. bovis challenge, is recognized by antibodies and T lymphocytes from cattle that have recovered from infection and are immune to subsequent challenge. RAP-1 is a 60-kDa protein with an N-terminal (NT) region that contains four cysteine residues conserved among all Babesia RAP-1 family members and a C-terminal (CT) region that contains multiple, degenerate, tandem 23-amino-acid (aa) repeats. To define the location of CD4(+)-T-cell epitopes for vaccine development using a recombinant protein or minigene construct, a series of truncated recombinant RAP-1 proteins and peptides were tested for stimulation of T-cell lines derived from B. bovis-immune cattle. CD4(+)-T-cell lines from three B. bovis-immune cattle with different DRB3 haplotypes responded to the NT region of RAP-1, whereas T cells from only one animal responded weakly to the CT region. T-cell lines from the three individuals recognized two to six NT-region peptides spanning aa 134 to 316 and representing at least four dominant epitopes. Using RAP-1-specific CD4(+)-T-cell clones, two NT-region epitopes, EYLVNKVLYMATMNYKT (aa 187 to 203) and EAPWYKRWIKKFR (aa 295 to 307), and one CT-region repeat epitope, FREAPQATKHFL, which is present twice at aa positions 391 to 402 and 414 to 425, were identified. Several peptides representing degenerate repeats of the agonist CT-region peptide FREAPQATKHFL neither stimulated responses of T-cell clones specific for this peptide nor inhibited responses to the agonist peptide. Upon stimulation with specific antigen, T-cell clones specific for NT or CT epitopes produced gamma interferon. The presence of T-helper-cell epitopes in the NT domain of RAP-1, which is highly conserved among otherwise antigenically different strains of B. bovis, supports the inclusion of this region in vaccine constructs to be tested in cattle.