Identification of murine H2-Dd- and H2-Ab-restricted T-cell epitopes on a novel protective antigen, MPT51, of Mycobacterium tuberculosis.
Both CD4(+) type 1 helper T (Th1) cells and CD8(+) cytotoxic T lymphocytes (CTL) play pivotal roles in protection against Mycobacterium tuberculosis infection. Here, we identified Th1 and CTL epitopes on a novel protective antigen, MPT51, in BALB/c and C57BL/6 mice. Mice were immunized with plasmid DNA encoding MPT51 by using a gene gun, and gamma interferon (IFN-gamma) production from the immune spleen cells was analyzed in response to a synthetic overlapping peptide library covering the mature MPT51 sequence. In BALB/c mice, only one peptide, p21-40, appeared to stimulate the immune splenocytes to produce IFN-gamma. Flow cytometric analysis with intracellular IFN-gamma and the T-cell phenotype revealed that the p21-40 peptide contains an immunodominant CD8(+) T-cell epitope. Further analysis with a computer-assisted algorithm permitted identification of a T-cell epitope, p24-32. In addition, a major histocompatibility complex class I stabilization assay with TAP2-deficient RMA-S cells transfected with K(d), D(d), or L(d) indicated that the epitope is presented by D(d). Finally, we proved that the p24-32/D(d) complex is recognized by IFN-gamma-producing CTL. In C57BL/6 mice, we observed H2-A(b)-restricted dominant and subdominant Th1 epitopes by using T-cell subset depletion analysis and three-color flow cytometry. The data obtained are useful for analyzing the role of MPT51-specific T cells in protective immunity and for designing a vaccine against M. tuberculosis infection.
Suzuki
Mina
M
Department of Microbiology and Immunology, Hamamatsu University School of Medicine, 1-20-1 Handa-yama, Hamamatsu 431-3192, Japan.
Aoshi
Taiki
T
Nagata
Toshi
T
Koide
Yukio
Y
D000595:Amino Acid Sequence; D000818:Animals; D000942:Antigens, Bacterial / Q000276:immunology* / Q000378:metabolism; D001426:Bacterial Proteins / Q000276:immunology* / Q000378:metabolism; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology; D000939:Epitopes / Q000276:immunology*; D006183:H-2 Antigens / Q000276:immunology*; D015395:Histocompatibility Antigens Class I / Q000276:immunology; D007371:Interferon-gamma / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D009169:Mycobacterium tuberculosis / Q000276:immunology*; D010455:Peptides / Q000378:metabolism; D013601:T-Lymphocytes / Q000276:immunology*; D018417:Th1 Cells / Q000276:immunology
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D000939:Epitopes; D006183:H-2 Antigens; D015395:Histocompatibility Antigens Class I; C082911:MPT51 protein, Mycobacterium; D010455:Peptides; D007371:Interferon-gamma