MAPPING OF TH1 HELPER T-CELL EPITOPES ON MAJOR SECRETED MYCOBACTERIAL ANTIGEN 85A IN MICE INFECTED WITH LIVE MYCOBACTERIUM-BOVIS BCG

TH1 cytokine secretion was examined in response to synthetic peptides of the 85A component of the major secreted, fibronectin-binding antigen 85 complex from Mycobacterium tuberculosis in seven different mouse strains infected with live M. bovis BCG. Twenty-eight overlapping 20-mer peptides covering the complete mature 295-amino-acid (AA) protein mere synthesized. Significant interleukin-2 (IL-2) and gamma interferon (IFN-gamma) secretion could be measured following in vitro stimulation of spleen cells with these peptides. H-2(d) haplotype mice reacted preferentially against the amino-terminal half of the protein, i.e., against peptide 5 (AA 41 to 60) and especially against peptide 11 (AA 101 to 120), which contained an I-Ed binding motif. H-2(b) haplotype mice, on the other hand, reacted against peptides from both amino- and carboxy-terminal halves of the protein, peptide 25 (AA 241 to 260) being the most potent stimulator of IL-2 and IFN-gamma production. (BALB/c x C57BL/6)F, animals with the H-2(d/b) haplotype weakly recognized peptides specific for both parental lines. Finally, CBA/J (H-2(k)) and major histocompatibility complex class II mutant B6.C.bm12 mice, carrying a mutant I-A beta(bm12) allele on an H-2(b) background, reacted only very weakly to the 85A peptides. Reactive T cells isolated from lungs of BCG-infected H-2(b) haplotype mice recognized the same epitopes as spleen cells, especially peptide 25. These data confirm previous findings regarding the powerful IL-2 and IFN-gamma-inducing properties of antigen 85 during infection with live ill. bovis BCG.