Dual overlapping peptides recognized by insulin peptide B : 9-23 T cell receptor AV13S3 T cell clones of the NOD mouse

T cells isolated from islets of non-obese diabetic (NOD) mice are enriched for insulin-reactive cells. The great majority of these T cells recognize i nsulin B chain peptide (B:9-23). B:9-23 reactive T cell clones are diabetog enic and show a dramatic TCR alpha-chain restriction (predominant AV13S3). We have studied the reactivity of five different B:9-23 reactive T cell clo nes to truncated peptides and alanine substituted analogues of B:9-23. Amon gst these AV13S3 T cell clones, one reacted with peptide B:9-16 and four wi th B:13-23. The two peptides have in common only four amino acids (B:13-16; EALY). Having defined minimal peptide epitopes, we evaluated a mutant insu lin sequence (B:13 glutamine) which retains metabolic activity. As predicte d, this single amino acid change abrogated T cell reactivity. In addition, we have created a modified I-A(g7) gene with the B:9-23 peptide covalently linked to I-A(g7). Antigen presenting cells transfected with this construct were excellent presenting cells for all clones studied. The definition of dual peptide motifs and creation of bioactive covalent I-A(g7)-B:9-23 shoul d facilitate studies of the pathogenic significance and antigen recognition by B:9-23 reactive diabetogenic T cells. (C) 2000 Academic Press.