Diversity of alpha and beta subunits of T-cell receptors specific for the H4 minor histocompatibility antigen

The mouse H4 minor histocompatibility antigen (HA) includes a single H2K(b)-bound peptide that stimulates rejection of skin allografts and generation of cytolytic T lymphocytes (CTL). We evaluated the diversity of the CTL response to this single minor HA peptide by sequencing alpha and beta chains of T-cell receptors (Tcr) from H4-specific CTLs as a first step toward understanding the diversity of Tcrs specific for single minor HA. We selected H4-specific CTL clones from short-term lines that were restricted by K-b (19 clones), K-bm5 (7 clones), and K-bm11 (10 clones). Whereas multiple V-alpha and V-beta family members were identified in the panel of K-b-restricted CTLs, five VB genes and one VA subfamily were predominant in CTLs derived from multiple individuals. Similar distributions were observed with K-bm5- and K-bm11-restricted CTLs, suggesting that these two mutants did not alter Tcr gene usage observable at the VA and/or VB gene levels. Negatively charged residues were present in the CDR3 regions of 12/13 unique K-b-restricted beta chains. A comparable observation was made with Kbms-restricted CTLs, and the distributions of these residues among CDR3 positions were similar in the two CTL panels. However, the K-bm11 mutation dramatically altered the distribution of these residues resulting in their presence in positions 10-12 of CDR3 regions in all CTLs. These results indicate that the Tcrs expressed by CTLs specific for this single minor HA peptide are oligoclonal and characterized by the presence of negatively charged residues in beta CDR3 regions, the distribution of which is profoundly altered by the K-bm11 mutation.