T cell receptor diversity in CTLs specific for the CTT-1 and CTT-2 minor histocompatibility antigens

Minor histocompatibility Ags (HA) and their associated immunogenic peptides provide a formidable barrier for successful transplantation, but there is virtually no information regarding the diversity of minor HA-specific TCRs. We have investigated the diversity of α- and β-chains in TCRs specific for the CTT-1 and CTT-2 peptides that are immunodominant in CTL responses to multiple minor HA. CTLs were cloned after in vitro stimulation, and α- and β-chain transcripts were amplified and sequenced to identify utilized V genes, complementarity-determining regions 3 (CDR3s), and joining region genes. Twenty-one unique CTT-2-specific TCRs were identified in 31 clones, and 22 CTT-1-specific TCRs were identified in 29 clones. A relatively limited number of Vβ subfamilies were represented in these panels of TCRs, with Vβ5 and Vβ8 genes expressed in multiple TCRs in each panel. Similar diversity was observed with Va usage, and Vα4 subfamily usage was more prominent in CTT-2-specific TCRs than in CTT-1-specific TCRs. Neither α nor β CDR3 regions exhibited prominent motifs or length restriction. However, CTT-1- and CTT-2-specific β CDR3 regions included an excess of negatively over positively charged residues that were bimodally distributed among CDR3 positions. In fact, 50% of CTT-1-specific CDR3 regions included two negative charges separated by three to five amino acids. Despite the similarities in net charge of β CDR3 regions, TCRs specific for these two minor HA peptides are relatively diverse and would expectedly withstand attempts at anti-TCR Ab-mediated immunosuppression.