Spectratyping of TCR expressed by CTL-infiltrating male antigen (HY)- disparate allografts

Minor histocompatibility Ags (HA) play prominent roles in stimulating allograft rejection and are recognized by TLs that mediate this process. There is limited information regarding the sequences of minor HA peptides and the diversity of minor HA-specific TCRs. In the case of the male minor HA (HY), a peptide presented by H2D^b molecules has been sequenced. We have used spectratyping to study the diversities of Vβ usage and β complementarily- determining region 3 (CDR3) lengths of TCRs expressed by GTLs that infiltrate HY-disparate skin allografts during rejection. Spectratyping of RNA from second- and third-set male allografts on CD4-depleted, female recipients showed a reduction in Vβ usage and β CDR3 length diversity with prominent representation of Vβ8 genes. CDR3 sequences, as a group, were characterized by net negative charges resulting from negatively charged residues at positions 5-6 and 10-11. The effects of in vivo anti-Vβ8 Ab treatment on rejection of second-set male allographs were investigated. This Ab treatment had no effect on allograft rejection time and resulted in increased Vβ7 usage in recipients with complete Vβ8 depletion. More interestingly, the net charges of β CDR3s derived from Vβ8-depleted recipients were altered by the inclusion of positively charged and polar residues at positions 4-6. These results indicate that Vβ-specific T cell depletion has no effect on HY- disparate allograft survival, but it alters Vβ usage and changes the characteristics of β CDR3s that facilitate class I:peptide recognition.