PEPTIDE-INDEPENDENT RECOGNITION BY ALLOREACTIVE CYTOTOXIC T-LYMPHOCYTES (CTL)

We have isolated several H-2K(b)-alloreactive cytotoxic T cell clones and analyzed their reactivity for several forms of H2K(b). These cytot oxic T lymphocytes (GTL) were elicited by priming with a skin graft fo llowed by in vitro stimulation using stimulator cells that express an H-2K(b) molecule unable to bind CD8. In contrast to most alloreactive T cells, these CTL were able to recognize H-2K(b) on the surface of th e antigen processing defective cell lines RMA-S and T2. Furthermore, t his reactivity was not increased by the addition of an extract contain ing peptides from C57BL/6 (H-2(b)) spleen cells, nor was the reactivit y decreased by treating the target cells with acid to remove peptides bound to MHC molecules. The CTL were also capable of recognizing targe ts expressing the mutant H-2K(bm8) molecule. These findings suggested that the clones recognized determinants on H-2K(b) that were independe nt of peptide. Further evidence for this hypothesis was provided by ex periments in which H-2K(b) produced in Drosophila melanogaster cells a nd immobilized on the surface of a tissue culture plate was able to st imulate hybridomas derived from these alloreactive T cells. Precursor frequency analysis demonstrated that skin graft priming, whether with skin expressing the wild-type or the mutant H-2K(b) molecule, is a str ong stimulus to elicit peptide-independent CTL. Moreover, reconstituti on experiments demonstrated that the peptide-independent CTL clones we re capable of mediating rapid and complete rejection of H-2-incompatib le skin grafts. These findings provide evidence that not all allorecog nition is peptide dependent.