HLA-B27-specific CTL are heterogeneous in their capacity to lyse murine P815 cells transfected with HLA-B27. Failure to kill murine transfectants could be caused by insufficient avidity of the human effector cells towards murine targets. Alternatively, it may imply alteration of allospecific T cell epitopes upon expression of HLA-B27 on mouse cells. To discern between these alternatives, P815 cells were co-transfected with HLA-B27, human ICAM-1, and LFA-3, and the transfectants were used as target cells with a series of HLA-B27-specific alloreactive CTL. Thirty-seven percent of the CTL tested significantly lysed HLA-B27(+)-P815 cells, without requiring simultaneous expression of human adhesion molecules. Twenty-one percent of the CTL showed significant lysis of only the murine transfectants expressing ICAM-1 or ICAM-1 + LFA-3. These CTL were shown by mAb-blocking analysis to have lower avidity than CTL from the previous group. In addition, they recognized HLA-B27 on murine cells not expressing human adhesion molecules, as assessed by cold target competition assays. As many as 42% of the CTL were unable to kill, or did so very inefficiently, P815 transfectants regardless of the presence of human ICAM-1 and LFA-3. With one detected exception, these CTL did not recognize HLA-B27 on murine cells in cold target competition assays. In contrast, they were able to recognize HLA-B27(+)-M1 fibroblast transfectant cells in direct cytotoxicity or cold target competition assays. Failure to kill murine transfectants by CTL from this group did not correlate with lower avidity, relative to CTL from the other groups, as shown by blocking experiments with mAb against human T cell adhesion molecules and their counter-receptors. These results indicate that lack of lysis of murine transfectants expressing class I HLA molecules by alloreactive CTL can be accounted for by low avidity of interspecies cell interactions in some cases but, more often, it is caused by alteration of allospecific T cell epitopes. Most likely, the basis for such alteration is allorecognition of HLA-B27-bound peptides that are expressed on human but not on mouse cells, mainly as a consequence of phylogenetic protein divergence between both species.