Restoration of normal perforin expression in CD8⁺ T cells is necessary and sufficient to restore perforin-dependent immune regulation. (A) Prf^−/− mice were reconstituted with 90% RAG/prf^−/− (DKO) marrow and 10% mixed marrow (variable amounts of WT [GFP⁺] and prf^−/−) to give animals with variable perforin expression within the T-cell compartment and perforin expression in <10% of non–T cells. After engraftment, animals were challenged with LCMV as in Figure 1, and day 8 IFN-γ levels are plotted against WT chimerism within the T-cell compartment. (B) Prf^−/− mice were reconstituted with 90% RAG^−/− marrow and 10% mixed marrow (WT [GFP⁺] and prf^−/−) to give animals with variable perforin expression within the T-cell compartment and normal perforin expression in 90% to 100% of non–T cells. IFN-γ levels and chimerism are plotted as in panel A. (C) Purified naïve WT (CD45.1⁺) or prf^−/− CD4⁺ T cells were transferred into prf^−/− mice (after conditioning with cyclophosphamide and waiting 3 weeks for recovery/stable engraftment). Animals were challenged with LCMV; IFN-γ levels and WT CD4⁺ T cell chimerism were assessed. (D) Purified naïve WT (CD45.1⁺) CD8⁺ T cells were transferred into prf^−/− mice (as in panel C). Animals were challenged with LCMV; IFN-γ levels and WT CD8⁺ T-cell chimerism were assessed. In vivo IFN-γ production by endogenous (prf^−/−) T cells was assessed on day 8 after LCMV in animals receiving CD8⁺ T-cell transfers, as previously described.⁹  *P < .01.