Previous studies have indicated that blockade of signaling through the T-cell receptor (TCR)/calcineurin/nuclear factor of activated T cells (NFAT) pathway impairs transplantation tolerance induced with anti-CD154 antibody. By using an allogeneic bone marrow transplantation model, we examined the role of the TCR/calcineurin/NFAT pathway for tolerance induction with anti-CD154. Calcineurin blockade by cyclosporine A led to a failure of CD8 but not CD4 tolerance, and experiments in NFAT1^−/− mice replicated this effect. Studies in thymectomized mice demonstrated that blockade of the calcineurin/NFAT pathway after bone marrow transplantation led to a failure of peripheral CD8 tolerance. Moreover, CD8 adoptive transfer studies demonstrated that NFAT1 is cell-intrinsically required for peripheral CD8 tolerance. NFAT1 deficiency did not impair CD8 T-cell up-regulation of PD1, which is required for CD8 tolerance in this model. NFAT1 has previously been shown to have a role in CD4 cells for anergy induction and for programming CD4 cells to become regulatory cells. By generating mice lacking NFAT1 in CD4 but not CD8 cells, we demonstrate that NFAT1 is neither required for CD4 tolerance induction nor for their regulatory function on CD8 T cells. Thus, our study reveals a CD8 T cell–intrinsic NFAT1 requirement for CD8 tolerance in vivo.