Although FOXP3 is primarily expressed by regulatory CD4 T cells (Treg) in vivo, polyclonal activation of human CD8 T cells can result in the expression of FOXP3 in a fraction of CD8 T cells. However, the cellular lineage and mechanism of FOXP3 induction in CD8 T cells remain unclear. Here, we demonstrate that interleukin-2 (IL-2) induces FOXP3 expression in OKT3-stimulated or antigen-stimulated CD8 T cells, indicating that FOXP3 expression is neither limited to a unique subset of CD8 T cells nor dependent on the mode of T-cell receptor stimulation. In the absence of IL-2, antigen stimulation resulted in T-cell activation and acquisition of effector function without induction of FOXP3, indicating that acquisition of effector function is independent of induction of FOXP3 expression in CD8 T cells. Interestingly, IL-15, but not IL-7 or IL-21, also led to de novo induction of FOXP3 in antigen-specific CD8 T cells, suggesting that signaling by IL-2/IL-15Rbeta chain is pivotal for induction of FOXP3 in human CD8 T cells. These findings indicate that induction of FOXP3 is intrinsic to CD8 T cells that are activated in the presence of IL-2 or IL-15, and in vitro-induced expression of FOXP3 cannot be simply interpreted as an indicator of Treg activity or activation marker.