The suppressive capacity of naturally occurring mouse CD4⁺CD25⁺ T cells on T-cell activation has been well documented. The present study is focused on the interaction of CD4⁺CD25⁺ T cells and B cells. By coculturing preactivated CD4⁺CD25⁺ T cells with B cells in the presence of polyclonal B-cell activators, we found that B-cell proliferation was significantly suppressed. The suppression of B-cell proliferation was due to increased cell death caused by the CD4⁺CD25⁺ T cells in a cell-contact–dependent manner. The induction of B-cell death is not mediated by Fas–Fas ligand pathway, but surprisingly, depends on the up-regulation of perforin and granzymes in the CD4⁺CD25⁺ T cells. Furthermore, activated CD4⁺CD25⁺ T cells preferentially killed antigen-presenting but not bystander B cells. Our results demonstrate that CD4⁺CD25⁺ T cells can act directly on B cells and suggest that the prevention of autoimmunity by CD4⁺CD25⁺ T cells can be explained, at least in part, by the direct regulation of B-cell function.