To examine the effects of gamma irradiation on Tregs, changes in phenotype and suppression function in Tregs treated with or without gamma ray were analyzed. Purified CD4+CD25+ regulatory T cells were irradiated at different dosages with a 137Cs source gamma ray at 4.8 Gy/min. After culture, the phenotype and function changes were determined by flow cytometry and [3H]-thymidine incorporation, respectively. A dose-dependent reduction of Tregs proliferation in response to gamma irradiation was noted, which paralleled the apoptosis induction of Tregs. Gamma irradiation downregulated the Tregs expression of CD45RO, CD62L, FOXP3, membrane TGF-β, but upregulated Bax and GITR. High dose gamma irradiation (30 Gy) significantly abolished the suppression of Tregs on CD4+CD25− T cells proliferation. Thus Tregs not only influences the phenotype but also alters their suppressive capacities. Our findings suggest that radiotherapy may be an important strategy to alter the immunologic balance of Tregs and effector cells in cancer therapy.