Low-dose UV radiation (UVR) inhibits the induction of contact hypersensitivity and induces regulatory T cells (Tregs), which because of their antigen specificity harbor therapeutic potential. Topical application of 1α,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) is known to induce Tregs as well, which implies that 1,25(OH)(2)D(3) might be involved in UVR-induced immunosuppression. It was the aim of this study to clarify this issue, to further characterize 1,25(OH)(2)D(3)-induced Tregs and to determine whether they differ from UVR-induced Tregs. Our data demonstrate that 1,25(OH)(2)D(3)-induced Tregs act in an antigen-specific manner and belong to the Foxp3-expressing subtype of Tregs as demonstrated by diphtheria toxin (DT)-mediated depletion of Foxp3(+) Tregs in DEREG (depletion of Tregs) mice. Using Langerin-DTR (DT receptor) knock-in mice, it was shown that Langerhans cells (LCs) are required for the induction of Tregs by 1,25(OH)(2)D(3), as depletion of LCs but not Langerin(+) dermal dendritic cells abrogated the induction of Tregs. Taken together, 1,25(OH)(2)D(3) affects the immune system in a similar manner as UVR, probably using the same pathways. However, vitamin D receptor knockout mice were equally susceptible to UVR-induced immunosupppression as wild-type controls. This indicates that 1,25(OH)(2)D(3) exerts similar immunosuppressive effects as UVR but is dispensable for local UVR-induced immunosuppression.