Down‐regulation of autoreactive T cell responses in vivo includes cell‐contact‐dependent as well as contact‐independent mechanisms. Infectious tolerance is a contact‐dependent mechanism used by naturally occurring CD25⁺ T regulatory cells (Tregs) to confer suppressive activity upon conventional CD4⁺ T cells thereby generating secondary T helper suppressor cells(Th_sup), which inhibit T cell activation via soluble mediators. Here, we describe two distinct subsets of human Tregs, characterized by expression of either the α₄β₇ integrin or the α₄β₁ integrin. Upon activation, both subsets show an enhanced expression of FoxP3, recently described as a key transcription factor of murine Tregs. In addition, both are able to convey suppressive capacity to conventional CD4⁺ T cells. However, the properties of Treg subsets are rather distinct: α₄β₇⁺Tregs induce IL‐10‐producing Th_sup (Tr1‐like), whereas α₄β₁⁺ Tregs induce TGF‐β‐producing Th_sup (Th3‐like). Our findings reconcile conflicting results by clearly demonstrating that suppression through naturally occurring CD25⁺ Tregs is primary cell‐contact‐dependent but is subsequently followed by cell‐contact‐independent T cell inhibition mediated by second‐generation Tr1‐ and Th3‐like Th_sup via the soluble factors IL‐10 and TGF‐β.