Regulatory CD4⁺ T cells (T_r cells), the development of which is critically dependent on X-linked transcription factor Foxp3 (forkhead box P3), prevent self-destructive immune responses. Despite its important role, molecular and functional features conferred by Foxp3 to T_r precursor cells remain unknown. It has been suggested that Foxp3 expression is required for both survival of T_r precursors as well as their inability to produce interleukin (IL)-2 and independently proliferate after T-cell-receptor engagement, raising the possibility that such ‘anergy’ and T_r suppressive capacity are intimately linked^,,. Here we show, by dissociating Foxp3-dependent features from those induced by the signals preceding and promoting its expression in mice, that the latter signals include several functional and transcriptional hallmarks of T_r cells. Although its function is required for T_r cell suppressor activity, Foxp3 to a large extent amplifies and fixes pre-established molecular features of T_r cells, including anergy and dependence on paracrine IL-2. Furthermore, Foxp3 solidifies T_r cell lineage stability through modification of cell surface and signalling molecules, resulting in adaptation to the signals required to induce and maintain T_r cells. This adaptation includes Foxp3-dependent repression of cyclic nucleotide phosphodiesterase 3B, affecting genes responsible for T_r cell homeostasis.