Regulatory CD4+ T cells (T(R) cells) develop under the control of the X-linked transcription factor Foxp3, which prevents autoimmune diseases by suppressing immune responses. While Foxp3 is essential for T(R) cell function, its molecular and functional roles in T(R) precursor cells remain unclear. This study shows that Foxp3 enhances and stabilizes pre-existing molecular features of T(R) cells, including anergy and dependence on paracrine IL-2. Foxp3 also solidifies T(R) cell lineage stability by modifying cell surface and signaling molecules, enabling adaptation to signals required for T(R) cell development and maintenance. Foxp3-dependent repression of cyclic nucleotide phosphodiesterase 3B (PDE3B) affects genes involved in T(R) cell homeostasis. In mice, Foxp3 deficiency leads to severe autoimmune disease, but heterozygous females with one mutant Foxp3 allele are protected due to the presence of normal T(R) cells. T(R) cells were found to be distinct from T(FN) cells, which express Foxp3-null but lack Foxp3 protein. T(FN) cells showed an anergic phenotype, similar to T(R) cells, but were less proliferative and had reduced IL-7R expression. Both T(FN) and T(R) cells relied on exogenous IL-2, but T(FN) cells were less competitive due to intermediate CD25 levels. T(FN) cells could produce IL-10 or IL-17, resembling Tr1 or Th17 cells, and expressed RORγt, which promotes IL-17 and blocks IL-2 production. T(FN) cells lacked suppressive activity against CD4+CD25- T-cell proliferation and failed to control effector T-cell expansion. However, T(FN) cells competed efficiently with effector T cells during homeostatic expansion. Foxp3 transcription was retained in T(R) cells but not in T(FN) cells, indicating that Foxp3 function is necessary for stable expression. Global gene expression profiling revealed that Foxp3 amplifies and stabilizes genes associated with T(R) cell characteristics, including cell surface markers like CD25, GITR, and CTLA4. Foxp3-dependent genes were enriched for cell surface and extracellular proteins, and Foxp3 promotes cell cycle-related genes, enhancing T(R) cell proliferation. Foxp3 also regulates PDE3B expression, which affects cAMP signaling and T(R) cell homeostasis. Reduced PDE3B expression in T(R) cells supports normal homeostasis and gene expression. Ectopic PDE3B expression in T(R) cells reduced their numbers, indicating that PDE3B repression is essential for T(R) cell maintenance. These findings suggest that Foxp3 consolidates anergic and paracrine IL-2Regulatory CD4+ T cells (T(R) cells) develop under the control of the X-linked transcription factor Foxp3, which prevents autoimmune diseases by suppressing immune responses. While Foxp3 is essential for T(R) cell function, its molecular and functional roles in T(R) precursor cells remain unclear. This study shows that Foxp3 enhances and stabilizes pre-existing molecular features of T(R) cells, including anergy and dependence on paracrine IL-2. Foxp3 also solidifies T(R) cell lineage stability by modifying cell surface and signaling molecules, enabling adaptation to signals required for T(R) cell development and maintenance. Foxp3-dependent repression of cyclic nucleotide phosphodiesterase 3B (PDE3B) affects genes involved in T(R) cell homeostasis. In mice, Foxp3 deficiency leads to severe autoimmune disease, but heterozygous females with one mutant Foxp3 allele are protected due to the presence of normal T(R) cells. T(R) cells were found to be distinct from T(FN) cells, which express Foxp3-null but lack Foxp3 protein. T(FN) cells showed an anergic phenotype, similar to T(R) cells, but were less proliferative and had reduced IL-7R expression. Both T(FN) and T(R) cells relied on exogenous IL-2, but T(FN) cells were less competitive due to intermediate CD25 levels. T(FN) cells could produce IL-10 or IL-17, resembling Tr1 or Th17 cells, and expressed RORγt, which promotes IL-17 and blocks IL-2 production. T(FN) cells lacked suppressive activity against CD4+CD25- T-cell proliferation and failed to control effector T-cell expansion. However, T(FN) cells competed efficiently with effector T cells during homeostatic expansion. Foxp3 transcription was retained in T(R) cells but not in T(FN) cells, indicating that Foxp3 function is necessary for stable expression. Global gene expression profiling revealed that Foxp3 amplifies and stabilizes genes associated with T(R) cell characteristics, including cell surface markers like CD25, GITR, and CTLA4. Foxp3-dependent genes were enriched for cell surface and extracellular proteins, and Foxp3 promotes cell cycle-related genes, enhancing T(R) cell proliferation. Foxp3 also regulates PDE3B expression, which affects cAMP signaling and T(R) cell homeostasis. Reduced PDE3B expression in T(R) cells supports normal homeostasis and gene expression. Ectopic PDE3B expression in T(R) cells reduced their numbers, indicating that PDE3B repression is essential for T(R) cell maintenance. These findings suggest that Foxp3 consolidates anergic and paracrine IL-2