Foxp3+ regulatory T (T reg) cells play a key role in controlling immune pathological re actions. Many develop their regulatory activity in the thymus, but there is also evidence for development of Foxp3+ T reg cells from naive precursors in the periphery. Recent studies have shown that transforming growth factor (TGF)-β can promote T reg cell development in culture, but little is known about the cellular and molecular mechanisms that mediate this pathway under more physiological conditions. Here, we show that after antigen activation in the intestine, naive T cells acquire expression of Foxp3. Moreover, we identify a population of CD103+ mesenteric lymph node dendritic cells (DCs) that induce the devel opment of Foxp3+ T reg cells. Importantly, promotion of T reg cell responses by CD103+ DCs is dependent on TGF-β and the dietary metabolite, retinoic acid (RA). These results newly identify RA as a cofactor in T reg cell generation, providing a mechanism via which functionally specialized gut-associated lymphoid tissue DCs can extend the repertoire of T reg cells focused on the intestine.

To maintain immune homeostasis, the intestinal immune system has evolved redundant regulatory strategies. In this regard, the gut is home to a large number of regulatory T (T reg) cells, including the Foxp3+ T reg cell. Therefore, we hypothesized that the gut environment preferentially supports extrathymic T reg cell development. We show that peripheral conversion of CD4+ T cells to T reg cells occurs primarily in gut-associated lymphoid tissue (GALT) after oral exposure to antigen and in a lymphopenic environment. Dendritic cells (DCs) purified from the lamina propria (Lp; LpDCs) of the small intestine were found to promote a high level of T reg cell conversion relative to lymphoid organ–derived DCs. This enhanced conversion by LpDCs was dependent on TGF-β and retinoic acid (RA), which is a vitamin A metabolite highly expressed in GALT. Together, these data demonstrate that the intestinal immune system has evolved a self-contained strategy to promote T reg cell neoconversion.

The intestinal tract is in intimate contact with the commensal microflora. Nevertheless, how commensals communicate with cells to ensure immune homeostasis is still unclear. In this study, we found that gut flora DNA (gfDNA) plays a major role in intestinal homeostasis through toll like receptor 9 engagement. TLR9 deficient mice displayed increased frequencies of CD4+Foxp3+ regulatory T cells (Treg) within intestinal effector sites, and reduced constitutive IL-17 and IFN-γ producing effector T cells (Teff). Complementing this, gfDNA limited lamina propria dendritic cells induced Treg conversion in vitro. Further, Treg/Teff disequilibrium in TLR9−/− mice led to impaired immune responses to oral infection and to oral vaccination, which were rescued through neutralization of Treg. Impaired intestinal immune responses were recapitulated in mice treated with antibiotics and were reversible following reconstitution with gfDNA. Together these data point to gfDNA as a natural adjuvant for priming intestinal responses via modulation of Treg/Teff equilibrium.