At the heart of an immune response are antigen-presenting cells, sentinels of the immune system that are uniquely poised to initiate the activation of T cells and regulate the balance between inflammatory immune responses and immune tolerance. Until recently, the identity of the “tolerogenic” antigen-presenting cell type that instructs peripheral regulatory T (pTreg) cell differentiation was not known. Our earlier work uncovered a new lineage of antigen-presenting cells named Thetis cells, encompassing four subsets (TC I–IV). Using genetic approaches to ablate key molecules on TCs, we found that TC IV plays a critical role in peripheral Treg cell generation and tolerance to both gut microbiota and food antigens. A notable feature of TCs was their transcriptional similarity to Aire+ medullary thymic epithelial cells, highlighting parallels between pathways of tolerance to self and foreign antigens in the thymus and periphery, respectively. In recent work, we have defined the ontogeny of Thetis cells and the transcription factors that promote their differentiation and cellular heterogeneity. Collectively, our studies define a new lineage of antigen-presenting cells, distinct from conventional dendritic cells, that includes a subset of cells uniquely equipped with the ability to induce tolerance to food and microbial antigens. Understanding the cues that drive TC differentiation and the mechanisms by which they promote T cell tolerance may lead to novel therapeutic approaches in autoimmunity, food allergy and inflammatory bowel disease.