Gene Expression profile of RORt-lineage antigen presenting cells collected from secondary lymphoid tissues in adult mice.

A key feature of immune tolerance is the ability to distinguish pathogens from innocuous signals including dietary antigens, but the precise mechanisms of oral tolerance induction and the key cellular actors have remained unclear. Classical dendritic cells (cDCs) have previously been implicated in the generation of regulatory T cells (Tregs) and T-cell anergy in response to food antigens in gut-associated lymphoid tissue1, but here we demonstrate that RORt-lineage antigen-presenting cells (APCs), specifically RORt-lineage cells expressing the Autoimmune Regulator (Aire) gene referred to as RORt eTACs are the critical APC population required for oral tolerance induction. We reconcile the identity of these rare RORt-lineage APC populations across new and all known single-cell transcriptomic datasets to establish consensus identities and show that RORt eTACs2 are a unique myeloid-derived lineage distinct from innate lymphoid cells (ILCs) which are both necessary and sufficient for oral tolerance induction. We obtained mLN, spleen and other LNs (cervical, inguinal, axillary, brachial, and para-aortic) from adult RORt-lineage mice (RORt-CreR26-lsl-tdTmt) and FACS-sorted tdTmt+ MHCII+ cells for subsequent scRNA-seq