Thymic regulatory T cells arise via two distinct developmental programs

The developmental programs that generate a broad repertoire of regulatory T cells (T-reg cells) able to respond to both self antigens and non-self antigens remain unclear. Here we found that mature T-reg cells were generated through two distinct developmental programs involving CD25(+) T-reg cell progenitors (CD25(+) TregP cells) and Foxp3(lo) T-reg cell progenitors (Foxp3(lo) TregP cells). CD25(+) TregP cells showed higher rates of apoptosis and interacted with thymic self antigens with higher affinity than did Foxp3(lo) TregP cells, and had a T cell antigen receptor repertoire and transcriptome distinct from that of Foxp3(lo) TregP cells. The development of both CD25(+) TregP cells and Foxp3(lo) TregP cells was controlled by distinct signaling pathways and enhancers. Transcriptomics and histocytometric data suggested that CD25(+) TregP cells and Foxp3(lo) TregP cells arose by coopting negative-selection programs and positive-selection programs, respectively. T-reg cells derived from CD25(+) TregP cells, but not those derived from Foxp3(lo) TregP cells, prevented experimental autoimmune encephalitis. Our findings indicate that T-reg cells arise through two distinct developmental programs that are both required for a comprehensive T-reg cell repertoire capable of establishing immunotolerance.