AbstractThymic central tolerance eliminates most immature T cells with autoreactive T cell receptors (TCR) that recognize self MHC/peptide complexes. Regardless, an unknown number of autoreactive CD4⁺Foxp3⁻ T cells escape negative selection and in the periphery require continuous suppression by CD4⁺Foxp3⁺ regulatory cells (Tregs). Here, we compare immune repertoires of Treg-deficient and Treg-sufficient mice to find Tregs continuously constraining one-third of mature CD4⁺Foxp3⁻ cells from converting to pathogenic effectors in healthy mice. These dormant pathogenic clones frequently express TCRs activatable by ubiquitous autoantigens presented by class II MHCs on conventional dendritic cells, including self-peptides that select them in the thymus. Our data thus suggest that identification of most potentially autoreactive CD4⁺ T cells in the peripheral repertoire is critical to harness or redirect these cells for therapeutic advantage.