The Lower Limit of Regulatory CD4⁺ Foxp3⁺ TCRβ Repertoire Diversity Required To Control Autoimmunity

The TCR repertoire of regulatory T cells (Tregs) is highly diverse. The relevance of this diversity to maintain self-tolerance remains unknown. We established a model where the TCR repertoire of normal polyclonal Tregs was limited by serial transfers into IL-2Rβ^-/- mice, which lack functional Tregs. After a primary transfer, the donor Treg TCR repertoire was substantially narrowed, yet the recipients remained autoimmune-free. Importantly, upon purification and transfer of donor-derived Tregs from an individual primary recipient into neonatal IL-2Rβ^-/- mice, the secondary recipients developed autoimmunity. In this study, the Treg TCRβ repertoire was reshaped and further narrowed. In contrast, secondary IL-2Rβ recipients showed fewer symptoms of autoimmunity when they received donor Tregs that were premixed from several primary recipients to increase their TCRβ repertoire diversity. About 8-11% of the Treg TCRβ repertoire was estimated to be the minimum required to establish and maintain tolerance in primary IL-2Rβ^-/- recipients. Collectively, these data quantify where limitations imposed on the Treg TCRβ repertoire results in a population of Tregs that cannot fully suppress polyclonal autoreactive T cells. Our data favor a model where the high diversity of the Treg TCR provides a mechanism for Tregs to actively adapt and effectively suppress autoreactive T cells, which are not fixed, but are evolving as they encounter self-antigens.