The Molecular Basis of Self-Antigen Recognition by Regulatory T Cells

Foxp3-expressing CD4+ regulatory T (Treg) cells are critical for the prevention of autoimmunity and the maintenance of immune homeostasis. The recent identification of self-peptide/MHC class II ligands recognized by thymic-derived Treg cells by our group and the Savage group has set the foundation for new studies into self-antigen recognition by Treg cells. We analyzed the T cell receptor (TCR) repertoire of Treg cells and T conventional (Tconv) cells reactive to a peptide (named "C4") derived from the prostate-specific protein Tcaf3 complexed with I-Ab using a droplet-based sequencing approach. We conducted sequencing in both C4-sufficient wild-type mice and mice with a targeted deletion of the C4 peptide, allowing us to analyze a "foreign" TCR response to the same epitope. We found negligible overlap in Treg and Tconv TCR repertoires for this single antigen and identified an abundant public C4-specific Treg cell clone in wild-type mice that is absent in C4-deficient mice. Through biochemical studies of both Treg and Tconv TCRs found in this dataset, we found that C4/I-Ab-specific Treg TCRs recognized ligand with higher affinity and longer half-life compared to TCRs expressed by C4/I-Ab-specific Tconv cells, supporting a TCR-intrinsic model of Treg cell development in which TCR-peptide/MHC-II interactions of higher stability promote Treg cell development. Transcriptional analysis of C4-specific T cells also revealed genetic markers and properties of Treg and Tconv cells elicited in both wild-type and C4-deficient mice, revealing candidate genes that may be important for optimal Treg cell fitness and function. Thus, our cumulative evidence supports a model in which TCR-self-peptide/MHC-II interactions of high stability promote Treg cell differentiation in the thymic medulla, leading to the establishment of distinct peripheral pools of antigen-specific Tconv and Treg cells.