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Clonal Deletion versus Clonal Diversion: Footprints of Self-Tolerance in the T Cell Repertoire

Periodic Reporting for period 3 - TOLERANCE FOOTPRINT (Clonal Deletion versus Clonal Diversion: Footprints of Self-Tolerance in the T CellRepertoire)

Reporting period: 2020-10-01 to 2022-03-31

Self-tolerance is a key feature of the immune system; its failure causes autoimmune diseases such as Multiple Sclerosis or Type-1-Diabetes. Understanding the determinants and physiological mechanisms of T cell tolerance, but also of its breakdown, will pave the way to rationale and more specific approaches to the treatment of autoimmune diseases. Remarkably, T cell tolerance operates via two fundamentally different mechanisms: potentially dangerous cells are either eliminated (clonal deletion) or re-programmed to differentiate into regulatory T (Treg) cells (clonal diversion). Paradoxically, both tolerance modes can ensue from self-antigen-encounter in the thymus, and the parameters specifying these opposing cell-fates remain poorly understood. Moreover, the relative contribution of clonal deletion versus clonal diversion to tolerance at the level of diverse immune cell repertoires has not been determined. In particular, the paucity of antigen-specific cells is a major experimental obstacle to unravelling to what extent cells with shared autoreactive specificity, yet different T cell receptors (TCRs), are subject to either mode of tolerance.
Based upon breakthroughs in visualizing minute cohorts of antigen-specific cells, we are establishing methods to identify and characterize minute cohorts (around 100 cells) of autoreactive T cells within diverse repertoires of millions of T cells with other antigen specificities. Using this methodology, we aim to reveal where and how tolerance either generates ‘holes’ in the repertoire or diverts cells into a ‘benign’ sub-repertoire of regulatory T cells. Our main objectives are to identify the determinants that specify clonal deletion versus clonal diversion. Our research elucidates a fundamental aspect of vertebrate immunology, but also has major implications regarding the therapeutic promise of harnessing endogenous, antigen-specific Treg cells in autoimmunity.
We have combined MHC tetramer technology and single-cell TCR sequencing to generate inventories of autoreactive CD4 T cells in the naïve versus tolerant repertoire. This revealed that some TCRs were ‘lost’ whereas others mediated Treg cell differentiation. The antigen responsiveness of these TCRs strongly supports an affinity model of central tolerance. Moreover, the contribution of different ‘Treg diverter’ TCRs to the nascent thymic Treg cell population reflected their antigen reactivity rather than their frequency among precursors. Our findings reveal a multilayered affinity hierarchy that not only separates deleted from diverted TCRs, but also generates a Treg cell compartment containing cells of optimal antigen reactivity.
So far, experimental evidence in favor of an avidity model of thymic selection was in essence limited to observations related to variations of antigen dose in cell receptor (TCR) transgenic systems that harbor unphysiological frequencies of cells of a given antigen specificity. However, it has become clear that monoclonal models only poorly recapitulate physiological Treg cell ‘instruction’, and close to nothing was known with regards to how clonal deletion and Treg cell diversion together shape the quality and quantity of polyclonal cohorts of antigen-specific CD4 T cells. Our work on the fate of autoreactive cells within naturally diverse T cell repertoires has shed light on TCR affinity as a crucial determinant of the decision between clonal deletion and clonal diversion (Treg cell differentiation). On the basis of these findings, we will now proceed to ask how various stromal cell types of the thymus shape the diverse T cell repertoire and how perturbed tolerance induction by these distinct cell types may lead to autoimmunity.