αβ T‐cell receptors with a central CDR3 cysteine are enriched in CD8αα intraepithelial lymphocytes and their thymic precursors

Abstract

The thymus plays a crucial role in immune tolerance by exposing developing T cells (thymocytes) to a myriad of self-antigens. Strong T-cell receptor (TCR) engagement induces tolerance in self-reactive thymocytes by stimulating apoptosis or selection into specialized T-cell lineages, including intestinal TCRαβ CD8αα intraepithelial lymphocytes (IEL). TCR-intrinsic amino acid motifs that can be used to predict whether a TCR will be strongly self-reactive remain elusive. Here, a novel TCR sequence alignment approach revealed that T-cell lineages in C57BL/6 mice had divergent usage of cysteine within two positions of the amino acid at the apex of the complementarity-determining region 3 (CDR3) of the TCRα or TCRβ chain. Compared to pre-selection thymocytes, central CDR3 cysteine usage was increased in IEL and Type A IEL precursors (IELp) and markedly decreased in Foxp3 regulatory T cells (T-reg) and naïve T cells. These findings reveal a TCR-intrinsic motif that distinguishes Type A IELp and IEL from T-reg and naïve T cells.

Linking T cell receptor sequence to transcriptional profiles with clonotype neighbor graph analysis (CoNGA)

“…These sequence characteristics are strikingly similar to features identified in a comparison of MHC-independent versus MHC-restricted TCR sequences from an experimental study of TCR repertoires in MHC-knockout mice 20 . Similar trends were also seen in comparisons of simulated and measured TCR sequences from pre-versus post-selection repertoires [21][22][23] , and in CD8aa intraepithelial lymphocytes and their thymic precursors 24,25 .…”

Cysteine and hydrophobic residues in CDR3 serve as distinct T-cell self-reactivity indices

“…1,5 The frequency of TCRa and/or TCRb sequences with cysteine within 2 positions of the CDR3 apex (cysteine index) was also greater in type A IELps than in other thymic and peripheral T-cell subsets in human subjects (Fig 1, G) and mice (Fig 1, H). 4 This was not due to preferential use of TCR gene segments because none of the cysteine residues detected by using the cysteine index in this study were encoded by germline codons; rather, they were encoded by codons spanning the junctions of TCR gene segments. Unlike the hydrophobic index, the cysteine index was not greater in Treg cells than in CD4 1 Tconv cells, suggesting that T cells with cysteine within 2 positions of the CDR3 apex typically acquire tolerance in the thymic cortex at a stage preceding the onset of Treg cell selection.…”

“…Although most self-reactive T cells are deleted in the thymus, rare thymocytes survive strong self-antigen engagement and differentiate into type A precursors of CD8aa 1 intestinal intraepithelial lymphocytes (type A IELps) in the thymic cortex 1 or CD4 1 forkhead box P3-positive regulatory T (Treg) cells in the thymic medulla. 2 Because TCR sequencing revealed amino acid motifs in complementarity-determining region 3 (CDR3) that are enriched in type A IELps or Treg cells, 3,4 we reasoned that the expression of these self-reactive TCR motifs might serve as a biomarker to evaluate T-cell self-tolerance.…”

“…Human type A IELps and Treg cells expressed more Helios than CD4 1 and CD8 1 Tconv cells (Fig 1, B), which is indicative of stronger TCR signaling. TCR-sequencing (for polymerase chain reaction primers, see Table E3 in this article's Online Repository at www.jacionline.org) revealed that, compared with other lineages, type A IELps used more proximal TCRa variable (TRAV/Trav) and joining (TRAJ/Traj) gene segments in human subjects (Fig 1, C) and mice (Fig 1, D), 4 indicating that type A IELps tend to receive a TCR signal early after initiating TCRa recombination. 5 Compared with CD4 1 Tconv cells, Treg cells are enriched in hydrophobic amino acid doublets at positions 6 and 7 of the CDR3 in the TCRb chain (CDR3b), a biomarker of self-reactivity.…”

“…ns, P > .05. Mouse data inFig 1, H,have been published4 and are included here to enable comparison with human data.…”