A Diverse Lipid Antigen–Specific TCR Repertoire Is Clonally Expanded during Active Tuberculosis

Abstract

Human T cells that recognize lipid Ags presented by highly conserved CD1 proteins often express semi-invariant TCRs, but the true diversity of lipid Ag-specific TCRs remains unknown. We use CD1b tetramers and high-throughput immunosequencing to analyze thousands of TCRs from ex vivo-sorted or in vitro-expanded T cells specific for the mycobacterial lipid Ag, glucose monomycolate. Our results reveal a surprisingly diverse repertoire resulting from editing of germline-encoded gene rearrangements analogous to MHC-restricted TCRs. We used a distance-based metric (TCRDist) to show how this diverse TCR repertoire builds upon previously reported conserved motifs by including subject-specific TCRs. In a South African cohort, we show that TCRDist can identify clonal expansion of diverse glucose monomycolate-specific TCRs and accurately distinguish patients with active tuberculosis from control subjects. These data suggest that similar mechanisms govern the selection and expansion of peptide and lipid Ag-specific T cells despite the nonpolymorphic nature of CD1.

T Cell Responses to Mycobacterial Glycolipids: On the Spectrum of “Innateness”

“…On the other hand, CD1-restricted TCRs are expected to be public due to the non-polymorphic nature of CD1. Indeed, our group found that both shared and private TCR repertoires of GMM-specific T cells were associated with active TB disease in a South African cohort (35). Thus, TCRs specific for mycolipids could be developed into a blood-based diagnostic tool.…”

“…The development of CD1b tetramers for GMM, SGLs, and MA have enabled the isolation of mycolipid-specific T cells to study the T cell receptor (TCR) repertoire and functional diversity directly ex vivo (35)(36)(37)(38)(39)(40). TCR diversity among T cells specific for mycolipids has been best studied using GMM as the model antigen.…”

“…This TCR is termed the germlineencoded mycolyl-reactive (GEM) TCR and includes a conserved TCR-α that consists of a TRAV1-2/TRAJ9 gene rearrangement (38). However, recent data suggest that GMM-specific T cells expressing this conserved TCR may actually constitute the minority of GMM-specific T cells, which have been shown to express a more diverse TCR repertoire (35,39). GMM-specific TCRs have also been shown to display varying levels of affinity and specificity for GMM.…”

Techniques to Study Antigen-Specific B Cell Responses

“…One weakness of the flow cytometric approach is the reliance on antigens that can be readily conjugated to a fluorochrome or biotinylated. In addition to recombinant proteins and synthesized peptides, labeled polysaccharides, lipids, haptens, virus-like particles, and pseudo viruses have also been used to identify antigen-specific cells by flow cytometry (33,(43)(44)(45)(46)(47)(48)(49)(50)(51)(52)(53)(54)(55)(56)(57)(58)(59). Further, epitope-specific B cells have been identified by screening bacteriophage-displays or microarray peptide libraries with polyclonal antibodies targeting the native antigen to select conformational epitopes that can be fused to fluorescent proteins for use in flow cytometry (47,60).…”

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

“…Single-chain variants of the iMHC score were fit with L1-regularized logistic regression just as for the paired iMHC score. Subset labels are as follows: 'CD1b', GMM:CD1b-tetramer sorted T cells from DeWitt et al 44 ; 'VDJdb-MHC1', TCRs reported to bind to MHC class 1 presented epitopes in the VDJdb database 45 ; 'VDJdb-MHC2', TCRs reported to bind to MHC class 2 presented epitopes in the VDJdb database; 'dMAIT', diverse MAIT TCR sequences from Gherardin et al 46 2016); 'hobit', T cells belonging to the HOBIT+/HELIOS+ population in 10x 200k donors 1, 3, and 4.…”