Single-cell TCR sequencing of gut intraepithelial γδ T cells reveals a vast and diverse repertoire in celiac disease

A hallmark of celiac disease (CeD), a chronic condition driven by cereal gluten exposure, is increase of gut intraepithelial gamma delta T cells. This may indicate pathogenic involvement of gamma delta T cells and existence of disease-specific gamma delta T-cell receptors (TCRs) recognizing defined antigen(s). We performed high-throughput and paired gamma delta TCR sequencing of single intraepithelial gamma delta T cells of untreated CeD patients (n = 8; 1821 cells), CeD patients treated with a gluten-free diet (n = 5; 436 cells) and controls (n = 7; 1068 cells). We found that CeD patients, both untreated and treated, had larger and more diverse gamma delta TCR repertoires, more frequent usage of TRDV1 and TRDV3 and different patterns of TCR gamma/TCR delta-pairing compared with controls. Although we observed no public CDR3 delta sequences, there were several public CDR3 gamma sequences-many of which were shared by not only the CeD patients, but also by the controls. These public CDR3s were characterized by few N/P nucleotide insertions with germline and near-germline configuration, hence being easy to generate. Previous findings of CeD-specific CDR3 motifs were not replicated. Thus, being unable to raise evidence for CeD-specific gamma delta TCRs in this first large, paired gamma delta TCR single-cell sequencing study, we project challenges for identification of CeD-relevant gamma delta TCR ligands.