T cells are defined by a heterodimeric surface receptor (the T cell receptor or TCR) that mediates recognition of pathogen-associated epitopes via interactions with peptide-major histocompatibility complexes (pMHC). TCRs are generated by genomic rearrangements of the germline TCR locus, a process termed V(D)J recombination that has the potential to generate a staggering diversity of TCRs (estimated to range from 1015 1 to as high as 1061 2 possible receptors). Despite this potential diversity, TCRs from T cells that recognize the same pMHC epitope often share conserved sequence features, suggesting that it may be possible to predictively model epitope specificity. Here we report the in-depth characterization of ten epitope-specific CD8+ TCR repertoires from mice and humans representing 4600+ in-frame, single cell-derived TCRαβ sequence pairs from 110 subjects. We developed novel analytical tools to characterize these epitope-specific repertoires: a distance measure on the space of TCRs that permits clustering and visualization (TCRdist), a robust repertoire diversity metric (TCRdiv) that accommodates the low number of paired public receptors observed when compared to single chain analyses, and a distance-based classifier capable of assigning previously unobserved TCRs to characterized repertoires with robust sensitivity and specificity. Our analysis demonstrates that each epitope-specific repertoire contains a clustered group of receptors that share core sequence similarities, together with a dispersed set of diverse “outlier” sequences. By identifying shared motifs in core sequences, we were able to highlight key conserved residues driving essential elements of TCR recognition. These analyses provide insights into the generalizable, underlying features of epitope-specific repertoires and adaptive immune recognition.

The ability to decode antigen specificities encapsulated in the sequences of rearranged T-cell receptor (TCR) genes is critical for our understanding of the adaptive immune system and promises significant advances in the field of translational medicine. Recent developments in high-throughput sequencing methods (immune repertoire sequencing technology, or RepSeq) and single-cell RNA sequencing technology have allowed us to obtain huge numbers of TCR sequences from donor samples and link them to T-cell phenotypes. However, our ability to annotate these TCR sequences still lags behind, owing to the enormous diversity of the TCR repertoire and the scarcity of available data on T-cell specificities. In this paper, we present VDJdb, a database that stores and aggregates the results of published T-cell specificity assays and provides a universal platform that couples antigen specificities with TCR sequences. We demonstrate that VDJdb is a versatile instrument for the annotation of TCR repertoire data, enabling a concatenated view of antigen-specific TCR sequence motifs. VDJdb can be accessed at https://vdjdb.cdr3.net and https://github.com/antigenomics/vdjdb-db.

Influenza A, B and C viruses (IAV, IBV, ICV) circulate globally and infect humans, with IAV/IBV causing most severe disease. While CD8 + T-cells confer cross-protection against different IAV strains, CD8 + T-cell responses to IBV/ICV are understudied. We dissected the CD8 + T-cell cross-reactome against influenza viruses and provided the first evidence of CD8 + T-cell cross-reactivity across IAV, IBV and ICV. Using immunopeptidomics, we identified immunodominant CD8 + T-cell epitopes from IBV, protective in mice, and found prominent memory CD8 + T-cells towards both universal and influenza type-specific epitopes in blood and lungs of healthy humans, with lung-derived CD8 + T-cells displaying a tissue-resident phenotype. Importantly, effector CD38 + Ki67 + CD8 + T-cells against novel epitopes were readily detected in IAV-and IBV-infected pediatric and adult patients. Our study introduces a new paradigm, whereby CD8 + T-cells confer unprecedented cross-reactivity across all influenza viruses, a key finding for designing universal vaccines.