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Hypervariable T cell receptors (TCRs) play a key role in adaptive immunity, recognizing a vast diversity of pathogen-derived antigens. Our ability to extract clinically relevant information from large high-throughput sequencing of TCR repertoires (RepSeq) data is limited, because little is known about TCR–disease associations. We present Antigen-specific Lymphocyte Identification by Clustering of Expanded sequences (ALICE), a statistical approach that identifies TCR sequences actively involved in current immune responses from a single RepSeq sample and apply it to repertoires of patients with a variety of disorders—patients with autoimmune disease (ankylosing spondylitis [AS]), under cancer immunotherapy, or subject to an acute infection (live yellow fever [YF] vaccine). We validate the method with independent assays. ALICE requires no longitudinal data collection nor large cohorts, and it is directly applicable to most RepSeq datasets. Its results facilitate the identification of TCR variants associated with diseases and conditions, which can be used for diagnostics and rational vaccine design. [ABSTRACT FROM AUTHOR]

T cell receptor (TCR) repertoire data contain information about infections that could be used in disease diagnostics and vaccine development, but extracting that information remains a major challenge. Here we developed a statistical framework to detect TCR clone proliferation and contraction from longitudinal repertoire data. We applied this framework to data from three pairs of identical twins immunized with the yellow fever vaccine. We identified 600 to 1,700 responding TCRs in each donor and validated them using three independent assays. While the responding TCRs were mostly private, albeit with higher overlap between twins, they could be well-predicted using a classifier based on sequence similarity. Our method can also be applied to samples obtained postinfection, making it suitable for systematic discovery of new infection-specific TCRs in the clinic. [ABSTRACT FROM AUTHOR]

Objective. The risk of AS is associated with genomic variants related to antigen presentation and specific cytokine signalling pathways, suggesting the involvement of cellular immunity in disease initiation/progression. The aim of the present study was to explore the repertoire of TCR sequences in healthy donors and AS patients to uncover AS-linked TCR variants. Methods. Using quantitative molecular-barcoded 50-RACE, we performed deep TCR β repertoire profiling of peripheral blood (PB) and SF samples for 25 AS patients and 108 healthy donors. AS-linked TCR variants were identified using a new computational approach that relies on a probabilistic model of the VDJ rearrangement process. Results. Using the donor-agnostic probabilistic model, we reveal a TCR β motif characteristic for PB of AS patients, represented by eight highly homologous amino acid sequence variants. Some of these variants were previously reported in SF and PB of patients with ReA and in PB of AS patients. We demonstrate that identified AS-linked clones have a CD8+ phenotype, present at relatively low frequencies in PB, and are significantly enriched in matched SF samples of AS patients. Conclusion. Our results suggest the involvement of a particular antigen-specific subset of CD8+ T cells in AS pathogenesis, confirming and expanding earlier findings. The high similarity of the clonotypes with the ones found in ReA implies common mechanisms for the initiation of the diseases. [ABSTRACT FROM AUTHOR]

Vaccination against influenza is widely used to protect against seasonal flu epidemic although its effectiveness is debated. Here we performed deep quantitative T cell receptor repertoire profiling in peripheral blood of a healthy volunteer in response to trivalent subunit influenza vaccine. We did not observe significant rebuilding of peripheral blood T cell receptors composition in response to vaccination. However, we found several clonotypes in memory T cell fraction that were undetectable before the vaccination and had a maximum concentration at day 45 after vaccine administration. These cells were found in lower concentration in the course of repertoire monitoring for two years period. Our observation suggests a potential for recruitment of only a limited number of new T cells after each seasonal influenza vaccination. [ABSTRACT FROM AUTHOR]

The diversity of T-cell receptors recognizing foreign pathogens is generated through a highly stochastic recombination process, making the independent production of the same sequence rare. Yet unrelated individuals do share receptors, which together constitute a “public” repertoire of abundant clonotypes. The TCR repertoire is initially formed prenatally, when the enzyme inserting random nucleotides is downregulated, producing a limited diversity subset. By statistically analyzing deep sequencing T-cell repertoire data from twins, unrelated individuals of various ages, and cord blood, we show that T-cell clones generated before birth persist and maintain high abundances in adult organisms for decades, slowly decaying with age. Our results suggest that large, low-diversity public clones are created during pre-natal life, and survive over long periods, providing the basis of the public repertoire. [ABSTRACT FROM AUTHOR]

Background: The Immunoglobulins (IG) and the T cell receptors (TR) play the key role in antigen recognition during the adaptive immune response. Recent progress in next-generation sequencing technologies has provided an opportunity for the deep T cell receptor repertoire profiling. However, a specialised software is required for the rational analysis of massive data generated by next-generation sequencing. Results: Here we introduce tcR, a new R package, representing a platform for the advanced analysis of T cell receptor repertoires, which includes diversity measures, shared T cell receptor sequences identification, gene usage statistics computation and other widely used methods. The tool has proven its utility in recent research studies. Conclusions: tcR is an R package for the advanced analysis of T cell receptor repertoires after primary TR sequences extraction from raw sequencing reads. The stable version can be directly installed from The Comprehensive R Archive Network (http://cran.r-project.org/mirrors.html). The source code and development version are available at tcR GitHub (http://imminfo.github.io/tcr/) along with the full documentation and typical usage examples. [ABSTRACT FROM AUTHOR]

Adaptive immunity in humans is provided by hypervariable Ig-like molecules on the surface of B and T cells. The final set of these molecules in each organism is formed under the influence of two forces: individual genetic traits and the environment, which includes the diverse spectra of alien and self-antigens. Here we assess the impact of individual genetic factors on the formation of the adaptive immunity by analyzing the T-cell receptor (TCR) repertoires of three pairs of monozygous twins by next-generation sequencing. Surprisingly, we found that an overlap between the TCR repertoires of monozygous twins is similar to an overlap between the TCR repertoires of nonrelated individuals. However, the number of identical complementary determining region 3 sequences in two individuals is significantly increased for twin pairs in the fraction of highly abundant TCR molecules, which is enriched by the antigen-experienced T cells. We found that the initial recruitment of particular TCR V genes for recombination and subsequent selection in the thymus is strictly determined by individual genetic factors. J genes of TCRs are selected randomly for recombination; however, the subsequent selection in the thymus gives preference to some α but not β J segments. These findings provide a deeper insight into the mechanism of TCR repertoire generation. [ABSTRACT FROM AUTHOR]

Background: Adaptive immune responses to newly encountered pathogens depend on the mobilization of antigen-specific clonotypes from a vastly diverse pool of naive T cells. Using recent advances in immune repertoire sequencing technologies, models of the immune receptor rearrangement process, and a database of annotated T cell receptor (TCR) sequences with known specificities, we explored the baseline frequencies of T cells specific for defined human leukocyte antigen (HLA) class I-restricted epitopes in healthy individuals. Methods: We used a database of TCR sequences with known antigen specificities and a probabilistic TCR rearrangement model to estimate the baseline frequencies of TCRs specific to distinct antigens epitopespecificT-cells. We verified our estimates using a publicly available collection of TCR repertoires from healthy individuals. We also interrogated a database of immunogenic and non-immunogenic peptides is used to link baseline T-cell frequencies with epitope immunogenicity. Results: Our findings revealed a high degree of variability in the prevalence of T cells specific for different antigens that could be explained by the physicochemical properties of the corresponding HLA class I-bound peptides. The occurrence of certain rearrangements was influenced by ancestry and HLA class I restriction, and umbilical cord blood samples contained higher frequencies of common pathogen-specific TCRs. We also identified a quantitative link between specific T cell frequencies and the immunogenicity of cognate epitopes presented by defined HLA class I molecules. Conclusions: Our results suggest that the population frequencies of specific T cells are strikingly non-uniform across epitopes that are known to elicit immune responses. This inference leads to a new definition of epitope immunogenicity based on specific TCR frequencies, which can be estimated with a high degree of accuracy in silico, thereby providing a novel framework to integrate computational and experimental genomics with basic and translational research efforts in the field of T cell immunology. [ABSTRACT FROM AUTHOR]

SARS-CoV-2 mRNA vaccines induce robust anti-spike (S) antibody and CD4+ T cell responses. It is not yet clear whether vaccine-induced follicular helper CD4+ T (T FH) cell responses contribute to this outstanding immunogenicity. Using fine-needle aspiration of draining axillary lymph nodes from individuals who received the BNT162b2 mRNA vaccine, we evaluated the T cell receptor sequences and phenotype of lymph node T FH. Mining of the responding T FH T cell receptor repertoire revealed a strikingly immunodominant HLA-DPB1∗04-restricted response to S 167–180 in individuals with this allele, which is among the most common HLA alleles in humans. Paired blood and lymph node specimens show that while circulating S-specific T FH cells peak one week after the second immunization, S-specific T FH persist at nearly constant frequencies for at least six months. Collectively, our results underscore the key role that robust T FH cell responses play in establishing long-term immunity by this efficacious human vaccine. [Display omitted] • SARS-CoV-2 vaccines induce robust human T FH cell responses in draining lymph nodes • HLA-DPB1∗04 restricts the immunodominant SARS-CoV-2 S 167–180 epitope • S 167–180 is recognized by T cell receptors with a public α-chain motif • S-specific T FH cells are maintained in draining lymph nodes 6 months after vaccination Analysis of draining lymph nodes of individuals vaccinated with BNT162b2 mRNA vaccine against SARS-CoV-2 identifies viral-spike-specific follicular helper CD4+ T cells that persist for months and contribute to long-term immunity. [ABSTRACT FROM AUTHOR]

COVID-19 is a global pandemic caused by the SARS-CoV-2 coronavirus. T cells play a key role in the adaptive antiviral immune response by killing infected cells and facilitating the selection of virus-specific antibodies. However, neither the dynamics and cross-reactivity of the SARS-CoV-2-specific T-cell response nor the diversity of resulting immune memory is well understood. In this study, we use longitudinal high-throughput T-cell receptor (TCR) sequencing to track changes in the T-cell repertoire following two mild cases of COVID-19. In both donors, we identified CD4+ and CD8+ T-cell clones with transient clonal expansion after infection. We describe characteristic motifs in TCR sequences of COVID-19-reactive clones and show preferential occurrence of these motifs in publicly available large dataset of repertoires from COVID-19 patients. We show that in both donors, the majority of infection-reactive clonotypes acquire memory phenotypes. Certain T-cell clones were detected in the memory fraction at the pre- infection time point, suggesting participation of pre-existing cross-reactive memory T cells in the immune response to SARS-CoV-2. [ABSTRACT FROM AUTHOR]

Immune repertoires provide a unique fingerprint reflecting the immune history of individuals, with potential applications in precision medicine. However, the question of how personal that information is and how it can be used to identify individuals has not been explored. Here, we show that individuals can be uniquely identified from repertoires of just a few thousands lymphocytes. We present "Immprint," a classifier using an information-theoretic measure of repertoire similarity to distinguish pairs of repertoire samples coming from the same versus different individuals. Using published T-cell receptor repertoires and statistical modeling, we tested its ability to identify individuals with great accuracy, including identical twins, by computing false positive and false negative rates < 10−6 from samples composed of 10,000 T-cells. We verified through longitudinal datasets that the method is robust to acute infections and that the immune fingerprint is stable for at least three years. These results emphasize the private and personal nature of repertoire data. Author summary: Immune repertoires are a trove of personal information: unique to each individual, they are also windows into their past and future health. Thanks to their potential for personalized medicine and progress of sequencing technologies, these repertoires are now routinely sequenced. As a consequence they raise the question of identifiability of samples. In this paper, we estimate the quantity of immune cells needed to associate two samples from the same individual: as little as a finger prick worth of blood can serve as an immune fingerprint that can distinguish even identical twins, without giving away genetic information about non-consenting relatives. We show that this fingerprint is stable through time, and is not erased during infections or vaccinations. [ABSTRACT FROM AUTHOR]

T cell maturation and activation depend upon T cell receptor (TCR) interactions with a wide variety of antigenic peptides displayed in a given major histocompatibility complex (MHC) context. Complementarity-determining region 3 (CDR3) is the most variable part of the TCRα and -β chains, which govern interactions with peptide-MHC complexes. However, it remains unclear how the CDR3 landscape is shaped by individual MHC context during thymic selection of naïve T cells. We established two mouse strains carrying distinct allelic variants of H2-A and analyzed thymic and peripheral production and TCR repertoires of naïve conventional CD4+ T (Tconv) and naïve regulatory CD4+ T (Treg) cells. Compared with tuberculosis-resistant C57BL/6 (H2-Ab) mice, the tuberculosis-susceptible H2-Aj mice had fewer CD4+ T cells of both subsets in the thymus. In the periphery, this deficiency was only apparent for Tconv and was compensated for by peripheral reconstitution for Treg. We show that H2-Aj favors selection of a narrower and more convergent repertoire with more hydrophobic and strongly interacting amino acid residues in the middle of CDR3α and CDR3β, suggesting more stringent selection against a narrower peptide-MHC-II context. H2-Aj and H2-Ab mice have prominent reciprocal differences in CDR3α and CDR3β features, probably reflecting distinct modes of TCR fitting to MHC-II variants. These data reveal the mechanics and extent of how MHC-II shapes the naïve CD4+ T cell CDR3 landscape, which essentially defines adaptive response to infections and self-antigens. [ABSTRACT FROM AUTHOR]

The diversity, architecture, and dynamics of the TCR repertoire largely determine our ability to effectively withstand infections and malignancies with minimal mistargeting of immune responses. In this study, we have employed deep TCRβ repertoire sequencing with normalization based on unique molecular identifiers to explore the long-term dynamics of T cell immunity. We demonstrate remarkable stability of repertoire, where approximately half of all T cells in peripheral blood are represented by clones that persist and generally preserve their frequencies for 3 y.We further characterize the extremes of lifelong TCR repertoire evolution, analyzing samples ranging from umbilical cord blood to centenarian peripheral blood. We show that the fetal TCR repertoire, albeit structurally maintained within regulated borders due to the lower numbers of randomly added nucleotides, is not limited with respect to observed functional diversity. We reveal decreased efficiency of nonsense-mediated mRNA decay in umbilical cord blood, which may reflect specific regulatory mechanisms in development. Furthermore, we demonstrate that human TCR repertoires are functionally more similar at birth but diverge during life, and we track the lifelong behavior of CMV- and EBV- specific T cell clonotypes. Finally, we reveal gender differences in dynamics of TCR diversity constriction, which come to naught in the oldest age. Based on our data, we propose a more general explanation for the previous observations on the relationships between longevity and immunity. [ABSTRACT FROM AUTHOR]

Despite the growing number of immune repertoire sequencing studies, the field still lacks software for analysis and comprehension of this high-dimensional data. Here we report VDJtools, a complementary software suite that solves a wide range of T cell receptor (TCR) repertoires post-analysis tasks, provides a detailed tabular output and publication-ready graphics, and is built on top of a flexible API. Using TCR datasets for a large cohort of unrelated healthy donors, twins, and multiple sclerosis patients we demonstrate that VDJtools greatly facilitates the analysis and leads to sound biological conclusions. VDJtools software and documentation are available at . [ABSTRACT FROM AUTHOR]

As the establishment of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific T cell memory in children remains largely unexplored, we recruited convalescent COVID-19 children and adults to define their circulating memory SARS-CoV-2-specific CD4+ and CD8+ T cells prior to vaccination. We analyzed epitope-specific T cells directly ex vivo using seven HLA class I and class II tetramers presenting SARS-CoV-2 epitopes, together with Spike-specific B cells. Unvaccinated children who seroconverted had comparable Spike-specific but lower ORF1a- and N-specific memory T cell responses compared with adults. This agreed with our TCR sequencing data showing reduced clonal expansion in children. A strong stem cell memory phenotype and common T cell receptor motifs were detected within tetramer-specific T cells in seroconverted children. Conversely, children who did not seroconvert had tetramer-specific T cells of predominantly naive phenotypes and diverse TCRαβ repertoires. Our study demonstrates the generation of SARS-CoV-2-specific T cell memory with common TCRαβ motifs in unvaccinated seroconverted children after their first virus encounter. [Display omitted] • SARS-CoV-2-specific T cells detected ex vivo in unvaccinated seroconverted children • Comparable Spike-T cell memory responses but lower ORF1a/N-responses found in children • Reduced clonal expansions within tetramer+ T cells seen in children compared with adults • Prevalent Tscm phenotype and TCR motif occur in SARS-CoV-2-specific T cells in children Children are at lower risk of developing severe COVID-19, yet their primary and memory immune responses are understudied. Rowntree et al. define ex vivo CD8+ and CD4+ T cell responses toward SARS-CoV-2 using peptide-HLA tetramers. They find that children have comparable Spike-specific but lower ORF1a- and N-specific memory T cell magnitude with less clonal expansion in comparison with adults. [ABSTRACT FROM AUTHOR]

Retroelement activity is a common source of polymorphisms in human genome. The mechanism whereby retroelements contribute to the intraindividual genetic heterogeneity by inserting into the DNA of somatic cells is gaining increasing attention. Brain tissues are suspected to accumulate genetic heterogeneity as a result of the retroelements somatic activity. This study aims to expand our understanding of the role retroelements play in generating somatic mosaicism of neural tissues. Whole-genome Alu and L1 profiling of genomic DNA extracted from the cerebellum, frontal cortex, subventricular zone, dentate gyrus, and the myocardium revealed hundreds of somatic insertions in each of the analyzed tissues. Interestingly, the highest concentration of such insertions was detected in the dentate gyrus—the hotspot of adult neurogenesis. Insertions of retroelements and their activity could produce genetically diverse neuronal subsets, which can be involved in hippocampal-dependent learning and memory. [ABSTRACT FROM AUTHOR]