Analysis of T-Cell Receptor Repertoire in Transplantation: Fingerprint of T Cell-mediated Alloresponse

T cells play a key role in determining allograft function by mediating allogeneic immune responses to cause rejection, and recent work pointed their role in mediating tolerance in transplantation. The unique T-cell receptor (TCR) expressed on the surface of each T cell determines the antigen specificity of the cell and can be the specific fingerprint for identifying and monitoring. Next-generation sequencing (NGS) techniques provide powerful tools for deep and high-throughput TCR profiling, and facilitate to depict the entire T cell repertoire profile and trace antigen-specific T cells in circulation and local tissues. Tailing T cell transcriptomes and TCR sequences at the single cell level provides a full landscape of alloreactive T-cell clones development and biofunction in alloresponse. Here, we review the recent advances in TCR sequencing techniques and computational tools, as well as the recent discovery in overall TCR profile and antigen-specific T cells tracking in transplantation. We further discuss the challenges and potential of using TCR sequencing-based assays to profile alloreactive TCR repertoire as the fingerprint for immune monitoring and prediction of rejection and tolerance.

Identification of CDRH3 Loops in the Naive B Cell Receptor Repertoire that Can Be Engaged by Candidate Immunogens

B cell lineages that are the current focus of vaccine development efforts against HIV-1, influenza or coronaviruses, often contain rare features, such as long heavy chain complementarity determining regions (CDRH3) loops. These unusual characteristics may limit the number of available B cells in the natural immunoglobulin repertoire that can respond to pathogen vaccinations. To measure the ability of a given immunogen to engage naturally occurring B cell receptors of interest, here we describe a mixed experimental and bioinformatic approach for determining the frequency and sequence of CDRH3 loops in the immune repertoire that can be recognized by a vaccine candidate. By combining deep mutational scanning and B cell receptor database analysis, CDRH3 loops were found that can be engaged by two HIV-1 germline-targeting immunogens, thus illustrating how the methods described here can be used to evaluate candidate immunogens based on their ability to engage diverse B cell lineage precursors.

T-Cell Receptor Repertoire Analysis with Computational Tools—An Immunologist’s Perspective

Over the last few years, there has been a rapid expansion in the application of information technology to biological data. Particularly the field of immunology has seen great strides in recent years. The development of next-generation sequencing (NGS) and single-cell technologies also brought forth a revolution in the characterization of immune repertoires. T-cell receptor (TCR) repertoires carry comprehensive information on the history of an individual’s antigen exposure. They serve as correlates of host protection and tolerance, as well as biomarkers of immunological perturbation by natural infections, vaccines or immunotherapies. Their interrogation yields large amounts of data. This requires a suite of highly sophisticated bioinformatics tools to leverage the meaning and complexity of the large datasets. Many different tools and methods, specifically designed for various aspects of immunological research, have recently emerged. Thus, researchers are now confronted with the issue of having to choose the right kind of approach to analyze, visualize and ultimately solve their task at hand. In order to help immunologists to choose from the vastness of available tools for their data analysis, this review addresses and compares commonly used bioinformatics tools for TCR repertoire analysis and illustrates the advantages and limitations of these tools from an immunologist’s perspective.

Viral infection engenders bona fide and bystander lung memory B cell subsets through permissive selection

Lung-resident memory B cells (MBCs) provide localized protection against reinfection in the respiratory airways. Currently, the biology of these cells remains largely unexplored. Here, we combined influenza and SARS-CoV-2 infection with fluorescent-reporter mice to identify MBCs regardless of antigen specificity. scRNA-seq analysis and confocal imaging revealed that two main transcriptionally distinct subsets of MBCs colonize the lung peribronchial niche after infection. These subsets arise from different progenitors and are both class-switched, somatically mutated and intrinsically biased in their differentiation fate towards plasma cells. Combined analysis of antigen-specificity and B cell receptor repertoire unveiled a highly permissive selection process that segregates these subsets into bona fide virus-specific MBCs and bystander MBCs with no apparent specificity for eliciting viruses. Thus, diverse transcriptional programs in MBCs are not linked to specific effector fates but rather to divergent strategies of the immune system to simultaneously provide rapid protection from reinfection while diversifying the initial B cell repertoire.

Coevolution of the olfactory organ and its receptor repertoire in ray-finned fishes

Ray-finned fishes (Actinopterygii) perceive their environment through a range of sensory modalities, including olfaction 1,2. Anatomical diversity of the olfactory organ suggests that olfaction is differentially important among species 1,3,4. To explore this topic, we studied the evolutionary dynamics of the four main gene families (OR, TAAR, ORA/VR1 and OlfC/VR2) 5 coding for olfactory receptors in 185 species of ray-finned fishes. The large variation in the number of functional genes, between 28 in the Ocean Sunfish Mola mola and 1317 in the Reedfish Erpetoichthys calabaricus, is the result of parallel expansions and contractions of the four main gene families. Several ancient and independent simplifications of the olfactory organ are associated with massive gene losses. In contrast, polypteriforms, which have a unique and complex olfactory organ, have almost twice as many olfactory receptor genes as any other ray-finned fish. These observations suggest a functional link between morphology of the olfactory organ and richness of the olfactory receptor repertoire. Further, our results demonstrate that the genomic underpinning of olfaction in ray-finned fishes is heterogeneous and presents a dynamic pattern of evolutionary expansions, simplifications and reacquisitions.

The Direct Influence of Cytomegalovirus Lysate on the Natural Killer Cell Receptor Repertoire

Natural killer (NK) cells are essential for controlling certain viral infections, including cytomegalovirus (CMV). In particular, the importance of NK cells in the context of CMV infection is underscored by the adaptive capabilities of these cells. Evidence suggests that some viruses can directly interfere with NK cell compartments and their activation and lead to shape-shifting the NK cell receptor repertoire. Still, it remains unknown whether the CMV can interact with NK cells without intermediaries. Here, we examined whether the direct effects of CMV lysate alter phenotypical properties of NK cells. To investigate this issue, NK cells were isolated from the blood of CMV seropositive healthy donors by negative magnetic separation. Isolated NK cells were cultured in the presence of CMV lysate and analyzed for the expression of NKG2A, NKG2C, and CD57 by FACS caliber. The results showed that NKG2C expression is significantly upregulated in the presence of CMV lysate compared to without stimulated group (mean increase, 6.65 %; 95% CI, 0.2582 to 13.02; p=0.043; R square: 0.38). Likewise, results have shown a significant decrease in the frequency of NKG2A+CD57- NK cell subsets (p=0.005; 95% CI, -13.49 to -3.151; R square: 0.5957) in the stimulated group compared to without stimulated ones. According to these results, CMV may drive a direct influence on NK cell receptor repertoire, including the expansion of NK cells expressing NKG2C receptor, which is needed for further studies.

T cell receptor repertoire signatures associated with COVID-19 severity

T cell receptor (TCR) repertoires are critical for antiviral immunity. Determining the TCR repertoires composition, diversity, and dynamics and how they change during viral infection can inform the molecular specificity of viral infection such as SARS-CoV-2. To determine signatures associated with COVID-19 disease severity, here we performed a large-scale analysis of over 4.7 billion sequences across 2,130 TCR repertoires from COVID-19 patients and healthy donors. TCR repertoire analyses from these data identified and characterized convergent COVID-19 associated CDR3 gene usages, specificity groups, and sequence patterns. T cell clonal expansion was found to be associated with upregulation of T cell effector function, TCR signaling, NF-kB signaling, and Interferon-gamma signaling pathways. Machine learning approaches accurately predicted disease severity for patients based on TCR sequence features, with certain high-power models reaching near-perfect AUROC scores across various predictor permutations. These analyses provided an integrative, systems immunology view of T cell adaptive immune responses to COVID-19.

Immunosenescence and Autoimmunity: Exploiting the T-Cell Receptor Repertoire to Investigate the Impact of Aging on Multiple Sclerosis

T-cell receptor (TCR) repertoire diversity is a determining factor for the immune system capability in fighting infections and preventing autoimmunity. During life, the TCR repertoire diversity progressively declines as a physiological aging progress. The investigation of TCR repertoire dynamics over life represents a powerful tool unraveling the impact of immunosenescence in health and disease. Multiple Sclerosis (MS) is a demyelinating, inflammatory, T-cell mediated autoimmune disease of the Central Nervous System in which age is crucial: it is the most widespread neurological disease among young adults and, furthermore, patients age may impact on MS progression and treatments outcome. Crossing knowledge on the TCR repertoire dynamics over MS patients’ life is fundamental to investigate disease mechanisms, and the advent of high- throughput sequencing (HTS) has significantly increased our knowledge on the topic. Here we report an overview of current literature about the impact of immunosenescence and age-related TCR dynamics variation in autoimmunity, including MS.