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Summary: HIV controllers can control viral replication and remain healthy, but the mechanism behind this control is unknown. Despite human leukocyte antigen (HLA) diversity in the population, almost 50% of HIV controllers express the HLA-B∗57:01 molecule, which presents, among others, the Gag-derived epitope TW10. Given TW10’s presentation in early infection, TW10-specific T cells could participate in the control of HIV. Here, we study the strength and functionality of TW10-specific T cells from HLA-B∗57:01+/HIV+ controller and non-controller individuals. We determine the TW10-specific T cell receptor (TCR) repertoire, revealing a bias in TCR gene usage with the presence of a public TCR. We determine that the T cell response is polyfunctional regardless of the viral load, despite the low affinity of TW10-specific TCRs. We solve the crystal structure of HLA-B∗57:01-TW10 in complex with a TCR, providing the basis of recognition that underpins the strong TRBV5 bias observed in TW10-specific clonotypes.

Abstract Objectives Seasonal influenza viruses cause roughly 650 000 deaths annually despite available vaccines. CD8+ T cells typically recognise influenza‐derived peptides from internal structural and non‐structural influenza proteins and are an attractive avenue for future vaccine design as they could reduce the severity of disease following infection with diverse influenza strains. CD8+ T cells recognise peptides presented by the highly polymorphic Human Leukocyte Antigens class I molecules (HLA‐I). Each HLA‐I variant has distinct peptide binding preferences, representing a significant obstacle for designing vaccines that elicit CD8+ T cell responses across broad populations. Consequently, the rational design of a CD8+ T cell‐mediated vaccine would require the identification of highly immunogenic peptides restricted to a range of different HLA molecules. Methods Here, we assessed the immunogenicity of six recently published novel influenza‐derived peptides identified by mass‐spectrometry and predicted to bind to the prevalent HLA‐B*18:01 molecule. Results Using CD8+ T cell activation assays and protein biochemistry, we showed that 3/6 of the novel peptides were immunogenic in several HLA‐B*18:01+ individuals and confirmed their HLA‐B*18:01 restriction. We subsequently compared CD8+ T cell responses towards the previously identified highly immunogenic HLA‐B*18:01‐restricted NP219 peptide. Using X‐ray crystallography, we solved the first crystal structures of HLA‐B*18:01 presenting immunogenic influenza‐derived peptides. Finally, we dissected the first TCR repertoires specific for HLA‐B*18:01 restricted pathogen‐derived peptides, identifying private and restricted repertoires against each of the four peptides. Conclusion Overall the characterisation of these novel immunogenic peptides provides additional HLA‐B*18:01‐restricted vaccine targets derived from the Matrix protein 1 and potentially the non‐structural protein and the RNA polymerase catalytic subunit of influenza viruses.

CD8+ T cells are crucial for viral elimination and recovery from viral infection. Nonetheless, the current understanding of the T cell response to SARS-CoV-2 at the antigen level remains limited. The Spike protein is an external structural protein that is prone to mutations, threatening the efficacy of current vaccines. Therefore, we have characterised the immune response towards the immunogenic Spike-derived peptide (S976-984, VLNDILSRL), restricted to the HLA-A*02:01 molecule, which is mutated in both Alpha (S982A) and Omicron BA.1 (L981F) variants of concern. We determined that the mutation in the Alpha variant (S982A) impacted both the stability and conformation of the peptide, bound to HLA-A*02:01, in comparison to the original S976-984. We identified a longer and overlapping immunogenic peptide (S975-984, SVLNDILSRL) that could be presented by HLA-A*02:01, HLA-A*11:01 and HLA-B*13:01 allomorphs. We showed that S975-specific CD8+ T cells were weakly cross-reactive to the mutant peptides despite their similar conformations when presented by HLA-A*11:01. Altogether, our results show that the impact of SARS-CoV-2 mutations on peptide presentation is HLA allomorph-specific, and that post vaccination there are T cells able to react and cross-react towards the variant of concern peptides.

Abstract Seasonal influenza viruses continue to cause severe medical and financial complications annually. Although there are many licenced influenza vaccines, there are billions of cases of influenza infection every year, resulting in the death of over half a million individuals. Furthermore, these figures can rise in the event of a pandemic, as seen throughout history, like the 1918 Spanish influenza pandemic (50 million deaths) and the 1968 Hong Kong influenza pandemic (~4 million deaths). In this review, we have summarised many of the currently licenced influenza vaccines available across the world and current vaccines in clinical trials. We then briefly discuss the important role of CD8+ T cells during influenza infection and why future influenza vaccines should consider targeting CD8+ T cells. Finally, we assess the current landscape of known immunogenic CD8+ T‐cell epitopes and highlight the knowledge gaps required to be filled for the design of rational future influenza vaccines that incorporate CD8+ T cells.

The T cell response to SARS-CoV-2 is important in protection from infection. Here the authors show by concentrating on a specific HLA haplotype that mutations in SARS-CoV-2 as new variants emerge can affect T cell recognition and reduce T cell responses to the virus.

Differentiation and activation of T cells are normally modulated by non-covalent interactions between T cell receptor (TCR) and antigenic peptides. Here the authors use step-wise mutations, biochemical characterization and structural insights to describe the contributions of natural covalent bonds between TCR and antigenic peptides during these processes.

ABSTRACT Objective Class III obesity (body mass index [BMI] ≥ 40 kg m−2) significantly impairs the immune response to SARS‐CoV‐2 vaccination. However, the effect of an elevated BMI (≥ 25 kg m−2) on humoral immunity to SARS‐CoV‐2 infection and COVID‐19 vaccination remains unclear. Methods We collected blood samples from people who recovered from SARS‐CoV‐2 infection approximately 3 and 13 months of post‐infection (noting that these individuals were not exposed to SARS‐CoV‐2 or vaccinated in the interim). We also collected blood samples from people approximately 5 months of post‐second dose COVID‐19 vaccination (the majority of whom did not have a prior SARS‐CoV‐2 infection). We measured their humoral responses to SARS‐CoV‐2, grouping individuals based on a BMI greater or less than 25 kg m−2. Results Here, we show that an increased BMI (≥ 25 kg m−2), when accounting for age and sex differences, is associated with reduced antibody responses after SARS‐CoV‐2 infection. At 3 months of post‐infection, an elevated BMI was associated with reduced antibody titres. At 13 months of post‐infection, an elevated BMI was associated with reduced antibody avidity and a reduced percentage of spike‐positive B cells. In contrast, no significant association was noted between a BMI ≥ 25 kg m−2 and humoral immunity to SARS‐CoV‐2 at 5 months of post‐secondary vaccination. Conclusions Taken together, these data showed that elevated BMI is associated with an impaired humoral immune response to SARS‐CoV‐2 infection. The impairment of infection‐induced immunity in individuals with a BMI ≥ 25 kg m−2 suggests an added impetus for vaccination rather than relying on infection‐induced immunity.

Detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) specific CD4+ and CD8+ T cells in SARS-CoV-2-unexposed donors has been explained by the presence of T cells primed by other coronaviruses. However, based on the relatively high frequency and prevalence of cross-reactive T cells, we hypothesized cytomegalovirus (CMV) may induce these cross-reactive T cells. Stimulation of pre-pandemic cryo-preserved peripheral blood mononuclear cells (PBMCs) with SARS-CoV-2 peptides revealed that frequencies of SARS-CoV-2-specific T cells were higher in CMV-seropositive donors. Characterization of these T cells demonstrated that membrane-specific CD4+ and spike-specific CD8+ T cells originate from cross-reactive CMV-specific T cells. Spike-specific CD8+ T cells recognize SARS-CoV-2 spike peptide FVSNGTHWF (FVS) and dissimilar CMV pp65 peptide IPSINVHHY (IPS) presented by HLA-B*35:01. These dual IPS/FVS-reactive CD8+ T cells were found in multiple donors as well as severe COVID-19 patients and shared a common T cell receptor (TCR), illustrating that IPS/FVS-cross-reactivity is caused by a public TCR. In conclusion, CMV-specific T cells cross-react with SARS-CoV-2, despite low sequence homology between the two viruses, and may contribute to the pre-existing immunity against SARS-CoV-2.

Summary: Directly ex vivo, peptide-specific CD8+ T cells are present at relatively low frequency and are typically in a resting state. This protocol details the expansion of memory peptide-specific CD8+ T cells by in vitro stimulation, which can be subsequently characterized using a range of assays including tetramer staining and intracellular cytokine staining.For complete details on the use and execution of this protocol, please refer to Lineburg et al. (2021). : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

The SARS-CoV-2 virus responsible for the COVID-19 pandemic has caused significant morbidity and mortality worldwide. With the remarkable advances in medical research, vaccines were developed to prime the human immune system and decrease disease severity. Despite these achievements, the fundamental basis of immunity to the SARS-CoV-2 virus is still largely undefined. Here, we solved the crystal structure of three spike-derived peptides presented by three different HLA molecules, and determined the stability of the overall peptide–HLA complexes formed. The peptide presentation of spike-derived peptides can influence the way in which CD8+ T cells can recognise infected cells, clear infection, and therefore, control the outcome of the disease.

The immunology of Indigenous populations is generally understudied outside the context of diseases that are prevalent in these communities. Here the authors identify prevalence of influenza CD8+ T cell epitopes in an Indigenous Australian population expressing the susceptibility allomorph HLA A*24:02 and validate immunodominance of some of these epitopes in mice.

The immunological parameters that define severe influenza disease are not clear within human real time infections. Here the authors compare a severe influenza infection cohort with an influenza vaccinated cohort to understand correlates of severe influenza disease.

HLA-A*11:01 is one of the most prevalent human leukocyte antigens (HLAs), especially in East Asian and Oceanian populations. It is also highly expressed in Indigenous people who are at high risk of severe influenza disease. As CD8+ T cells can provide broadly cross-reactive immunity to distinct influenza strains and subtypes, including influenza A, B and C viruses, understanding CD8+ T cell immunity to influenza viruses across prominent HLA types is needed to rationally design a universal influenza vaccine and generate protective immunity especially for high-risk populations. As only a handful of HLA-A*11:01-restricted CD8+ T cell epitopes have been described for influenza A viruses (IAVs) and epitopes for influenza B viruses (IBVs) were still unknown, we embarked on an epitope discovery study to define a CD8+ T cell landscape for HLA-A*11:01-expressing Indigenous and non-Indigenous Australian people. Using mass-spectrometry, we identified IAV- and IBV-derived peptides presented by HLA-A*11:01 during infection. 79 IAV and 57 IBV peptides were subsequently screened for immunogenicity in vitro with peripheral blood mononuclear cells from HLA-A*11:01-expressing Indigenous and non-Indigenous Australian donors. CD8+ T cell immunogenicity screening revealed two immunogenic IAV epitopes (A11/PB2320-331 and A11/PB2323-331) and the first HLA-A*11:01-restricted IBV epitopes (A11/M41-49, A11/NS1186-195 and A11/NP511-520). The immunogenic IAV- and IBV-derived peptides were >90% conserved among their respective influenza viruses. Identification of novel immunogenic HLA-A*11:01-restricted CD8+ T cell epitopes has implications for understanding how CD8+ T cell immunity is generated towards IAVs and IBVs. These findings can inform the development of rationally designed, broadly cross-reactive influenza vaccines to ensure protection from severe influenza disease in HLA-A*11:01-expressing individuals.

ABSTRACT Understanding the immune response to severe acute respiratory syndrome coronavirus (SARS-CoV-2) is critical to overcome the current coronavirus disease (COVID-19) pandemic. Efforts are being made to understand the potential cross-protective immunity of memory T cells, induced by prior encounters with seasonal coronaviruses, in providing protection against severe COVID-19. In this study we assessed T-cell responses directed against highly conserved regions of SARS-CoV-2. Epitope mapping revealed 16 CD8+ T-cell epitopes across the nucleocapsid (N), spike (S), and open reading frame (ORF)3a proteins of SARS-CoV-2 and five CD8+ T-cell epitopes encoded within the highly conserved regions of the ORF1ab polyprotein of SARS-CoV-2. Comparative sequence analysis showed high conservation of SARS-CoV-2 ORF1ab T-cell epitopes in seasonal coronaviruses. Paradoxically, the immune responses directed against the conserved ORF1ab epitopes were infrequent and subdominant in both convalescent and unexposed participants. This subdominant immune response was consistent with a low abundance of ORF1ab encoded proteins in SARS-CoV-2 infected cells. Overall, these observations suggest that while cross-reactive CD8+ T cells likely exist in unexposed individuals, they are not common and therefore are unlikely to play a significant role in providing broad preexisting immunity in the community. IMPORTANCE T cells play a critical role in protection against SARS-CoV-2. Despite being highly topical, the protective role of preexisting memory CD8+ T cells, induced by prior exposure to circulating common coronavirus strains, remains less clear. In this study, we established a robust approach to specifically assess T cell responses to highly conserved regions within SARS-CoV-2. Consistent with recent observations we demonstrate that recognition of these highly conserved regions is associated with an increased likelihood of milder disease. However, extending these observations we observed that recognition of these conserved regions is rare in both exposed and unexposed volunteers, which we believe is associated with the low abundance of these proteins in SARS-CoV-2 infected cells. These observations have important implications for the likely role preexisting immunity plays in controlling severe disease, further emphasizing the importance of vaccination to generate the immunodominant T cells required for immune protection.

Abstract Objective Influenza A, B and C viruses (IAV, IBV and ICV, respectively) circulate globally, infecting humans and causing widespread morbidity and mortality. Here, we investigate the T cell response towards an immunodominant IAV epitope, NP265‐273, and its IBV and ICV homologues, presented by HLA‐A*03:01 molecule expressed in ~ 4% of the global population (~ 300 million people). Methods We assessed the magnitude (tetramer staining) and quality of the CD8+ T cell response (intracellular cytokine staining) towards NP265‐IAV and described the T cell receptor (TCR) repertoire used to recognise this immunodominant epitope. We next assessed the immunogenicity of NP265‐IAV homologue peptides from IBV and ICV and the ability of CD8+ T cells to cross‐react towards these homologous peptides. Furthermore, we determined the structures of NP265‐IAV and NP323‐IBV peptides in complex with HLA‐A*03:01 by X‐ray crystallography. Results Our study provides a detailed characterisation of the CD8+ T cell response towards NP265‐IAV and its IBV and ICV homologues. The data revealed a diverse repertoire for NP265‐IAV that is associated with superior anti‐viral protection. Evidence of cross‐reactivity between the three different influenza virus strain‐derived epitopes was observed, indicating the discovery of a potential vaccination target that is broad enough to cover all three influenza strains. Conclusion We show that while there is a potential to cross‐protect against distinct influenza virus lineages, the T cell response was stronger against the IAV peptide than IBV or ICV, which is an important consideration when choosing targets for future vaccine design.

Summary: Understanding T-cell responses requires identifying viral peptides presented by human leukocyte antigens (HLAs). X-ray crystallography can be used to visualize their presentation. This protocol describes the expression, purification, and crystallization of HLA-A∗02:01, one of the most frequent HLA in the global population in complex with peptides derived from the SARS-CoV-2 nucleocapsid protein. This protocol can be applied to different HLA class I molecules bound to other peptides.For complete details on the use and execution of this protocol, please refer to Szeto et al. (2021).

CD1 proteins present lipid antigens to T cells via the T cell receptor. Here the authors describe T cell reactivity to human membrane lipid moieties and provide structural data to define a molecular mechanism of promiscuous recognition of self-derived phospholipids.

Mutations within immunological epitope containing regions of influenza A virus can impair the established immune response between influenza strains and could impact rational vaccine design. Here Grant et al. examine the presence, structural impact and cross reactivity of two human immunodominant influenza epitope variants.

Summary: CD8+ T cells are crucial for anti-viral immunity; however, understanding T cell responses requires the identification of epitopes presented by human leukocyte antigens (HLA). To date, few SARS-CoV-2-specific CD8+ T cell epitopes have been described. Internal viral proteins are typically more conserved than surface proteins and are often the target of CD8+ T cells. Therefore, we have characterized eight peptides derived from the internal SARS-CoV-2 nucleocapsid protein predicted to bind HLA-A∗02:01, the most common HLA molecule in the global population. We determined not all peptides could form a complex with HLA-A∗02:01, and the six crystal structures determined revealed that some peptides adopted a mobile conformation. We therefore provide a molecular understanding of SARS-CoV-2 CD8+ T cell epitopes. Furthermore, we show that there is limited pre-existing CD8+ T cell response toward these epitopes in unexposed individuals. Together, these data show that SARS-CoV-2 nucleocapsid might not contain potent epitopes restricted to HLA-A∗02:01.

Seasonal influenza virus infections cause 290,000-650,000 deaths annually and severe morbidity in 3-5 million people. CD8+ T-cell responses towards virus-derived peptide/human leukocyte antigen (HLA) complexes provide the broadest cross-reactive immunity against human influenza viruses. Several universally-conserved CD8+ T-cell specificities that elicit prominent responses against human influenza A viruses (IAVs) have been identified. These include HLA-A*02:01-M158-66 (A2/M158), HLA-A*03:01-NP265-273, HLA-B*08:01-NP225-233, HLA-B*18:01-NP219-226, HLA-B*27:05-NP383-391 and HLA-B*57:01-NP199-207. The immunodominance hierarchies across these universal CD8+ T-cell epitopes were however unknown. Here, we probed immunodominance status of influenza-specific universal CD8+ T-cells in HLA-I heterozygote individuals expressing two or more universal HLAs for IAV. We found that while CD8+ T-cell responses directed towards A2/M158 were generally immunodominant, A2/M158+CD8+ T-cells were markedly diminished (subdominant) in HLA-A*02:01/B*27:05-expressing donors following ex vivo and in vitro analyses. A2/M158+CD8+ T-cells in non-HLA-B*27:05 individuals were immunodominant, contained optimal public TRBV19/TRAV27 TCRαβ clonotypes and displayed highly polyfunctional and proliferative capacity, while A2/M158+CD8+ T cells in HLA-B*27:05-expressing donors were subdominant, with largely distinct TCRαβ clonotypes and consequently markedly reduced avidity, proliferative and polyfunctional efficacy. Our data illustrate altered immunodominance patterns and immunodomination within human influenza-specific CD8+ T-cells. Accordingly, our work highlights the importance of understanding immunodominance hierarchies within individual donors across a spectrum of prominent virus-specific CD8+ T-cell specificities prior to designing T cell-directed vaccines and immunotherapies, for influenza and other infectious diseases.

Human leukocyte antigen (HLA) presents peptides to activate T cells, but many aspects in the T cell receptor (TCR)/HLA interaction remain unclear. Here the authors show, via structural data, that two TCRs differentially recognize the same tumour peptide/HLA complex and induce contrasting conformation changes of the peptide.

The data currently available on how the immune system recognises the SARS-CoV-2 virus is growing rapidly. While there are structures of some SARS-CoV-2 proteins in complex with antibodies, which helps us understand how the immune system is able to recognise this new virus; however, we lack data on how T cells are able to recognise this virus. T cells, especially the cytotoxic CD8+ T cells, are critical for viral recognition and clearance. Here we report the X-ray crystallography structure of a T cell receptor, shared among unrelated individuals (public TCR) in complex with a dominant spike-derived CD8+ T cell epitope (YLQ peptide). We show that YLQ activates a polyfunctional CD8+ T cell response in COVID-19 recovered patients. We detail the molecular basis for the shared TCR gene usage observed in HLA-A*02:01+ individuals, providing an understanding of TCR recognition towards a SARS-CoV-2 epitope. Interestingly, the YLQ peptide conformation did not change upon TCR binding, facilitating the high-affinity interaction observed.

HLA-C expression levels correlate with immune responses to pathogens and autoimmunity, and vary in an allele-specific manner across individuals. Here the authors identify factors that drive differential expression of HLA-C allomorphs at the cell surface, and influence the structure of the peptide-binding cleft and diversity of peptides bound by HLA-C molecules.

T cells are a critical part of the adaptive immune system that are able to distinguish between healthy and unhealthy cells. Upon recognition of protein fragments (peptides), activated T cells will contribute to the immune response and help clear infection. The major histocompatibility complex (MHC) molecules, or human leukocyte antigens (HLA) in humans, bind these peptides to present them to T cells that recognise them with their surface T cell receptors (TCR). This recognition event is the first step that leads to T cell activation, and in turn can dictate disease outcomes. The visualisation of TCR interaction with pMHC using structural biology has been crucial in understanding this key event, unravelling the parameters that drive this interaction and their impact on the immune response. The last five years has been the most productive within the field, wherein half of current unique TCR–pMHC-I structures to date were determined within this time. Here, we review the new insights learned from these recent TCR–pMHC-I structures and their impact on T cell activation.

As a major arm of the cellular immune response, CD4+ T cells are important in the control and clearance of infections. Primarily described as helpers, CD4+ T cells play an integral role in the development and activation of B cells and CD8+ T cells. CD4+ T cells are incredibly heterogeneous, and can be divided into six main lineages based on distinct profiles, namely T helper 1, 2, 17 and 22 (Th1, Th2, Th17, Th22), regulatory T cells (Treg) and T follicular helper cells (Tfh). Recent advances in structural biology have allowed for a detailed characterisation of the molecular mechanisms that drive CD4+ T cell recognition. In this review, we discuss the defining features of the main human CD4+ T cell lineages and their role in immunity, as well as their structural characteristics underlying their detection of pathogens.

Germline-encoded mycolyl lipid-reactive (GEM) T cells recognize CD1b proteins presenting mycobacterial mycolates via their T-cell receptors (TCRs). Here, the authors present the structure of this interaction and provide a molecular basis for the co-recognition of CD1b and a mycobacterial glycolipid.

The invariant αβTCR of type I NKT cells recognizes a lipid α-GalCer presented by CD1d. Here the authors describe atypical α-GalCer-reactive NKT cells with diverse TCRs, which bind to CD1d-α-GalCer in a manner distinct from type I NKT cells, thus unveiling greater diversity in lipid antigen recognition.

The interaction between T cell receptor (TCR) and peptide (p)-Human Leukocyte Antigen (HLA) complexes is the critical first step in determining T cell responses. X-ray crystallographic studies of pHLA in TCR-bound and free states provide a structural perspective that can help understand T cell activation. These structures represent a static “snapshot”, yet the nature of pHLAs and their interactions with TCRs are highly dynamic. This has been demonstrated for HLA class I molecules with in silico techniques showing that some interactions, thought to stabilise pHLA-I, are only transient and prone to high flexibility. Here, we investigated the dynamics of HLA class II molecules by focusing on three allomorphs (HLA-DR1, -DR11 and -DR15) that are able to present the same epitope and activate CD4+ T cells. A single TCR (F24) has been shown to recognise all three HLA-DR molecules, albeit with different affinities. Using molecular dynamics and crystallographic ensemble refinement, we investigate the molecular basis of these different affinities and uncover hidden roles for HLA polymorphic residues. These polymorphisms were responsible for the widening of the antigen binding cleft and disruption of pHLA-TCR interactions, underpinning the hierarchy of F24 TCR binding affinity, and ultimately T cell activation. We expanded this approach to all available pHLA-DR structures and discovered that all HLA-DR molecules were inherently rigid. Together with in vitro protein stability and peptide affinity measurements, our results suggest that HLA-DR1 possesses inherently high protein stability, and low HLA-DM susceptibility.

CD8+ T cells recognizing antigenic peptides derived from conserved internal viral proteins confer broad protection against distinct influenza viruses. As memory CD8+ T cells change throughout the human lifetime and across tissue compartments, we investigated how T cell receptor (TCR) composition and diversity relate to memory CD8+ T cells across anatomical sites and immunological phases of human life. We used ex vivo peptide-HLA tetramer magnetic enrichment, single-cell multiplex RT-PCR for both the TCR-alpha (TCRα) and TCR-beta (TCRβ) chains, and new TCRdist and grouping of lymphocyte interactions by paratope hotspots (GLIPH) algorithms to compare TCRs directed against the most prominent human influenza epitope, HLA-A*02:01-M158–66 (A2+M158). We dissected memory TCR repertoires directed toward A2+M158 CD8+ T cells within human tissues and compared them to human peripheral blood of young and elderly adults. Furthermore, we compared these memory CD8+ T cell repertoires to A2+M158 CD8+ TCRs during acute influenza disease in patients hospitalized with avian A/H7N9 virus. Our study provides the first ex vivo comparative analysis of paired antigen-specific TCR-α/β clonotypes across different tissues and peripheral blood across different age groups. We show that human A2+M158 CD8+ T cells can be readily detected in human lungs, spleens, and lymph nodes, and that tissue A2+M158 TCRαβ repertoires reflect A2+M158 TCRαβ clonotypes derived from peripheral blood in healthy adults and influenza-infected patients. A2+M158 TCRαβ repertoires displayed distinct features only in elderly adults, with large private TCRαβ clonotypes replacing the prominent and public TRBV19/TRAV27 TCRs. Our study provides novel findings on influenza-specific TCRαβ repertoires within human tissues, raises the question of how we can prevent the loss of optimal TCRαβ signatures with aging, and provides important insights into the rational design of T cell-mediated vaccines and immunotherapies.