Quantification of epitope abundance reveals the effect of direct and cross-presentation on influenza CTL responses

The magnitude of T cell responses to infection is a function of the naïve T cell repertoire combined with the context and duration of antigen presentation. Using mass spectrometry, we identify and quantify 21 class 1 MHC-restricted influenza A virus (IAV)-peptides following either direct or cross-presentation. All these peptides, including seven novel epitopes, elicit T cell responses in infected C57BL/6 mice. Directly presented IAV epitopes maintain their relative abundance across distinct cell types and reveal a broad range of epitope abundances. In contrast, cross-presented epitopes are more uniform in abundance. We observe a clear disparity in the abundance of the two key immunodominant IAV antigens, wherein direct infection drives optimal nucleoprotein (NP)366–374 presentation, while cross-presentation is optimal for acid polymerase (PA)224–233 presentation. The study demonstrates how assessment of epitope abundance in both modes of antigen presentation is necessary to fully understand the immunogenicity and response magnitude to T cell epitopes

Reversed T cell receptor docking on a major histocompatibility class I complex limits involvement in the immune response

The anti-viral T cell response is drawn from the naive T cell repertoire. During influenza infection, the CD8+ T cell response to an H-2Db-restricted nucleoprotein epitope (NP366) is characterized by preferential expansion of T cells bearing TRBV13+ T cell receptors (TCRs) and avoidance of TRBV17+ T cells, despite the latter dominating the naive precursor repertoire. We found two TRBV17+ TCRs that bound H-2Db-NP366 with a 180° reversed polarity compared to the canonical TCR-pMHC-I docking. The TRBV17 β-chain dominated the interaction and, whereas the complementarity determining region-3 (CDR3) loops exclusively mediated contacts with the MHC-I, peptide specificity was attributable to germline-encoded recognition. Nevertheless, the TRBV17+ TCR exhibited moderate affinity toward H-2Db-NP366 and was capable of signal transduction. Thus, the naive CD8+ T cell pool can comprise TCRs adopting reversed pMHC-I docking modes that limit their involvement in the immune response

Heightened self-reactivity associated with selective survival, but not expansion, of naïve virus-specific CD8⁺ T cells in aged mice

In advanced age, decreased CD8(+) cytotoxic T-lymphocyte (CTL) responses to novel pathogens and cancer is paralleled by a decline in the number and function of naïve CTL precursors (CTLp). Although the age-related fall in CD8(+) T-cell numbers is well established, neither the underlying mechanisms nor the extent of variation for different epitope specificities have been defined. Furthermore, naïve CD8(+) T cells expressing high levels of CD44 accumulate with age, but it is unknown whether this accumulation reflects their preferential survival or an age-dependent driver of CD8(+) T-cell proliferation. Here, we track the number and phenotype of four influenza A virus (IAV)-specific CTLp populations in naïve C57BL/6 (B6) mice during aging, and compare T-cell receptor (TCR) clonal diversity for the CD44hi and CD44lo subsets of one such population. We show differential onset of decline for several IAV-specific CD8(+) T-cell populations with advanced age that parallel age-associated changes in the B6 immunodominance hierarchy, suggestive of distinct impacts of aging on different epitope-specific populations. Despite finding no evidence of clonal expansions in an aged, epitope-specific TCR repertoire, nonrandom alterations in TCR usage were observed, along with elevated CD5 and CD8 coreceptor expression. Collectively, these data demonstrate that naïve CD8(+) T cells expressing markers of heightened self-recognition are selectively retained, but not clonally expanded, during aging

Overlapping peptides elicit distinct CD8+ T cell responses following Influenza A virus infection

The presentation of pathogen-derived peptides on MHC class I molecules is essential for the initiation of adaptive CD8+ T cell immunity, which in turn is critical for effective control of many significant human infections. The identification of immunogenic pathogen-derived epitopes and a detailed understanding of how they are recognized by TCRs is essential for the design of effective T cell–based vaccines. In this study, we have characterized the T cell recognition and immune responses in mice to two naturally presented influenza A virus–derived peptides previously identified from virally infected cells via mass spectrometry. These neuraminidase-derived peptides, NA181–190 (SGPDNGAVAV) and NA181–191 (SGPDNGAVAVL), are completely overlapping with the exception of a 1 aa extension at the C terminus of the longer peptide. This minor peptidic difference results in the induction of two completely independent and non–cross-reactive T cell populations that show distinct functional characteristics after influenza A virus infection of B6 mice. We show that the unique TCR reactivity to the overlapping peptides is present in the naive repertoire prior to immune expansion in B6 mice. Moreover, we provide a structural explanation underlying the distinct CD8+ T cell reactivities, which reinforces the concept that peptide length is a key determinant of Ag specificity in CD8+ T cell responses