Mechanisms Of Age-related Changes in Transcriptional Regulation, TCR Repertoire, and Cytokine Expression

不同的T细胞受体（TCR）库对于控制病毒感染至关重要。然而，关于人类TCR库的一般信息和定义的病毒抗原有限。我们利用高通量测序（HTS）和单细胞配对TCR分析对CD4+和CD8+的TCR谱进行了综合分析，并对巨细胞病毒的pp65495503 （NLV）和甲型流感病毒的M15866 （GIL）这两个显性病毒表位的CD8+ TCR谱进行了特异性分析。我们对CD4+和CD8+ T细胞的TCR谱进行了比较分析。CD4+ T细胞的TCR多样性在1.8-8.2 x105之间，平均是CD8+ T细胞的3-4倍。此外，CD4+(0.3%)和CD8+ (1.3%) T细胞之间的TCR序列几乎没有重叠。进一步的分析表明，CD4+和CD8+ T细胞对CDR3中的某些氨基酸表现出明显的偏好，这一点通过支持向量机分类器进一步证实，表明CD4+和CD8+ T细胞中的TCR CDR3存在明显的可识别的差异。最后，我们确定了6-12%的独特tcr与不同的V基因共享相同的CDR3。总之，我们的发现揭示了CD4+和CD8+ T细胞之间TCR库的独特特征，并可能用于评估T细胞免疫能力。

A diverse T cell receptor (TCR) repertoire is essential for controlling viral infections. However, information on human TCR repertoires in general and to defined viral antigens is limited. We performed a comprehensive analysis of TCR repertoires of CD4+ and CD8+, and CD8+ TCR repertoires specific for two dominant viral epitopes: pp65495503 (NLV) of cytomegalovirus and M15866 (GIL) of influenza A virus using the high-throughput sequencing (HTS) and single-cell paired TCR analysis. We provided a comparative analysis of the TCR repertoires of CD4+ and CD8+ T cells. TCR diversity of CD4+ T cells ranges from 1.8-8.2 x105 and is 3-4 times greater on average than that of CD8+ T cells in each study subject. Furthermore, there was little overlap in TCR sequences between CD4+ (0.3%) and CD8+ (1.3%) T cells. Further analysis showed that CD4+ and CD8+ T cells exhibited distinct preferences for certain amino acids in the CDR3, and this was further confirmed by a support vector machine classifier, suggesting there are distinct and discernible differences between TCR CDR3 in CD4+ and CD8+ T cells. Finally, we identified 6-12% of the unique TCRs that share an identical CDR3 with different V genes. Together, our findings reveal the distinct features of the TCR repertoire between CD4+ and CD8+ T cells and could potentially be used to evaluate the competency of T cell immunity. For antigen-specific CD8+ TCR repertoires, we identified thousands of new NLV- and GIL-specific TCR and TCR sequences, as well as dozens of distinct CDR3 and CDR3 consensus motifs. This diversity is substantially greater than previously described for T cell responses to single viral epitopes, both for private and public TCR clonotypes, and exhibited a high degree of individual variations (12028,000 clonotypes per subject). However, diversity is effectively restricted by preferential V-J combinations, CDR3 lengths, and CDR3/CDR3 pairings. We further found that GIL-specific, but not NLV-specific, TCRs exhibited a surprisingly wide range of binding affinities, with dissociation constants (KDs) from 2 to 200 M. Finally, we determined the crystal structures of two unrelated GIL-specific TCRs in complex with GILHLA-A2 and two TCR-NLV- HLA-A2 complexes. These structures provide an explanation of the lower diversity of GIL-specific than NLV-specific repertoires. Remarkably, these two anti-viral TCR repertoires occupied 0.220% of the total CD8+ TCR repertoire, ensuring broad and robust T cell responses to single epitopes. Our comprehensive genetic, biochemical, and structural portrait of two different anti-viral T cell responses may contribute to the future development of predictors of immunity or disease at the personal level.