Mechanisms Of Age-related Changes in Transcriptional Regulation in lympphocytes

T细胞受体(TCR)为T细胞提供抗原识别的特异性。一个有能力的T细胞免疫系统依赖于不同的TCR谱系。然而，关于人类和小鼠TCR谱系的信息以及对已定义的病毒抗原的信息相对有限。我们使用高通量测序(HTS)和单细胞配对TCR分析以及小鼠CD4T细胞亚群包括NAVE、记忆性和调节性T细胞亚群，对两个主要病毒表位：巨细胞病毒的PP65(NLV)和甲型流感病毒的M1(GIL)的人CD8+TCR谱系进行了全面的分析。 我们对抗原特异性CD8+TCR谱系的分析鉴定了数千个新的NLV和GIL特异性的α和βTCR序列，以及数十个不同的CDR3和CDR3共识基序。这种多样性大大高于之前描述的T细胞对单个病毒表位的反应，无论是私人的还是公共的TCR克隆型，并显示出高度的个体差异(每个受试者875,533个TCR-α或β)。然而，多样性受到优先的V-J组合、CDR3长度和CDR3/CDR3配对的有效限制。两个与GILHL-A2结合的GIL特异性TCR的结构为GIL特异性TCR的多样性低于NLV特异性谱系提供了潜在的解释。这些抗病毒的TCR占据了CD8+TCR谱系的3.4%，确保了T细胞对单一表位的广泛反应。我们对两种不同的抗病毒T细胞反应的基因、生化和结构的综合描述可能有助于未来在个人水平上开发免疫或疾病的预测指标。 小鼠是研究最多的免疫系统模型，但有关α/βTCR谱系的信息有限。在这里，我们应用高通量测序和独特的分子识别符，从C57BL/6小鼠体内大约14%的CD4+T细胞中确定了TCR-α和TCR-β序列。我们已经鉴定了1.67×105个独特的TCR-α和2.34×105个独特的TCR-β序列，预测的TCR-α和TCR-β分别为2.79×106和5.09×106。TCR-β是TCR-α的1.6~1.8倍，Nave TCR-α和TCR-β分别是记忆CD4+T细胞的2.0和1.7倍。NAVE和Memory CD4+T细胞有大量重叠的TCR序列，反映出谱系的相对较小。CD5hi记忆CD4+T细胞比CD5lo记忆CD4+T细胞具有更高的亲和力，但它们的Alpha/βTCR谱系没有显着差异。调节性T细胞(Treg)的α/βTCR谱系介于初代T细胞和记忆性T细胞之间，其中15%的Treg由不同的CDR3氨基酸序列TCR-α(5.2×103)和TCR-β(9.7×103)组成，可能代表来自胸腺的天然Treg。综上所述，这些发现首次提供了小鼠CD4+T细胞α/βTCR谱系的深度信息，为进一步阐明CD4+T细胞谱系的广泛性和特异性奠定了基础。

T cell receptor (TCR) provides the specificity of antigen-recognition for T cells. A competent T cell immune system depends on a diverse TCR repertoire. However, information on human and mouse TCR repertoires in general and to defined viral antigens is relatively limited. We performed a comprehensive analysis of TCR repertoires of human CD8+ TCR repertoires specific for two dominant viral epitopes: pp65 (NLV) of cytomegalovirus and M1 (GIL) of influenza A virus using the high-throughput sequencing (HTS) and single-cell paired TCR analysis and mouse CD4 T cell subsets including nave, memory, and regulatory T cells. Our analysis of antigen-specific CD8+ TCR repertoires resulted in identification of thousands of new NLV- and GIL-specific alpha and beta TCR sequences and 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 (875,533 TCR-alpha or beta per subject). However, diversity is effectively restricted by preferential V-J combinations, CDR3 lengths, and CDR3/CDR3 pairings. Structures of two GIL-specific TCRs bound to GILHLA-A2 provided a potential explanation for the lower diversity of GIL-specific than NLV-specific repertoires. These anti-viral TCRs occupied up to 3.4% of the CD8+ TCR repertoire, ensuring broad 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. Mouse is the most studied model of immune system and yet the information of alpha/beta TCR repertoire is limited. Here we applied high throughput sequencing with unique molecular identifier to determine TCR-alpha and TCR-beta sequences from approximately 14% of total CD4+ T cells in a C57BL/6 mouse. We have identified 1.67 x 105 unique TCR-alpha and 2.34 x 105 unique TCR-beta sequences with projected TCR-alpha and TCR-beta repertoires 2.79 x 106 and 5.09 x 106, respectively. The repertoire of TCR-beta was 1.6-1.8 time larger than that of TCR-alpha, and nave TCR-alpha and TCR-beta repertoires were 2.0 and 1.7 larger than those of memory CD4+ T cells, respectively. Nave and memory CD4+ T cells had substantial overlapping TCR sequences, reflecting relative small size of the repertoires. CD5hi memory CD4+ T cells have higher affinity than CD5lo memory CD4+ T cells but their Alpha/beta TCR repertoires were not significantly different. Alpha/beta TCR repertoires of regulatory (Treg) CD4+ T cells was between nave and memory T cells and a fraction of Treg (15%) consists of distinct CDR3 amino acid sequences of TCR-alpha (5.2 x 103) and TCR-beta (9.7 x 103), may represent the natural Treg derived from thymus. Together, these findings provide the depth information of alpha/betaTCR repertoire of mouse CD4+ T cells for the first time and serve as a foundation for further elucidating the broadness and specificity of CD4+ T cell repertoire.