Linking the T cell receptor to the single cell transcriptome in antigen-specific human T cells
Linking the T cell receptor to the single cell transcriptome in antigen-specific human T cells
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DOI：
10.1038/icb.2016.16
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发表时间：
2016-07-01
影响因子：
4
通讯作者：
Luciani, Fabio
中科院分区：
文献类型：
--
作者：
Eltahla, Auda A.;Rizzetto, Simone;Luciani, Fabio
T细胞的异质性是成功的适应性免疫反应的标志，利用抗原特异性T细胞的巨大多样性来协调效应和记忆结果的进化。T细胞受体（TCR）库是高度多样化的，以解释高度异质的抗原世界。在对病毒的反应过程中，抗原特异性CD8+ （ag特异性）T细胞的多个个体克隆可以针对单个表位进行识别，并且可以识别多个表位。单细胞技术的进步提供了在表面表型和转录组水平上研究ag特异性T细胞异质性的潜力，从而允许研究同一表观亚群内的多样性。我们提出了一种新的方法（VDJPuzzle），从ag特异性T细胞的单细胞RNA-seq数据中重建天然TCR α - β，然后将这些数据与单个细胞的基因表达谱联系起来。我们使用从清除丙型肝炎病毒感染的受试者外周血中分离的罕见ag特异性T细胞应用这种方法。我们成功地在63个细胞中的56个（89%）中重建了生产性TCR α β，其中双a和双β分别在18个和7%中重建，双TCR α β在2个细胞中重建。该方法通过TCR的标准单细胞PCR测序进行验证。我们证明，单细胞转录组分析可以成功区分抗原特异性T细胞群直接从静息记忆细胞在外周血和体外刺激后分类。这种方法可以详细分析TCR多样性及其与不同克隆转录谱的关系。
Heterogeneity of T cells is a hallmark of a successful adaptive immune response, harnessing the vast diversity of antigen-specific T cells into a coordinated evolution of effector and memory outcomes. The T cell receptor (TCR) repertoire is highly diverse to account for the highly heterogeneous antigenic world. During the response to a virus multiple individual clones of antigen specific CD8+ (Ag-specific) T cells can be identified against a single epitope and multiple epitopes are recognised. Advances in single-cell technologies have provided the potential to study Ag-specific T cell heterogeneity at both surface phenotype and transcriptome levels, thereby allowing investigation of the diversity within the same apparent sub-population. We propose a new method (VDJPuzzle) to reconstruct the native TCR alpha beta from single cell RNA-seq data of Ag-specific T cells and then to link these with the gene expression profile of individual cells. We applied this method using rare Ag-specific T cells isolated from peripheral blood of a subject who cleared hepatitis C virus infection. We successfully reconstructed productive TCR alpha beta in 56 of a total of 63 cells (89%), with double a and double beta in 18, and 7% respectively, and double TCR alpha beta in 2 cells. The method was validated via standard single cell PCR sequencing of the TCR. We demonstrate that single-cell transcriptome analysis can successfully distinguish Ag-specific T cell populations sorted directly from resting memory cells in peripheral blood and sorted after ex vivo stimulation. This approach allows a detailed analysis of the TCR diversity and its relationship with the transcriptional profile of different clones.