Linking the T cell receptor to the single cell transcriptome in antigen‐specific human T cells

• Published in: Immunology and cell biology 2016-07, Vol.94 (6), p.604-611
• Main Authors: Eltahla, Auda A, Rizzetto, Simone, Pirozyan, Mehdi R, Betz‐Stablein, Brigid D, Venturi, Vanessa, Kedzierska, Katherine, Lloyd, Andrew R, Bull, Rowena A, Luciani, Fabio
• Format: Article
• Language: English
• Subjects: Epitopes - genetics; Epitopes, T-Lymphocyte - immunology; Gene Expression Profiling; Gene Expression Regulation; Hepatitis C virus; Humans; Receptors, Antigen, T-Cell - metabolism; Receptors, Antigen, T-Cell, alpha-beta - genetics; Sequence Analysis, RNA; Single-Cell Analysis - methods; T-Lymphocytes - metabolism; Transcriptome - genetics; V(D)J Recombination - genetics
• ISSN: 0818-9641; 1440-1711
• DOI: 10.1038/icb.2016.16

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 TCRαβ 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 TCRαβ in 56 of a total of 63 cells (89%), with double α and double β in 18, and 7% respectively, and double TCRαβ 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.