Sequence analysis of T-cell repertoires in health and disease

Abstract

T-cell antigen receptor (TCR) variability enables the cellular immune system to discriminate between self and non-self. High-throughput TCR sequencing (TCR-seq) involves the use of next generation sequencing platforms to generate large numbers of short DNA sequences covering key regions of the TCR coding sequence, which enables quantification of T-cell diversity at unprecedented resolution. TCR-seq studies have provided new insights into the healthy human T-cell repertoire, such as revised estimates of repertoire size and the understanding that TCR specificities are shared among individuals more frequently than previously anticipated. In the context of disease, TCR-seq has been instrumental in characterizing the recovery of the immune repertoire after hematopoietic stem cell transplantation, and the method has been used to develop biomarkers and diagnostics for various infectious and neoplastic diseases. However, T-cell repertoire sequencing is still in its infancy. It is expected that maturation of the field will involve the introduction of improved, standardized tools for data handling, deposition and statistical analysis, as well as the emergence of new and equivalently large-scale technologies for T-cell functional analysis and antigen discovery. In this review, we introduce this nascent field and TCR-seq methodology, we discuss recent insights into healthy and diseased TCR repertoires, and we examine the applications and challenges for TCR-seq in the clinic.

Figure 1

T-cell receptor-antigen-peptide-MHC interaction and TCR gene recombination. (a) A T-cell (pink) encountering an antigen-presenting cell (APC; blue). The APC presents peptide antigen (Ag; yellow) in complex with the larger major histocompatibility complex (MHC; turquoise). The T-cell receptor (TCR; multi-colored) binds to both the antigen and MHC, and if the binding avidity is sufficiently high the T-cell is activated. (b) A TCR heterodimer, composed of an α and β chain, engaging peptide-MHC (pMHC). Moving outward from the T cell, the constant region (green) of the TCR is anchored to the cell membrane, followed by the J region (red). In TCR α chains the J region is followed by the V region (orange), whereas in TCR β chains, a D region is located between the V and J regions. The complementarity determining region 3 (CDR3) domain, approximately 45 nucleotides long, comprises the VJ (for TCR-α) or VDJ (for TCR-β) junction. Color gradients at junctions represent the regions encoded by arbitrary, untemplated nucleotides introduced during somatic recombination, and which represent a primary source of sequence diversification and TCR variability (see (c) for details). The CDR3 regions are the main domains of the TCR that are in contact with peptide antigen, and largely determine TCR specificity. (c) Simplified representation of TCR-β VDJ gene recombination resulting in TCR diversity. The TCR-β locus is located on chromosome 7 and is approximately 620 kb in length. Initially one of the two D regions is joined with one of 13 J regions (both randomly selected), followed by joining of the DJ region to one of more than 50 V regions (also randomly selected), yielding a final VDJ region that is approximately 500 bp in length. The mechanism by which gene segments are joined also introduces base pair variability, which together with the combinatorial selection of these segments results in TCR diversity. A completely analogous process occurs for the TCR α chain, without the D gene segment included.