High throughput sequencing reveals a complex pattern of dynamic interrelationships among human T cell subsets

Abstract

Developing T cells face a series of cell fate choices in the thymus and in the periphery. The role of the individual T cell receptor (TCR) in determining decisions of cell fate remains unresolved. The stochastic/selection model postulates that the initial fate of the cell is independent of TCR specificity, with survival dependent on additional TCR/coreceptor "rescue" signals. The "instructive" model holds that cell fate is initiated by the interaction of the TCR with a cognate peptide-MHC complex. T cells are then segregated on the basis of TCR specificity with the aid of critical coreceptors and signal modulators [Chan S, Correia-Neves M, Benoist C, Mathis (1998) Immunol Rev 165: 195-207]. The former would predict a random representation of individual TCR across divergent T cell lineages whereas the latter would predict minimal overlap between divergent T cell subsets. To address this issue, we have used highthroughput sequencing to evaluate the TCR distribution among key T cell developmental and effector subsets from a single donor. We found numerous examples of individual subsets sharing identical TCR sequence, supporting a model of a stochastic process of cell fate determination coupled with dynamic patterns of clonal expansion of T cells bearing the same TCR sequence among both CD4 + and CD8+ populations.F ollowing production of their T cell receptors (TCRs), T cells experience several developing stages. An encounter with a cognate peptide-MHC complex can induce naïve T (Tn) cells expressing the CD45RA isomer to begin to express CD45RO. Cells expressing both isomers are considered transitional in nature (Tt), thus cells identified on the basis of CD45RA expression alone include Tn and Tt and can thus be referred to as Tn+t. Cells expressing only CD45RO have passed into the memory (Tm) compartment, where they can lay quiescent awaiting repeat stimulation by the same or similar peptide-MHC complexes. Activated T cells (Ta) driven to effector function lose expression of both CD45RA and RO and express CD69. During different developing stages, T cells also face a series of cell fate choices: CD4 + CD8+ cells commit to either the CD4 + helper (Th) or CD8+ cytotoxic (Tc) lineages, a choice closely associated with binding to MHC class II or class I peptide complexes, respectively. Subsequently, CD4 + T cells can develop into regulatory (Tr) CD25+ cells, or into CD25−CD294− Th1 (IFN-γ producing) or CD25−CD294+ Th2 (IL-4 producing) effector subsets. Other choices are also available (1, 2).Although it is generally accepted that the TCR expressed by the developing T lineage cell will determine the response to a specific peptide-MHC complex, the role of the individual TCR in determining decisions of cell fate remains unresolved. To address these issues, we have coupled high-throughput sequencing techniques (3, 4) to high volume antibody covered superparamagnetic polystyrene bead isolation of defined T cell subsets with semiquantitative PCR amplification of the complementarity determining region 3 regions (CDR3) from mR...

Determinants of public T cell responses

“…Detailed analyses on recombinatorial biases were facilitated by recent high-throughput sequencings [33,[49][50][51][52] (and our unpublished data), which enable comparison between the empirical TCR β repertoires and the simu- npg lated model being made, so that biases during recombination could be revealed. A simulated TCR β repertoire should incorporate the effect of random convergent recombination, which assumes random nucleotide deletion at the coding ends of those germline segments, and random nucleotide addition at the junctions within different V β -J β combination.…”

Functional TCR Retrieval from Single Antigen-Specific Human T Cells Reveals Multiple Novel Epitopes

“…During the last years, several PCR-based methods have been developed, allowing TCR repertoire analyses at the single-cell level (8)(9)(10)(11)(12) as well as the in vitro reconstitution of full-length a/b TCRs (13-16) from single cells. Likewise, next-generation sequencing methods have evolved that generate millions of short sequence reads, which enable high-throughput profiling of TCR repertoires but have not addressed single T cells, thus precluding analysis of paired TCR-a/b chains (17)(18)(19). Only recently three different approaches addressing the latter limitation were reported.…”

Distinctive properties of identical twins' TCR repertoires revealed by high-throughput sequencing

“…Some of these studies reported the common use of particular V genes and common clonotypes. In recent years, the high-throughput sequencing technologies paved the way to whole-repertoire studies of individual TCRs that led to new findings in the field of adaptive immunity (1,5,6,(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22). In this study, for the first time to the best of our knowledge, we obtain and compare the α and β chain TCR repertoires of three pairs of MZ twins using next-generation sequencing (NGS).…”