The shape of the lymphocyte receptor repertoire: lessons from the B cell receptor

Abstract

Both the B cell receptor (BCR) and the T cell receptor (TCR) repertoires are generated through essentially identical processes of V(D)J recombination, exonuclease trimming of germline genes, and the random addition of non-template encoded nucleotides. The naïve TCR repertoire is constrained by thymic selection, and TCR repertoire studies have therefore focused strongly on the diversity of MHC-binding complementarity determining region (CDR) CDR3. The process of somatic point mutations has given B cell studies a major focus on variable (IGHV, IGLV, and IGKV) genes. This in turn has influenced how both the naïve and memory BCR repertoires have been studied. Diversity (D) genes are also more easily identified in BCR VDJ rearrangements than in TCR VDJ rearrangements, and this has allowed the processes and elements that contribute to the incredible diversity of the immunoglobulin heavy chain CDR3 to be analyzed in detail. This diversity can be contrasted with that of the light chain where a small number of polypeptide sequences dominate the repertoire. Biases in the use of different germline genes, in gene processing, and in the addition of non-template encoded nucleotides appear to be intrinsic to the recombination process, imparting "shape" to the repertoire of rearranged genes as a result of differences spanning many orders of magnitude in the probabilities that different BCRs will be generated. This may function to increase the precursor frequency of naïve B cells with important specificities, and the likely emergence of such B cell lineages upon antigen exposure is discussed with reference to public and private T cell clonotypes.

Keywords: BCR repertoire; TCR repertoire; V(D)J recombination; combinatorial diversity; junctional diversity; private clonotypes; public clonotypes.

Figure 1

The receptor repertoire has “shape,” as biases and constraints in the recombination process vary the probabilities of generating particular V(D)J rearrangements. (A) Higher probability rearrangements are generated through utilization of more frequently rearranged gene segments. These segments are joined with minimal gene processing and N-addition, increasing the chance of independent rearrangements with identical or near identical CDR3s. (B) Lower probability rearrangements utilize rarely rearranged germline gene segments and the CDR3s are more diverse owing to increased nucleotide removals and additions at the joins. (C) The many order of magnitude differences in the likelihood of generating particular rearrangements shape the repertoire, with higher probability rearrangements being frequently generated and as a consequence being carried by many identical B-cells. Only a relatively small number of unique rearrangements will be generated with probabilities high enough to be carried by a large number of B cells, but this should ensure that they are always present in the repertoire at significant levels. Conversely, lower probability rearrangements may be so rare that they are carried only by a single B cell, or are entirely absent from the repertoire. The lower probability rearrangements that are carried by just one or at most a few B-cells likely represent many millions of unique rearrangements.

Figure 2

Inference of IGH haplotypes from VDJ rearrangements. The availability of large datasets of VDJ rearrangements from single individuals permit the inference of all germline V, D, and J genes within the genome, from analysis of apparent gene utilization within a dataset. As IGH VDJ rearrangement is an intra-chromosomal event, gene pairing in VDJ rearrangements can be used to infer which gene segments are carried by the same chromosome. Leveraging the heterozygous IGHJ6 or IGHJ4 locus allows the reconstruction of IGH gene segments on each chromosome and for the IGH haplotype to be inferred.

Figure 3

Germline gene segment sequences constrain junctional diversity. This is illustrated by processing of the codon TAC, which encodes tyrosine as the most 3′ amino acid in the preferred reading frame of many IGHD genes. A highly probable outcome of the VDJ rearrangement process is the loss of a single nucleotide from the IGHD coding end. If this occurs, the productive repertoire for these Ds is strongly skewed toward tyrosine, as two of the four possible nucleotide replacement events generate stop codons, and the other two ensure the maintenance of tyrosine. These nucleotides may be sourced from either N-addition or from the 5′ end of an IGHJ segment. The outcome of N-addition is depicted in the bottom panel. N-additions are colored in orange and below each of the four possible outcomes is shown the likelihood of the enzyme TdT adding each nucleotide base.