The Inference of Phased Haplotypes for the Immunoglobulin H Chain V Region Gene Loci by Analysis of VDJ Gene Rearrangements

Abstract

The existence of many highly similar genes in the lymphocyte receptor gene loci makes them difficult to investigate, and the determination of phased “haplotypes” has been particularly problematic. However, V(D)J gene rearrangements provide an opportunity to infer the association of Ig genes along the chromosomes. The chromosomal distribution of H chain genes in an Ig genotype can be inferred through analysis of VDJ rearrangements in individuals who are heterozygous at points within the IGH locus. We analyzed VDJ rearrangements from 44 individuals for whom sufficient unique rearrangements were available to allow comprehensive genotyping. Nine individuals were identified who were heterozygous at the IGHJ6 locus and for whom sufficient suitable VDJ rearrangements were available to allow comprehensive haplotyping. Each of the 18 resulting IGHV|IGHD|IGHJ haplotypes was unique. Apparent deletion polymorphisms were seen that involved as many as four contiguous, functional IGHV genes. Two deletion polymorphisms involving multiple contiguous IGHD genes were also inferred. Three previously unidentified gene duplications were detected, where two sequences recognized as allelic variants of a single gene were both inferred to be on a single chromosome. Phased genomic data brings clarity to the study of the contribution of each gene to the available repertoire of rearranged VDJ genes. Analysis of rearrangement frequencies suggests that particular genes may have substantially different yet predictable propensities for rearrangement within different haplotypes. Together with data highlighting the extent of haplotypic variation within the population, this suggests that there may be substantial variability in the available Ab repertoires of different individuals.

Chromatin conformation governs T-cell receptor Jβ gene segment usage

“…An intriguing possibility is that genetic changes such as insertions or deletions in noncoding regions, as well as deletions or duplications of genes (40), could change distances between VDJ gene segments, thus altering their frequencies in the repertoire. Such changes can in turn tune the composition of the set of public clonotypes in accordance with threats posed by common pathogens, and potentially also with evolving needs for self-maintenance (41,42).…”

Automated analysis of high-throughput B-cell sequencing data reveals a high frequency of novel immunoglobulin V gene segment alleles

“…Application of this approach to the seven subjects in Table 1 found that each subject had a unique genotype, a result similar to that found in ref. 6. Most (63%) IGHV genes were homozygous, and duplications of IGHV1-69 or IGHV2-70 were found in three cases (Table S1).…”

“…Existing approaches for inferring Ig genotypes are based on identifying the set of alleles that appear above a specified frequency (6,19). This frequency is calculated using the set of unmutated sequences, as existing V(D)J segment identification tools are most accurate at identifying and aligning these sequences to their germline sequences (2).…”

“…The IMGT (18) database of germline Ig alleles is the most widely used, and the National Center for Biotechnology Information refers to IMGT for its reference genome. However, recent studies have discovered the presence of numerous V segments and alleles not reported in any published databases (6,(19)(20)(21), as well as a several novel D and J alleles (19). Some of these V alleles have been incorporated into the IMGT database or alternative databases of germline alleles [such as VBASE2 (22) or the UNSWIg human heavy chain repertoire (23)].…”

“…Previous studies have, for example: revealed the association of certain germline genotypes with susceptibility to such diseases as rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), and multiple sclerosis (MS) (5,6); found correlations of age with a reduced Ig clonal diversity and less intense response to immune challenge (7,8); found overly expanded clones in cases of lymphoma (9,10); and discovered convergent Ig evolution across subjects in response to certain immune challenges (11,12). These repertoire-sequencing (Rep-Seq) studies have benefitted from improvements in sequencing technologies, which allow for the generation of millions of reads per run (13).…”