Determination of gene organization in the human IGHV region on single chromosomes

Abstract

Organization of the IGHV genes (n ¼ 108) on single human chromosomes has been determined by detecting these sequences in single sperm using multiplex PCR amplification followed by microarray detection. A total of 374 single sperm samples from five Caucasian males were studied. Three deletion/insertion polymorphisms (Del I-Del III) with deletion allele frequencies ranging from 0.1 to 0.3 were identified. Del I is a previously reported polymorphism affecting three IGHV genes (IGHV1-8, IGHV3-9, and IGHV2-10). Del II affects a region 2-18 kb containing two pseudogenes IGHV(II)-28.1 and IGHV3-29, and Del III spans B21-53 kb involving genes IGHV4-39, IGHV7-40, IGHV(II)-40-1, and IGHV3-41. Deletion alleles of both Dels II and III were found in a heterozygous state, and therefore, could not be easily detected if haploid samples were not used in the study. Results of the present study indicate that deletions/insertions together with other possible chromosomal rearrangements may play an important role in forming the genetic structure of the IGHV region, and may significantly contribute to antibody diversity. Since these three polymorphisms are located within or next to the 3 0 half of the IGHV region, they may have an important role in the expressed IGHV gene repertoire during immune response.

The immunoglobulin heavy chain locus: genetic variation, missing data, and implications for human disease

“…64 Importantly, four of these five germline IGHV genes contributing to these antibodies are known to vary in copy number. 33,38,40,41,51,52 Taken together, such observations provide a potential example for which complete descriptions of IGHV gene allelic and genomic structural variation may prove essential for understanding infectious disease susceptibility, and useful in the administration of vaccines aimed at eliciting specific antibodies.…”

“…There are several examples of homozygous deletions containing functional IGHV genes (Table 1). 28,33,38,39,51,52 In addition, an analysis of naive B-cell IGHV gene repertoires using next-generation 454 sequencing revealed that 15 of the 56 IGHV genes described from these repertoires were observed in only a subset of the 12 individuals screened. 6 Seven of these fifteen genes were previously reported to occur in insertion/deletion polymorphisms, and thus these observed 'holes' in the naive repertoire are likely due to homozygous deletions.…”

“…6 For example, the proportion of the gene IGHV4-39 in expressed repertoires ranged from 2-8%. Given that IGHV4-39 is known to be part of a deletion haplotype that occurs at a frequency of 17-25% in Caucasians, 38,51,52 it is possible that interindividual variation in IGHV4-39 expression is related to differences in copy number at this locus. Boyd et al 6 also showed skewed usage ratios in individual repertoires for multiple alleles expressed from the same locus; in some cases certain alleles were expressed twofold to threefold higher than other alleles in the same individual.…”

“…The presence of alternate IGHV haplotypes in the IGHV cluster has been explored extensively using a variety of techniques, including restriction-fragment length polymorphisms, [25][26][27][28][29][30] analysis of largeinsert clones, [31][32][33][34][35] and PCR and microarray-based methods utilizing haploid sperm samples. [36][37][38][39] These approaches have identified a number of CNV polymorphisms, including insertions Figure 1. The number of known alleles for each of the mapped and unmapped IGHV genes (functional and ORF).…”

“…39 For example, an B50-Kb insertion with at least four structural variants, and up to four additional haplotypes of various sizes containing deletions and/or nucleotide (nt) polymorphisms compared with the NCBI reference assembly, is known to occur in the IGHV3-30/IGHV4-31 gene region. 30,33,[38][39][40][41] Another insertion polymorphism that is known to include the gene, IGHV1-69, has also been identified in the distal part of the locus. 31 At least 12 haplotypes in this region that vary with respect to IGHV1-69 copy number and allelic polymorphism have been identified in Caucasians.…”