Antibody repertoire analysis in polygenic autoimmune diseases

Bashford-Rogers, Rachael; Smith, Kenneth G. C.; Thomas, David

doi:10.1111/imm.12927

Cited by 61 publications

(50 citation statements)

References 132 publications

(237 reference statements)

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“…The usage of immunoglobulin heavy chain variable ( IGHV) genes and their mutational status are most frequently studied in relation to cancer 13, 14 , responses to vaccines 15, 16 , or in autoimmune diseases 17–19 . Most IGHV genes have several allelic variants and more alleles are being discovered as a result of adaptive immune receptor repertoire-sequencing (AIRR-seq) 20, 21 .…”

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confidence: 99%

Polymorphisms in immunoglobulin heavy chain variable genes and their upstream regions

Mikocziova

Gidoni

Lindeman

et al. 2020

Preprint

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14Germline variations in immunoglobulin genes influence the repertoire of B cell receptors and 15 antibodies, and such polymorphisms may impact disease susceptibility. However, the knowledge of 16 the genomic variation of the immunoglobulin loci is scarce. Here, we report 25 novel germline IGHV 17 alleles as inferred from rearranged naïve B cell cDNA repertoires of 98 individuals. Thirteen novel 18 alleles were selected for validation, out of which ten were successfully confirmed by targeted 19 amplification and Sanger sequencing of non-B cell DNA. Moreover, we detected a high degree of 20 variability upstream of the V-region in the 5'UTR, leader 1, and leader 2 sequences, and found that These loci remain incompletely characterized due to the fact that they contain many repetitive 35 sequence segments with many duplicated genes 4 , which makes it difficult to correctly assemble short 36 reads from whole genome sequencing. Single nucleotide polymorphisms as well as copy number 37 variations are in linkage disequilibrium and make up distinct haplotypes 4 . To this date, a limited 38 number of genomically sequenced 5-7 and inferred 8,9 haplotypes of the heavy chain and the two light 39 chain loci have been described. Different databases exist for genomic immune receptor DNA 40 sequences (IMGT/GENE-DB 10 ), putative novel variants from inferred data (IgPdb 11 ) or entire immune 41 receptor repertoires (OGRDB 12 ). 42The usage of immunoglobulin heavy chain variable (IGHV) genes and their mutational status are most 43 frequently studied in relation to cancer 13,14 , responses to vaccines 15,16 , or in autoimmune diseases [17][18][19] . 44Most IGHV genes have several allelic variants and more alleles are being discovered as a result of 45 adaptive immune receptor repertoire-sequencing (AIRR-seq) 20,21 . Software tools such as TIgGER 22,23 , 46IgDiscover 24 and partis 25 allow to infer germline alleles from such repertoire data. Based on these 47 inferred alleles, the data can then be input to other tools that infer haplotypes and repertoire 48 deletions 26 . Incorrect annotation could possibly lead to inferring wrong deletions and biased 49 assessments. Therefore, having a full overview of germline variants is essential for studying the 50 adaptive immune response with high accuracy.51 Some allelic variants have been associated with increased disease susceptibility 27,28 , yet the impact of 52 immunoglobulin gene variation on disease risks is still unknown 29 . These regions have not been 53 sufficiently covered in the numerous genome wide association studies performed to date. More 54 comprehensive maps of polymorphisms are required for proper analysis. 55Here, we have used previously generated AIRR-seq data 30 from naïve B-cells of 98 Norwegian 56 individuals to identify novel IGHV alleles, a selection of which we then validated from genomic DNA 57 (gDNA) of non-B cells, i.e. T cells and monocytes. We also analyzed the sequences upstream of the 58 V-region, and constructed consensus sequences for the upstream variant...

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confidence: 99%

Polymorphisms in immunoglobulin heavy chain variable genes and their upstream regions

Mikocziova

Gidoni

Lindeman

et al. 2020

Preprint

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“…There is also enhanced focus on T and B lymphocyte profiling in infections [7], or in patients treated with vaccines [8] or antibody-based immunotherapies [9]. Additionally, studies have also investigated antibody repertoires in patients with autoimmune disorders [10, 11]. Inspired by these early efforts, large disease-focused consortia are increasingly investing in SC transcriptomics on human biological samples due to the broad readout that this technology can provide using only a small amount of tissue as input (cf.…”

Section: Introductionmentioning

confidence: 99%

An optimized workflow for single-cell transcriptomics and repertoire profiling of purified lymphocytes from clinical samples

Hanamsagar

Reizis

Chamberlain

et al. 2019

Preprint

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Establishing clinically relevant single-cell (SC) transcriptomic workflows from cryopreserved tissue is essential to move this emerging immune monitoring technology from the bench to the bedside. Improper sample preparation leads to detrimental cascades, resulting in loss of precious time, money and finally compromised data. There is an urgent need to establish protocols specifically designed to overcome the inevitable variations in sample quality resulting from uncontrollable factors in a clinical setting. Here, we explore sample preparation techniques relevant to a range of clinically relevant scenarios, where SC gene expression and repertoire analysis are applied to a cryopreserved sample derived from a small amount of blood, with unknown or partially known preservation history. We compare a total of ten cell-counting, viability-improvement, and lymphocyte-enrichment methods to highlight a number of unexpected findings. Trypan blue-based automated counters, typically recommended for single-cell sample quantitation, consistently overestimate viability. Advanced sample clean-up procedures significantly impact total cell yield, while only modestly increasing viability. Finally, while pre-enrichment of B cells from whole peripheral blood mononuclear cells (PBMCs) results in the most reliable BCR repertoire data, comparable T-cell enrichment strategies distort the ratio of CD4+ and CD8+ cells. Furthermore, we provide high-resolution analysis of gene expression and clonotype repertoire of different B cell subtypes. Together these observations provide both qualitative and quantitative sample preparation guidelines that increase the chances of obtaining high-quality single-cell transcriptomic and repertoire data from human PBMCs in a variety of clinical settings.

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“…Detailed characterization of B cells and their respective BCR sequences offers important information on B-cell generation and selection as well as immune competence in health and disease. High-throughput sequencing of antibody genes (Ig-seq) has become a widely used tool in human translational research (4,5). Abnormal B-cell responses can be explored by investigating BCR repertoires from patients and comparing their characteristics to those of healthy controls.…”

Section: Introductionmentioning

confidence: 99%

Maturation of the human B-cell receptor repertoire with age

Ghraichy

Galson

Kovaltsuk

et al. 2019

Preprint

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B cells play a central role in adaptive immune processes, mainly through the production of antibodies. The maturation of the B-cell system with age is poorly studied. We extensively investigated age-related alterations of naïve and antigen-experienced B-cell receptor (BCR) repertoires. The most significant changes were observed in the first 10 years of life, and were characterized by altered immunoglobulin gene usage and an increased frequency of mutated antibodies structurally diverging from their germline precursors. Older age was associated with an increased usage of downstream constant region genes and fewer antibodies with self-reactive properties. As mutations accumulated with age, the frequency of germline-encoded self-reactive antibodies decreased, indicating a possible beneficial role of self-reactive B-cells in the developing immune system. Our results suggest a continuous process of change through childhood across a broad range of parameters characterizing BCR repertoires and stress the importance of using wellselected, age-appropriate controls in BCR studies.(149 words)

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Antibody repertoire analysis in polygenic autoimmune diseases

Cited by 61 publications

References 132 publications

Polymorphisms in immunoglobulin heavy chain variable genes and their upstream regions

Polymorphisms in immunoglobulin heavy chain variable genes and their upstream regions

An optimized workflow for single-cell transcriptomics and repertoire profiling of purified lymphocytes from clinical samples

Maturation of the human B-cell receptor repertoire with age

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