Sequence Transfers between Variable Regions in a Mouse Antibody Transgene Can Occur by Gene Conversion

D'Avirro, Nicole; Truong, David M.; Xu, Bo; Selsing, Erik

doi:10.4049/jimmunol.175.12.8133

Cited by 9 publications

(5 citation statements)

References 36 publications

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“…The chimeras we report do not resemble the mosaic gene conversion products reported by others (8,26,27). Rather, entire CDR1 regions are replaced, not segments of them as in the case of converted V H genes.…”

Section: Discussioncontrasting

confidence: 79%

Antibody Repertoire Development in Fetal and Neonatal Piglets. XIII. Hybrid VH Genes and the Preimmune Repertoire Revisited

Butler

Weber

Wertz

2006

The Journal of Immunology

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The expressed porcine VH genes belong to the VH3 family (clan), four of which, VHA, VHB, VHC, and VHE, alone comprise ∼80% of the preimmune repertoire. However, so-called “hybrid” VH genes that use CDR1 of one VH gene and the CDR2 of another are frequently encountered. We studied >3000 cloned VDJs and found that such hybrids can contribute up to 10% of the preimmune repertoire. Based on the 1) recovery of hybrid VH genes from bacterial artificial chromosome clones, 2) frequency of occurrence of certain hybrids in the preimmune repertoire, and 3) failure to recover equal numbers of reciprocal hybrids, we concluded that some chimeric genes are present in the genome and are not PCR artifacts. Two chimeric germline genes (VHZ and VHY), together with VHF and the four genes mentioned above, constitute the major VH genes and these account for >95% of the preimmune repertoire. Diversification of the preimmune IgG and IgM repertoires after environmental exposure was mainly due to somatic hypermutation of major VH genes with no evidence of gene conversion. Somatic hypermutation was 3- to 10-fold higher in CDRs than in framework regions, most were R mutations and transversions and transitions equally contributed. Data were used to 1) develop an index to quantify the degree of VH repertoire diversification and 2) establish a library of 29 putative porcine VH genes. One-third of these genes are chimeric genes and their sequences suggest that the porcine VH genome developed by duplication and splicing from a small number of prototypic genes.

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Section: Discussioncontrasting

confidence: 79%

Antibody Repertoire Development in Fetal and Neonatal Piglets. XIII. Hybrid VH Genes and the Preimmune Repertoire Revisited

Butler

Weber

Wertz

2006

The Journal of Immunology

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“…In rabbits, 23% of IgG sequences and 32% of Igκ sequences were the products of gene conversion. There have been previous, although somewhat controversial, indications suggesting gene conversion occurs in humans and mice as well, albeit at a much lower frequency [47], [48], [49]. We find that, in the mouse, putative gene conversion events are nearly absent, with an estimated frequency of 0.1% of all unique IgG sequences.…”

Section: Resultscontrasting

confidence: 43%

Systematic Characterization and Comparative Analysis of the Rabbit Immunoglobulin Repertoire

et al. 2014

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Rabbits have been used extensively as a model system for the elucidation of the mechanism of immunoglobulin diversification and for the production of antibodies. We employed Next Generation Sequencing to analyze Ig germline V and J gene usage, CDR3 length and amino acid composition, and gene conversion frequencies within the functional (transcribed) IgG repertoire of the New Zealand white rabbit (Oryctolagus cuniculus). Several previously unannotated rabbit heavy chain variable (VH) and light chain variable (VL) germline elements were deduced bioinformatically using multidimensional scaling and k-means clustering methods. We estimated the gene conversion frequency in the rabbit at 23% of IgG sequences with a mean gene conversion tract length of 59±36 bp. Sequencing and gene conversion analysis of the chicken, human, and mouse repertoires revealed that gene conversion occurs much more extensively in the chicken (frequency 70%, tract length 79±57 bp), was observed to a small, yet statistically significant extent in humans, but was virtually absent in mice.

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“…There is also evidence of GCV for the diversification of Vλ repertoire of cattle (Parng et al, 1996; Lucier et al, 1998). Some good evidence of GCV has been found in mice (Dildrop et al, 1982; Krawinkel et al, 1983; David et al, 1992; Selsing et al, 1996; D’Avirro et al, 2002, 2005; Tsai et al, 2002). GCV in human rearranged immunoglobulin genes have not been identified except for one recent study (Darlow and Stott, 2006).…”

Section: Introductionmentioning

confidence: 98%

Gene Conversion-Like Events in the Diversification of Human Rearranged IGHV3-23*01 Gene Sequences

Duvvuri¹,

Wu²

2012

Front. Immun.

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Gene conversion (GCV), a mechanism mediated by activation-induced cytidine deaminase (AID) is well established as a mechanism of immunoglobulin diversification in a few species. However, definitive evidence of GCV-like events in human immunoglobulin genes is scarce. The lack of evidence of GCV in human rearranged immunoglobulin gene sequences is puzzling given the presence of highly similar germline donors and the presence of all the enzymatic machinery required for GCV. In this study, we undertook a computational analysis of rearranged IGHV3-23*01 gene sequences from common variable immunodeficiency (CVID) patients, AID-deficient patients, and healthy individuals to survey “GCV-like” activities. We analyzed rearranged IGHV3-23*01 gene sequences obtained from total PBMC RNA and single-cell polymerase chain reaction of individual B cell lysates. Our search identified strong evidence of GCV-like activity. We observed that GCV-like tracts are flanked by AID hotspot motifs. Structural modeling of IGHV3-23*01 gene sequence revealed that hypermutable bases flanking GCV-like tracts are in the single stranded DNA (ssDNA) of stable stem-loop structures (SLSs). ssDNA is inherently fragile and also an optimal target for AID. We speculate that GCV could have been initiated by the targeting of hypermutable bases in ssDNA state in stable SLSs, plausibly by AID. We have observed that the frequency of GCV-like events is significantly higher in rearranged IGHV3-23-*01 sequences from healthy individuals compared to that of CVID patients. We did not observe GCV-like events in rearranged IGHV3-23-*01 sequences from AID-deficient patients. GCV, unlike somatic hypermutation (SHM), can result in multiple base substitutions that can alter many amino acids. The extensive changes in antibody affinity by GCV-like events would be instrumental in protecting humans against pathogens that diversify their genome by antigenic shift.

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Sequence Transfers between Variable Regions in a Mouse Antibody Transgene Can Occur by Gene Conversion

Cited by 9 publications

References 36 publications

Antibody Repertoire Development in Fetal and Neonatal Piglets. XIII. Hybrid VH Genes and the Preimmune Repertoire Revisited

Antibody Repertoire Development in Fetal and Neonatal Piglets. XIII. Hybrid VH Genes and the Preimmune Repertoire Revisited

Systematic Characterization and Comparative Analysis of the Rabbit Immunoglobulin Repertoire

Gene Conversion-Like Events in the Diversification of Human Rearranged IGHV3-23*01 Gene Sequences

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