Genetic Control of DH Reading Frame and Its Effect on B-Cell Development and Antigen-Specifc Antibody Production

Schroeder, Harry W.; Zemlin, Michael; Khass, Mohamed; Nguyen, Huan Huu; Schelonka, Robert L.

doi:10.1615/critrevimmunol.v30.i4.20

Cited by 42 publications

(86 citation statements)

References 74 publications

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“…In the ΔD-iD strain, the center of the DFL16.1 segment was replaced with the DSP2.2 D H gene segment in inverted form. This leads to CDR-H3 enriched for charged arginine, asparagine, and histidine in place of tyrosine and glycine (6, 8). …”

Section: Resultsmentioning

confidence: 99%

“…To test the role of natural selection of D gene segment sequence on humoral immune function, we previously created a panel of BALB/c-derived D-altered mutant mouse strains (6–8). ΔD-DμFS and ΔD-iD B cells produce two alternative, polyclonal Ig repertoires with a normal and intact set of V H , J H , and C H exons that are fully capable of undergoing somatic hypermutation and class switching (6, 8).…”

Section: Introductionmentioning

confidence: 99%

“…ΔD-DμFS and ΔD-iD B cells produce two alternative, polyclonal Ig repertoires with a normal and intact set of V H , J H , and C H exons that are fully capable of undergoing somatic hypermutation and class switching (6, 8). The only change that has been made is the simplification of D H locus to contain only one D of alternative sequence.…”

Section: Introductionmentioning

confidence: 99%

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Clonal Progression during the T Cell-Dependent B Cell Antibody Response Depends on the Immunoglobulin DH Gene Segment Repertoire

et al. 2014

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The diversity of the third complementarity determining region of the IgH chain is constrained by natural selection of immunoglobulin diversity (DH) sequence. To test the functional significance of this constraint in the context of thymus-dependent (TD) immune responses, we immunized BALB/c mice with WT or altered DH sequence with 2-phenyloxazolone-coupled chicken serum albumin (phOx-CSA). We chose this antigen because studies of the humoral immune response to the hapten phOx were instrumental in the development of the current theoretical framework on which our understanding of the forces driving TD responses is based. To allow direct comparison, we used the classic approach of generating monoclonal Ab (mAb) from various stages of the immune response to phOx to assess the effect of changing the sequence of the DH on clonal expansion, class switching, and affinity maturation, which are hallmarks of TD responses. Compared to WT, TD-induced humoral IgM as well as IgG antibody production in the D-altered ΔD-DμFS and ΔD-iD strains were significantly reduced. An increased prevalence of IgM-producing hybridomas from late primary, secondary, and tertiary memory responses suggested either impaired class switch recombination (CSR) or impaired clonal expansion of class switched B cells with phOx reactivity. Neither of the D-altered strains demonstrated the restriction in the VH/VL repertoire, the elimination of VH1 family-encoded antibodies, the focusing of the distribution of CDR-H3 lengths, or the selection for the normally dominant Ox1 clonotype, which all are hallmarks of the anti-phOx response in WT mice. These changes in clonal selection and expansion, as well as CSR indicate that the genetic constitution of the DH locus, which has been selected by evolution, can strongly influence the functional outcome of a TD humoral response.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Clonal Progression during the T Cell-Dependent B Cell Antibody Response Depends on the Immunoglobulin DH Gene Segment Repertoire

et al. 2014

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“…However, it is clear that structural and functional constraints, as well as clonal selection operating at different developmental stages influence its ultimate size and shape. 35 Owing to the capacity to screen a larger sample of the lymphocyte repertoire, HTS is offering the possibility to approach its complexity, and how it is affected during normal and pathological immune response. 5 Additionally, the information generated by HTS on antibody repertoires can be exploited to address higher order statistical properties of biological structures in general.…”

Section: Discussionmentioning

confidence: 99%

Reconstructing and mining the B cell repertoire with ImmunediveRsity

Cortina-Ceballos

Godoy-Lozano

Sámano-Sánchez

et al. 2015

mAbs

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(2015) Reconstructing and mining the B cell repertoire with ImmunediveRsity , mAbs, 7:3, 516-524, DOI: 10.1080/19420862.2015 To link to this article: https://doi.org/10. 1080/19420862.2015 Keywords: high-throughput sequencing, Ig repertoire, CDR3, data mining Abbreviations: HEL, hen egg lysozyme; CDRH3, heavy chain complementarity determining region 3; Rep-Seq, repertoire sequencing; SHM, somatic hypermutation.The B cell antigen receptor repertoire is highly diverse and constantly modified by clonal selection. High-throughput DNA sequencing (HTS) of the lymphocyte repertoire (Rep-Seq) represents a promising technology to explore such diversity ex-vivo and assist in the identification of antigen-specific antibodies based on molecular signatures of clonal selection. Therefore, integrative tools for repertoire reconstruction and analysis from antibody sequences are needed. We developed ImmunediveRity, a stand-alone pipeline primarily based in R programming for the integral analysis of B cell repertoire data generated by HTS. The pipeline integrates GNU software and in house scripts to perform quality filtering, sequencing noise correction and repertoire reconstruction based on V, D and J segment assignment, clonal origin and unique heavy chain identification. Post-analysis scripts generate a wealth of repertoire metrics that in conjunction with a rich graphical output facilitates sample comparison and repertoire mining. Its performance was tested with raw and curated human and mouse 454-Roche sequencing benchmarks providing good approximations of repertoire structure. Furthermore, ImmunediveRsity was used to mine the B cell repertoire of immunized mice with a model antigen, allowing the identification of previously validated antigen-specific antibodies, and revealing different and unexpected clonal diversity patterns in the post-immunization IgM and IgG compartments. Although ImmunediveRsity is similar to other recently developed tools, it offers significant advantages that facilitate repertoire analysis and repertoire mining. ImmunediveRsity is open source and free for academic purposes and it runs on 64 bit GNU/Linux and MacOS. Available at: https://bitbucket.org/ImmunediveRsity/immunediversity/

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“…Each B cell expresses a unique BCR, and each individual mouse or human has the capacity to generate up to 10 13 or more distinct BCRs in the primary repertoire (13), with a large fraction of these being generated by junctional diversification of IgH and IgL CDR3s (14). In this regard, the ability to quantitatively identify the IgH and IgL variable region exons that contribute to the primary antibody repertoire is of great interest in elucidating contributions of this repertoire to immune responses and to immune diseases (15).…”

Section: Significancementioning

confidence: 99%

Highly sensitive and unbiased approach for elucidating antibody repertoires

Lin

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Developing B lymphocytes undergo V(D)J recombination to assemble germ-line V, D, and J gene segments into exons that encode the antigen-binding variable region of Ig heavy (H) and light (L) chains. IgH and IgL chains associate to form the B-cell receptor (BCR), which, upon antigen binding, activates B cells to secrete BCR as an antibody. Each of the huge number of clonally independent B cells expresses a unique set of IgH and IgL variable regions. The ability of V(D)J recombination to generate vast primary B-cell repertoires results from a combinatorial assortment of large numbers of different V, D, and J segments, coupled with diversification of the junctions between them to generate the complementary determining region 3 (CDR3) for antigen contact. Approaches to evaluate in depth the content of primary antibody repertoires and, ultimately, to study how they are further molded by secondary mutation and affinity maturation processes are of great importance to the B-cell development, vaccine, and antibody fields. We now describe an unbiased, sensitive, and readily accessible assay, referred to as high-throughput genome-wide translocation sequencing-adapted repertoire sequencing (HTGTS-Repseq), to quantify antibody repertoires. HTGTS-Rep-seq quantitatively identifies the vast majority of IgH and IgL V(D)J exons, including their unique CDR3 sequences, from progenitor and mature mouse B lineage cells via the use of specific J primers. HTGTS-Rep-seq also accurately quantifies DJ H intermediates and V(D)J exons in either productive or nonproductive configurations. HTGTS-Rep-seq should be useful for studies of human samples, including clonal B-cell expansions, and also for following antibody affinity maturation processes. 1). In this process, the V, D, and J coding ends are generated as covalent hairpins that must be opened and that are often further processed, before being joined by classical nonhomologous end joining (2). Processing of V, D, J coding ends can involve generation of deletions or insertions of nucleotides at the junction regions (2), including the frequent de novo addition of nucleotides by the terminal deoxynucleotidyl transferase component of the V(D)J recombination process (3). Notably the V(D)J junctional region encodes a major antigen contact region of the antibody variable region, known as complementarity determining region 3 (CDR3), and thus these junctional diversification processes make a huge contribution to antibody diversity.The mouse IgH locus spans 2.7 megabases (Mb). There are 100s of V H s in the several megabase distal portion of the IgH, with the number varying substantially in certain mouse strains (4). The V H s lie ∼100 kb upstream from a 50-kb region containing 13 D H s, which is followed several kilobases downstream by a 2-kb region containing four J H s. The IgH constant region (C H ) exons lie downstream of the J H s. After assembly of a V H DJ H exon, transcription initiates upstream of the V H and terminates downstream of the C H exons, with V(D)J and C H portions being fused...

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Genetic Control of DH Reading Frame and Its Effect on B-Cell Development and Antigen-Specifc Antibody Production

Cited by 42 publications

References 74 publications

Clonal Progression during the T Cell-Dependent B Cell Antibody Response Depends on the Immunoglobulin DH Gene Segment Repertoire

Clonal Progression during the T Cell-Dependent B Cell Antibody Response Depends on the Immunoglobulin DH Gene Segment Repertoire

Reconstructing and mining the B cell repertoire with ImmunediveRsity

Highly sensitive and unbiased approach for elucidating antibody repertoires

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