Long-Range Regulation of V(D)J Recombination

Proudhon, Charlotte; Hao, Bingtao; Raviram, Ramya; Chaumeil, Julie; Skok, Jane A.

doi:10.1016/bs.ai.2015.07.003

Cited by 66 publications

(72 citation statements)

References 143 publications

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“…The Vκ locus organization is distinct from that of the V H locus. Besides not having D segments and, therefore, undergoing direct Vκ-to-Jκ rearrangements, the Vκ locus contains V segments organized in both direct and inverted orientation relative to the Jκ segments (6) (Fig. 3A).…”

Section: Resultsmentioning

confidence: 99%

“…IgL chain variable region exons are assembled from just V and J segments but otherwise follow similar basic principles to those of IgH. The mouse Igκ light chain locus spans 3.2 Mb with 100s of Vκs in a 3.1-Mb region separated by 20 kb from five Jκs downstream whereas the Igλ light chain locus is smaller and less complex (6). RNA splicing again joins assembled VJ L exons to corresponding C L exons.…”

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confidence: 99%

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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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Section: Resultsmentioning

confidence: 99%

mentioning

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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“…In that regard, separating compacted from non-compacted chromatin fibers (Bantignies et al, 2011; Clowney et al, 2012; Markenscoff-Papadimitriou et al, 2014) could be a more urgent task for the maintenance of a cell-type specific expression program than the convergence of co-regulated loci. In this regard, interchromosomal association of co-regulated loci (Clowney et al, 2012; Markenscoff-Papadimitriou et al, 2014) due to its stochastic nature may be suitable only for biological systems that tolerate, or even seek, stochasticity like the olfactory and the immune systems (Dekker and Mirny, 2016; Monahan and Lomvardas, 2015; Proudhon et al, 2015; Raviram et al, 2016). Nevertheless, non-stereotypic convergence of multiple gene loci (Osborne et al, 2007) and regulatory sequences in transcription factories (Li et al, 2012a) may also contribute to their co-regulation and their transcription at similar rates at a cellular population level.…”

Section: Discussionmentioning

confidence: 99%

Soft X-Ray Tomography Reveals Gradual Chromatin Compaction and Reorganization during Neurogenesis In Vivo

et al. 2016

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Summary The realization that nuclear distribution of DNA, RNA and proteins differs between cell types and developmental stages suggests that nuclear organization serves regulatory functions. Understanding the logic of nuclear architecture and how it contributes to differentiation and cell fate commitment remains challenging. Here, we use Soft X-ray Tomography (SXT) to image chromatin organization, distribution and biophysical properties during neurogenesis in vivo. Our analyses reveal that chromatin with similar biophysical properties forms an elaborate connected network throughout the entire nucleus. Although this interconnectivity is present in every developmental stage, differentiation proceeds with concomitant increase in chromatin compaction and redistribution of condensed chromatin towards the nuclear core. HP1β, but not nucleosome spacing or phasing, regulates chromatin rearrangements since it governs both the compaction of chromatin and its interactions with the nuclear envelope. Our experiments introduce SXT as a powerful imaging technology for nuclear architecture.

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“…GATA3 binds to the (A/T)GATAA sequence (29,57,58) in chromatin, and while not ubiquitously expressed, GATA3 has been shown to function as either a transcriptional activator or a repressor in many different tissues and cell types (29,30,(59)(60)(61)(62)(63). A second viable hypothesis is that GATA3 directly controls the accessibility or nuclear localization of the Tcrb loci in chromatin (64), and it is this feature of the nuclear architecture that is regulated in the GATA3 pathway. Computationally, more than 10 high-affinity binding sites for GATA3 have been identified (by chromatin immunoprecipitation sequencing [ChIP-seq]) within the Tcrb locus (58) (Fig.…”

Section: Discussionmentioning

confidence: 99%

GATA3 Abundance Is a Critical Determinant of T Cell Receptor β Allelic Exclusion

Sekiguchi

Panwar

et al. 2017

Molecular and Cellular Biology

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Allelic exclusion describes the essential immunological process by which feedback repression of sequential DNA rearrangements ensures that only one autosome expresses a functional T or B cell receptor. In wild-type mammals, approximately 60% of cells have recombined the DNA of one T cell receptor ␤ (TCR␤) V-to-DJ-joined allele in a functional configuration, while the second allele has recombined only the DJ sequences; the other 40% of cells have recombined the V to the DJ segments on both alleles, with only one of the two alleles predicting a functional TCR␤ protein. Here we report that the transgenic overexpression of GATA3 leads predominantly to biallelic TCR␤ gene (Tcrb) recombination. We also found that wild-type immature thymocytes can be separated into distinct populations based on intracellular GATA3 expression and that GATA3 LO cells had almost exclusively recombined only one Tcrb locus (that predicted a functional receptor sequence), while GATA3 HI cells had uniformly recombined both Tcrb alleles (one predicting a functional and the other predicting a nonfunctional rearrangement). These data show that GATA3 abundance regulates the recombination propensity at the Tcrb locus and provide new mechanistic insight into the historic immunological conundrum for how Tcrb allelic exclusion is mediated.KEYWORDS allelic exclusion, T cell receptor beta locus, GATA3, monoallelic-tobiallelic switch O ne enduring mystery in cellular immunology regards the underlying mechanisms that control antigen receptor allelic exclusion (1, 2), the process whereby B or T lymphocytes are programmed to express only one functional allele for each chain of their respective antigen receptors (B cell receptor [BCR] or T cell receptor [TCR]), thus avoiding the coexistence of multiple antigen specificities in a single immune cell. Lymphocytes acquire the diversity of antigen recognition (3) as well as a unique monospecificity for particular antigens (4) during development in the bone marrow (B cells) or the thymus (T cells).T lymphocyte development is generally characterized by division into multiple stages based on developmental timing and the location and expression of specific cell surface markers (5). T cell development begins when multipotential hematopoietic progenitor cells in the bone marrow migrate through the bloodstream to the thymus, where early T lineage progenitors (ETPs) are generated and later specified to become T cells (6-10). ETPs differentiate into double-negative (DN) cells (DN2 to DN4 stages) that express neither the CD4 nor the CD8 coreceptor, then into double-positive (DP)

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Long-Range Regulation of V(D)J Recombination

Cited by 66 publications

References 143 publications

Highly sensitive and unbiased approach for elucidating antibody repertoires

Highly sensitive and unbiased approach for elucidating antibody repertoires

Soft X-Ray Tomography Reveals Gradual Chromatin Compaction and Reorganization during Neurogenesis In Vivo

GATA3 Abundance Is a Critical Determinant of T Cell Receptor β Allelic Exclusion

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