Transcription-Dependent Mobilization of Nucleosomes at Accessible TCR Gene Segments In Vivo

Kondilis-Mangum, Hrisavgi D.; Cobb, Robin Milley; Osipovich, Oleg; Srivatsan, Sruti; Oltz, Eugene M.; Krangel, Michael S.

doi:10.4049/jimmunol.0903923

Cited by 30 publications

(23 citation statements)

References 64 publications

(93 reference statements)

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“…In isolation, the consensus nonamer sequence within the RSS can influence nucleosome position of in vitro assemblies (33), but, when longer flanking DNA, with a variety of other sequences, is included, this nonamer effect is found to be relatively weak and easily overcome (20,23). When the nucleosome positioning and occupancy of a few, specific TCR gene segments was compared between recombinationally competent and recombinationally incompetent TCRβ and TCRα alleles in primary thymocytes, no consistent positioning of nucleosomes with respect to the RSS was observed (34). However, because the recombinationally incompetent alleles were missing key enhancers and promoters,…”

Section: Significancementioning

confidence: 98%

Regulated large-scale nucleosome density patterns and precise nucleosome positioning correlate with V(D)J recombination

Pulivarthy

Lion

Kuzu

et al. 2016

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

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We show that the physical distribution of nucleosomes at antigen receptor loci is subject to regulated cell type-specific and lineagespecific positioning and correlates with the accessibility of these gene segments to recombination. At the Ig heavy chain locus (IgH), a nucleosome in pro-B cells is generally positioned over each IgH variable (VH) coding segment, directly adjacent to the recombination signal sequence (RSS), placing the RSS in a position accessible to the recombination activating gene (RAG) recombinase. These changes result in establishment of a specific chromatin organization at the RSS that facilitates accessibility of the genomic DNA for the RAG recombinase. In contrast, in mouse embryonic fibroblasts the coding segment is depleted of nucleosomes, which instead cover the RSS, thereby rendering it inaccessible. Pro-T cells exhibit a pattern intermediate between pro-B cells and mouse embryonic fibroblasts. We also find large-scale variations of nucleosome density over hundreds of kilobases, delineating chromosomal domains within IgH, in a cell typedependent manner. These findings suggest that developmentally regulated changes in nucleosome location and occupancy, in addition to the known chromatin modifications, play a fundamental role in regulating V(D)J recombination. Nucleosome positioning-which has previously been observed to vary locally at individual enhancers and promoters-may be a more general mechanism by which cells can regulate the accessibility of the genome during development, at scales ranging from several hundred base pairs to many kilobases.T he diverse repertoire of antigen receptors in vertebrates is generated by an ordered series of somatic site-specific DNA rearrangement events, collectively termed V(D)J recombination. Antigen receptor genes are assembled from V, D, and J gene segments organized into seven different loci [T-cell receptor (TCR) α, β, γ, and δ, and Ig H, κ, and λ], each of which undergoes a series of recombination reactions to generate a functional TCR or B-cell receptor (BCR) (1, 2) Rearrangement is lineage-specific, such that BCR genes are fully rearranged only in B cells and TCR genes are assembled only in T cells. Rearrangement also occurs in a preferred temporal order. For example, at the human and murine IgH loci, joining of D to J segments precedes V to DJ joining, and IgH joining precedes joining at Igκ (or Igλ) (3). In addition, recombination at several loci shows "allelic exclusion": Functional VDJH or VJκ/Jλ joining occurs only on one allele (4).Recombination is initiated by the lymphocyte-specific recombination activating gene (RAG)1/RAG2 endonuclease. RAG1/2 cleaves at the recombination signal sequences (RSSs) flanking all antigen receptor gene segments. The consensus RSS consists of a heptamer sequence directly adjacent to the coding element and an A/T-rich nonamer separated from the heptamer by a spacer region of conserved length (12 or 23 bp) but relatively nonconserved sequence. The broken ends are then joined with the aid of DNA repair proteins,...

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Section: Significancementioning

confidence: 98%

Regulated large-scale nucleosome density patterns and precise nucleosome positioning correlate with V(D)J recombination

Pulivarthy

Lion

Kuzu

et al. 2016

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

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“…Transcription-based acquisition of H3K4-trimethyl histones leads to recruitment of Rag2 and stimulates the activity of the RAG complex (34). Also, transcription displaces histones from the RSS flanking the V, D, and J segments and provides accessibility to the RAG complex (42,43). These observations seem to provide the link between transcription and recombination.…”

Section: Pdβ1-djcβ1mentioning

confidence: 99%

Influence of a CTCF-Dependent Insulator on Multiple Aspects of Enhancer-Mediated Chromatin Organization

Varma

Rawat

Jalan

et al. 2015

Molecular and Cellular Biology

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Developmental stage-specific enhancer-promoter-insulator interactions regulate the chromatin configuration necessary for transcription at various loci and additionally for VDJ recombination at antigen receptor loci that encode immunoglobulins and T-cell receptors. To investigate these regulatory interactions, we analyzed the epigenetic landscape of the murine T-cell receptor ␤ (TCR␤) locus in the presence and absence of an ectopic CTCF-dependent enhancer-blocking insulator, H19-ICR, in genetically manipulated mice. Our analysis demonstrated the ability of the H19-ICR insulator to restrict several aspects of enhancerbased chromatin alterations that are observed during activation of the TCR␤ locus for transcription and recombination. The H19-ICR insulator abrogated enhancer-promoter contact-dependent chromatin alterations and additionally prevented E␤-mediated histone modifications that have been suggested to be independent of enhancer-promoter interaction. Observed enhancerpromoter-insulator interactions, in conjunction with the chromatin structure of the E␤-regulated domain at the nucleosomal level, provide useful insights regarding the activity of the regulatory elements in addition to supporting the accessibility hypothesis of VDJ recombination. Analysis of H19-ICR in the heterologous context of the developmentally regulated TCR␤ locus suggests that different mechanisms proposed for CTCF-dependent insulator action might be manifested simultaneously or selectively depending on the genomic context and the nature of enhancer activity being curtailed.T ranscriptional insulators regulate the enhancer-promoter communication that orchestrates the epigenetic landscape of specific loci to activate or repress genes in metazoan genomes. Enhancers can regulate their cognate promoters by diverse mechanisms (1, 2). These may involve direct contact with the promoter by looping and/or alteration of the epigenetic landscape of large domains that render them "open," i.e., associated with chromatin modifications that make them accessible to trans-acting factors. Alteration of the large domains could be due to "tracking" of some proteins initially bound at the enhancer or, as proposed during "facilitated tracking," the movement of enhancer itself along the chromatin. Insulators in the vertebrate genomes have been proposed to modulate these interactions by binding to CTCF, a multiZn-finger protein (3). Genome-wide analysis of CTCF demonstrates its crucial role in higher-order chromatin organization that can facilitate enhancer-promoter interactions as well as curtail them depending on the relative positions of these regulatory elements (4). Importantly, insulators block enhancer-promoter communication only when present between them. Several models have been proposed for enhancer blocking, keeping in view the varied mechanisms underlying enhancer activity (5). It has been postulated that the CTCF-bound insulator partitions the enhancer and promoter to topologically distinct chromatin loops and thus prevents their interaction. The i...

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“…Conversely, 5'PDβ2 repression may account for nucleosome occlusion of the Dβ2 23-RSS (9). We now show that this repression of 5'PDβ2 activity is mediated by binding of USF-1 to a non-canonical E box within the Dβ2 12-RSS spacer sequence, and that DSBs can induce a DNA-PKcs-dependent loss of USF-1 that relieves repression.…”

Section: Discussionmentioning

confidence: 99%

“…Activation of promoters associated with D or J segments (so-called germline transcription, reflecting the unrearranged nature of the transcribed template) augments the accessibility of transcribed segments to recombinase. Chromatin remodeling (8), nucleosome repositioning (9) and transcriptional elongation associated with germline promoter activation (10) facilitate the recombinational accessibility of individual gene segments. However, the mechanism by which promoter-mediated accessibility is modulated during lymphocyte development is unclear.…”

Section: Introductionmentioning

confidence: 99%

“…Though Dβ RSS sequence strongly influences the order of gene segment assembly (5, 13, 14), the recombinational accessibility of individual RSSs is dependent on their chromosomal location (15) and the activity of associated germline promoters. Deletion of the Dβ1-associated promoter, PDβ1, alters nucleosomal phasing across the Dβ1 5'RSS (9) and specifically impairs Tcrb Dβ1-to-Jβ recombination (16, 17) without affecting recombination at the downstream DJβ2 gene segment cluster (17). Though the mechanism of PDβ1's influence over DJβ1 assembly is unclear, the promoter's position immediately upstream of Dβ1(18), and its recruitment of SWI/SNF chromatin remodeling complexes are critical for efficient DJβ1 assembly (8).…”

Section: Introductionmentioning

confidence: 99%

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DNA Double-Strand Breaks Relieve USF-Mediated Repression of Dβ2 Germline Transcription in Developing Thymocytes

Stone

McMillan

Skaar

et al. 2012

The Journal of Immunology

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Activation of germline promoters is central to V(D)J recombinational accessibility, driving chromatin remodeling, nucleosome repositioning and transcriptional readthrough of associated DNA. We have previously shown that of the 2 Tcrb D segments, Dβ1 is flanked by an upstream promoter that directs its transcription and recombinational accessibility. In contrast, transcription within the DJβ2 segment cluster is initially restricted to the J segments and only redirected upstream of Dβ2 after D-to-J joining. The repression of upstream promoter activity prior to Tcrb assembly correlates with evidence that suggests DJβ2 recombination is less efficient than that of DJβ1. Since inefficient DJβ2 assembly offers the potential for V-to-DJβ2 recombination to rescue frameshifted V-to-DJβ1 joints, we wished to determine how Dβ2 promoter activity is modulated upon Tcrb recombination. Here, we show that repression of the otherwise transcriptionally primed 5'Dβ2 promoter (5'PDβ2) requires binding of USF-1 to a non-canonical E-box within the Dβ2 12-RSS spacer prior to Tcrb recombination. USF binding is lost from both rearranged and germline Dβ2 sites in DNA-PKcs-competent thymocytes. Finally, genotoxic double-stranded DNA breaks lead to rapid loss of USF binding and gain of 5'PDβ2 activity in a DNA-PKcs-dependent manner. Together, these data suggest a mechanism by which V(D)J recombination may feedback to regulate local Dβ2 recombinational accessibility during thymocyte development.

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Transcription-Dependent Mobilization of Nucleosomes at Accessible TCR Gene Segments In Vivo

Abstract: Material

Cited by 30 publications

References 64 publications

Regulated large-scale nucleosome density patterns and precise nucleosome positioning correlate with V(D)J recombination

Regulated large-scale nucleosome density patterns and precise nucleosome positioning correlate with V(D)J recombination

Influence of a CTCF-Dependent Insulator on Multiple Aspects of Enhancer-Mediated Chromatin Organization

DNA Double-Strand Breaks Relieve USF-Mediated Repression of Dβ2 Germline Transcription in Developing Thymocytes

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