CCCTC-binding factor (CTCF) and cohesin influence the genomic architecture of the
            <i>Igh</i>
            locus and antisense transcription in pro-B cells

Degner, Stephanie C.; Verma-Gaur, Jiyoti; Wong, Timothy P.; Bossen, Claudia; Iverson, G M; Torkamani, Ali; Vettermann, Christian; Lin, Yin; Zhang, Ju; Schulz, Danae; Murre, Caroline S.; Birshtein, Barbara K.; Schork, Nicholas J.; Schlissel, Mark S.; Riblet, Roy; Murre, Cornelis; Feeney, Ann J.

doi:10.1073/pnas.1019391108

Cited by 190 publications

(293 citation statements)

References 35 publications

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“…ChIP experiments for H3K4me2, H3K27ac, and P300 were performed with chromatin from RAG-deficient thymocytes (C57BL/6) as described previously (59). The ChIP DNA was purified using a Qiagen DNA purification kit and subjected to whole genome amplification (Sigma), labeled, and hybridized to custom Nimblegen microarrays according to the manufacturer's protocol by Mogene.…”

Section: Methodsmentioning

confidence: 99%

Unifying model for molecular determinants of the preselection Vβ repertoire

Gopalakrishnan

Majumder

Predeus

et al. 2013

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

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The primary antigen receptor repertoire is sculpted by the process of V(D)J recombination, which must strike a balance between diversification and favoring gene segments with specialized functions. The precise determinants of how often gene segments are chosen to complete variable region coding exons remain elusive. We quantified Vβ use in the preselection Tcrb repertoire and report relative contributions of 13 distinct features that may shape their recombination efficiencies, including transcription, chromatin environment, spatial proximity to their DβJβ targets, and predicted quality of recombination signal sequences (RSSs). We show that, in contrast to functional Vβ gene segments, all pseudo-Vβ segments are sequestered in transcriptionally silent chromatin, which effectively suppresses wasteful recombination. Importantly, computational analyses provide a unifying model, revealing a minimum set of five parameters that are predictive of Vβ use, dominated by chromatin modifications associated with transcription, but largely independent of precise spatial proximity to DβJβ clusters. This learned model-building strategy may be useful in predicting the relative contributions of epigenetic, spatial, and RSS features in shaping preselection V repertoires at other antigen receptor loci. Ultimately, such models may also predict how designed or naturally occurring alterations of these loci perturb the preselection use of variable gene segments.lymphocytes | T-cell receptor | gene regulation G ene activity is regulated at multiple levels to coordinate expression during development. At a most basic level, the collection of cis-acting elements for a genetic locus recruits transcription factors that alter its chromatin environment to either induce or repress gene activity. Emerging studies indicate that the 3D conformation of a locus also plays an important role in the regulation of its composite genes (1). At most genes, many levels of control are integrated to achieve the requisite gene expression state. For example, transcriptional promoters interact with their cognate enhancers over considerable distances in the linear genome to generate "hubs" where the two cis elements are in spatial proximity (1, 2).All of these regulatory strategies are used to generate functional Ig (Ig) and T-cell receptor (Tcr) genes during lymphocyte development (3). Each antigen receptor (AgR) locus is composed of multiple variable (V), joining (J), and sometimes diversity (D) gene segments that are assembled by the process of V (D)J recombination, creating a potential variable region exon (4). Recombination is mediated by the RAG-1/2 enzymatic complex, which is expressed in all developing lymphocytes and recognizes semiconserved recombination signal sequences (RSSs) flanking all AgR gene segments (5). On selection of two compatible gene segments by RAG-1/2, recombination proceeds via a DNA break/repair mechanism, ultimately fusing the two selected segments (4, 5).The assembly of AgR genes is strictly regulated despite a common collection of ge...

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Section: Methodsmentioning

confidence: 99%

Unifying model for molecular determinants of the preselection Vβ repertoire

Gopalakrishnan

Majumder

Predeus

et al. 2013

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

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“…Global mapping of CTCF-binding sites and CTCF-associated chromatin loops also suggests that CTCF can facilitate long-distance interactions for coordinated gene expression, demarcate chromatin boundaries, or facilitate enhancer-promoter communication (19-23). However, it remains difficult to predict how CTCF might function at any individual locus.Recently, several studies have suggested a role for CTCF in the regulation of V(D)J recombination at antigen receptor loci (9,(24)(25)(26)(27)(28). For example, knockdown of CTCF in a pro-B-cell line reduced DNA contacts between regulatory elements that flank the D H J H C H region, increased antisense transcription through the D H and V H regions, and partially suppressed contraction of the Igh locus (25).…”

mentioning

confidence: 99%

“…Recently, several studies have suggested a role for CTCF in the regulation of V(D)J recombination at antigen receptor loci (9,(24)(25)(26)(27)(28). For example, knockdown of CTCF in a pro-B-cell line reduced DNA contacts between regulatory elements that flank the D H J H C H region, increased antisense transcription through the D H and V H regions, and partially suppressed contraction of the Igh locus (25).…”

mentioning

confidence: 99%

“…For example, knockdown of CTCF in a pro-B-cell line reduced DNA contacts between regulatory elements that flank the D H J H C H region, increased antisense transcription through the D H and V H regions, and partially suppressed contraction of the Igh locus (25). Deletion of a pair of CTCF-binding sites located between V H and D H segments caused high-frequency rearrangement of the D H -proximal V H gene segments and led to developmentally inappropriate V H gene recombination in B cells and thymocytes (24).…”

mentioning

confidence: 99%

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Tcragene recombination is supported by aTcraenhancer- and CTCF-dependent chromatin hub

Shih

Verma-Gaur

Torkamani

et al. 2012

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

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153

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Antigen receptor locus V(D)J recombination requires interactions between widely separated variable (V), diversity (D), and joining (J) gene segments, but the mechanisms that generate these interactions are not well understood. Here we assessed mechanisms that direct developmental stage-specific long-distance interactions at the Tcra/Tcrd locus. The Tcra/Tcrd locus recombines Tcrd gene segments in CD4 − CD8 − double-negative thymocytes and Tcra gene segments in CD4 + CD8 + double-positive thymocytes. Initial V α -to-J α recombination occurs within a chromosomal domain that displays a contracted conformation in both thymocyte subsets. We used chromosome conformation capture to demonstrate that the Tcra enhancer (E α ) interacts directly with V α and J α gene segments distributed across this domain, specifically in double-positive thymocytes. Moreover, E α promotes interactions between these V α and J α segments that should facilitate their synapsis. We found that the CCCTC-binding factor (CTCF) binds to E α and to many locus promoters, biases E α to interact with these promoters, and is required for efficient V α -J α recombination. Our data indicate that E α and CTCF cooperate to create a developmentally regulated chromatin hub that supports V α -J α synapsis and recombination.T-cell development | T-cell receptor | thymus T and B cells produce diverse antigen receptors through the recombination of variable (V), diversity (D), and joining (J) gene segments at the T-cell receptor (Tcra, Tcrb, Tcrg, and Tcrd) and Ig (Igh, Igκ, and Igλ) loci. This V(D)J recombination is initiated by the lymphoid-specific recombination-activating gene-1 (RAG-1) and RAG-2 proteins, which recognize the recombination signal sequences (RSSs) that flank all V, D, and J gene segments and then cleave the DNA between the RSSs and the adjacent coding gene segments (1). A critical feature of the reaction is the assembly of a synaptic complex composed of two RSSs before the generation of RAG-dependent DNA doublestrand breaks (DSBs). As such, lineage-and developmental stagespecific V(D)J recombination events can be regulated not only by changes in RAG protein expression and RSS accessibility to RAG proteins but also by the ability of those RSSs to undergo synapsis (2).Conformational changes of antigen receptor loci are believed to support V(D)J recombination events because they can bring distant RSSs into proximity and therefore increase the probability of RSS synapsis (2, 3). Studies using 3D-FISH have demonstrated that lineage-and development stage-specific locus contraction marks the recombination windows at antigen receptor loci (3). For example, the 3-Mb Igh locus contracts specifically in pro-B cells to support V H -to-D H J H recombination (4-7). This contracted conformation brings distal and proximal V H segments, which are separated by megabases in the linear DNA sequence, to the vicinity of the D H J H cluster, presumably allowing all V H segments a similar opportunity for recombination (8). In addition, the mapping of Igh locus DNA-DNA...

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“…High-resolution chromatin conformation studies have revealed that cohesin forms long-range interactions between its binding sites (Figure 2) [13-18, [67][68][69][70]. Current models support the idea 6 that cohesin controls gene expression by contributing to the organisation of higher-order chromatin structure.…”

Section: Cohesin Shapes Interphase Chromatin By Forming Long-range Inmentioning

confidence: 95%