Facultative CTCF sites moderate mammary super-enhancer activity and regulate juxtaposed gene in non-mammary cells
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE92932 | GEO | 2017/05/25
SECONDARY ACCESSION(S): PRJNA359102
REPOSITORIES: GEO
Similar Datasets
Project description:Precise spatiotemporal gene regulation is paramount for the establishment and maintenance of cell-specific programmes. Although there is evidence that chromatin neighbourhoods, formed by the zinc-finger protein CTCF, can sequester enhancers and their target genes, there is limited in vivo evidence for CTCF demarcating super-enhancers and preventing cross talk between distinct regulatory elements. Here, we address these questions in the Wap locus with its mammary-specific super-enhancer separated by CTCF sites from widely expressed genes. Mutational analysis demonstrates that the Wap super-enhancer controls Ramp3, despite three separating CTCF sites. Their deletion in mice results in elevated expression of Ramp3 in mammary tissue through augmented promoter-enhancer interactions. Deletion of the distal CTCF-binding site results in loss of Ramp3 expression in non-mammary tissues. This suggests that CTCF sites are porous borders, allowing a super-enhancer to activate a secondary target. Likewise, CTCF sites shield a widely expressed gene from suppressive influences of a silent locus.
| S-EPMC5520053 | biostudies-literature
Project description:Precise spatiotemporal regulation of genetic programs, driven by cellspecific super-enhancers, is paramount for the function of cell lineages. Studies have suggested that insulated neighborhoods, formed by the zincfinger protein CTCF, sequester genes and their associated enhancers thus preventing them from trespassing on off-target genes. Although this could explain the enhancer-gene-specificity conundrum, there is limited genetic evidence that the search space of cell-specific super-enhancers is constrained by CTCF. We have addressed this question in the Wap locus with its exceptional mammary-specific super-enhancer, which is separated by five CTCF sites from neighboring genes. Three of these sites are positioned between the Wap super-enhancer and the widely expressed Ramp3. Enhancer deletions demonstrated that the Wap super-enhancer controls Ramp3 expression despite the presence of three parting CTCF sites. Individual and combinatorial deletions of these CTCF sites revealed cell-specific functions of the conserved anchor site. Although unable to block super-enhancer activity, it muffled its impact on Ramp3 in mammary tissue. Unexpectedly, this CTCF site was obligatory for Ramp3 expression in cerebellum, suggesting the coinciding presence of regulatory elements. While our results suggest a surprisingly limited in vivo role for a CTCF anchor in blocking a mammary-specific super-enhancer, they also implicate this site in cerebellum-specific gene activation. Our study illustrates additional complexities of CTCF sites supporting tissue-specific functions.
2017-05-25 | GSE92931 | GEO
Project description:Precise spatiotemporal regulation of genetic programs, driven by cellspecific super-enhancers, is paramount for the function of cell lineages. Studies have suggested that insulated neighborhoods, formed by the zincfinger protein CTCF, sequester genes and their associated enhancers thus preventing them from trespassing on off-target genes. Although this could explain the enhancer-gene-specificity conundrum, there is limited genetic evidence that the search space of cell-specific super-enhancers is constrained by CTCF. We have addressed this question in the Wap locus with its exceptional mammary-specific super-enhancer, which is separated by five CTCF sites from neighboring genes. Three of these sites are positioned between the Wap super-enhancer and the widely expressed Ramp3. Enhancer deletions demonstrated that the Wap super-enhancer controls Ramp3 expression despite the presence of three parting CTCF sites. Individual and combinatorial deletions of these CTCF sites revealed cell-specific functions of the conserved anchor site. Although unable to block super-enhancer activity, it muffled its impact on Ramp3 in mammary tissue. Unexpectedly, this CTCF site was obligatory for Ramp3 expression in cerebellum, suggesting the coinciding presence of regulatory elements. While our results suggest a surprisingly limited in vivo role for a CTCF anchor in blocking a mammary-specific super-enhancer, they also implicate this site in cerebellum-specific gene activation. Our study illustrates additional complexities of CTCF sites supporting tissue-specific functions.
2017-05-25 | GSE92587 | GEO
Project description:Precise spatiotemporal regulation of genetic programs, driven by cellspecific super-enhancers, is paramount for the function of cell lineages. Studies have suggested that insulated neighborhoods, formed by the zincfinger protein CTCF, sequester genes and their associated enhancers thus preventing them from trespassing on off-target genes. Although this could explain the enhancer-gene-specificity conundrum, there is limited genetic evidence that the search space of cell-specific super-enhancers is constrained by CTCF. We have addressed this question in the Wap locus with its exceptional mammary-specific super-enhancer, which is separated by five CTCF sites from neighboring genes. Three of these sites are positioned between the Wap super-enhancer and the widely expressed Ramp3. Enhancer deletions demonstrated that the Wap super-enhancer controls Ramp3 expression despite the presence of three parting CTCF sites. Individual and combinatorial deletions of these CTCF sites revealed cell-specific functions of the conserved anchor site. Although unable to block super-enhancer activity, it muffled its impact on Ramp3 in mammary tissue. Unexpectedly, this CTCF site was obligatory for Ramp3 expression in cerebellum, suggesting the coinciding presence of regulatory elements. While our results suggest a surprisingly limited in vivo role for a CTCF anchor in blocking a mammary-specific super-enhancer, they also implicate this site in cerebellum-specific gene activation. Our study illustrates additional complexities of CTCF sites supporting tissue-specific functions.
2017-05-25 | GSE97803 | GEO
Project description:Facultative CTCF sites moderate mammary super-enhancer activity and regulate juxtaposed gene in non-mammary cells
| PRJNA359102 | ENA
Project description:Super-enhancers comprise dense transcription factor platforms highly enriched for active chromatin marks. A paucity of functional data led us to investigate the role of super-enhancers in the mammary gland, an organ characterized by exceptional gene regulatory dynamics during pregnancy. ChIP-seq analysis for the master regulator STAT5A, the glucocorticoid receptor, H3K27ac and MED1 identified 440 mammary-specific super-enhancers, half of which were associated with genes activated during pregnancy. We interrogated the Wap super-enhancer, generating mice carrying mutations in STAT5-binding sites within its constituent enhancers. Individually, the most distal site displayed the greatest enhancer activity. However, combinatorial mutation analysis showed that the 1,000-fold induction in gene expression during pregnancy relied on all enhancers. Disabling the binding sites of STAT5, NFIB and ELF5 in the proximal enhancer incapacitated the entire super-enhancer. Altogether, these data suggest a temporal and functional enhancer hierarchy. The identification of mammary-specific super-enhancers and the mechanistic exploration of the Wap locus provide insights into the regulation of cell-type-specific expression of hormone-sensing genes.
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Project description:Chromosomal rearrangements are a frequent cause of oncogene deregulation in human malignancies. Overexpression of EVI1 is found in a subgroup of acute myeloid leukemia (AML) with 3q26 chromosomal rearrangements, which is often therapy resistant. In AMLs harboring a t(3;8)(q26;q24), we observed the translocation of a MYC super-enhancer (MYC SE) to the EVI1 locus. We generated an in vitro model mimicking a patient-based t(3;8)(q26;q24) using CRISPR-Cas9 technology and demonstrated hyperactivation of EVI1 by the hijacked MYC SE. This MYC SE contains multiple enhancer modules, of which only one recruits transcription factors active in early hematopoiesis. This enhancer module is critical for EVI1 overexpression as well as enhancer-promoter interaction. Multiple CTCF binding regions in the MYC SE facilitate this enhancer-promoter interaction, which also involves a CTCF binding site upstream of the EVI1 promoter. We hypothesize that this CTCF site acts as an enhancer-docking site in t(3;8) AML. Genomic analyses of other 3q26-rearranged AML patient cells point to a common mechanism by which EVI1 uses this docking site to hijack enhancers active in early hematopoiesis.
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Project description:We have ablated the cellular RNA degradation machinery in differentiated B cells and pluripotent embryonic stem cells (ESCs) by conditional mutagenesis of core (Exosc3) and nuclear RNase (Exosc10) components of RNA exosome and identified a vast number of long non-coding RNAs (lncRNAs) and enhancer RNAs (eRNAs) with emergent functionality. Unexpectedly, eRNA-expressing regions accumulate R-loop structures upon RNA exosome ablation, thus demonstrating the role of RNA exosome in resolving deleterious DNA/RNA hybrids arising from active enhancers. We have uncovered a distal divergent eRNA-expressing element (lncRNA-CSR) engaged in long-range DNA interactions and regulating IgH 3' regulatory region super-enhancer function. CRISPR-Cas9-mediated ablation of lncRNA-CSR transcription decreases its chromosomal looping-mediated association with the IgH 3' regulatory region super-enhancer and leads to decreased class switch recombination efficiency. We propose that the RNA exosome protects divergently transcribed lncRNA expressing enhancers by resolving deleterious transcription-coupled secondary DNA structures, while also regulating long-range super-enhancer chromosomal interactions important for cellular function.
| S-EPMC4428671 | biostudies-literature
Project description:CTCF and the associated cohesin complex play a central role in insulator function and higher-order chromatin organization of mammalian genomes. Recent studies identified a correlation between the orientation of CTCF-binding sites (CBSs) and chromatin loops. To test the functional significance of this observation, we combined CRISPR/Cas9-based genomic-DNA-fragment editing with chromosome-conformation-capture experiments to show that the location and relative orientations of CBSs determine the specificity of long-range chromatin looping in mammalian genomes, using protocadherin (Pcdh) and ?-globin as model genes. Inversion of CBS elements within the Pcdh enhancer reconfigures the topology of chromatin loops between the distal enhancer and target promoters and alters gene-expression patterns. Thus, although enhancers can function in an orientation-independent manner in reporter assays, in the native chromosome context, the orientation of at least some enhancers carrying CBSs can determine both the architecture of topological chromatin domains and enhancer/promoter specificity. These findings reveal how 3D chromosome architecture can be encoded by linear genome sequences.
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Project description:The mammary luminal lineage relies on the common cytokine-sensing transcription factor STAT5 to establish super-enhancers during pregnancy and initiate a genetic program that activates milk production. As pups grow, the greatly increasing demand for milk requires progressive differentiation of mammary cells with advancing lactation. Here we investigate how persistent hormonal exposure during lactation shapes an evolving enhancer landscape and impacts the biology of mammary cells. Employing ChIP-seq, we uncover a changing transcription factor occupancy at mammary enhancers, suggesting that their activities evolve with advancing differentiation. Using mouse genetics, we demonstrate that the functions of individual enhancers within the Wap super-enhancer evolve as lactation progresses. Most profoundly, a seed enhancer, which is mandatory for the activation of the Wap super-enhancer during pregnancy, is not required during lactation, suggesting compensatory flexibility. Combinatorial deletions of structurally equivalent constituent enhancers demonstrated differentiation-specific compensatory activities during lactation. We also demonstrate that the Wap super-enhancer, which is built on STAT5 and other common transcription factors, retains its exquisite mammary specificity when placed into globally permissive chromatin, suggesting a limited role of chromatin in controlling cell specificity. Our studies unveil a previously unrecognized progressive enhancer landscape where structurally equivalent components serve unique and differentiation-specific functions.
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