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High-resolution mapping defines the cooperative architecture of Polycomb Response Elements

ABSTRACT: Polycomb-mediated chromatin repression modulates gene expression during development in metazoans. Binding of multiple sequence-specific factors at discrete Polycomb Response Elements (PREs) is thought to recruit repressive complexes that spread across an extended chromatin domain. To dissect the structure of PREs, we applied high-resolution mapping of non-histone chromatin proteins in native chromatin of Drosophila cells. Analysis of occupied sites reveal cooperative interactions between transcription factors that stabilize Polycomb anchoring to DNA, and implicate the general transcription factor Adf1 as a novel PRE component. By comparing two Drosophila cell lines with differential chromatin states, we provide evidence that repression is accomplished at multiple steps in transcription, including inactivation of distant enhancers, enhanced Polycomb recruitment to PREs and target promoters, and elevated stalling of RNAPII in repressed genes. These results suggest that the stability of complexes bridging promoters, enhancers, and PREs is a crucial aspect of developmentally regulated gene expression.

ORGANISM(S): Drosophila melanogaster

PROVIDER: GSE47829 | GEO | 2014/02/08

SECONDARY ACCESSION(S): PRJNA208052

REPOSITORIES: GEO

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High-resolution mapping defines the cooperative architecture of Polycomb Response Elements

Project description:Polycomb-mediated chromatin repression modulates gene expression during development in metazoans. Binding of multiple sequence-specific factors at discrete Polycomb Response Elements (PREs) is thought to recruit repressive complexes that spread across an extended chromatin domain. To dissect the structure of PREs, we applied high-resolution mapping of non-histone chromatin proteins in native chromatin of Drosophila cells. Analysis of occupied sites reveal cooperative interactions between transcription factors that stabilize Polycomb anchoring to DNA, and implicate the general transcription factor Adf1 as a novel PRE component. By comparing two Drosophila cell lines with differential chromatin states, we provide evidence that repression is accomplished at multiple steps in transcription, including inactivation of distant enhancers, enhanced Polycomb recruitment to PREs and target promoters, and elevated stalling of RNAPII in repressed genes. These results suggest that the stability of complexes bridging promoters, enhancers, and PREs is a crucial aspect of developmentally regulated gene expression. Native chromatin immunoprecipitation of histones, transcription factors and Polycomb protein in Drosophila cell lines.

2014-02-08 | E-GEOD-47829 | biostudies-arrayexpress

An epigenetic mark of polycomb response elements implemented by Trx/MLL/COMPASS

Project description:In Drosophila, Polycomb Response Elements (PREs) are identified as genomic sequences allowing the maintenance of transcriptional repression in the absence of the initiating signal. Although PREs in Drosophila are well characterized, the existence of mammalian PRE-like elements remains debated. Accumulating evidence supports a model in which CpG islands function to recruit Polycomb-Group complexes (PcG), however, it is not evident which subclasses of CpG islands serve as PREs. Trithorax (Trx), which is required for positive regulation of gene expression in Drosophila, is known to co-bind Drosophila PREs where it is thought to antagonize polycomb-dependent silencing of nearby genes. Here, we demonstrate the existence of Trx-dependent H3K4 dimethylation loci that specifically mark Drosophila PREs and are required for the maintenance of expression of the nearby genes. Similarly, in human cells, we find ~ 3000 MLL1 (human Trx homologue)-dependent H3K4 dimethylation loci, which correlate strongly with CpG island density. In the absence of MLL1 and H3K4 dimethylation at these loci, there is an increase in H3K27 trimethylation levels, suggesting these sites can recruit Polycomb Repressive Complex 2 (PRC2). By inhibiting PRC2-dependent silencing in the absence of MLL1, we establish that a balance exists between MLL1 and PRC2, and their respective capacity to maintain or repress transcription. Thus, by investigating a conserved function between Trx and MLL1, we provide rules for the identification of CpG island subclasses serving as PRE-like sequences within the human genome. To examine changes in histone-modification profiles and gene expression after depletion of Trx in Drosophila S2 cells, and MLL1 in human HCT116 cells. We also treated MLL1-NULL HCT116 cells with GSK126 (5uM) for 4 days and measured changes in gene expression.

2016-07-26 | E-GEOD-81795 | biostudies-arrayexpress

An epigenetic mark of polycomb response elements implemented by Trx/MLL/COMPASS

2016-07-26 | GSE81795 | GEO

RNAi-independent role for Argonaute2 in CTCF/CP190 chromatin insulator function

Project description:Chromatin insulators and Polycomb group (PcG) complexes control nuclear organization to effect changes in gene expression. In Drosophila, RNA silencing pathways influence long range interactions mediated by PcG proteins and nuclear localization of the gypsy insulator; however, the underlying mechanisms are unknown. Here, we identify a singular requirement for Argonaute2 (AGO2) for the activity of the CCCTC-binding factor (CTCF)/Centrosomal protein 190 (CP190) dependent Fab-8 insulator. AGO2 and CP190 interact physically, and genome wide localization of AGO2 by chromatin immunoprecipitation and sequencing (ChIP-seq) reveals extensive colocalization of AGO2 with insulators and Polycomb Response Elements (PREs) but minimal overlap with regions of endogenous small interfering RNA (endo-siRNA) production. Finally, depletion of either CTCF or CP190 results in loss of AGO2 association with insulators, PREs, and other cis-regulatory regions. Our findings suggest that Dicer-independent recruitment of AGO2 to chromatin by insulator proteins promotes the definition of transcriptional domains throughout the genome. ChIP-seq of AGO2 in two Drosophila cell types (S2 and S3)

2011-08-15 | E-GEOD-22623 | biostudies-arrayexpress

A Region of Human HoxD that Confers Polycomb-Group Responsiveness

Project description:Polycomb group (PcG) proteins are essential for accurate axial body patterning during embryonic development. PcG-mediated repression is conserved in metazoans and is targeted in Drosophila by Polycomb response elements (PREs). Targeting sequences in humans have not been described. While analyzing chromatin architecture in the context of human embryonic stem cell (hESC) differentiation, we discovered a 1.8kb region between HOXD11 and HOXD12 (D11.12) that is associated with PcG proteins, is nuclease hypersensitive, and shows alteration as hESCs differentiate. D11.12 repressed luciferase expression from a reporter construct both before and after differentiation of mesenchymal stem cells into adipocytes. Full repression by D11.12 required a highly conserved region and YY1 binding sites. Repression relied upon PcG proteins Bmi1 and Eed and a YY1-interacting partner, RYBP. We conclude that D11.12 is a Polycomb-dependent regulatory region with similarities to Drosophila PREs, indicating conservation in the mechanisms that target PcG function in mammals and flies. Data contains microarray measurements of the MNase-digested mononucleosomal fragments in hES cells, derived MSCs, osteoblasts and adipocytes. The ChIP-chip data includes Suz12, Bmi1 and H3K27me3 measurements in MSCs and adipocytes.

2010-06-19 | E-GEOD-19046 | biostudies-arrayexpress

A Region of Human HoxD that Confers Polycomb-Group Responsiveness

2009-11-19 | GSE19046 | GEO

A PRE loop at the dac locus acts as a topological chromatin structure that restricts and specifies enhancer promoter communication

Project description:3D genome folding plays a fundamental role for the regulation of developmental genes by facilitating or constraining chromatin interactions between cis-regulatory elements (CREs). Polycomb response elements (PREs) are a specific kind of CREs involved in the memory of transcriptional states in Drosophila melanogaster. PREs act as nucleation sites for Polycomb group (PcG) proteins, which deposit the repressive histone mark H3K27me3, leading to the formation of a class of topologically associating domain (TADs) called Polycomb domains. PREs can establish looping contacts that stabilize gene repression of key developmental genes during development. However, the mechanism by which PRE loops fine tune gene expression is unknown. Using CRISPR/Cas9 genome engineering, we specifically perturbed PRE contacts or enhancer function and used complementary approaches including 4C-seq, Hi-C, and Hi-M to analyze how chromatin architecture perturbation affects gene expression. Our results suggest that the PRE loop at the dac gene locus acts as a constitutive 3D chromatin scaffold during Drosophila development that forms independently of gene expression states and has a versatile function: it restricts enhancer promoter communication and contributes to enhancer specificity.

2024-06-13 | GSE247377 | GEO

DNA elements tether canonical Polycomb Repressive Complex 1 to human genes

Project description:Development of multicellular animals requires epigenetic repression by Polycomb group proteins. The latter assemble in multi-subunit complexes, of which two kinds, Polycomb Repressive Complex 1 (PRC1) and Polycomb Repressive Complex 2 (PRC2), act together to effect the repression of key developmental genes. How PRC1 and PRC2 recognize specific genes remains an open question. Here we report that small adjustment in the next generation sequencing of the Immunoprecipitated Chromatin (ChIP-seq) allows identification of several hundreds of DNA elements that tether canonical PRC1 to human developmental genes. Their analysis indicates that sequence features associated with canonical PRC1 tethering differ from those that favour the retention of PRC2. Throughout the genome, the two kinds of sequence features mix in different proportions to yield a gamut of DNA elements that range from those tethering predominantly PRC1 to ones capable of tethering both PRC1 and PRC2 or the elements that retain exclusively PRC2. The emerging picture is similar to paradigmatic targeting of Polycomb complexes by Polycomb Response Elements (PREs) of Drosophila but providing for unexpected degree of plasticity.

2023-01-13 | GSE207401 | GEO

Distinct Polycomb Response Elements control chromatin state and gene activation of the vestigial gene

Project description:Patterned expression of many developmental genes is thought to rely on chromatin-mediated repression. Regulatory DNA sequences called Polycomb Response Elements (PREs) play critical roles in repression. While PREs in transgenes can nucleate trimethylation on lysine 27 of the histone H3 tail (H3K27me3), none have been demonstrated to be necessary at endogenous chromatin domains. This is thought to be due to the fact that most endogenous H3K27me3 domains contain many PREs, and individual PREs may be redundant. We show here that PREs near the wing selector gene vestigial have distinctive roles at their endogenous locus, even though both PREs are repressors in transgenes. First, a PRE near the promoter is required for vestigial activation and not for repression. Second, only the distal PRE contributes to H3K27me3, but even removal of both PREs does not eliminate H3K27me3 across the vestigial domain. Thus, endogenous chromatin domains appear to be intrinsically marked by H3K27me3, and PREs can enhance this chromatin modification at inactive genes.

2019-05-01 | GSE121028 | GEO

Comparative analysis of PRE DNA-binding factors interactomes: purification of Combgap, Zeste, Psq and Adf1 associated proteins

Project description:The Polycomb group (PcG) and Trithorax group (TrxG) proteins are key epigenetic regulators controlling silenced and active states of genes in multicellular organisms, respectively. In Drosophila PcG/TrxG proteins are recruited to chromatin via binding to specialized DNA-elements termed Polycomb Response Elements (PREs). While precise mechanisms of PcG/TrxG proteins recruitment remains unknown, the important role is suggested to belong to sequence specific DNA-binding factors. Here, using affinity purification coupled with high throughput mass spectrometry (IP/MS), we isolated factors associated with Combgap, Psq, Zeste and Adf1 PRE DNA-binding Drosophila proteins. As a control we used unspecific IgG. For affinity purification the nuclear extract from Drosophila S2 cells and polyclonal antibodies against Combgap, Psq, Zeste and Adf1 were used.

2024-05-22 | PXD029459 | Pride

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