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A Region of Human HoxD that Confers Polycomb-Group Responsiveness

ABSTRACT: 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.

ORGANISM(S): Homo sapiens

PROVIDER: GSE19046 | GEO | 2009/11/19

SECONDARY ACCESSION(S): PRJNA120539

REPOSITORIES: GEO

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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

Context dependent role of Pho binding sites in Polycomb complex recruitment in Drosophila

Project description:Polycomb group (PcG) proteins maintain the silenced state of key developmental genes, but how these proteins are recruited to specific regions of the genome is still not completely understood. In Drosophila, PcG proteins are recruited to Polycomb response elements (PREs) comprised of a flexible array of sites for sequence-specific DNA binding proteins, “PcG recruiters”, including Pho, Spps, Cg, GAF and many others. Pho is thought to play a central role in PcG recruitment. Early data showed that mutation of Pho binding sites in PREs in transgenes abrogated the ability of those PREs to repress gene expression. In contrast, genome-wide experiments in pho mutants or by Pho knockdown showed that PcG proteins can bind to PREs in the absence of Pho. Here we directly addressed the importance of Pho binding sites in two engrailed (en) PREs at the endogenous locus and in transgenes. Our results show that Pho binding sites are required for PRE activity in transgenes with a single PRE. In a transgene, two PREs together lead to stronger, more stable repression and confer some resistance to the loss of Pho binding sites. Making the same mutation in Pho binding sites has little effect on PcG-protein binding at the endogenous en gene. Overall, our data support the model that Pho is important for PcG binding but emphasize how multiple PREs and chromatin environment increase the ability of PREs to function in the absence of Pho.

2023-05-08 | GSE227553 | GEO

Identification of new regulators of the three dimensional Polycomb organization by a microscopy-based genome-wide RNAi screen

Project description:Polycomb group (PcG) proteins dynamically define cellular identities through epigenetic repression of key developmental genes. PcG target gene repression can be stabilized through the interaction in the nucleus at PcG foci. Here, we report the results of a high-resolution microscopy genome-wide RNAi screen that identifies 129 genes that regulate the nuclear organization of Pc foci. Candidate genes include PcG components and chromatin factors, as well as many novel protein-modifying enzymes, including components of the SUMOylation pathway. In the absence of SUMO Pc foci coagulate into larger aggregates. Conversely, loss of function of the SUMO peptidase velo disperses Pc foci. Moreover, SUMO and velo colocalize with PcG proteins at PREs and Pc SUMOylation affects its chromatin targeting, suggesting that the dynamic regulation of Pc SUMOylation regulates PcG-mediated silencing by modulating the kinetics of Pc binding to chromatin as well as its ability to form Polycomb foci. ChIP-Seq mapping of Polycomb (PC), SUMO and Velo on Drosophila Melanogaster

2014-04-03 | E-GEOD-55303 | biostudies-arrayexpress

p300/CBP sustains Polycomb silencing by non-enzymatic functions (ChIP-seq on inhibitor-treated mouse ES cells)

Project description:Maintenance of appropriate cell states involves epigenetic mechanisms, including Polycomb group (PcG)-mediated transcriptional repression. While PcG proteins are known to induce chromatin compaction, how PcG proteins gain access to DNA in compact chromatin to achieve long-term silencing is poorly understood. Here we show that the p300/CBP co-activator is associated with two-thirds of PcG-regions and required for PcG occupancy at many of these in Drosophila and mouse cells. CBP stabilizes RNA polymerase II (Pol II) at PcG-bound repressive sites and promotes Pol II pausing independently of its histone acetyltransferase activity. CBP and Pol II pausing are necessary for RNA-DNA hybrid (R-loop) formation and nucleosome depletion at Polycomb Response Elements (PREs), whereas transcription beyond the pause region is not. These results suggest that non-enzymatic activities of the CBP co-activator have been repurposed to support PcG-mediated silencing, revealing how chromatin regulator interplay maintains transcriptional states.

2022-10-06 | GSE186722 | GEO

p300/CBP sustains Polycomb silencing by non-enzymatic functions (ChIP-seq on RNAi-treated Drosophila S2 cells)

2022-10-06 | GSE186721 | GEO

p300/CBP sustains Polycomb silencing by non-enzymatic functions (ATAC-seq on inhibitor-treated Drosophila S2 cells)

2022-10-06 | GSE186720 | GEO

p300/CBP sustains Polycomb silencing by non-enzymatic functions (ChIP-seq on inhibitor-treated Drosophila S2 cells)

2022-10-06 | GSE173473 | GEO

p300/CBP sustains Polycomb silencing by non-enzymatic functions (RNA-seq on inhibitor-treated Drosophila S2 cells)

2022-10-06 | GSE205627 | GEO

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

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

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