Project description:Polycomb group (PcG) proteins maintain the silenced state of key developmental genes in animals, but how these proteins are recruited to specific regions of the genome is still poorly understood. In Drosophila, PcG proteins are recruited to Polycomb response elements (PREs) that include combinations of sites for sequence specific DNA binding “PcG recruiters,” including Pho, Cg, and Spps. To understand their roles in PcG recruitment, we compared Pho-, Cg-, and Spps-binding sites against H3K27me3 and key PcG proteins by ChIP-seq in wild-type and mutant third instar larvae. H3K27me3 in canonical Polycomb domains is decreased after the reduction of any recruiter. Reduction of Spps and Pho, but not Cg, causes the redistribution of H3K27me3 to heterochromatin. Regions with dramatically depleted H3K27me3 after Spps knockout are usually accompanied by decreased Pho binding, suggesting their cooperative binding. PcG recruiters, the PRC2 component E(z), and the PRC1 components Psc and Ph cobind thousands of active genes outside of H3K27me3 domains. This study demonstrates the importance of distinct PcG recruiters for the establishment of unique Polycomb domains. Different PcG recruiters can act both cooperatively and independently at specific PcG target genes, highlighting the complexity and diversity of PcG recruitment mechanisms.
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.
Project description:Genome-wide binding profile of PcG proteins Pho and Ph, H3K27me3 and Engrailed in Drosophila third instar larval brains and disks, in duplicate, using Illumina HiSeq 2500. The sequence reads that passed quality filters were analyzed. Drosophila third instar larval brains and disks were dissected and fixed with 2% HCHO before sonication. After sonication, chromatin immunoprecipitation using anti-Pho, -Ph, -H3K27me3, and -Engrailed antibodies was carried out.
Project description:Genome-wide binding profile of PcG proteins Pho and Ph, H3K27me3 and Engrailed in Drosophila third instar larval brains and disks, in duplicate, using Illumina HiSeq 2500. The sequence reads that passed quality filters were analyzed.
Project description:Genome-wide binding profile of PcG proteins- Pho and Ph, H3K27me3 and Engrailed in Drosophila third instar larval brains and disks, in duplicate, using Illumina HiSeq 2500. The sequence reads that passed quality filters were analyzed.
Project description:Drosophila Polycomb group (PcG) and Trithorax group (TrxG) proteins are responsible for the maintenance of stable transcription patterns of many developmental regulators, such as the homeotic genes. We have used a ChIP-on- Chip approach to map the distribution of several PcG/TrxG proteins as well as histone modifications across the two homeotic complexes ANT-C and BX-C. Our data indicate the colocalization of the Polycomb repressive complex 1 (PRC1) with Trx and the DNA binding protein Pleiohomeotic (Pho) at discrete sequence elements as well as significant chromatin assembly differences in active and inactive regions. Trx binds to the promoters of active genes and noncoding transcripts. Most strikingly, in the active state Pho covers extended chromatin domains over many kilobases. The histone modifications investigated, seem to be mutually exclusive, with histone H3 trimethylated at lysine 27 covering inactive chromatin domains whereas hyperacetylated histone H4 is present only at active genes. Keywords: ChIP-chip, cell line comparison
Project description:To understand the combinatorial binding of different PcG proteins and their binding dynamics across cell types, we performed multi-omics profiling in three Drosophila cell lines, including S2, Kc167 and/or D17-c3 cells. The profiled data included: 1) RNA-seq; 2) ATAC-seq; 3) ChIP-seq of multiple PcG proteins and other related proteins, including E(z), GAGA, Pc, Pcl, Ph, Psc, Scm, Sfmbt, Pho, Spps; 4) Multiple active and repressive histone modifications, including H3K27ac, H3K27me3, H3Krme2, H3K4me3, H3K36me2, H3K36me3.
Project description:To understand the combinatorial binding of different PcG proteins and their binding dynamics across cell types, we performed multi-omics profiling in three Drosophila cell lines, including S2, Kc167 and/or D17-c3 cells. The profiled data included: 1) RNA-seq; 2) ATAC-seq; 3) ChIP-seq of multiple PcG proteins and other related proteins, including E(z), GAGA, Pc, Pcl, Ph, Psc, Scm, Sfmbt, Pho, Spps; 4) Multiple active and repressive histone modifications, including H3K27ac, H3K27me3, H3Krme2, H3K4me3, H3K36me2, H3K36me3.
Project description:To understand the combinatorial binding of different PcG proteins and their binding dynamics across cell types, we performed multi-omics profiling in three Drosophila cell lines, including S2, Kc167 and/or D17-c3 cells. The profiled data included: 1) RNA-seq; 2) ATAC-seq; 3) ChIP-seq of multiple PcG proteins and other related proteins, including E(z), GAGA, Pc, Pcl, Ph, Psc, Scm, Sfmbt, Pho, Spps; 4) Multiple active and repressive histone modifications, including H3K27ac, H3K27me3, H3Krme2, H3K4me3, H3K36me2, H3K36me3.
Project description:To understand the combinatorial binding of different PcG proteins and their binding dynamics across cell types, we performed multi-omics profiling in three Drosophila cell lines, including S2, Kc167 and/or D17-c3 cells. The profiled data included: 1) RNA-seq; 2) ATAC-seq; 3) ChIP-seq of multiple PcG proteins and other related proteins, including E(z), GAGA, Pc, Pcl, Ph, Psc, Scm, Sfmbt, Pho, Spps; 4) Multiple active and repressive histone modifications, including H3K27ac, H3K27me3, H3Krme2, H3K4me3, H3K36me2, H3K36me3; 5) Micro-C data.
