Polycomb Repressive Complex 1 Generates Discrete Compacted Domains that Change during Differentiation.

Master regulatory genes require stable silencing by the polycomb group (PcG) to prevent misexpression during differentiation and development. Some PcG proteins covalently modify histones, which contributes to heritable repression. The role for other effects on chromatin structure is less understood. We characterized the organization of PcG target genes in ESCs and neural progenitors using 5C and super-resolution microscopy. The genomic loci of repressed PcG targets formed discrete, small (20-140 Kb) domains of tight interaction that corresponded to locations bound by canonical polycomb repressive complex 1 (PRC1). These domains changed during differentiation as PRC1 binding changed. Their formation depended upon the Polyhomeotic component of canonical PRC1 and occurred independently of PRC1-catalyzed ubiquitylation. PRC1 domains differ from topologically associating domains in size and boundary characteristics. These domains have the potential to play a key role in transmitting epigenetic silencing of PcG targets by linking PRC1 to formation of a repressive higher-order structure.

[1]  M. Vidal,et al.  Role of histone H2A ubiquitination in Polycomb silencing , 2004, Nature.

[2]  D. Reinberg,et al.  Epigenetic inheritance: histone bookmarks across generations. , 2014, Trends in cell biology.

[3]  L. Mirny,et al.  Iterative Correction of Hi-C Data Reveals Hallmarks of Chromosome Organization , 2012, Nature Methods.

[4]  Jing Liang,et al.  Chromatin architecture reorganization during stem cell differentiation , 2015, Nature.

[5]  Guo-Cheng Yuan,et al.  EZH1 mediates methylation on histone H3 lysine 27 and complements EZH2 in maintaining stem cell identity and executing pluripotency. , 2008, Molecular cell.

[6]  J. Rougemont,et al.  The Dynamic Architecture of Hox Gene Clusters , 2011, Science.

[7]  James A. Cuff,et al.  A Bivalent Chromatin Structure Marks Key Developmental Genes in Embryonic Stem Cells , 2006, Cell.

[8]  Jean-Marie Rouillard,et al.  Versatile design and synthesis platform for visualizing genomes with Oligopaint FISH probes , 2012, Proceedings of the National Academy of Sciences.

[9]  G. Schroth,et al.  Cohesin-mediated interactions organize chromosomal domain architecture , 2013, The EMBO journal.

[10]  P. Park,et al.  Design and analysis of ChIP-seq experiments for DNA-binding proteins , 2008, Nature Biotechnology.

[11]  Peng Yin,et al.  Single-molecule super-resolution imaging of chromosomes and in situ haplotype visualization using Oligopaint FISH probes , 2015, Nature Communications.

[12]  M. Tolstorukov,et al.  Multiplexed Illumina sequencing libraries from picogram quantities of DNA , 2013, BMC Genomics.

[13]  Robert S Illingworth,et al.  The E3 ubiquitin ligase activity of RING1B is not essential for early mouse development , 2015, Genes & development.

[14]  N. Brockdorff,et al.  Variant PRC1 Complex-Dependent H2A Ubiquitylation Drives PRC2 Recruitment and Polycomb Domain Formation , 2014, Cell.

[15]  T. Jessell,et al.  Pax6 Controls Progenitor Cell Identity and Neuronal Fate in Response to Graded Shh Signaling , 1997, Cell.

[16]  Job Dekker,et al.  Determining spatial chromatin organization of large genomic regions using 5C technology. , 2009, Methods in molecular biology.

[17]  Jesse R. Dixon,et al.  Topological Domains in Mammalian Genomes Identified by Analysis of Chromatin Interactions , 2012, Nature.

[18]  Wendy A Bickmore,et al.  Ring1B compacts chromatin structure and represses gene expression independent of histone ubiquitination. , 2010, Molecular cell.

[19]  D. Duboule,et al.  A Switch Between Topological Domains Underlies HoxD Genes Collinearity in Mouse Limbs , 2013, Science.

[20]  J. Rubenstein,et al.  Homeobox gene Nkx2.2 and specification of neuronal identity by graded Sonic hedgehog signalling , 1999, Nature.

[21]  Louise S. Matheson,et al.  Polycomb repressive complex PRC1 spatially constrains the mouse embryonic stem cell genome , 2015, Nature Genetics.

[22]  C. Nusbaum,et al.  Chromosome Conformation Capture Carbon Copy (5C): a massively parallel solution for mapping interactions between genomic elements. , 2006, Genome research.

[23]  L. Cozzuto,et al.  RYBP and Cbx7 define specific biological functions of polycomb complexes in mouse embryonic stem cells. , 2013, Cell reports.

[24]  J. Dekker,et al.  The long-range interaction landscape of gene promoters , 2012, Nature.

[25]  Howard Y. Chang,et al.  A long noncoding RNA maintains active chromatin to coordinate homeotic gene expression , 2011, Nature.

[26]  Leonid A. Mirny,et al.  Super-resolution imaging reveals distinct chromatin folding for different epigenetic states , 2015, Nature.

[27]  B. Bernstein,et al.  SAM domain polymerization links subnuclear clustering of PRC1 to gene silencing. , 2013, Developmental cell.

[28]  S. Pollard,et al.  Neural stem cells, neurons, and glia. , 2006, Methods in enzymology.

[29]  C. Ponting,et al.  KDM2B links the Polycomb Repressive Complex 1 (PRC1) to recognition of CpG islands , 2012, eLife.

[30]  B. Garcia,et al.  Inhibition of PRC2 Activity by a Gain-of-Function H3 Mutation Found in Pediatric Glioblastoma , 2013, Science.

[31]  Patrick Schorderet,et al.  Chromatin topology is coupled to Polycomb group protein subnuclear organization , 2016, Nature Communications.

[32]  C. Peterson,et al.  Chromatin immunoprecipitation (ChIP). , 2009, Cold Spring Harbor protocols.

[33]  Hengbin Wang,et al.  Role of Histone H3 Lysine 27 Methylation in Polycomb-Group Silencing , 2002, Science.

[34]  Oliver Weichenrieder,et al.  Structure and E3‐ligase activity of the Ring–Ring complex of Polycomb proteins Bmi1 and Ring1b , 2006, The EMBO journal.

[35]  D. Reinberg,et al.  Histone methyltransferase activity associated with a human multiprotein complex containing the Enhancer of Zeste protein. , 2002, Genes & development.

[36]  L. Ringrose,et al.  What are memories made of? How Polycomb and Trithorax proteins mediate epigenetic memory , 2014, Nature Reviews Molecular Cell Biology.

[37]  R. Kingston,et al.  Compaction of chromatin by diverse Polycomb group proteins requires localized regions of high charge. , 2011, Genes & development.

[38]  Cole Trapnell,et al.  Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation. , 2010, Nature biotechnology.

[39]  David K. Gifford,et al.  Saltatory remodeling of Hox chromatin in response to rostro-caudal patterning signals , 2013, Nature Neuroscience.

[40]  Robert E. Kingston,et al.  Occupying chromatin: Polycomb mechanisms for getting to genomic targets, stopping transcriptional traffic, and staying put. , 2013, Molecular cell.

[41]  Lior Pachter,et al.  Sequence Analysis , 2020, Definitions.

[42]  Edith Heard,et al.  MicroRNA Regulation of Cbx7 Mediates a Switch of Polycomb Orthologs during ESC Differentiation , 2012, Cell stem cell.

[43]  D. Duboule,et al.  Topological Domains, Metagenes, and the Emergence of Pleiotropic Regulations at Hox Loci. , 2016, Current topics in developmental biology.

[44]  D. Duboule,et al.  Structure, function and evolution of topologically associating domains (TADs) atHOX loci , 2015, FEBS letters.

[45]  R. Kingston,et al.  H3K27 modifications define segmental regulatory domains in the Drosophila bithorax complex , 2014, eLife.

[46]  D. Reinberg,et al.  Putting a halt on PRC2 in pediatric glioblastoma , 2013, Nature Genetics.

[47]  Job Dekker,et al.  My5C: web tools for chromosome conformation capture studies , 2009, Nature Methods.

[48]  Benjamin Leblanc,et al.  Polycomb-Dependent Regulatory Contacts between Distant Hox Loci in Drosophila , 2011, Cell.

[49]  Yuval Kluger,et al.  PCGF homologs, CBX proteins, and RYBP define functionally distinct PRC1 family complexes. , 2012, Molecular cell.

[50]  A. Tanay,et al.  Probabilistic modeling of Hi-C contact maps eliminates systematic biases to characterize global chromosomal architecture , 2011, Nature Genetics.

[51]  D. Duboule,et al.  Epigenetic Temporal Control of Mouse Hox Genes in Vivo , 2009, Science.

[52]  T. Bestor,et al.  FBXL10 protects Polycomb-bound genes from hypermethylation , 2015, Nature Genetics.

[53]  Jeannie T. Lee,et al.  The Xist RNA-PRC2 complex at 20-nm resolution reveals a low Xist stoichiometry and suggests a hit-and-run mechanism in mouse cells , 2015, Proceedings of the National Academy of Sciences.

[54]  Jennifer E. Phillips-Cremins,et al.  Architectural Protein Subclasses Shape 3D Organization of Genomes during Lineage Commitment , 2013, Cell.

[55]  Giacomo Cavalli,et al.  Polycomb silencing: from linear chromatin domains to 3D chromosome folding. , 2014, Current opinion in genetics & development.

[56]  N. Brockdorff,et al.  RYBP-PRC1 Complexes Mediate H2A Ubiquitylation at Polycomb Target Sites Independently of PRC2 and H3K27me3 , 2012, Cell.

[57]  Kiran C. Bobba,et al.  Discovery of Novel Recurrent Mutations in Childhood Early T-Cell Precursor Acute Lymphoblastic Leukemia by Whole Genome Sequencing - a Report From the St Jude Children's Research Hospital - Washington University Pediatric Cancer Genome Project , 2011 .

[58]  Zhaohui S. Qin,et al.  Gene density, transcription, and insulators contribute to the partition of the Drosophila genome into physical domains. , 2012, Molecular cell.

[59]  David G Hendrickson,et al.  Differential analysis of gene regulation at transcript resolution with RNA-seq , 2012, Nature Biotechnology.

[60]  Li Ding,et al.  Somatic Histone H3 Alterations in Paediatric Diffuse Intrinsic Pontine Gliomas and Non-Brainstem Glioblastomas , 2012, Nature Genetics.

[61]  V. Pirrotta,et al.  Drosophila Enhancer of Zeste/ESC Complexes Have a Histone H3 Methyltransferase Activity that Marks Chromosomal Polycomb Sites , 2002, Cell.

[62]  Neva C. Durand,et al.  A 3D Map of the Human Genome at Kilobase Resolution Reveals Principles of Chromatin Looping , 2014, Cell.

[63]  Giacomo Cavalli,et al.  Regulation of Genome Architecture and Function by Polycomb Proteins. , 2016, Trends in cell biology.

[64]  Pedro P. Rocha,et al.  CTCF establishes discrete functional chromatin domains at the Hox clusters during differentiation , 2015, Science.

[65]  H. Jäckle,et al.  A Histone Mutant Reproduces the Phenotype Caused by Loss of Histone-Modifying Factor Polycomb , 2013, Science.

[66]  C. Creppe,et al.  A Cbx8-Containing Polycomb Complex Facilitates the Transition to Gene Activation during ES Cell Differentiation , 2014, PLoS genetics.

[67]  David T. W. Jones,et al.  Driver mutations in histone H3.3 and chromatin remodelling genes in paediatric glioblastoma , 2012, Nature.

[68]  R. Kingston,et al.  Chromatin Compaction by a Polycomb Group Protein Complex , 2004, Science.

[69]  David A. Orlando,et al.  Mediator and Cohesin Connect Gene Expression and Chromatin Architecture , 2010, Nature.

[70]  Richard Durbin,et al.  Sequence analysis Fast and accurate short read alignment with Burrows – Wheeler transform , 2009 .

[71]  L. Cozzuto,et al.  Nonoverlapping functions of the Polycomb group Cbx family of proteins in embryonic stem cells. , 2012, Cell stem cell.

[72]  Brigitte Wild,et al.  Histone Methyltransferase Activity of a Drosophila Polycomb Group Repressor Complex , 2002, Cell.

[73]  K. Helin,et al.  Transcriptional regulation by Polycomb group proteins , 2013, Nature Structural &Molecular Biology.