Publications by authors named "Robert E Thurman"

32 Publications

Mapping and Dynamics of Regulatory DNA in Maturing Siliques.

Front Plant Sci 2019 14;10:1434. Epub 2019 Nov 14.

Department of Genome Sciences, University of Washington, Seattle, WA, United States.

The genome is reprogrammed during development to produce diverse cell types, largely through altered expression and activity of key transcription factors. The accessibility and critical functions of epidermal cells have made them a model for connecting transcriptional events to development in a range of model systems. In and many other plants, fertilization triggers differentiation of specialized epidermal seed coat cells that have a unique morphology caused by large extracellular deposits of polysaccharides. Here, we used DNase I-seq to generate regulatory landscapes of seeds at two critical time points in seed coat maturation (4 and 7 DPA), enriching for seed coat cells with the INTACT method. We found over 3,000 developmentally dynamic regulatory DNA elements and explored their relationship with nearby gene expression. The dynamic regulatory elements were enriched for motifs for several transcription factors families; most notably the TCP family at the earlier time point and the MYB family at the later one. To assess the extent to which the observed regulatory sites in seeds added to previously known regulatory sites in we compared our data to 11 other data sets generated with 7-day-old seedlings for diverse tissues and conditions. Surprisingly, over a quarter of the regulatory, i.e. accessible, bases observed in seeds were novel. Notably, plant regulatory landscapes from different tissues, cell types, or developmental stages were more dynamic than those generated from bulk tissue in response to environmental perturbations, highlighting the importance of extending studies of regulatory DNA to single tissues and cell types during development.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fpls.2019.01434DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6868056PMC
November 2019

Integrative analysis of 111 reference human epigenomes.

Nature 2015 Feb;518(7539):317-30

1] Department of Cellular and Molecular Medicine, Institute of Genomic Medicine, Moores Cancer Center, Department of Chemistry and Biochemistry, University of California San Diego, 9500 Gilman Drive, La Jolla, California 92093, USA. [2] Ludwig Institute for Cancer Research, 9500 Gilman Drive, La Jolla, California 92093, USA.

The reference human genome sequence set the stage for studies of genetic variation and its association with human disease, but epigenomic studies lack a similar reference. To address this need, the NIH Roadmap Epigenomics Consortium generated the largest collection so far of human epigenomes for primary cells and tissues. Here we describe the integrative analysis of 111 reference human epigenomes generated as part of the programme, profiled for histone modification patterns, DNA accessibility, DNA methylation and RNA expression. We establish global maps of regulatory elements, define regulatory modules of coordinated activity, and their likely activators and repressors. We show that disease- and trait-associated genetic variants are enriched in tissue-specific epigenomic marks, revealing biologically relevant cell types for diverse human traits, and providing a resource for interpreting the molecular basis of human disease. Our results demonstrate the central role of epigenomic information for understanding gene regulation, cellular differentiation and human disease.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/nature14248DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4530010PMC
February 2015

Mouse regulatory DNA landscapes reveal global principles of cis-regulatory evolution.

Science 2014 Nov;346(6212):1007-12

Howard Hughes Medical Institute. Division of Hematology/Oncology, Children's Hospital Boston and Department of Pediatric Oncology, Dana-Farber Cancer Institute, Harvard Stem Cell Institute, Harvard Medical School, Boston, MA 02115, USA.

To study the evolutionary dynamics of regulatory DNA, we mapped >1.3 million deoxyribonuclease I-hypersensitive sites (DHSs) in 45 mouse cell and tissue types, and systematically compared these with human DHS maps from orthologous compartments. We found that the mouse and human genomes have undergone extensive cis-regulatory rewiring that combines branch-specific evolutionary innovation and loss with widespread repurposing of conserved DHSs to alternative cell fates, and that this process is mediated by turnover of transcription factor (TF) recognition elements. Despite pervasive evolutionary remodeling of the location and content of individual cis-regulatory regions, within orthologous mouse and human cell types the global fraction of regulatory DNA bases encoding recognition sites for each TF has been strictly conserved. Our findings provide new insights into the evolutionary forces shaping mammalian regulatory DNA landscapes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/science.1246426DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4337786PMC
November 2014

Topologically associating domains are stable units of replication-timing regulation.

Nature 2014 Nov;515(7527):402-5

Department of Biological Science, 319 Stadium Drive, Florida State University, Tallahassee, Florida 32306, USA.

Eukaryotic chromosomes replicate in a temporal order known as the replication-timing program. In mammals, replication timing is cell-type-specific with at least half the genome switching replication timing during development, primarily in units of 400-800 kilobases ('replication domains'), whose positions are preserved in different cell types, conserved between species, and appear to confine long-range effects of chromosome rearrangements. Early and late replication correlate, respectively, with open and closed three-dimensional chromatin compartments identified by high-resolution chromosome conformation capture (Hi-C), and, to a lesser extent, late replication correlates with lamina-associated domains (LADs). Recent Hi-C mapping has unveiled substructure within chromatin compartments called topologically associating domains (TADs) that are largely conserved in their positions between cell types and are similar in size to replication domains. However, TADs can be further sub-stratified into smaller domains, challenging the significance of structures at any particular scale. Moreover, attempts to reconcile TADs and LADs to replication-timing data have not revealed a common, underlying domain structure. Here we localize boundaries of replication domains to the early-replicating border of replication-timing transitions and map their positions in 18 human and 13 mouse cell types. We demonstrate that, collectively, replication domain boundaries share a near one-to-one correlation with TAD boundaries, whereas within a cell type, adjacent TADs that replicate at similar times obscure replication domain boundaries, largely accounting for the previously reported lack of alignment. Moreover, cell-type-specific replication timing of TADs partitions the genome into two large-scale sub-nuclear compartments revealing that replication-timing transitions are indistinguishable from late-replicating regions in chromatin composition and lamina association and accounting for the reduced correlation of replication timing to LADs and heterochromatin. Our results reconcile cell-type-specific sub-nuclear compartmentalization and replication timing with developmentally stable structural domains and offer a unified model for large-scale chromosome structure and function.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/nature13986DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4251741PMC
November 2014

Conservation of trans-acting circuitry during mammalian regulatory evolution.

Nature 2014 Nov;515(7527):365-70

1] Department of Genome Sciences, University of Washington, Seattle, Washington 98195, USA [2] Department of Medicine, University of Washington, Seattle, Washington 98195, USA.

The basic body plan and major physiological axes have been highly conserved during mammalian evolution, yet only a small fraction of the human genome sequence appears to be subject to evolutionary constraint. To quantify cis- versus trans-acting contributions to mammalian regulatory evolution, we performed genomic DNase I footprinting of the mouse genome across 25 cell and tissue types, collectively defining ∼8.6 million transcription factor (TF) occupancy sites at nucleotide resolution. Here we show that mouse TF footprints conjointly encode a regulatory lexicon that is ∼95% similar with that derived from human TF footprints. However, only ∼20% of mouse TF footprints have human orthologues. Despite substantial turnover of the cis-regulatory landscape, nearly half of all pairwise regulatory interactions connecting mouse TF genes have been maintained in orthologous human cell types through evolutionary innovation of TF recognition sequences. Furthermore, the higher-level organization of mouse TF-to-TF connections into cellular network architectures is nearly identical with human. Our results indicate that evolutionary selection on mammalian gene regulation is targeted chiefly at the level of trans-regulatory circuitry, enabling and potentiating cis-regulatory plasticity.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/nature13972DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4405208PMC
November 2014

A comparative encyclopedia of DNA elements in the mouse genome.

Nature 2014 Nov;515(7527):355-64

Bioinformatics and Genomics, Centre for Genomic Regulation (CRG) and UPF, Doctor Aiguader, 88, 08003 Barcelona, Catalonia, Spain.

The laboratory mouse shares the majority of its protein-coding genes with humans, making it the premier model organism in biomedical research, yet the two mammals differ in significant ways. To gain greater insights into both shared and species-specific transcriptional and cellular regulatory programs in the mouse, the Mouse ENCODE Consortium has mapped transcription, DNase I hypersensitivity, transcription factor binding, chromatin modifications and replication domains throughout the mouse genome in diverse cell and tissue types. By comparing with the human genome, we not only confirm substantial conservation in the newly annotated potential functional sequences, but also find a large degree of divergence of sequences involved in transcriptional regulation, chromatin state and higher order chromatin organization. Our results illuminate the wide range of evolutionary forces acting on genes and their regulatory regions, and provide a general resource for research into mammalian biology and mechanisms of human diseases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/nature13992DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4266106PMC
November 2014

Mapping and dynamics of regulatory DNA and transcription factor networks in A. thaliana.

Cell Rep 2014 Sep 15;8(6):2015-2030. Epub 2014 Sep 15.

Department of Genome Sciences, University of Washington, Seattle, WA 98195, USA. Electronic address:

Our understanding of gene regulation in plants is constrained by our limited knowledge of plant cis-regulatory DNA and its dynamics. We mapped DNase I hypersensitive sites (DHSs) in A. thaliana seedlings and used genomic footprinting to delineate ∼ 700,000 sites of in vivo transcription factor (TF) occupancy at nucleotide resolution. We show that variation associated with 72 diverse quantitative phenotypes localizes within DHSs. TF footprints encode an extensive cis-regulatory lexicon subject to recent evolutionary pressures, and widespread TF binding within exons may have shaped codon usage patterns. The architecture of A. thaliana TF regulatory networks is strikingly similar to that of animals in spite of diverged regulatory repertoires. We analyzed regulatory landscape dynamics during heat shock and photomorphogenesis, disclosing thousands of environmentally sensitive elements and enabling mapping of key TF regulatory circuits underlying these fundamental responses. Our results provide an extensive resource for the study of A. thaliana gene regulation and functional biology.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.celrep.2014.08.019DOI Listing
September 2014

Reduced local mutation density in regulatory DNA of cancer genomes is linked to DNA repair.

Nat Biotechnol 2014 Jan 15;32(1):71-5. Epub 2013 Dec 15.

1] Division of Genetics, Department of Medicine, Brigham & Women's Hospital, Boston, Massachusetts, USA. [2] Harvard Medical School, Boston, Massachusetts, USA. [3] The Broad Institute of MIT and Harvard, Cambridge, Massachusetts, USA.

Carcinogenesis and neoplastic progression are mediated by the accumulation of somatic mutations. Here we report that the local density of somatic mutations in cancer genomes is highly reduced specifically in accessible regulatory DNA defined by DNase I hypersensitive sites. This reduction is independent of any known factors influencing somatic mutation density and is observed in diverse cancer types, suggesting a general mechanism. By analyzing individual cancer genomes, we show that the reduced local mutation density within regulatory DNA is linked to intact global genome repair machinery, with nearly complete abrogation of the hypomutation phenomenon in individual cancers that possess mutations in components of the nucleotide excision repair system. Together, our results connect chromatin structure, gene regulation and cancer-associated somatic mutation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/nbt.2778DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4116484PMC
January 2014

Developmental fate and cellular maturity encoded in human regulatory DNA landscapes.

Cell 2013 Aug;154(4):888-903

Department of Genome Sciences, University of Washington, Seattle, WA 98109, USA.

Cellular-state information between generations of developing cells may be propagated via regulatory regions. We report consistent patterns of gain and loss of DNase I-hypersensitive sites (DHSs) as cells progress from embryonic stem cells (ESCs) to terminal fates. DHS patterns alone convey rich information about cell fate and lineage relationships distinct from information conveyed by gene expression. Developing cells share a proportion of their DHS landscapes with ESCs; that proportion decreases continuously in each cell type as differentiation progresses, providing a quantitative benchmark of developmental maturity. Developmentally stable DHSs densely encode binding sites for transcription factors involved in autoregulatory feedback circuits. In contrast to normal cells, cancer cells extensively reactivate silenced ESC DHSs and those from developmental programs external to the cell lineage from which the malignancy derives. Our results point to changes in regulatory DNA landscapes as quantitative indicators of cell-fate transitions, lineage relationships, and dysfunction.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.cell.2013.07.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3962256PMC
August 2013

Genome-scale mapping of DNase I hypersensitivity.

Curr Protoc Mol Biol 2013 Jul;Chapter 27:Unit 21.27

Department of Genome Sciences, University of Washington, Seattle, Washington, USA.

DNase I-seq is a global and high-resolution method that uses the nonspecific endonuclease DNase I to map chromatin accessibility. These accessible regions, designated as DNase I hypersensitive sites (DHSs), define the regulatory features, (e.g., promoters, enhancers, insulators, and locus control regions) of complex genomes. In this unit, methods are described for nuclei isolation, digestion of nuclei with limiting concentrations of DNase I, and the biochemical fractionation of DNase I hypersensitive sites in preparation for high-throughput sequencing. DNase I-seq is an unbiased and robust method that is not predicated on an a priori understanding of regulatory patterns or chromatin features.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/0471142727.mb2127s103DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4405172PMC
July 2013

Patterns of regulatory activity across diverse human cell types predict tissue identity, transcription factor binding, and long-range interactions.

Genome Res 2013 May 12;23(5):777-88. Epub 2013 Mar 12.

Program in Computational Biology and Bioinformatics, Duke University, Durham, North Carolina 27710, USA.

Regulatory elements recruit transcription factors that modulate gene expression distinctly across cell types, but the relationships among these remains elusive. To address this, we analyzed matched DNase-seq and gene expression data for 112 human samples representing 72 cell types. We first defined more than 1800 clusters of DNase I hypersensitive sites (DHSs) with similar tissue specificity of DNase-seq signal patterns. We then used these to uncover distinct associations between DHSs and promoters, CpG islands, conserved elements, and transcription factor motif enrichment. Motif analysis within clusters identified known and novel motifs in cell-type-specific and ubiquitous regulatory elements and supports a role for AP-1 regulating open chromatin. We developed a classifier that accurately predicts cell-type lineage based on only 43 DHSs and evaluated the tissue of origin for cancer cell types. A similar classifier identified three sex-specific loci on the X chromosome, including the XIST lincRNA locus. By correlating DNase I signal and gene expression, we predicted regulated genes for more than 500K DHSs. Finally, we introduce a web resource to enable researchers to use these results to explore these regulatory patterns and better understand how expression is modulated within and across human cell types.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1101/gr.152140.112DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3638134PMC
May 2013

Personal and population genomics of human regulatory variation.

Genome Res 2012 Sep;22(9):1689-97

Department of Genome Sciences, University of Washington, Seattle, Washington 98195, USA.

The characteristics and evolutionary forces acting on regulatory variation in humans remains elusive because of the difficulty in defining functionally important noncoding DNA. Here, we combine genome-scale maps of regulatory DNA marked by DNase I hypersensitive sites (DHSs) from 138 cell and tissue types with whole-genome sequences of 53 geographically diverse individuals in order to better delimit the patterns of regulatory variation in humans. We estimate that individuals likely harbor many more functionally important variants in regulatory DNA compared with protein-coding regions, although they are likely to have, on average, smaller effect sizes. Moreover, we demonstrate that there is significant heterogeneity in the level of functional constraint in regulatory DNA among different cell types. We also find marked variability in functional constraint among transcription factor motifs in regulatory DNA, with sequence motifs for major developmental regulators, such as HOX proteins, exhibiting levels of constraint comparable to protein-coding regions. Finally, we perform a genome-wide scan of recent positive selection and identify hundreds of novel substrates of adaptive regulatory evolution that are enriched for biologically interesting pathways such as melanogenesis and adipocytokine signaling. These data and results provide new insights into patterns of regulatory variation in individuals and populations and demonstrate that a large proportion of functionally important variation lies beyond the exome.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1101/gr.134890.111DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3431486PMC
September 2012

Widespread plasticity in CTCF occupancy linked to DNA methylation.

Genome Res 2012 Sep;22(9):1680-8

Department of Genome Sciences, University of Washington, Seattle, Washington 98195, USA.

CTCF is a ubiquitously expressed regulator of fundamental genomic processes including transcription, intra- and interchromosomal interactions, and chromatin structure. Because of its critical role in genome function, CTCF binding patterns have long been assumed to be largely invariant across different cellular environments. Here we analyze genome-wide occupancy patterns of CTCF by ChIP-seq in 19 diverse human cell types, including normal primary cells and immortal lines. We observed highly reproducible yet surprisingly plastic genomic binding landscapes, indicative of strong cell-selective regulation of CTCF occupancy. Comparison with massively parallel bisulfite sequencing data indicates that 41% of variable CTCF binding is linked to differential DNA methylation, concentrated at two critical positions within the CTCF recognition sequence. Unexpectedly, CTCF binding patterns were markedly different in normal versus immortal cells, with the latter showing widespread disruption of CTCF binding associated with increased methylation. Strikingly, this disruption is accompanied by up-regulation of CTCF expression, with the result that both normal and immortal cells maintain the same average number of CTCF occupancy sites genome-wide. These results reveal a tight linkage between DNA methylation and the global occupancy patterns of a major sequence-specific regulatory factor.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1101/gr.136101.111DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3431485PMC
September 2012

Systematic localization of common disease-associated variation in regulatory DNA.

Science 2012 Sep 5;337(6099):1190-5. Epub 2012 Sep 5.

Department of Genome Sciences, University of Washington, Seattle, WA 98195, USA.

Genome-wide association studies have identified many noncoding variants associated with common diseases and traits. We show that these variants are concentrated in regulatory DNA marked by deoxyribonuclease I (DNase I) hypersensitive sites (DHSs). Eighty-eight percent of such DHSs are active during fetal development and are enriched in variants associated with gestational exposure-related phenotypes. We identified distant gene targets for hundreds of variant-containing DHSs that may explain phenotype associations. Disease-associated variants systematically perturb transcription factor recognition sequences, frequently alter allelic chromatin states, and form regulatory networks. We also demonstrated tissue-selective enrichment of more weakly disease-associated variants within DHSs and the de novo identification of pathogenic cell types for Crohn's disease, multiple sclerosis, and an electrocardiogram trait, without prior knowledge of physiological mechanisms. Our results suggest pervasive involvement of regulatory DNA variation in common human disease and provide pathogenic insights into diverse disorders.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/science.1222794DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3771521PMC
September 2012

An expansive human regulatory lexicon encoded in transcription factor footprints.

Nature 2012 Sep;489(7414):83-90

Department of Genome Sciences, University of Washington, Seattle, Washington 98195, USA.

Regulatory factor binding to genomic DNA protects the underlying sequence from cleavage by DNase I, leaving nucleotide-resolution footprints. Using genomic DNase I footprinting across 41 diverse cell and tissue types, we detected 45 million transcription factor occupancy events within regulatory regions, representing differential binding to 8.4 million distinct short sequence elements. Here we show that this small genomic sequence compartment, roughly twice the size of the exome, encodes an expansive repertoire of conserved recognition sequences for DNA-binding proteins that nearly doubles the size of the human cis-regulatory lexicon. We find that genetic variants affecting allelic chromatin states are concentrated in footprints, and that these elements are preferentially sheltered from DNA methylation. High-resolution DNase I cleavage patterns mirror nucleotide-level evolutionary conservation and track the crystallographic topography of protein-DNA interfaces, indicating that transcription factor structure has been evolutionarily imprinted on the human genome sequence. We identify a stereotyped 50-base-pair footprint that precisely defines the site of transcript origination within thousands of human promoters. Finally, we describe a large collection of novel regulatory factor recognition motifs that are highly conserved in both sequence and function, and exhibit cell-selective occupancy patterns that closely parallel major regulators of development, differentiation and pluripotency.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/nature11212DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3736582PMC
September 2012

The accessible chromatin landscape of the human genome.

Nature 2012 Sep;489(7414):75-82

Department of Genome Sciences, University of Washington, Seattle, Washington 98195, USA.

DNase I hypersensitive sites (DHSs) are markers of regulatory DNA and have underpinned the discovery of all classes of cis-regulatory elements including enhancers, promoters, insulators, silencers and locus control regions. Here we present the first extensive map of human DHSs identified through genome-wide profiling in 125 diverse cell and tissue types. We identify ∼2.9 million DHSs that encompass virtually all known experimentally validated cis-regulatory sequences and expose a vast trove of novel elements, most with highly cell-selective regulation. Annotating these elements using ENCODE data reveals novel relationships between chromatin accessibility, transcription, DNA methylation and regulatory factor occupancy patterns. We connect ∼580,000 distal DHSs with their target promoters, revealing systematic pairing of different classes of distal DHSs and specific promoter types. Patterning of chromatin accessibility at many regulatory regions is organized with dozens to hundreds of co-activated elements, and the transcellular DNase I sensitivity pattern at a given region can predict cell-type-specific functional behaviours. The DHS landscape shows signatures of recent functional evolutionary constraint. However, the DHS compartment in pluripotent and immortalized cells exhibits higher mutation rates than that in highly differentiated cells, exposing an unexpected link between chromatin accessibility, proliferative potential and patterns of human variation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/nature11232DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3721348PMC
September 2012

An encyclopedia of mouse DNA elements (Mouse ENCODE).

Genome Biol 2012 Aug 13;13(8):418. Epub 2012 Aug 13.

To complement the human Encyclopedia of DNA Elements (ENCODE) project and to enable a broad range of mouse genomics efforts, the Mouse ENCODE Consortium is applying the same experimental pipelines developed for human ENCODE to annotate the mouse genome.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/gb-2012-13-8-418DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3491367PMC
August 2012

BEDOPS: high-performance genomic feature operations.

Bioinformatics 2012 Jul 9;28(14):1919-20. Epub 2012 May 9.

Department of Genome Sciences and Department of Medicine, University of Washington, Seattle, Washington, DC 98195, USA.

Unlabelled: The large and growing number of genome-wide datasets highlights the need for high-performance feature analysis and data comparison methods, in addition to efficient data storage and retrieval techniques. We introduce BEDOPS, a software suite for common genomic analysis tasks which offers improved flexibility, scalability and execution time characteristics over previously published packages. The suite includes a utility to compress large inputs into a lossless format that can provide greater space savings and faster data extractions than alternatives.

Availability: http://code.google.com/p/bedops/ includes binaries, source and documentation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/bioinformatics/bts277DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3389768PMC
July 2012

Transcription factor AP1 potentiates chromatin accessibility and glucocorticoid receptor binding.

Mol Cell 2011 Jul;43(1):145-55

Laboratory of Receptor Biology and Gene Expression, B602, Building 41, 41 Library Drive, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.

Ligand-dependent transcription by the nuclear receptor glucocorticoid receptor (GR) is mediated by interactions with coregulators. The role of these interactions in determining selective binding of GR to regulatory elements remains unclear. Recent findings indicate that a large fraction of genomic GR binding coincides with chromatin that is accessible prior to hormone treatment, suggesting that receptor binding is dictated by proteins that maintain chromatin in an open state. Combining DNaseI accessibility and chromatin immunoprecipitation with high-throughput sequencing, we identify the activator protein 1 (AP1) as a major partner for productive GR-chromatin interactions. AP1 is critical for GR-regulated transcription and recruitment to co-occupied regulatory elements, illustrating an extensive AP1-GR interaction network. Importantly, the maintenance of baseline chromatin accessibility facilitates GR recruitment and is dependent on AP1 binding. We propose a model in which the basal occupancy of transcription factors acts to prime chromatin and direct inducible transcription factors to select regions in the genome.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.molcel.2011.06.016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3138120PMC
July 2011

DNA methylation status predicts cell type-specific enhancer activity.

EMBO J 2011 Jun 24;30(15):3028-39. Epub 2011 Jun 24.

Center for Cancer Research, Laboratory of Receptor Biology and Gene Expression, National Cancer Institute, NIH, Bethesda, MD, USA.

Cell-selective glucocorticoid receptor (GR) binding to distal regulatory elements is associated with cell type-specific regions of locally accessible chromatin. These regions can either pre-exist in chromatin (pre-programmed) or be induced by the receptor (de novo). Mechanisms that create and maintain these sites are not well understood. We observe a global enrichment of CpG density for pre-programmed elements, and implicate their demethylated state in the maintenance of open chromatin in a tissue-specific manner. In contrast, sites that are actively opened by GR (de novo) are characterized by low CpG density, and form a unique class of enhancers devoid of suppressive effect of agglomerated methyl-cytosines. Furthermore, treatment with glucocorticoids induces rapid changes in methylation levels at selected CpGs within de novo sites. Finally, we identify GR-binding elements with CpGs at critical positions, and show that methylation can affect GR-DNA interactions in vitro. The findings present a unique link between tissue-specific chromatin accessibility, DNA methylation and transcription factor binding and show that DNA methylation can be an integral component of gene regulation by nuclear receptors.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/emboj.2011.210DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3160184PMC
June 2011

Dynamic reprogramming of chromatin accessibility during Drosophila embryo development.

Genome Biol 2011 11;12(5):R43. Epub 2011 May 11.

Department of Genome Sciences, University of Washington, Foege S310A, 1705 NE Pacific Street, Box 355065, Seattle, WA 98195, USA.

Background: The development of complex organisms is believed to involve progressive restrictions in cellular fate. Understanding the scope and features of chromatin dynamics during embryogenesis, and identifying regulatory elements important for directing developmental processes remain key goals of developmental biology.

Results: We used in vivo DNaseI sensitivity to map the locations of regulatory elements, and explore the changing chromatin landscape during the first 11 hours of Drosophila embryonic development. We identified thousands of conserved, developmentally dynamic, distal DNaseI hypersensitive sites associated with spatial and temporal expression patterning of linked genes and with large regions of chromatin plasticity. We observed a nearly uniform balance between developmentally up- and down-regulated DNaseI hypersensitive sites. Analysis of promoter chromatin architecture revealed a novel role for classical core promoter sequence elements in directing temporally regulated chromatin remodeling. Another unexpected feature of the chromatin landscape was the presence of localized accessibility over many protein-coding regions, subsets of which were developmentally regulated or associated with the transcription of genes with prominent maternal RNA contributions in the blastoderm.

Conclusions: Our results provide a global view of the rich and dynamic chromatin landscape of early animal development, as well as novel insights into the organization of developmentally regulated chromatin features.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/gb-2011-12-5-r43DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3219966PMC
February 2012

Diverse gene reprogramming events occur in the same spatial clusters of distal regulatory elements.

Genome Res 2011 May 6;21(5):697-706. Epub 2011 Apr 6.

Laboratory of Receptor Biology and Gene Expression, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892-5055, USA.

The spatial organization of genes in the interphase nucleus plays an important role in establishment and regulation of gene expression. Contradicting results have been reported to date, with little consensus about the dynamics of nuclear organization and the features of the contact loci. In this study, we investigated the properties and dynamics of genomic loci that are in contact with glucocorticoid receptor (GR)-responsive loci. We took a systematic approach, combining genome-wide interaction profiling by the chromosome conformation capture on chip (4C) technology with expression, protein occupancy, and chromatin accessibility profiles. This approach allowed a comprehensive analysis of how distinct features of the linear genome are organized in the three-dimensional nuclear space in the context of rapid gene regulation. We found that the transcriptional response to GR occurs without dramatic nuclear reorganization. Moreover, contrary to the view of transcription-driven organization, even genes with opposite transcriptional responses colocalize. Regions contacting GR-regulated genes are not particularly enriched for GR-regulated loci or for any functional group of genes, suggesting that these subnuclear environments are not organized to respond to a specific factor. The contact regions are, however, highly enriched for DNase I-hypersensitive sites that comprehensively mark cell-type-specific regulatory sites. These findings indicate that the nucleus is pre-organized in a conformation allowing rapid transcriptional reprogramming, and this organization is significantly correlated with cell-type-specific chromatin sites accessible to regulatory factors. Numerous open chromatin loci may be arranged in nuclear domains that are poised to respond to diverse signals in general and to permit efficient gene regulation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1101/gr.111153.110DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3083086PMC
May 2011

Chromatin accessibility pre-determines glucocorticoid receptor binding patterns.

Nat Genet 2011 Mar 23;43(3):264-8. Epub 2011 Jan 23.

Laboratory of Receptor Biology and Gene Expression, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, USA.

Development, differentiation and response to environmental stimuli are characterized by sequential changes in cellular state initiated by the de novo binding of regulated transcriptional factors to their cognate genomic sites. The mechanism whereby a given regulatory factor selects a limited number of in vivo targets from a myriad of potential genomic binding sites is undetermined. Here we show that up to 95% of de novo genomic binding by the glucocorticoid receptor, a paradigmatic ligand-activated transcription factor, is targeted to preexisting foci of accessible chromatin. Factor binding invariably potentiates chromatin accessibility. Cell-selective glucocorticoid receptor occupancy patterns appear to be comprehensively predetermined by cell-specific differences in baseline chromatin accessibility patterns, with secondary contributions from local sequence features. The results define a framework for understanding regulatory factor-genome interactions and provide a molecular basis for the tissue selectivity of steroid pharmaceuticals and other agents that intersect the living genome.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/ng.759DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6386452PMC
March 2011

Comprehensive analysis of the chromatin landscape in Drosophila melanogaster.

Nature 2011 Mar 22;471(7339):480-5. Epub 2010 Dec 22.

Center for Biomedical Informatics, Harvard Medical School, Boston, Massachusetts 02115, USA.

Chromatin is composed of DNA and a variety of modified histones and non-histone proteins, which have an impact on cell differentiation, gene regulation and other key cellular processes. Here we present a genome-wide chromatin landscape for Drosophila melanogaster based on eighteen histone modifications, summarized by nine prevalent combinatorial patterns. Integrative analysis with other data (non-histone chromatin proteins, DNase I hypersensitivity, GRO-Seq reads produced by engaged polymerase, short/long RNA products) reveals discrete characteristics of chromosomes, genes, regulatory elements and other functional domains. We find that active genes display distinct chromatin signatures that are correlated with disparate gene lengths, exon patterns, regulatory functions and genomic contexts. We also demonstrate a diversity of signatures among Polycomb targets that include a subset with paused polymerase. This systematic profiling and integrative analysis of chromatin signatures provides insights into how genomic elements are regulated, and will serve as a resource for future experimental investigations of genome structure and function.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/nature09725DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3109908PMC
March 2011

Sequencing newly replicated DNA reveals widespread plasticity in human replication timing.

Proc Natl Acad Sci U S A 2010 Jan 4;107(1):139-44. Epub 2009 Dec 4.

Department of Medicine, Division of Medical Genetics, University of Washington School of Medicine, Seattle, WA 98195, USA.

Faithful transmission of genetic material to daughter cells involves a characteristic temporal order of DNA replication, which may play a significant role in the inheritance of epigenetic states. We developed a genome-scale approach--Repli Seq--to map temporally ordered replicating DNA using massively parallel sequencing and applied it to study regional variation in human DNA replication time across multiple human cell types. The method requires as few as 8,000 cytometry-fractionated cells for a single analysis, and provides high-resolution DNA replication patterns with respect to both cell-cycle time and genomic position. We find that different cell types exhibit characteristic replication signatures that reveal striking plasticity in regional replication time patterns covering at least 50% of the human genome. We also identified autosomal regions with marked biphasic replication timing that include known regions of monoallelic expression as well as many previously uncharacterized domains. Comparison with high-resolution genome-wide profiles of DNaseI sensitivity revealed that DNA replication typically initiates within foci of accessible chromatin comprising clustered DNaseI hypersensitive sites, and that replication time is better correlated with chromatin accessibility than with gene expression. The data collectively provide a unique, genome-wide picture of the epigenetic compartmentalization of the human genome and suggest that cell-lineage specification involves extensive reprogramming of replication timing patterns.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.0912402107DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2806781PMC
January 2010

Global mapping of protein-DNA interactions in vivo by digital genomic footprinting.

Nat Methods 2009 Apr 22;6(4):283-9. Epub 2009 Mar 22.

Department of Genome Sciences, University of Washington, Seattle, USA.

The orchestrated binding of transcriptional activators and repressors to specific DNA sequences in the context of chromatin defines the regulatory program of eukaryotic genomes. We developed a digital approach to assay regulatory protein occupancy on genomic DNA in vivo by dense mapping of individual DNase I cleavages from intact nuclei using massively parallel DNA sequencing. Analysis of >23 million cleavages across the Saccharomyces cerevisiae genome revealed thousands of protected regulatory protein footprints, enabling de novo derivation of factor binding motifs and the identification of hundreds of new binding sites for major regulators. We observed striking correspondence between single-nucleotide resolution DNase I cleavage patterns and protein-DNA interactions determined by crystallography. The data also yielded a detailed view of larger chromatin features including positioned nucleosomes flanking factor binding regions. Digital genomic footprinting should be a powerful approach to delineate the cis-regulatory framework of any organism with an available genome sequence.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/nmeth.1313DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2668528PMC
April 2009

Human mutation rate associated with DNA replication timing.

Nat Genet 2009 Apr 15;41(4):393-5. Epub 2009 Mar 15.

Department of Genome Sciences and Medicine, University of Washington, Seattle, WA, USA.

Eukaryotic DNA replication is highly stratified, with different genomic regions shown to replicate at characteristic times during S phase. Here we observe that mutation rate, as reflected in recent evolutionary divergence and human nucleotide diversity, is markedly increased in later-replicating regions of the human genome. All classes of substitutions are affected, suggesting a generalized mechanism involving replication time-dependent DNA damage. This correlation between mutation rate and regionally stratified replication timing may have substantial evolutionary implications.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/ng.363DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2914101PMC
April 2009

Predicting human nucleosome occupancy from primary sequence.

PLoS Comput Biol 2008 Aug 22;4(8):e1000134. Epub 2008 Aug 22.

Department of Genome Sciences, University of Washington, Seattle, Washington, United States of America.

Nucleosomes are the fundamental repeating unit of chromatin and comprise the structural building blocks of the living eukaryotic genome. Micrococcal nuclease (MNase) has long been used to delineate nucleosomal organization. Microarray-based nucleosome mapping experiments in yeast chromatin have revealed regularly-spaced translational phasing of nucleosomes. These data have been used to train computational models of sequence-directed nuclesosome positioning, which have identified ubiquitous strong intrinsic nucleosome positioning signals. Here, we successfully apply this approach to nucleosome positioning experiments from human chromatin. The predictions made by the human-trained and yeast-trained models are strongly correlated, suggesting a shared mechanism for sequence-based determination of nucleosome occupancy. In addition, we observed striking complementarity between classifiers trained on experimental data from weakly versus heavily digested MNase samples. In the former case, the resulting model accurately identifies nucleosome-forming sequences; in the latter, the classifier excels at identifying nucleosome-free regions. Using this model we are able to identify several characteristics of nucleosome-forming and nucleosome-disfavoring sequences. First, by combining results from each classifier applied de novo across the human ENCODE regions, the classifier reveals distinct sequence composition and periodicity features of nucleosome-forming and nucleosome-disfavoring sequences. Short runs of dinucleotide repeat appear as a hallmark of nucleosome-disfavoring sequences, while nucleosome-forming sequences contain short periodic runs of GC base pairs. Second, we show that nucleosome phasing is most frequently predicted flanking nucleosome-free regions. The results suggest that the major mechanism of nucleosome positioning in vivo is boundary-event-driven and affirm the classical statistical positioning theory of nucleosome organization.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1371/journal.pcbi.1000134DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2515632PMC
August 2008

Nucleosome positioning signals in genomic DNA.

Genome Res 2007 Aug 9;17(8):1170-7. Epub 2007 Jul 9.

Bioinformatics Program, Boston University, Boston, MA 02215, USA.

Although histones can form nucleosomes on virtually any genomic sequence, DNA sequences show considerable variability in their binding affinity. We have used DNA sequences of Saccharomyces cerevisiae whose nucleosome binding affinities have been experimentally determined (Yuan et al. 2005) to train a support vector machine to identify the nucleosome formation potential of any given sequence of DNA. The DNA sequences whose nucleosome formation potential are most accurately predicted are those that contain strong nucleosome forming or inhibiting signals and are found within nucleosome length stretches of genomic DNA with continuous nucleosome formation or inhibition signals. We have accurately predicted the experimentally determined nucleosome positions across a well-characterized promoter region of S. cerevisiae and identified strong periodicity within 199 center-aligned mononucleosomes studied recently (Segal et al. 2006) despite there being no periodicity information used to train the support vector machine. Our analysis suggests that only a subset of nucleosomes are likely to be positioned by intrinsic sequence signals. This observation is consistent with the available experimental data and is inconsistent with the proposal of a nucleosome positioning code. Finally, we show that intrinsic nucleosome positioning signals are both more inhibitory and more variable in promoter regions than in open reading frames in S. cerevisiae.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1101/gr.6101007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1933512PMC
August 2007

Identification and analysis of functional elements in 1% of the human genome by the ENCODE pilot project.

Nature 2007 Jun;447(7146):799-816

We report the generation and analysis of functional data from multiple, diverse experiments performed on a targeted 1% of the human genome as part of the pilot phase of the ENCODE Project. These data have been further integrated and augmented by a number of evolutionary and computational analyses. Together, our results advance the collective knowledge about human genome function in several major areas. First, our studies provide convincing evidence that the genome is pervasively transcribed, such that the majority of its bases can be found in primary transcripts, including non-protein-coding transcripts, and those that extensively overlap one another. Second, systematic examination of transcriptional regulation has yielded new understanding about transcription start sites, including their relationship to specific regulatory sequences and features of chromatin accessibility and histone modification. Third, a more sophisticated view of chromatin structure has emerged, including its inter-relationship with DNA replication and transcriptional regulation. Finally, integration of these new sources of information, in particular with respect to mammalian evolution based on inter- and intra-species sequence comparisons, has yielded new mechanistic and evolutionary insights concerning the functional landscape of the human genome. Together, these studies are defining a path for pursuit of a more comprehensive characterization of human genome function.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2212820PMC
http://dx.doi.org/10.1038/nature05874DOI Listing
June 2007