title: Identification of transcription factor HLH-8 binding sites in C.elegans

summary:
We, being members of modENCODE consortium, have established an experimetal pipeline in C.elegans that allows global identification of the binding sites for transcription factors using chromatin immunoprecipitation followed by illumina high-throughput sequencing (ChIP-seq). In current study, we identified the binding sites for HLH-8 at L3 stage, which is required for normal muscle development and normal defecation and egg-laying.

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: HLH-8 was tagged with a dual GFP:3xFLAG tag at the carboxy terminus, whose expression pattern was confirmed through both fluorescence imaging and immunoblot analysis. The binding sites were determined using ChIP-seq. Two biological replicates (one is early early L3) were used in this study and non-immunoprecipitated chromatin (input) from the same sample served as a control. The concordance between two replicates is just above 50% when p-value is 0.05


title: Proteomic analysis of murine Piwi proteins reveals a role for arginine methylation in specifying interaction with Tudor family members

status: Public on Jul 25 2009

city: Cold Spring Harbor

first: Vasily

last: Vagin

state: NY

address: 1 Bungtown Rd

country: USA

institute: Cold Spring Harbor

type: Non-coding RNA profiling by high throughput sequencing

summary:
In germ cells, Piwi proteins interact with a specific class of small non-coding RNAs, piwi-interacting RNAs (piRNAs). Together, these form a pathway that represses transposable elements, thus safeguarding germ cell genomes.  While basic models describe the operation of piRNA pathways, neither the protein compositions of Piwi complexes, the critical protein-protein interactions that drive small RNA production and target recognition, or the precise molecular consequences of conserved localization to germline structures, call nuage, is well understood. We purified the three murine Piwi family proteins, Mili, Miwi, and Miwi2, from mouse germ cells and characterized their interacting protein partners.  Piwi proteins were found in complex with Prmt5/Wdr77, an enzyme that di-methylates arginine residues.  By immunoprecipitation with specific antibodies and by mass spectrometry, we found that Piwi proteins are arginine methylated at conserved positions in their amino termini.  These modifications are essential to direct complex formation with specific Tudor-domain proteins, whose interactions with Piwis can be required for localization of RNP complexes in cytoplasmic nuage, proper piRNA expression, and transposon silencing.  Considered together, our findings indicate that arginine methylation drives the assembly of multi-protein machines whose integrity and specific sub-cellular localization is necessary for efficient function of the piRNA pathway.

Keywords: gene regulation study

overall_design: Total small RNA in embryonic and post-birth mouse testes of tdrd1 and tdrd6 mutants

middle: V

first: James

last: Wohlschlegel

first: Jun

last: Qu

first: Zophonias

last: Jonsson

first: Xinhua

last: Huang

first: Shinichiro

last: Chuma

first: Angelique

last: Girard

first: Ravi

last: Sachidanandam

first: Gregory

middle: J

last: Hannon

first: Alexei

middle: A

last: Aravin


title: Small regulatory RNA in the bovine genome

status: Public on Jul 29 2009

city: Brisbane

first: Evgeny

last: Glazov

address: Ipswich Road

department: Diamantina Institute

country: Australia

institute: University of Queensland

summary:
MicroRNA and other types of small regulatory RNAs play a crucial role in the regulation of gene expression in eukaryotes. Several distinct classes of small regulatory RNAs have been discovered in recent years. To extend the repertoire of small regulatory RNAs characterized in mammals and identify miRNA expressed upon viral infection we used Illumina’s ultrahigh throughput sequencing approach. Results presented in this study extend our knowledge of the biology and evolution of small regulatory RNAs in mammals and illuminate mechanisms of small RNA biogenesis and function.

Keywords: Transcriptome analysis

overall_design: We sequenced three small RNA libraries prepared from cell line derived from the adult bovine kidney under normal conditions and upon infection of the cell line with the bovine herpesvirus 1.

first: Wanchai

last: Assavalapsakul

first: Paul

middle: F

last: Horwood

first: Kritaya

last: Kongsuwan

first: Neena

last: Mitter

first: Timothy

last: Mahony


title: Identification of genes that are regulated by TAK1 in human cutaneous T cell lymphoma HuT-102 cells

status: Public on Aug 18 2009

city: Toyama

first: Yoshiaki

last: Tabuchi

state: Toyama

address: 2630 Sugitani

department: Life Science Research Center

country: Japan

institute: University of Toyama

summary: We previously reported that human T cell lymphotropic virus 1 (HTLV-1) Tax oncoprotein constitutively activates TAK1. Here, we established Tax-positive HuT-102 cells stably downregulated TAK1 expression by short-hairpin RNA (HuT-shTAK1 cells), and investigated the physiological function of TAK1. Microarray analysis demonstrated that several interferon (IFN)-inducible genes including chemokines such as CXCL10 and CCL5 were significantly downregulated in HuT-shTAK1 cells. In contrast, Tax-mediated constitutive activation of NF-kB was intact in HuT-shTAK1 cells. IRF3, a critical transcription factor in innate immunity to viral infection, was constitutively activated in a Tax-dependent manner. Activation of IRF3 and IRF3-dependent gene expression were dependent on TAK1 and TBK1. On the other hand, IRF4, another IRF family of transcription factor overexpressed in a Tax-independent manner, negatively regulated the TAK1-dependent IRF3 transcriptional activity. Together, HTLV-1 manipulates IFN signaling by regulating both positive and negative IRFs.

overall_design: HuT-102 cells, a human cutaneous T cell lymphoma, were stably transfected with shRNA expression vectors against human MAP3K7 (TAK1) or firefly luciferase (Luc). The cells were maintained in media containing 0.5 mg/ml G418. For the experiment, the cells were incubated in media without G418 for 36 h at 37°C. Total RNA samples were prepared from the cells. Gene expression was analyzed by an Affymetrix GeneChip® system with a Human Genome U133-plus 2.0 array for analysis of over 47,000 transcripts. Sample preparation for array hybridization was carried out as described in the manufacturer’s instructions. Two replicates per sample type.

first: Shunsuke

last: Suzuki

first: Ichiro

last: Takasaki

first: Hiroaki

last: Sakurai


title: Processing of Drosophila endo-siRNAs depends on a specific Loquacious isoform

status: Public on Jul 31 2009

first: Benjamin

last: Czech

institute: Cold Spring Harbor Laboratory

summary:
Drosophila melanogaster expresses three classes of small RNAs, which are classified according to their mechanisms of biogenesis. MicroRNAs are ~22-23-nt, ubiquitously expressed small RNAs that are sequentially processed from hairpin-like precursors by Drosha/Pasha and Dcr-1/Loquacious complexes. MicroRNAs usually associate with AGO1 and regulate the expression of protein-coding genes. Piwi-interacting RNAs (piRNAs) of ~24-28-nt associate with Piwi-family proteins and can arise from single-stranded precursors. piRNAs function in transposon silencing and are mainly restricted to gonadal tissues. Endo-siRNAs are found in both germline and somatic tissues. These ~21-nt RNAs are produced by a distinct Dicer, Dcr-2, and do not depend on Drosha/Pasha complexes. They predominantly bind to AGO2 and target both mobile elements and protein-coding genes. Surprisingly, a subset of endo-siRNAs strongly depend for their production on the dsRNA-binding protein Loquacious (Loqs), thought generally to be a partner for Dcr-1 and a co-factor for miRNA biogenesis. Endo-siRNA production depends on a specific Loqs isoform, Loqs-PD, which is distinct from the one, Loqs-PB, required for the production of microRNAs. Paralleling their roles in the biogenesis of distinct small RNA classes, Loqs-PD and Loqs-PB bind to different Dicer proteins, with Dcr-1/Loqs-PB complexes and Dcr-2/Loqs-PD complexes driving microRNA and endo-siRNA biogenesis, respectively.

Small RNA profiling by high throughput sequencing

overall_design: Total RNA was isolated using Trizol reagent (Invitrogen) and size-fractionated by PAGE into 19-24nt. These were independently processed and sequenced using the Illumina GAII platform. In total, six libraries were analyzed.

first: Rui

last: Zhou

first: Julius

last: Brennecke

last: Sachindanandam

first: James

last: Wohlschlegel

first: Norbert

last: Perrimon


title: Chromatin architecture organized by histone variants H3.3 and H2A.Z in the human genome

status: Public on Jul 26 2009

city: Charlottesville

first: Chongzhi

last: Zang

state: VA

institute: University of Virginia

type: Genome binding/occupancy profiling by high throughput sequencing

summary: To understand how chromatin structure is organized by different histone variants, we have measured the genome-wide distribution of nucleosome core particles (NCPs) containing the histone variants H3.3 and H2A.Z in human cells. We find that a special class of NCPs containing both variants is enriched at ‘nucleosome-free regions’ of active promoters, enhancers and insulator regions. We show that preparative methods used previously in studying nucleosome structure result in the loss of these unstable double-variant NCPs. It seems likely that this instability facilitates the access of transcription factors to promoters and other regulatory sites in vivo. Other combinations of variants have different distributions, consistent with distinct roles for histone variants in the modulation of gene expression.

overall_design: genome-wide analysis of histone variants H2AZ, H3.3, and H3.3-H2A.Z double ChIP, plus input and genomic DNA controls in HeLa cells. H2A.Z samples are prepared under two different salt concentration conditions. (6 samples in total)

first: Chunyuan

last: Jin

first: Gang

last: Wei

first: Kairong

last: Cui

first: Weiqun

last: Peng

first: Keji

last: Zhao

first: Gary

last: Felsenfeld


title: Identification of transcription factor CEH-14 binding sites in C.elegans

city: New Haven

first: Flora

last: Vaccarino

state: CT

address: 230 South Frontage Road

department: Child Study Center

institute: Yale University

summary:
We, being members of modENCODE consortium, have established an experimetal pipeline in C.elegans that allows global identification of the binding sites for transcription factors using chromatin immunoprecipitation followed by illumina high-throughput sequencing (ChIP-seq). In current study, we identified the binding sites for CEH-14 at L2 stage, which is required for specification of the AFD thermosensory neurons and for normal thermotactic behavior.

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: CEH-14 was tagged with a dual GFP:3xFLAG tag at the carboxy terminus, whose expression pattern was confirmed through both fluorescence imaging and immunoblot analysis. The binding sites were determined using ChIP-seq. Two biological replicates were used in this study, but replicate1 is at a later L2 stage. Only one Input sample (non-immunoprecipitated chromatin (input) from replicate2 was used as a control. The concordance between two replicates is just above 50% when p-value is 0.05.

first: Wei

last: Niu


title: modENCODE RNA-Seq of Drosophila cell line Kc167

status: Public on Jul 15 2009

city: St Louis

first: Marijke

middle: Jeltje

last: van Baren

state: MO

address: One Brookings Drive

department: CSE

institute: Washington University

type: Expression profiling by high throughput sequencing

summary:
Deep Sequencing of mRNA from the Drosophila melanogaster Kc167 cell line.

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: Analysis of poly(A)+ RNA from Kc167 cell line, technical replicates.

first: Michael

middle: R

last: Brent

first: Laura

last: Langton

first: Charles

middle: L

last: Comstock

first: Randall

middle: H

last: Brown

first: Brian

middle: C

last: Koebbe


title: Intrinsic histone-DNA interactions are not the major determinant of nucleosome positions in vivo

city: Palo Alto

first: Ghia

last: Euskirchen

state: CA

address: 1501 S. California Ave.

department: Genetics

institute: Stanford University

summary: We assess the role of intrinsic histone-DNA interactions by mapping nucleosomes assembled in vitro on genomic DNA. Nucleosomes strongly prefer yeast DNA over E. coli DNA, indicating that the yeast genome evolved to favor nucleosome formation. Many yeast promoter and terminator regions intrinsically disfavor nucleosome formation, and nucleosomes assembled in vitro display strong rotational positioning. Nucleosome arrays generated by the ACF assembly factor display fewer nucleosome-free regions, reduced rotational positioning, and less translational positioning than obtained by intrinsic histone-DNA interactions. Importantly, in vitro assembled nucleosomes display only a limited preference for specific translational positions and do not show the pattern observed in vivo. Our results argue against a genomic code for nucleosome positioning, and they suggest that the nucleosomal pattern in coding regions arises primarily from statistical positioning from a barrier near the promoter that involves some aspect of transcriptional initiation by RNA polymerase II.

sample_organism:
Escherichia coli
Saccharomyces cerevisiae

overall_design: This data series consists of two experimental samples, "Nucleosomes assembled on yeast and E. coli DNA by salt dialysis" and "Nucleosomes assembled on yeast and E. coli DNA by ACF" plus one control sample, "Yeast-Ecoli control, sonicated".

first: Yong

last: Zhang

first: Zarmik

last: Moqtaderi

first: Barbara

last: Rattner

last: Snyder

first: James

middle: T

last: Kadonaga

first: X

middle: S

last: Liu

first: Kevin

last: Struhl


title: High-throughput pyrosequencing of miRNAs from Medicago truncatula nodules and roots

status: Public on Jul 07 2009

city: Castanet Tolosan

first: Jerome

last: Gouzy

address: 24 Chemin de Borde Rouge, BP52627

department: LIPM

country: France

institute: INRA/CNRS

summary:
We have used deep sequencing of small RNAs from nodules and root apexes of the model legume Medicago truncatula, to identify 113 novel candidate miRNAs. These miRNAs (legume or Mt-specific) are encoded by 278 putative hairpin precursors in the M. truncatula genome. Several miRNAs are differentially expressed in nodules and root tips and large variety of targets could be predicted for these genes. Specific miRNA isoforms showed contrasting expression patterns in these tissues

Keywords: Transcriptome analysis

overall_design: 3 samples examined: nodules, root tips, and root tips + NaCl

first: Christine

last: Lelandais

first: Loreto

last: Naya

first: Erika

last: Sallet

first: Fanny

last: Calenge

first: Florian

last: Frugier

first: Caroline

last: Hartmann

first: Martin

last: Crespi


title: Genome-wide mapping of HATs and HDACs in human CD4+ T cells

status: Public on Aug 20 2009

first: Chongzhi

last: Zang

summary: Histone acetyltransferases (HATs) and deacetylases (HDACs) function antagonistically to control histone acetylation. As acetylation is a histone mark for active transcription, HATs have been associated with active and HDACs with inactive genes. We describe here genome-wide mapping of HATs and HDACs binding on chromatin and ﬁnd that both are found at active genes with acetylated histones. Our data provide evidence that HATs and HDACs are both targeted to transcribed regions of active genes by phosphorylated RNA Pol II. Furthermore, the majority of HDACs in the human genome function to reset chromatin by removing acetylation at active genes. Inactive genes that are primed by MLL-mediated histone H3K4 methylation are subject to a dynamic cycle of acetylation and deacetylation by transient HAT/HDAC binding, preventing Pol II from binding to these genes but poising them for future activation. Silent genes without any H3K4 methylation signal show no evidence of being bound by HDACs.

overall_design:
high throughput sequencing: genome-wide analysis of 5 HATs, 4 HDACs in human CD4+ cells, Tip60 and HDAC6 in activated CD4 cells, genome-wide analysis of 2 histone acetylations and RNA Polymerase II after HDAC inhibitor treatment in CD4, histone modification with WDR5 knock-down and HDAC inhibitor treatment in HeLa cells

expression profiling: Global change in gene expression in human CD4+ T cells after HDAC inhibitor treatment for 2hours and 12 hours. (9 samples in total)

first: Zhibin

last: Wang

first: Kairong

last: Cui

first: Dustin

middle: E

last: Schones

first: Artem

last: Barski

first: Weiqun

last: Peng

first: Keji

last: Zhao


title: High definition profiling of mammalian DNA methylation by array capture and single molecule bisulfite sequencing

status: Public on Jul 08 2009

first: Emily

last: Hodges

address: 1 Bungtown Road

type: Methylation profiling by high throughput sequencing

summary:
DNA methylation stabilizes developmentally programmed gene expression states.  Aberrant methylation is associated with disease progression and is a common feature of cancer genomes. Presently, few methods enable quantitative, large-scale, single-base resolution mapping of DNA methylation states in desired regions of a complex mammalian genome. Here, we present an approach that combines array-based hybrid selection and massively parallel bisulfite sequencing to profile DNA methylation in genomic regions spanning hundreds of thousands of bases. This single molecule strategy enables methylation variable positions to be quantitatively examined with high sampling precision. Using bisulfite capture, we assessed methylation patterns across 324 randomly selected CpG islands (CGI) representing more than 25,000 CpG sites. A single lane of Illumina sequencing permitted methylation states to be definitively called for >90% of target sties. The accuracy of the hybrid-selection approach was verified using conventional bisulfite capillary sequencing of cloned PCR products amplified from a subset of the selected regions. This confirmed that even partially methylated states could be successfully called. A comparison of human primary and cancer cells revealed multiple differentially methylated regions.  More than 25% of islands showed complex methylation patterns either with partial methylation states defining the entire CGI or with contrasting methylation states appearing in specific regional blocks within the island. We observed that transitions in methylation state often correlate with genomic landmarks, including transcriptional start sites and intron-exon junctions.  Methylation, along with specific histone marks, was enriched in exonic regions, suggesting that chromatin states can foreshadow the content of mature mRNAs.

Keywords: DNA methylation profiling by massively parallel sequencing
Keywords: Epigenetics

overall_design: Targeted examination of DNA methylation in two human cell types by combining array capture and bisulfite sequencing. In addition, this study examined two histone marks in the breast tumor cell line MDA-MB-231.

first: Andrew

middle: D

last: Smith

first: Jude

last: Kendall

first: Zhenyu

last: Xuan

first: Kandasamy

last: Ravi

first: Michelle

last: Rooks

first: Michael

middle: Q

last: Zhang

first: Kenny

last: Ye

first: Arindam

last: Bhattacharjee

first: Leonardo

last: Brizuela

first: W

last: McCombie

last: Wigler

first: Gregory

last: Hannon

first: James

middle: B

last: Hicks


title: 454 Life Sciences/Roche and Illumina sequencing of small RNAs from planarian samples

status: Public on Jul 01 2009

city: Berlin

first: Marc

middle: Riemer

last: Friedlaender

state: Berlin

address: Robert-Rössle-Str. 10

country: Germany

institute: MDC, Berlin-Buch

summary: Freshwater planarian flatworms possess uncanny regenerative capacities mediated by abundant and collectively totipotent adult stem cells. Key functions of these cells during regeneration and tissue homeostasis have been shown to depend on PIWI, a molecule required for piRNA expression in planarians. Nevertheless, the full complement of piRNAs, and microRNAs (miRNAs) in this organism has yet to be defined. Here, we report on the large scale cloning and sequencing of small RNAs from the planarian Schmidtea mediterranea, yielding altogether millions of sequenced unique small RNAs. We show that piRNAs are in part organized in genomic clusters and that they share characteristic features with mammalian and fly piRNAs. We further identify 61 novel miRNA genes and thus double the number of known planarian miRNAs. Sequencing, as well as quantitative PCR of small RNAs uncovered ten miRNAs enriched in planarian stem cells. These miRNAs are down-regulated in animals in which stem cells have been abrogated by irradiation, and thus constitute miRNAs likely associated with specific stem cell functions. Altogether, we present the first comprehensive small RNA analysis in animals belonging to the third animal superphylum, the Lophotrochozoa, and single out a number of miRNAs that may function in regeneration. Several of these miRNAs are deeply conserved in animals.

overall_design: Small RNAs were sequenced from FACS-sorted neoblast stem cells, untreated planarians and irradiated planarians, depleted of neoblasts

last: Friedländer

first: Catherine

last: Adamidi

first: Ting

last: Han

first: Svetlana

last: Lebedeva

first: Thomas

last: Isenbarger

first: Martin

last: Hirst

first: Marco

last: Marra

first: Chad

last: Nusbaum

first: William

last: Lee

last: Jenkin

first: Alejandro

last: Alvarado

first: John

middle: K

last: Kim

first: Nikolaus

last: Rajewsky


title: Global identification of yeast chromosome interactions using genome conformation capture

status: Public on Jul 21 2009

city: Auckland

first: Justin

middle: M.

last: O'Sullivan

address: 85 Park Road, Grafton

department: Liggins Institute

country: New Zealand

institute: University of Auckland

summary: The objective of this study was to determine the global chromosomal interaction map for exponentially growing Saccharomyces cerevisiae cells using Genome Conformation Capture. Interactions between chromosomes were identified within a population of yeast cells growing exponentially in a semi-defined medium containing glucose. The series contains the sequences of the ligated restriction fragments that identified the interactions.

overall_design:
A GCC library was prepared, using MspI, from 1.5 x 10^5 cells and sequenced.

Data was processed using the Flp_spin_man program suite in conjunction with SOAP (Rodley C.D., Bertels F., Jones B., and O'Sullivan J.M., submitted). The software package initially sorted sequences according to the presence or absence of the MspI restriction site. Sequences containing the restriction site were broken, in silico, and their positions mapped onto the yeast genome (using SOAP). Thus regions of the genome that were in close enough proximity to be cross-linked and subsequently ligated together were identified.

first: Chris

last: Rodley

first: F

last: Bertels

first: B

last: Jones

first: J

middle: M


title: Chromatin Poises miRNA- and Protein-coding Genes for Expression

status: Public on Aug 10 2009

city: Bethesda

first: Keji

last: Zhao

state: MD

address: Rockville Pike

institute: NHLBI, NIH

summary:
Chromatin modifications have been implicated in regulation of gene expression. While association of certain modifications with expressed or silent genes has been established, it remains unclear how changes in chromatin environment relate to changes in gene expression. In this report, we used ChIP-Seq to analyze the genome-wide changes in chromatin modifications during activation of total human CD4+ T cells by T cell receptor (TCR) signaling. Surprisingly, we found that the chromatin modification patterns at many induced and silenced genes are relatively stable during the short-term activation of resting T cells. Active chromatin modifications were already in place for a majority of inducible protein-coding genes even while the genes were silent in resting cells. Similarly, genes that were silenced upon T cell activation retained positive chromatin modifications even after being silenced. To investigate if these observations are also valid for miRNA-coding genes, we systematically identified promoters for known miRNA genes using epigenetic marks and profiled their expression patterns using deep sequencing. We found that chromatin modifications can poise miRNA-coding genes as well. Our data suggest that miRNA- and protein-coding genes share similar mechanisms of regulation by chromatin modifications, which poise inducible genes for activation in response to environmental stimuli.
Keyword(s): Epigenetics

overall_design: Examination of several chromatin modifications in human CD4+ T cells

first: Artem

last: Barski

first: Raja

last: Jothi

first: Suresh

last: Cuddapah


title: The Transcriptional Programme Controlled by Scl/Tal1 Gene during Early Embryonic Haematopoietic Development

status: Public on Jun 19 2009

city: Cambridge

first: Samuel

middle: David

last: Foster

state: Cambridgeshire

address: Wellcome Trust/MRC Building, Addenbrooke's Hospital, Hills Road

department: Department of Haematology

country: United Kingdom

institute: Cambridge Institute for Medical Research

summary:
The basic-helix-loop-helix transcription factor Scl/Tal1 controls the development and subsequent differentiation of haematopoietic stem cells (HSCs).  However, since few Scl target genes have been validated to date, the underlying mechanisms have remained largely unknown.  Here we have employed ChIP-Seq technology to generate a genome-wide catalogue of Scl binding events in a stem/progenitor cell line followed by validation using primary fetal liver cells and comprehensive transgenic mouse assays.  Transgenic analysis provided in vivo validation of multiple new direct Scl target genes and allowed us to reconstruct an in vivo validated network consisting of 17 factors and their respective regulatory elements.  By coupling ChIP-Seq in model cell lines with in vivo transgenic validation and sophisticated bioinformatic analysis, we have identified a widely applicable strategy for the reconstruction of stem cell regulatory networks where biological material is otherwise limiting.  Moreover, in addition to revealing multiple previously unrecognised links to known HSC regulators as well as novel links to genes not previously implicated in HSC function, comprehensive transgenic analysis of regulatory elements provided substantial new insights into the transcriptional control of several important haematopoietic regulators including Cbfa2t3h/Eto2, Cebpe, Nfe2, Zfpm1/Fog1, Erg, Mafk, Gfi1b and Myb.

Keywords: Gene regulation study

overall_design:
Examination of one transcription factor in a haematopoietic progenitor cell line
The samples provided were all sourced from the same ChIP experiment. They were then split into three and sent for sequencing. The samples labelled Scl_lane7 and Scl_lane8 were sequenced in parallel using an Illumina Genome Analyzer and then aligned using Solexa alignment software. There only variation was that the Scl_lane8 sample contained twice as much of the DNA as Scl_lane7. The sample labelled Scl_vancouver was sent to an alternative site for sequencing, again on an Illumina Genome Analyzer, and was aligned using the Eland aligner software.

first: Nicola

last: Wilson

first: Diego

last: Miranda-Saavedra

first: Sarah

last: Kinston

first: Nicolas

last: Bonadies

first: Fernando

last: Nieto

first: Mark

middle: A

last: Dawson

first: Ian

middle: J

last: Donaldson

first: Stephanie

last: Dumon

first: Jonathan

last: Frampton

first: Rekin’s

last: Janky

first: Xiao-Hong

last: Sun

last: Teichmann

first: Andrew

last: Bannister

first: Berthold

last: Göttgens


status: Public on Jul 10 2009

first: Mike

last: Gilchrist

address: Tennis Court Road

department: Gurdon Institute

institute: Cambridge University

summary:
High-throughput sequencing of small RNAs from Xenopus tropicalis (adult liver, adult skin, oocytes stage I, II, III, IV, V, VI).
total RNA, ~18-42 nt RNAs isolated using PAGE, ligation to adapters requires 5' monophosphate and 3' OH

overall_design: Illumina/Solexa sequencing of adult liver, adult skin, oocytes stage I, II, III, IV, V, VI


title: Extensive demethylation of repetitive elements during seed development underlies gene imprinting

status: Public on Jun 12 2009

city: Seattle

first: Jorja

last: Henikoff

state: WA

address: 1100 Fairview AV N, A1-162

department: Basic Sciences

institute: Fred Hutchinson Cancer Research Center

type: Methylation profiling by genome tiling array

summary:
DNA methylation is an epigenetic mark associated with transposable element silencing and gene imprinting in flowering plants and mammals. In plants, imprinting occurs in the endosperm, which nourishes the embryo during seed development. We have profiled Arabidopsis DNA methylation genome-wide in the embryo and endosperm. Large-scale methylation changes accompany endosperm development and endosperm-specific gene expression. Transposable element fragments are extensively demethylated in the endosperm. We discovered new imprinted genes by identifying candidate genes associated with the top differentially methylated regions. Our data suggest that imprinting in plants evolved from genome defense against transposable elements.

Keywords: Affinity-purification on microarray

overall_design: All experiments were done using two channels per chip. Immunoprecipitated methylated DNA (IP) was compared to control genomic DNA (input). Both the IP and input represent Cy5 and Cy3 labeled Illumina GA libraries.

first: Mary

last: Gehring

first: Steve


title: Mycobacterium marinum chronic infection of adult zebrafish

status: Public on Jun 30 2009

city: Leiden

first: Anna

middle: Magdalena

last: Zakrzewska

address: Wassenaarseweg 64

department: IBL

country: Netherlands

institute: University of Leiden

summary: Novel high-throughput deep sequencing technology has dramatically changed the way that the functional complexity of transcriptomes can be studied. Here we report on the first use of this technology to gain insight into the wide range of transcriptional alterations that are associated with an infectious disease process. Using Solexa/Illumina’s digital gene expression (DGE) system, a tag-based transcriptome sequencing method, we investigated mycobacterium-induced transcriptome changes in a model vertebrate species, the zebrafish. Our DGE data substantiate recent RNA-seq results from other models indicating a much larger extent of genome transcription than previously thought, and demonstrate that the host response to bacterial infection adds a further degree of complexity to the transcriptome. We obtained a sequencing depth of over 5 million tags per sample with strong correlation between replicates. Tag mapping indicated that mycobacterium-infected adult zebrafish express over 70% of all genes represented in transcript databases. Comparison of our DGE data with a previous multiplatform microarray analysis showed that both types of technologies identified regulation of similar functional groups of genes, more specifically the up-regulation of different classes of immune response genes concomitant with a broad down-regulation of metabolic genes. However, the unbiased nature of DGE analysis provided insights that microarray analysis could not have achieved. As demonstrated here, DGE data are useful for the verification of predicted gene models and allowed us to detect mycobacterium-regulated switching between different transcript isoforms. Moreover, genomic mapping of infection-induced DGE tags revealed novel transcript forms for which any previous EST-based evidence of expression was lacking.

overall_design:
Adult male zebrafish were infected by intraperitoneal inoculation with approximately 1x 10-3 M. marinum bacteria. All four infected fish were sacrificed when they showed overt signs of fish tuberculosis, including lethargy, skin ulcers and extensive granuloma formation in organs such as liver and kidney. Histological examination of fish from the same experiments confirmed that the pathology of infected fish corresponded to fish tuberculosis and that no characteristics of the disease were present in the control fish.
For DGE analysis RNA samples from the four control adult zebrafish (c1,c2,c3,c4) were pooled, and RNA samples from the four M.marinum-infected adult zebrafish (i1,i2,i3,i4) were pooled. Before pooling the individual RNA samples had been checked by microarray analysis for correlation between biological replicates.

first: Zoltan

last: Hegedus

first: Vilmos

last: Agoston

first: Anita

last: Ordas

first: Peter

last: Racz

first: Matyas

last: Mink

first: Herman

middle: P

last: Spaink

first: Annemarie

last: Meijer


title: modENCODE RNA-Seq of Drosophila developmental stage 16-18 hr

status: Public on Aug 13 2009

city: Woodbury

first: Chris

last: Zaleski

address: 500 Sunnyside Blvd

institute: Cold Spring Harbor Labs

summary:
Deep Sequencing of mRNA from the Drosophila melanogaster developmental stage 16-18 hr.

Keywords: Expression profiling by high throughput sequencing

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: Seq of Poly-A+ RNA from D. melanogaster 16-18hr embryo

first: Carrie

last: Davis

first: Tom

last: Gingeras

first: Sue

last: Celniker


title: Molecular profiling of CD3- CD4+ T-cells from patients with the lymphocytic variant of hypereosinophilic syndrome

status: Public on Jul 17 2009

city: Brussels

first: Karen

last: Willard-Gallo

address: 127 Blvd de Waterloo

department: Institut Bordet

country: Belgium

institute: Université Libre de Bruxelles

summary: The clonal CD3- CD4+ Th2 cell population characterizing some hypereosinophilic syndrome patients stably endures for years provoking a chronic inflammatory skin disease, with a subgroup of patients ultimately progressing to T-cell lymphoma. The aim of this study is the identification of the molecular changes (1) associated with the persistence of the pre-malignant clone (2) associated with the activation of co-stimulatory receptors and (3) associated with the emergence of malignant T-cell subclones.

overall_design:
(1) Molecular changes associated with the persistence of the pre-malignant clone.
The gene expression profile of CD3- CD4+ T-cells isolated at diagnosis of three LV-HES patients (chronic disease phase) was compared to the gene expression profile of CD4+ T-cells isolated from four healthy controls. Patients samples were P1 (yr.0), P2 (yr.0) and P3 (yr.0).
For P1 (yr.0), three experimental replicates were processed: P1.a (yr.0), P1.b (yr.0) and P1.c (yr.0). For P2 (yr.0), two experimental replicates were processed: P2.a (yr.0) and P2.b (yr.0). For P3 (yr.0), two experimental replicates were processed: P3.a (yr.0) and P3.b (yr.0).
Healthy controls were D1, D2, D3 and D4.

(2) Molecular changes associated with the activation of co-stimulatory receptors.
The gene expression profile of CD3- CD4+ T-cells isolated at diagnosis from three LV-HES patients (chronic disease phase) and activated in vitrowith a combination of rhIL2 and anti-CD2/CD28 antibodies was compared to the gene expression profile of the same CD3- CD4+ T-cell populations, cultured without activation.
In vitro stimulated patients samples were P1 (yr.0) Stim, P2 (yr.0) Stim and P3 (yr.0) Stim. For P1 (yr.0) Stim, three experimental replicates were processed: P1.a (yr.0) Stim, P1.b (yr.0) Stim and P1.c (yr.0) Stim. For P2 (yr.0) Stim, two experimental replicates were processed: P2.a (yr.0) Stim and P2.b (yr.0) Stim. For P3 (yr.0) Stim, two experimental replicates were processed: P3.a (yr.0) Stim and P3.b (yr.0) Stim.
Unstimulated patients samples were P1 (yr.0), P2 (yr.0) and P3 (yr.0). For P1 (yr.0), three experimental replicates were processed: P1.a (yr.0), P1.b (yr.0) and P1.c (yr.0). For P2 (yr.0), two experimental replicates were processed: P2.a (yr.0) and P2.b (yr.0). For P3 (yr.0), two experimental replicates were processed: P3.a (yr.0) and P3.b (yr.0).

(3) Molecular changes associated with the emergence of malignant T-cell subclones.
The gene expression profiles of CD3- CD4+ T-cells isolated at three stages in patient 1’s clinical progression were assessed: two chronic disease stages were at diagnosis P1 (yr.0) and follow up P1 (yr.+4) and one T-lymphoma stage was at follow up P1 (yr.+6).
For P1 (yr.0), three experimental replicates were processed: P1.a (yr.0), P1.b (yr.0) and P1.c (yr.0). For P1 (yr.+4), three experimental replicates were processed: P1.a (yr.+4), P1.b (yr.+4) and P1.c (yr.+4). For P1 (yr.+6), three experimental replicates were processed: P1.a (yr.+6), P1.b (yr.+6) and P1.c (yr.+6).

first: Marie

last: Ravoet

first: Catherine

last: Sibille

first: Chunyan

last: Gu

first: Myriam

last: Libin

first: Benjamin

last: Haibe-Kains

first: Christos

last: Sotiriou

first: Michel

last: Goldman

first: Florence

last: Roufosse


title: Solexa sequencing of small RNAs in Chinese Amphioxus

city: Shenzhen

first: li

last: qibin

address: Beishan, Yantian

country: China

institute: Beijing Genomic Institute

summary:
We applied Solexa sequencing technology to identify Amphioxus microRNA genes. We identified 113 amphioxus microRNA genes, 55 were conserved across species and coded for 45 nonredundant mature miRNAs, whereas 58 were amphioxus-specific and accounted for 53 mature miRNAs. Analyzing the evolutionary history of amphioxus miRNAs, we found amphioxus possessed many miRNAs that are also found in chordates and vertebrates, suggesting these miRNAs may represent cephalochordate lineage innovation. By detailed comparison of the miRNA phylogenetic histories, we further found that amphioxus is more vertebrate-like than tunicates.

Keywords: Small RNA

overall_design: 18-30 nt small RNAs from 12 adult Chinese Amphioxus were sequenced in one Solexa lane.

first: Xi

last: Chen

first: Qibin

last: Li


title: Genome-wide demethylation of Arabidopsis endosperm

status: Public on Jun 16 2009

city: Berkeley

first: Toshiro

last: Nishimura

address: 211 Koshland Hall

department: Plant and Microbial Biology

institute: University of California at Berkeley

summary:
Parent-of-origin-specific (imprinted) gene expression is regulated in Arabidopsis thaliana endosperm by cytosine demethylation of the maternal genome mediated by the DNA glycosylase DEMETER, but the extent of the methylation changes is not known.  Here we show that virtually the entire endosperm genome is demethylated, coupled with extensive local non-CG hypermethylation of siRNA-targeted sequences.  Mutation of DEMETER partially restores endosperm CG methylation to levels found in other tissues, indicating that CG demethylation is specific to maternal sequences.  Endosperm demethylation is accompanied by CHH hypermethylation of embryo transposable elements.  Our findings demonstrate extensive reconfiguration of the endosperm methylation landscape that likely reinforces transposon silencing in the embryo.

Keywords: Epigenetics; bisulfite sequencing

overall_design:
Examination of DNA methylation in four Arabidopsis tissues
Description of processed data and raw data file contents can be found in the attached README.txt file

first: Tzung-Fu

last: Hsieh

first: Christian

last: Ibarra

first: Pedro

last: Silva

first: Assaf

last: Zemach

first: Leor

last: Williams

first: Robert

middle: L

last: Fisher

first: Daniel

last: Zilberman


title: Detection of single nucleotide variations in expressed exons of the human genome using RNA-Seq

status: Public on Jun 15 2009

city: Cincinnati

first: Iouri

last: Chepelev

state: OH

address: 3200 Vine St

institute: US Department of Veterans Affairs Medical Center

summary: Whole genome re-sequencing is still a costly method to detect genetic mutations that lead to altered forms of proteins and may be associated with disease development. Since the majority of disease-related single nucleotide variations (SNVs) are found in protein-coding regions, we propose to identify SNVs in expressed exons of the human genome using the recently developed RNA-Seq technique. We identify 12,176 and 10,621 SNVs, respectively, in Jurkat T cells and CD4+ T cells from a healthy donor. Interestingly, our data show that one copy of the TAL-1 protooncogene has a point mutation in 3’ UTR and only the mutant allele is expressed in Jurkat cells. We provide a comprehensive dataset for further understanding the cancer biology of Jurkat cells. Our results indicate that this is a cost-effective and efficient strategy to systematically identify SNVs in the expressed regions of the human genome.

overall_design: RNA-Seq experiments for two samples: CD4+ T cells from a healthy donor and Jurkat T cells. There are 8 lanes of data for each sample.

first: Gang

last: Wei

first: Qiangsong

last: Tang

first: Keji

last: Zhao


title: Drosophila OSS cell small RNA libraries

status: Public on Jun 09 2009

city: New York

first: Nicolas

last: Robine

address: 1275 York Avenue, Box 252

department: Developmental Biology

institute: Memorial Sloan Kettering Cancer Center

summary:
High-throughput sequencing of Drosophila melanogaster small RNAs from OSS cells.
total RNA, ~18-26nt RNAs isolated using PAGE, ligation to adapters requires 5' monophosphate and 3' OH

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: Small RNAs were sequenced from D. melanogaster OSS cells. Raw sequences were clipped by 3' linker sequences recognition, and select clipped sequences longer than 18 nt.

first: Nelson

last: Lau

first: Eric

middle: C

last: Lai


title: An Expression Alteration of Small RNA Profiling in Autotriploids Derived from Rice Twin-Seedlings

city: Wuhan

first: hai

last: peng

address: College of Life Sciences, Jianghan University

institute: Jianghan University

summary: Although closely related to plant evolution, polyploid sRNAs (small RNAs) were seldom studied with experiments, especially on genome-wide range. In this study, a rice twin-seeding (two seedlings from the same grain) line SARII-658 was employed to isolate autotriploids. Those autotriploids possess unique merits to study the two interesting topics: (i) natural polyploids; (ii) the autonomy of epigenetic variations (i.e. sRNAs) from genome doubling per se. sRNA libraries were prepared from those diploid-autotriploid twin-seedlings and then were sequenced to produce 13,308,226 short sequence reads in total. Out of the 35,429 miRNA genes and siRNA clusters, 1,547 (4.36%) changed their expression levels for two folds or above after genome doubling (thereinafter, referred those sRNAs as ploidy-sensitive). The expression-unregulated sRNAs (3.71%) were far more than the downregulated ones (0.64%), suggesting that the negative regulation of the coding gene by sRNA increase is more prevalent than  the positive regulation during genome doubling. The expression of sRNAs were obviously increased and biased to accumulate toward centromeric and heterochromatic regions, suggesting that sRNAs should play a role in repressing transposition activity during genome doubling. Except transposition activity, the targets of those ploidy-sensitive sRNAs involved many kinds of gene function categories, suggesting an overall regulation of sRNAs to polyploid biological processes. Findings from this study will provide theoretical bases for elucidating epigenetic mechanism of plant and sRNA evolution via genome doubling.

overall_design: Examination of 2 different small RNA expression profilings in 2 ploidy plants.

first: Hai

last: Peng


title: High resolution analysis of epigenetic changes associated with X inactivation

status: Public on Jun 06 2009

city: Nijmegen

last: Stunnenberg

address: Geert Grooteplein 28

department: Department of Molecular Biology (274)

institute: Radboud University

type: Expression profiling by genome tiling array

type: Genome binding/occupancy profiling by genome tiling array

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Transcriptome of wild type N2 worms at embronic stage and starved L1 stage

status: Public on Jun 11 2009

city: New Haven

first: Flora

last: Vaccarino

address: 230 South Frontage Road

institute: Yale University

summary:
Transcription factors are key components of regulatory networks that control development, as well as the response to environmental stimuli.  We have established an experimental pipeline in C. elegans that permits global identification of the binding sites for transcription factors using chromatin immunoprecipitation and deep sequencing.  We describe and validate this strategy, and apply it to the transcription factor PHA-4, which plays critical roles in organ development and starvation stress response.  We defined binding sites for PHA-4 during formation of the embryonic pharynx, and in starved larvae.  PHA-4 binds to thousands of locations throughout the genome.  These sites shift dramatically between embryos and larvae, from developmentally regulated genes to genes involved in metabolism. We acquired transcriptome of wild type N2 worms at embyonic and starved L1 stages using RNA-seq method in comparing to the ChIP-seq experiment to study transcription factor binding. The wild type worms were cultured the same as the transgenic worms for transcription factor binding experiment.

Keywords: Transcriptome Analysis

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: Two development stages embryo and starved larvae each contains 3 technical replicates are reported

first: Mei

last: Zhong

first: Debasish

last: Raha


title: Distinctive Chromatin in Human Sperm Packages Genes that Guide Embryo Development

city: Salt Lake City

first: Bradley

middle: R.

last: Cairns

state: UT

address: 2000 Circle of Hope

department: Oncological Sciences

institute: HHMI and Huntsman Cancer Institute


title: Loss of H3K27 monomethylation leads to DNA-methylation-independent release of gene silencing

status: Public on Jun 01 2009

city: Los Angeles

first: matteo

last: Pellegrini

address: 621 Charles Young Drive South

department: MCD Biology

institute: UCLA

summary: Constitutive heterochromatin in Arabidopsis thaliana is marked by repressive chromatin modifications including DNA methylation, histone 3 dimethylation at lysine 9 (H3K9me2), and monomethylation at lysine 27 (H3K27me1). The enzymes catalyzing DNA methylation and H3K9me2 have been identified and mutations in these proteins lead to the reactivation of silenced heterochromatic elements. The enzymes responsible for heterochromatic H3K27me1, in contrast, remain unknown. Here we show that the divergent SET-domain proteins ARABIDOPSIS TRITHORAX-RELATED PROTEIN5 (ATXR5) and ATXR6 exhibit H3K27 monomethyltransferase activity and double mutants have reduced H3K27me1 in vivo and show partial heterochromatin decondensation. atxr5 atxr6 plants also show transcriptional activation of repressed heterochromatic elements. Interestingly, H3K9me2 and DNA methylation are unaffected in the double mutant. These results indicate that ATXR5 and ATXR6 form a novel class of H3K27 methyltransferases and that H3K27me1 represents a new pathway required for transcriptional repression in Arabidopsis.

overall_design: Comparison of methylation in wild type and ATXR6/ATXR6 mutants

first: Scott

last: Michaels

first: Matteo


title: Genome-wide Analysis of Histone Methylations in Memory CD8+ T Cells

city: Charlottesville

first: Chongzhi

last: Zang

institute: University of Virginia

summary: Histone methylations play a major role in regulating the chromatin state and gene expression, yet little is known about their involvement in differential gene expression and function of memory CD8 T cells.  Here, we report a genome-wide analysis of two histone H3 methylations (H3K4me3 and H3K27me3) and gene expression in naïve, central (TCM) and effector (TEM) memory CD8 T cells.  Analysis of 16,314 annotated genes in CD8 T cell subsets revealed that gene expression were positively correlated with the levels of H3K4me3 and negatively correlated with the levels of H3K27me3 in these gene loci.  The correlation between differential H3K4me3 orH3K27me3 levels with gene expressions in memory CD8 T cells displayed four distinct modes: repressive, active, poised, and bivalent, reflecting their complex regulation and different function of these genes.  Furthermore, accessible chromatin states of different gene loci were preferentially influenced by different histone modifications as demonstrated here high levels of H3K9ac found in active gene loci without high levels of H3K4me3. These findings reveal a histone methylation based complex regulation of differential gene expression in memory CD8 T cells.  Thus, change of chromatin structure mediated by histone methylation may serve a fundamental basis for the rapid transcriptional response of memory CD8 T cells.

overall_design: genome-wide analysis of two histone H3 methylations (H3K4me3 and H3K27me3) in naïve, central (TCM) and effector (TEM) memory CD8 T cells.

first: Yasuto

last: Araki

first: Zhibin

last: Wang

first: William

last: Wood

first: Dustin

last: Schones

first: Kairong

last: Cui

first: Tae-Young

last: Roh

first: Brad

last: Lhotsky

first: Robert

last: Wersto

first: Weiqun

first: Kevin

last: Becker

first: Nan-ping

last: Weng


title: Novel microRNAs uncovered by deep sequencing of small RNA transcriptomes in T. aestivum and B. distachyon

status: Public on Jun 08 2009

city: beijing

first: bo

last: wei

institute: caas

summary:
The small RNA transcriptomes of bread wheat (Triticum aestivum L.) and its emerging model (Brachypodium distachyon (L.) Beauv) were obtained by using deep sequencing technology. Small RNA compositions were analyzed in these two species. In addition to 70 conserved microRNAs (miRNA) from 25 families, 23 novel wheat miRNAs were identified. For Brachypodium, 12 putative miRNAs were predicted from a limited number of ESTs, of which one was a potential novel miRNA. Also, 94 conserved miRNAs from 28 families were identified in this species. Expression validation was performed for several novel wheat miRNAs. RNA ligase-mediated 5' RACE experiments demonstrated their capability to cleave predicted target genes including three disease resistant gene analogs. Differential expression of miRNAs was observed between Brachypodium vegetative and reproductive tissues, suggesting their different roles at the two growth stages. Our work significantly increases the novel miRNA numbers in wheat and provides the first set of small RNAs in Brachypodium distachyon.

Keywords: Small RNA

sample_organism:
Triticum aestivum
Brachypodium distachyon

overall_design: One wheat small RNA library (Tae) and two Brachypodium small RNA libraries (BdR and BdV) were sequenced.


title: Distinctive Chromatin in Human Sperm Packages Genes that Guide Embryo Development: ChIP-seq

first: Bradley

summary: As nucleosomes are widely replaced by protamine in mature human sperm, epigenetic  contributions to embryo development appear limited.  However, our genome-wide approaches find nucleosomes at low levels genome-wide, but also significantly enriched at imprinted gene clusters, miRNA clusters, HOX gene clusters, and the promoters of other developmental transcription and signaling factors. Developmental promoters were often DNA hypomethylated, and bore histone modifications localized to discrete locations: H3K4me2 is enriched at certain developmental promoters, whereas large blocks of H3K4me3 localize to a subset of developmental promoters, regions in HOX loci, certain non-coding RNAs, and generally to paternally-expressed imprinted loci.  In contrast, H3K4me3 is generally absent at paternally-repressed imprinted loci. Interestingly, repressive H3K27me3 is enriched at many developmental promoters that lack early expression in embryos, with significant overlap with bivalent (H3K4me3/H3K27me3) promoters in ES cells. Taken together, epigenetic marking in sperm is extensive, and correlated with developmental regulators.

overall_design:
*Use of ChIP-seq to identify regions enriched for histone, and several histone variants in human sperm.

*Histone Localization: Chromatin was prepared from 40 million sperm as described previously (1) in the absence of crosslinking reagent, treated with sequential and increasing MNase (10U-160U), and centrifuged to sediment protamine-associated DNA, releasing mononucleosomes. The pooled mononucleosomes were gel purified (~140-155 bp) for sequencing and array analysis.  Mononucleosomes for ChIP-seq were sequenced from a single donor (3 replicas combined) or a donor pool (4 donors each ~6 million reads) and data was normalized to 20 million input control reads (pooled donor sperm genomic DNA).

*Chromatin IP and Preparation for Genomics Methods: ChIP methods were as described previously (2) but were performed without a cross linking agent and slight modifications to the salt levels, 250 mM NaCl, 200 mM LiCl, and replaced the TE wash with 150 mM PBS wash. A subset of the modifications H3K4me3, H3K27me3 and H2A.z were done from a donor pool and only analyzed by Solexa. Each IP was repeated three time and the three replicas were pooled. For sequencing, DNA lengths corresponding to mononucleosomes with adapters  (220-280 bp) were gel purified after the addition of the Illumina adaptors. The sequencing data was normalized to histone donor pool (D1 was subsampled to 5million and combined with the reads from D2-D4).

1. Zalenskaya, I.A., Bradbury, E.M. & Zalensky, A.O. Chromatin structure of telomere domain in human sperm. Biochem Biophys Res Commun 279, 213-8 (2000).
2. Gordon, M. et al. Genome-Wide Dynamics of SAPHIRE, an Essential Complex for Gene Activation and Chromatin Boundaries. Mol Cell Biol 27, 4058-69 (2007).

Samples corresponding to each GSE15690_Seq* processed data supplementary file:

GSE15690_Seq_H2AZ.txt:
GSM392695

GSE15690_Seq_H3K27Me3.txt:
GSM392699, GSM392700

GSE15690_Seq_H3K4Me3.txt:
GGSM392696-GSM392698

GSE15690_Seq_HistoneMnase_D1.txt:
GSM392701-GSM392707, GSM392714-GSM392716

GSE15690_Seq_HistoneMnase_PooledDonor.txt:
GSM392708-GSM392713, GSM392717-GSM392720

first: S

middle: S

last: Hammoud

first: David

last: Nix

first: Haying

last: Zhang

first: Douglas

middle: T

last: Carrell

middle: R


title: small RNA from S2 cells (S2-NP)

status: Public on Apr 30 2009

first: Nicolas

last: Robine

summary:
High-throughput sequencing of Drosophila melanogaster small RNAs from S2 cells.
total RNA, ~18-26nt RNAs isolated using PAGE, ligation to adapters requires 5' monophosphate and 3' OH

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: Small RNAs were sequenced from D. melanogaster S2 cells obtained from Norbert Perrimon's lab. Raw sequences were clipped by 3' linker sequences recognition, and select clipped sequences longer than 18 nt.

first: Eric

last: Lai


title: High resolution analysis of epigenetic changes associated with X inactivation, ChIP-Seq

last: Stunnenberg

summary: This study describes the epigenetic profiling of the X chromosome during X inactivation. It includes H3K4me3 and H3K27me3 ChIP-Seq profiles of male (E14) and female (LF2 and XT67E1) mouse ES cells, together with their differentiated derivatives (either 4d atRA or 10d EB). It also includes ChIP-chip profiles around the Xic on chromosome X of H3K4me3, H3K27me3, H3K9me2, H3K36me3, Pol II, TBP, H3-Core as well as expression, using male (E14) and female (LF2) mouse ES cells, together with their differentiated derivatives (either 4d atRA or 10d EB).

overall_design: Examination of two different histone modifications in 3 cell lines under 3 conditions using ChIP-Seq. Examination of five different histone modifications two transcription factors and gene expression under three conditions in 2 cell lines using ChIP-chip.

first: Hendrik

last: Marks

first: Jennifer

last: Chow

first: Kees-Jan

last: Françoijs

first: Sergei

last: Denissov

first: Neil

last: Brockdorff

first: Edith

last: Heard

middle: G


title: Specialized piRNA pathways act in germline and somatic tissues of the Drosophila ovary

first: Colin

middle: Dean

last: Malone

address: NYUMC, Skirball Institute, 540 1st Ave, 4-10

department: Skirball Institute - Developmental Genetics

institute: NYU Medical Center

summary:
Here, we analyzed nine Drosophila piRNA pathway mutants for their impacts on both small RNA populations and the sub-cellular localization patterns of Piwi proteins. We find that distinct piRNA pathways with differing components function in ovarian germ and somatic cells.

Keywords: Epigenetics

sample_organism:
Drosophila erecta
Drosophila melanogaster

overall_design: 20 libraries were analyzed, with 9 being control samples (heterozygotes).

middle: D

first: Gregory

last: Hannon


title: Drosophila melanogaster small RNA sequence: Kc167 cells (Rubin lab)

city: Toronto

first: DCC

last: modENCODE

state: Ontario

address: MaRS Centre, South Tower, 101 College Street, Suite 800

country: Canada

institute: Ontario Institute for Cancer Research

summary:
modENCODE_submission_2374
This submission comes from a modENCODE project of Eric Lai. For full list of modENCODE projects, see http://www.genome.gov/26524648
Project Goal: We plan to generate a comprehensive catalog of expressed and functional microRNAs, and generate biological evidence for their regulatory activity. We plan also to delineate the primary transcription units of microRNA genes. Finally, we plan to annotate other classes of non-miRNA expressed small RNAs, as least some of which may define novel classes of small RNA genes.
For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: EXPERIMENT TYPE: RNA-seq. BIOLOGICAL SOURCE: Cell Line: Kc-Rubin; Tissue: embryo-derived cell-line; Developmental Stage: late embryonic stage; Genotype: se/e; Sex: Female;  EXPERIMENTAL FACTORS: Cell Line Kc-Rubin; Replicate type: biological


title: B. anthracis transcriptome analysis using Illumina and SOLiD sequencing

status: Public on May 01 2009

city: Atlanta

first: Nicholas

last: Bergman

state: GA

address: 310 Ferst Dr., Rm 231

department: Biology

institute: Georgia Institute of Technology

summary: Although gene expression has been studied in bacteria for decades, many aspects of the bacterial transcriptome remain poorly understood.  Transcript structure, operon linkages, and absolute abundance information all provide valuable insights into gene function and regulation, but none has ever been determined on a genome-wide scale for any bacterium.  Here we report the first comprehensive and unbiased definition of a bacterial transcriptome under various growth conditions.  Using a high throughput sequencing approach, we mapped transcript start sites and operons throughout the genome, and enhanced existing genome annotations.  Further, our data provide estimates of absolute transcript abundance, and suggest there is significant transcriptional heterogeneity within a clonal, synchronized bacterial population.  Overall, our results offer an unprecedented view of gene expression and regulation in a bacterial cell.

overall_design:
Includes:

Comparison of transcriptomes under 2 different atmospheric conditions using Illumina sequencing (2 Samples)

and

Comparison of transcriptomes in different growth phases under 2 different atmospheric conditions using SOLiD sequencing (6 Samples).

first: Karla

last: Passalacqua

first: Anjana

middle: Varadarajan

first: Brian

last: Ondov

last: Okou

first: Michael

middle: E

last: Zwick

middle: H


title: Gene expression and UH2A ChIP-Seq binding analysis in Bmi1 knock-out and wild type MEFs

status: Public on May 20 2009

city: RTP

first: Raja

last: Jothi

state: NC

address: 111 TW Alexander Drive; A314

department: National Institute of Environmental Health Sciences

institute: National Institutes of Health


title: Genome-wide maps of chromatin state in staged Drosophila embryos, ChIP-seq

status: Public on May 12 2009

city: Chicago

first: Kevin

middle: P.

last: White

state: IL

address: 900 E. 57th STR. 10th FL.

department: Institute for Genomics and Systems Biology

institute: University of Chicago

summary:
This is a dataset generated by the Drosophila Regulatory Elements modENCODE Project led by Kevin P. White at the University of Chicago. It contains ChIP-seq data generated on Solexa Genome Analyzer for 6 Histone modifications (H3K9me3, H3K27me3, H3K4me3, H3K4me1, H3K27Ac, H3K9Ac), PolII and CBP/p300. Each factor has been studied for 12 different time-points of Drosophila development.

Keywords: Epigenetics

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: For each combination of time-point and antibody, triplicate ChIP experiments have been performed and hybridized on Agilent 244K arrays. The hybridizations have been verified by sequencing one replicate of IP and one replicate of Input following Solexa sequencing procedure.

first: Nicolas

last: Negre

first: Carolyn

last: Morrison

first: Parantu

middle: K

last: Shah

first: Nick

last: Bild


title: Loss of the Mili-interacting Tdrd1 activates transposons and alters Mili-associated small RNA profile

status: Public on May 24 2009

city: Gneveva

first: Ramesh

last: Pillai

address: 30, Quai Ernest-Ansermet

department: Department of Molecular Biology

country: Switzerland

institute: University of Geneva

summary: Piwi proteins and their associated Piwi-interacting RNAs (piRNAs) are implicated in transposon silencing in the murine germline. There is currently little information on additional proteins in the murine Piwi complex and how they might regulate the entry of transcripts that accumulate as piRNAs in the Piwi ribonucleoprotein (piRNP). We isolated Mili-containing complexes from adult mouse testes and identified Tudor domain-containing protein 1 (Tdrd1) as a factor specifically associated with the Mili piRNP throughout spermatogenesis. Complex formation is promoted by the recognition of symmetrically dimethylated arginines at the N-terminus of Mili by the Tudor domains of Tdrd1. Similar to a Mili mutant, mice lacking Tdrd1 show derepression of L1 transposons accompanied by a loss of DNA methylation of their regulatory elements, and delocalization of Miwi2 from the nucleus to the cytoplasm. Finally, we show that Mili piRNPs devoid of Tdrd1 accept the entry of abundant cellular transcripts into the piRNA pathway and accumulate piRNAs with a profile that is drastically different from the wild-type. Our data suggests that Mili recruits Tdrd1 to ensure only the entry of transcripts that contribute to its normal piRNA pool.

overall_design: Immunoprecipitation of Tdrd1 or Mili from mouse testes isolated from wild-type or tdrd1 ko animals. Associated RNAs were sequenced in this study.

last: Reuter

first: Alexander

last: Stark

first: Shinichiro

last: Chuma

first: Takashi

last: Tanaka

first: Thomas

last: Franz


title: High-throughput sequencing of small RNAs in chicken somite embryos

status: Public on Apr 28 2009

city: Oxford

first: Helio

last: Pais

address: Jonh Radcliffe Hospital

department: Weatherall Institute of Molecular Medicine

institute: University of Oxford

overall_design: Dissected tissue white leghorn chicken embryos was disaggregated and RNA extracted using the miRVana kit (Ambion). Small RNA fraction between 19 and 24 nt was isolated from 15% denaturing polyacrylamide gel and 15 micro gram was ligated to Solexa adaptors (Illumina) without de-phosphorylating and re-phosphorylating. The short RNAs were converted to DNA by RT-PCR following the Illumina protocol.

first: Tina

last: Rathjen

first: Dylan

last: Sweetman

first: Vincent

last: Moulton

first: Andrea

last: Munsterberg

first: Tamas

last: Dalmay


title: Genome wide uH2A localization analysis highlights Bmi1-dependent deposition of the mark at repressed genes.

first: Raja

last: Jothi

summary:
Polycomb group (PcG) proteins control organism development by regulating the expression of developmental genes. Transcriptional regulation by PcG proteins is achieved at least partly through the PRC2-mediated methylation on lysine 27 of histone H3 (H3K27) and PRC1-mediated ubiquitylation on lysine 119 of histone H2A (uH2A). As an integral component of PRC1, Bmi1 has been demonstrated to be critical for H2A ubiquitylation. Here we report genome wide localization of the Bmi1-dependent uH2A mark in MEF cells. Gene promoter averaging analysis indicates a peak of uH2A just inside the transcription start site (TSS) of well annotated genes.  This peak is enriched at promoters containing the H3K27me3 mark and represents the least expressed genes in WT MEF cells.  In addition, peak finding reveals regions of local uH2A enrichment throughout the mouse genome, including almost 700 gene promoters.  Genes with promoter peaks of uH2A exhibit lower level expression when compared to genes that do not contain promoter peaks of uH2A.  Moreover, we demonstrate that genes with uH2A peaks have increased expression upon Bmi1 knockout.  Importantly, local enrichment of uH2A is not limited to regions containing the H3K27me3 mark.  We describe the enrichment of H2A ubiquitylation at high density CpG promoters and provide evidence to suggest that DNA methylation may be linked to uH2A at these regions.  Thus, our work not only reveals Bmi1-dependent H2A ubiquitylation but also suggests that uH2A targeting in differentiated cells may employ a different mechanism from that in ES cells.

Keywords: Epigenetics

overall_design: Examination of uH2A binding in WT and Bmi1 null MEF cells.

first: Eric

middle: M

last: Kallin

first: Ru

last: Cao

first: Kai

last: Xia

first: Kairong

last: Cui

first: Keji

last: Zhao

first: Yi

last: Zhang


title: small RNA from Drosophila melanogaster - cell line Kc167

first: Nicolas

last: Robine

summary:
High-throughput sequencing of Drosophila melanogaster small RNAs from Kc167 cells. total RNA, ~18-26nt RNAs isolated using PAGE, ligation to adapters requires 5' monophosphate and 3' OH

Keywords: small RNA discovery and profiling

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: Small RNAs were sequenced from D. melanogaster Kc167 cells. Raw sequences were clipped by 3' linker sequences recognition, and select clipped sequences longer than 18 nt.

first: E

last: Lai


title: Identification of transcription factor MAB-5 binding sites

status: Public on Apr 11 2009

first: Flora

last: Vaccarino

department: Child Study Center

summary:
We identified the DNA binding sites of MAB-5 in C. elegans. MAB-5 was C-terminally tagged with GFP and the expression pattern was examined through fluorescent microscopy. The binding sites were determined using chromatin immunoprecipitation with anti-GFP antibody followed by illumina high-throughput sequencing (ChIP-seq).

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: Identification of MAB-5 binding sites at L3 stage

first: Mei

last: Zhong

first: Wei

last: Niu


title: Genome-wide identification of the occupancy of RNA polymerase II (POL II) at L4/young adult stage in C. elegans

summary:
We acquied the RNA polymerase II binding profiles in ama-1 transgenic worms. The large subuint of POL II, AMA-1, was C-terminally tagged with GFP, the binding regions were determined using chromatin immunoprecipitation with anti-GFP and anti-polymerase II antibodies followed by illumina high-throughput sequencing (ChIP-seq).

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: RNA polymerase II binding profiles at L4/young adult stage were acquied with anti-native protein antibody (8WG16) and anti-epitope (GFP) antibody


title: Allelic imbalance sequencing reveals that single-nucleotide polymorphisms frequently alter microRNA-directed repression

status: Public on Apr 26 2009

city: Seoul

first: Jinkuk

last: Kim

address: 50 Irwon-dong, Gangnam-gu

country: South Korea

institute: Samsung Medical Center

summary: Genetic changes that help explain the differences between two individuals might create or disrupt sites complementary to microRNAs (miRNAs), but the extent to which such polymorphic sites influence miRNA-mediated repression is unknown. Here, we describe a method to measure mRNA allelic imbalances associated with a regulatory site found in mRNA transcribed from one allele but not found in that transcribed from the other. Applying this method, called allelic imbalance sequencing, to sites for three miRNAs (miR-1, miR-133 and miR-122) provided quantitative measurements of repression in vivo without altering either the miRNAs or their targets. A substantial fraction of polymorphic sites mediated repression in tissues that expressed the cognate miRNA, and downregulation was correlated with site type and site context. Extrapolating these results to the other broadly conserved miRNAs suggests that when comparing two mouse strains (or two human individuals), polymorphic miRNA sites cause expression of many genes (often hundreds) to differ.

overall_design: There are four distinct samples (pool1-4), each of which is a pool of ~60 distinct PCR or RT-PCR products. The samples were loaded into different segments of the same pyrosequencing plate. Three sequencing runs (run1-3) were performed for each sample pool. Any amplicons of the four pools are for miR-1, miR-122, miR-133 or control sites. To control for allele-specific PCR bias, genomic DNA was also used as a template for generating amplicons.

first: David

last: Bartel


title: Computational and Analytical Framework for Small RNA Profiling by High-Throughput Sequencing (reproducibility)

status: Public on Jun 03 2009

city: Saint Louis

first: James

last: Carrington

address: 975 North Warson Road

institute: Donald Danforth Plant Science Center

summary: The purpose of this work was to describe a computational and analytical methodology for profiling small RNA by high-throughput sequencing.  The datasets here were used to assess the reproducibility of small RNA datasets produced using Illumina sequencing-by-synthesis technology (SBS).

overall_design: We analyzed the reproducibility of small RNA SBS datasets by comparing libraries generated for biological replicates (rep1 and rep2) and technical replicates (rep2 and rep3).

first: Noah

last: Fahlgren

first: Kristin

last: Kasschau


title: RNA polymerase occupancy at embryonic stage and starved L1 stage in pha-4 transgenic worms

summary:
We acquired RNA Polymerase II (POL II) binding profiles at embyonic and starved L1 stages in pha-4 transgenic worms. Transcription factor gene pha-4 was tagged with GFP and their expression examined throughout the C. elegans life cycle. The POL II occupancy were determined using chromatin immunoprecipitation with anti-POL II antibodies followed by illumina high-throughput sequencing (ChIP-seq).

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: Two growth stages of pha-4 transgenic worm were studied including embryo and starved L1.


title: Identification of transcription factor DAF-16 binding sites in C. elegans

summary:
We identified the DNA binding sites of DAF-16 in transgenic line TJ356 (daf-16::gfp) that carries high-copy number daf-16. The expression pattern was examined through fluorescent microscopy. The binding sites were determined using chromatin immunoprecipitation with anti-GFP antibody and anti-POL II antibody followed by illumina high-throughput sequencing (ChIP-seq).

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: Identification of DAF-16 binding sites at L4/Young adult stage when DAF-16 was highly overexpressed


title: modENCODE RNA-Seq of Drosophila cell lines Kc167, CME_W1_Cl.8+, S2-DRSC and ML-DmBG3-c2

status: Public on Apr 08 2009

city: Farmington

first: Brenton

last: Graveley

address: 400 Farmington Avenue

department: Genetics and Genome Sciences

institute: University of Connecticut Health Center

summary:
Deep Sequencing of mRNA from the Drosophila melanogaster cell lines Kc167, CME_W1_Cl.8+, S2-DRSC and ML-DmBG3-c2.

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: Analysis of poly(A)+ RNA from 2 biological replicates of Kc167, CME_W1_Cl.8+, S2-DRSC and ML-DmBG3-c2 cell lines.

first: Li

last: Yang

first: Michael

last: Duff

first: Peter

last: Cherbas

first: Lucy

first: Sue

last: Celniker


title: Computational and Analytical Framework for Small RNA Profiling by High-Throughput Sequencing

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Computational and Analytical Framework for Small RNA Profiling by High-Throughput Sequencing (standards)

summary: The purpose of this work was to describe a computational and analytical methodology for profiling small RNA by high-throughput sequencing.  The datasets here were used to develop synthetic oligoribonucleotides as spike-in standards.

overall_design: We assessed the use of synthetic oligoribonucleotide standards as spike-in controls.  These standards can be used to set an objective standard against which to compare samples.  Standards were added to the total RNA (100 ug) in the following amounts: Std2 (TATATGCAAGTCCGGCCATAC) 0.01 pmol, Std3 (TAGCTAACGCATATCCGCATC) 0.1 pmol, Std6 (TGAAGCTGACATCGGTCATCC) 1.0 pmol.


title: Small RNAs in Arabidopsis hybrid siliques

status: Public on Mar 24 2009

city: Cambridge

first: Rebecca

middle: A

last: Mosher

address: Downing Street

department: Plant Sciences

institute: Cambridge University

summary:
Five days post-fertilization siliques were collected for RNA extraction. After purification, small RNA libraries were generated for high throughput sequencing. Resulting sequences were analyzed to determine the contribution of the paternal genome to the small RNA transcriptome.

Keywords: Epigenetics

overall_design: 4 unique samples. small RNAs purified from Arabidopsis siliques 5 days after pollination. Samples were selfed or hybrid crosses between ecotypes Columbia (Col) and Landsberg erecta (Ler).

first: Charles

middle: W

last: Melnyk

first: Krystyna

last: Kelly

first: Ruth

last: Dunn

middle: J

last: Studholme

last: Baulcombe


title: Identification of transcription factor DPY-27 binding sites

status: Public on Apr 03 2009

first: Flora

last: Vaccarino

summary:
We identified the DNA binding sites of DPY-27 in C. elegans. DPY-27 was C-terminally tagged with GFP and the expression pattern was examined through fluorescent microscopy. The binding sites were determined using chromatin immunoprecipitation with anti-GFP antibody followed by illumina high-throughput sequencing (ChIP-seq).

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: Identification of DPY-27 binding sites at embryonic stage

first: Mei

last: Zhong

last: Niu


title: ChIP-Seq of ERalpha and RNA polymerase II defines genes differentially responding to ligands

status: Public on Mar 09 2009

last: Stunnenberg

institute: Radboud University

summary: We used ChIP-Seq to map ERalpha binding sites and to profile changes in RNA polymerase II (RNAPII) occupancy in MCF-7 cells in response to estradiol (E2), tamoxifen or fulvestrant. We identified 10,205 high confidence ERalpha binding sites in response to E2 of which 68% contained an estrogen response element (ERE) and only 7% contained a FOXA1 motif. Remarkably, 596 genes already change significantly in RNAPII occupancy (59% up and 41% down) following one hour of E2 exposure. Although pausing of RNA polymerase II occurs frequently in MCF-7 cells (17%) it is only observed on a minority of E2-regulated genes (4%). Tamoxifen and fulvestrant partially reduce ERalpha DNA binding and prevent RNAPII loading on the promoter and coding body on E2-upregulated genes. Both antagonists act differently on E2-downregulated genes. Tamoxifen acts as an agonist, also downregulating these genes while fulvestrant antagonizes E2 induced repression and often increases RNAPII occupancy. Furthermore our data identified genes preferentially regulated by tamoxifen but not by E2 or fulvestrant. Thus, antagonist loaded ERalpha acts mechanistically different on E2-activated and E2-repressed genes.

overall_design: Examination of ERalpha binding sites upon the binding of different ligands and association with transcription via RNAPII occupancy.

first: Henk

first: W

last: Welboren

first: P

last: Span

first: F

last: Sweep

first: M

last: Van Driel

first: E

last: Janssen-Megens

first: S

last: Van Heeringen


title: small RNA from Drosophila melanogaster - adult bodies

city: New York

first: Nicolas

last: Robine

address: 1275 York Avenue, Box 252

institute: Memorial Sloan Kettering Cancer Center

summary:
High-throughput sequencing of Drosophila melanogaster small RNAs from imaginal disc / brain, female body and male body. total RNA, ~18-26nt RNAs isolated using PAGE, ligation to adapters requires 5' monophosphate and 3' OH

Keywords: small RNA discovery and profiling

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: Small RNAs were sequenced from D. melanogaster adult bodies. Raw sequences were clipped by 3' linker sequences recognition, and select clipped sequences longer than 18 nt.

middle: C

last: Lai


title: Genome-wide profiles of STAT1 DNA association using chromatin immunoprecipitation and massively parallel sequencing

status: Public on Mar 23 2009

city: Vancouver

first: Eric

last: Chuah

state: BC

address: 570 West 7th Ave

institute: BC Genome Sciences Centre

summary: We developed a method, ChIP-sequencing (ChIP-seq), combining chromatin immunoprecipitation (ChIP) and massively parallel sequencing to identify mammalian DNA sequences bound by transcription factors in vivo. We used ChIP-seq to map STAT1 targets in interferon stimulated and unstimulated human HeLa S3 cells, and compared the method's performance to ChIP-PCR and to ChIP-chip for four chromosomes. By ChIP-seq, using 15.1 and 12.9 million uniquely mapped sequence reads, and an estimated false discovery rate of less than 0.001, we identified 41,582 and 11,004 putative STAT1-binding regions in stimulated and unstimulated cells, respectively. Of the 34 loci known to contain STAT1 interferon-responsive binding sites, ChIP-seq found 24 (71%). ChIP-seq targets were enriched in sequences similar to known STAT1 binding motifs. Comparisons with two ChIP-PCR data sets suggested that ChIP-seq sensitivity was between 70% and 92% and specificity was at least 95%.

overall_design: Use ChIP-seq to map STAT1 targets in interferon-gamma (IFN-gamma)-stimulated and unstimulated human HeLa S3 cells, and compared the method's performance to ChIP-PCR and to ChIP-chip for four chromosomes

first: G

last: Robertson

last: Hirst

last: Bainbridge

last: Bilenky

first: Y

last: Zhao

first: T

last: Zeng

last: Euskirchen

first: B

last: Bernier

first: R

last: Varhol

first: A

last: Delaney

first: N

last: Thiessen

first: O

middle: L

last: Griffith

last: He

last: Marra

last: Snyder

last: Jones


title: Nutritional control of gene expression during C. elegans L1 arrest and recovery

status: Public on Feb 27 2009

city: Durham

first: L.

middle: Ryan

last: Baugh

address: 4314 French Family Science Center

institute: Duke University


title: Genome-Wide Analysis In Vivo of Translation with Nucleotide Resolution Using Ribosome Profiling

status: Public on Feb 12 2009

city: Berkeley

first: Nicholas

middle: T

last: Ingolia

email: ingolia@berkeley.edu, nick@ingolia.org

address: 1 Barker Hall # 3202

department: Molecular and Cell Biology

institute: University of California, Berkeley

summary: Techniques for systematically monitoring protein translation have lagged far behind methods for measuring mRNA levels. Here we present a ribosome profiling strategy, based on deep sequencing of ribosome protected mRNA fragments, that enables genome-wide investigation of translation with sub-codon resolution. We used this technique to monitor translation in budding yeast under both rich and starvation conditions. These studies defined the protein sequences being translated and found extensive translational control both for determining absolute protein abundance and for responding to environmental stress. We also observed distinct phases during translation involving a large decrease in ribosome density going from early to late peptide elongation as well as wide-spread, regulated initiation at non-AUG codons. Ribosome profiling is readily adaptable to other organisms, making high-precision investigation of protein translation experimentally accessible.

overall_design: Examine replicates of ribosome footprints and mRNA abundance in biological replicates of log-phase growth and acute amino acid starvation

first: Jonathan

middle: S

last: Weissman

first: John

last: Newman

first: Sina

last: Ghaemmaghami


title: ChIP-seq Accurately Predicts Tissue-Specific Activity of Enhancers

status: Public on Feb 11 2009

first: Matthew

middle: James

last: Blow

address: 1 Cyclotron Road

department: Genomics Division

institute: Lawrence Berkeley National Laboratory

summary: Determining the spatial and temporal activity patterns of enhancers remains a challenge in the functional annotation of the human genome.  Here, we performed genome-wide mapping of tissue-specific in vivo binding sites for the enhancer-associated protein p300 and assessed in transgenic mice the utility of this information in identifying enhancers and predicting their activity patterns.  Chromatin immunoprecipitation followed by massively-parallel sequencing was used to identify p300-enriched sites in mouse embryonic day 11.5 (e11.5) forebrain, midbrain, and limb.  In total, 4,686 genomic regions were enriched for p300 in vivo in at least one of these tissues.  To determine whether p300-binding accurately identifies enhancers and predicts their activity patterns, we tested 86 of these regions in a transgenic mouse enhancer assay at e11.5.  In 88% of the cases, p300-enriched sequences were reproducible enhancers, and in 91% of these cases p300 enrichment correctly predicted the tissues in which in vivo activity was observed.  Our results indicate that in vivo mapping of p300 binding to non-coding DNA is a highly effective means for identifying enhancers and their associated spatial activity patterns.

overall_design: Examination of p300 binding in 3 embryonic stage 11.5 mouse tissues

first: Axel

last: Visel

first: Len

last: Pennacchio


title: Genome-wide annotation of small RNAs expressed in HeLa and HepG2 cells

status: Public on Jan 26 2009

city: Cold Spring Harbor

first: Katalin

last: Fejes Toth

state: NY

address: 1 Bungtown Rd

institute: Cold Spring Harbor Labs

summary: We report an applicaton of small RNA sequencing using high throughput next generation sequencing to identify the small RNA content of cell lines. By sequencing over 30 million reads we could identify a new class of small RNAs previousy observed with tiling arrays and mapping to promoter regions of coding genes. We also identified a large number of small RNAs corresponding to internal exons of coding genes. By using different enzymatic treatments and immunoprecipitation experiments, we have determined that both the promoter associated small RNAs as well as ones within the body of the genes bear 5' cap structures.

overall_design: Examination of the expression of small RNAs (<200nt).

first: Katlin

last: Fejes-Toth

first: Philipp

last: Kapranov


title: Identification of transcription factor Pha-4 binding regions

status: Public on Jan 24 2009

city: New Haven

first: Flora

last: Vaccarino

address: 230 South Frontage Road

institute: Yale University

summary:
We identified the DNA binding sites for transcription factors Pha-4 in C. elegans. Transcription factor genes were tagged with GFP and their expression examined throughout the C. elegans life cycle. The binding sites were determined using chromatin immunoprecipitation with anti-GFP antibody followed by illumina high-throughput sequencing (ChIP-seq).

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

first: Mei

last: Zhong


title: High-throughput pyrosequencing of endogenous small RNAs from Arabidopsis thaliana wild-type and ddm1 mutant.

status: Public on Feb 02 2009

city: PARIS

first: Alexis

last: Sarazin

address: 46 rue d'Ulm

department: Ecole Normale Supérieure, Département de Biologie

institute: CNRS UMR 8186 - Biologie Moléculaire des Organismes Photosynthétiques

summary: DNA methylation is essential for silencing transposable elements and some genes in higher eukaryotes, implying that this modification must be tightly controlled. However, accidental changes in DNA methylation can be transmitted through mitosis, as in cancer, or meiosis, leading to epiallelic variation. Here, we demonstrate the existence of an efficient and faithful mechanism that protects against transgenerational loss of DNA methylation in the plant Arabidopsis. This process is specific to the subset of heavily methylated genomic repeats that are targeted by the RNAi machinery, and does not spread into flanking regions. Remethylation is often progressive over two to four sexual generations. This differential and incremental correction of epigenetic defects may preserve genome stability while increasing adaptive opportunities.

overall_design: 2 samples examined: wild type, and ddm1 mutant.

first: Felipe

last: Teixeira

first: Fabiana

last: Heredia

first: François

last: Roudier

first: Martine

last: Boccara

first: Constance

last: Ciaudo

first: C

last: Cruaud

first: J

last: Poulain

first: Maria

last: Berdasco

first: Mario

middle: F

last: Fraga

first: Olivier

last: Voinnet

first: P

last: Wincker

first: Manel

last: Esteller

first: Vincent

last: Colot


title: Drosophila melanogaster S2-GMR cells, second biological replicate

status: Public on Jan 22 2009

first: Nicolas

last: Robine

summary:
High-throughput sequencing of Drosophila melanogaster small RNAs from S2 cells.
total RNA, ~18-26nt RNAs isolated using PAGE, ligation to adapters requires 5' monophosphate and 3' OH

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: Small RNAs were sequenced from D. melanogaster S2 cells obtained from Gerald Rubin's lab. Raw sequences were clipped by 3' linker sequences recognition, and select clipped sequences longer than 18 nt.

first: Eric

last: Lai


title: Epigenetic modifications and their relationships to smRNA and mRNA transcriptomes in maize

first: Xiangfeng

last: Wang

address: 352 OML, 165 Prospect st

department: MCDB

summary:
Genome-wide and organ-specific landscapes of epigenetic modifications and their relationships to mRNA and smRNA transcriptomes in maize
We report an integrated genome-wide analysis of DNA methylation, histone modifications, smRNAs and mRNA transcriptional activity, using maize as a model. We surveyed the epigenomes of the maize inbred line B73 in shoot and root tissue by Illumina/Solexa 1G parallel sequencing after digesting genomic DNA with a methylation-sensitive restriction enzyme and after conducting chromatin immunoprecipitations (ChIP) using antibodies that target specific histone modifications (H3K4me3, H3K9ac, H3K27me3, H3K36me3, respectively). Additionally, we profiled RNA pools (micro RNA (miRNA), siRNA and mRNA) using the same sequencing strategy.

Keywords: Epigenetics, mRNA transcription and small RNAs

overall_design: H3K4me3, H3K9ac, H3K27me3, H3K36me3, DNA methylation and mRNA, small RNA profiled from shoots and roots of 14 day-old maize B73 seedlings

first: Xing-Wang

last: Deng


title: Identification of novel Epstein-Barr Virus miRNA genes from Nasopharyngeal Carcinomas

status: Public on Feb 07 2009

city: Martinsried

first: Gunter

last: Meister

address: Am Klopferspitz 18

department: Center for Integrated Protein Science Munich (CIPSM)

institute: Max Planck Institute of Biochemistry

summary: MicroRNAs (miRNAs) represent a conserved class of small non-coding RNAs that are found in all higher eukaryotes as well as some DNA viruses. MiRNAs are 20-25 nucleotides (nt) in length and have important regulatory functions in biological processes such as embryonic development, cell differentiation, hormone secretion or metabolism. Furthermore, miRNAs have been implicated in the pathology of various diseases including cancer. MiRNA expression profiles not only classify different types of cancer but also may even help to characterize distinct tumor stages, therefore constituting a valuable tool for prognosis. Here we report the miRNA profile of Epstein-Barr Virus (EBV)-positive nasopharyngeal carcinoma (NPC) tissue samples characterized by cloning and sequencing. We find that all EBV miRNAs from the BART region are expressed in NPC tissues whereas ebv-miRNAs from the BHRF1 region are not found. Moreover, we identify two novel EBV miRNA genes originating from the BART region that have not been found in other tissues or cell lines before. We also identify three new human miRNAs, which might be specific for nasopharyngeal tissues. We further show that a number of different cellular miRNAs are upor down-regulated in NPC tissues compared to control tissue including miR-15a and miR-16. We find that the tumor suppressor BRCA-1 is a target of miR-15a as well as miR-16 suggesting a miRNA role in NPC pathogenesis.

overall_design: 2 pairs of NPC and control tissues from 2 patients (4 samples in total) were examined.

first: Jiayun

last: Zhu

first: Thorstern

last: Pfuhl

first: Natalie

last: Motsch

first: Stephanie

last: Barth

first: John

last: Nicholls

first: Friedrich

last: Graesser


title: A general mechanism for transcription regulation by Oct1 and Oct4 in response to genotoxic and oxidative stress

city: Salt Lake City

first: Dean

last: Tantin

address: 15 North Medical Drive East

department: Department of Pathology

institute: University of Utah

summary: We identify inducible binding of the transcription factor Oct1 to numerous targets following exposure to hydrogen peroxide.

overall_design:
Oct1 bound ChIP fragments were sequenced in the presence and absence of H2O2 exposure using HeLa cells. Both 26 and 36bp reads were generated.

Each processed file contains a list of genomic regions (NCBI 36.1, H_sapiens_Mar_2006, hg18) enriched for Oct1 binding.
These files were generated by running the EnrichedRegionMaker on ScanSeqs window scored data, see http://useq.sourceforge.net/ .
Adjacent and overlapping windows with a q-value FDR of 0.0001 were joined into larger enriched regions.
The best scoring window's scores are used to represent the enriched region.

first: Jinsuk

last: Kang


title: esBAF is an essential component of the core pluripotency transcriptional network

status: Public on Jan 20 2009

city: RTP

first: Raja

last: Jothi

address: 111 TW Alexander Drive; A314

institute: National Institutes of Health

summary: Distinctive SWI/SNF-like ATP-dependent chromatin remodeling esBAF complexes are indispensable for the maintenance and pluripotency of mouse embryonic stem (ES) cells. To understand the mechanism underlying the roles of these complexes in ES cells, we performed high-resolution genome-wide mapping of the core ATPase subunit, Brg, using ChIP-Seq technology. We find that that esBAF, as represented by Brg, binds to genes encoding components of the core ES transcriptional circuitry, including Polycomb group proteins. esBAF colocalizes extensively with Oct4, Sox2 and Nanog genome-wide, and shows distinct functional interactions with Oct4 and Sox2 at its target genes. Surprisingly, no significant colocalization of esBAF with PRC2 complexes, represented by Suz12, is observed. Lastly, esBAF co-binds with Stat3 and Smad1 genome-wide, consistent with a direct and critical role in LIF and BMP signaling essential to maintain pluripotency. Taken together, our studies indicate that esBAF is both an essential component of the core pluripotency transcriptional network, and might also be a critical component of the LIF and BMP signaling pathways essential for maintenance of self-renewal and pluripotency.

overall_design: Brg knockdown effect on expression, Brg ChIP-Seq

first: Lena

last: Ho

first: Jehnna

last: Ronan

first: Kairong

last: Cui

first: Keji

last: Zhao

first: Gerald

middle: R

last: Crabtree


title: A distinct class of small RNAs arises from pre-miRNA-proximal regions in a simple chordate

status: Public on Jan 18 2009

city: Corvallis

first: David

middle: Anthony

last: Hendrix

state: OR

address: 2011 Ag & Life Sciences Bldg

department: Biochemistry and Biophysics/EECS

institute: Oregon State University

summary: We report the comprehensive sequencing of small RNA libraries created from different developmental stages of the basal chordate, Ciona intestinalis. Surprisingly, half of the identified miRNA loci encode not two, but up to four distinct small RNAs.  These additional RNAs, termed “moRs”, are generated from sequences adjacent to the predicted ~60-nt pre-miRNA. moRs appear to be produced by RNAse-III-like processing, are ~20-nt in length, have 5’-monophosphate and 3’-hydroxl termini, and like miRNAs, are developmentally regulated.

sample_organism:
Drosophila melanogaster
Ciona intestinalis

overall_design:
Sequencing of small RNA libraries from 4 stages of Ciona intestinalis.

Ciona_LarvaMir309_RNASeq and Dmel_Toll10b_RNASeq are sequencing runs to detect if the expression of these RNA products is specific to Ciona or not. An ectopic transcript of the Drosophila  melanogaster miR-309 locus was expressed in Ciona to see if it processed the miRs differently than Drosophila melanogaster.  The Dmel sequence is from the endogenous case and is a control for comparison.

first: Weiyang

last: Shi

first: Michael

last: Levine

first: Ben

last: Haley


title: Global Mapping of Histone H3 K4 and K27 Trimethylation: Lineage Fate Determination of Differentiating CD4+ T Cells

status: Public on Jan 21 2009

city: DC

first: Weiqun

last: Peng

address: 725 21st street, NW

institute: The Georege Washington University

summary: Multipotential naïve CD4+ T cells differentiate into distinct lineages including Th1, Th2, Th17, and inducible T regulatory (iTreg) cells. The remarkable diversity of CD4+ T cells begs the question whether the observed changes reflect terminal differentiation with heritable epigenetic modifications or plasticity in T cell responses. We generated genome-wide histone H3 lysine 4 (H3K4) and lysine 27 (H3K27) trimethylation maps in naïve, Th1, Th2, Th17, iTreg, and natural (n)Treg cells. We find that while modifications of signature cytokine genes (Ifng, Il4, and Il17) partially conform to the expectation of lineage commitment, critical transcription factors such as Tbx21 exhibit a broad spectrum of epigenetic states, consistent with our demonstration of T-bet and IFN-γ induction in nTreg cells. Our data suggest an epigenetic mechanism underlying the specificity and plasticity of effector and regulatory T cells and also provide a framework for understanding complexity of CD4+ T helper cell differentiation.

overall_design: genome-wide analysis of histone H3 K4 and K27 trimethylation in different sub-lineages of mouse CD4+ T cells. (12 samples in total)

first: Gang

last: Wei

first: Lai

first: Jinfang

last: Zhu

first: Chongzhi

last: Zang

first: Jane

last: Hu-Li

first: Zhengju

last: Yao

first: Yuka

last: Kanno

first: Tae-Young

last: Roh

first: Wendy

last: Watford

first: Dustin

middle: E

last: Schones

first: Hong-Wei

last: Sun

first: William

last: Paul

last: O'Shea


title: High-throughput sequencing of small RNA transcriptomes in Locusta migratoria

status: Public on Jan 16 2009

city: Beijing

first: Yuanyuan

address: Datun Road, Chaoyang District

institute: Institute of Zoology, Chinese Academy of Sciences

summary: All the reports on insect small RNAs come from holometabolous insects. However, small RNAs of hemimetabolous insects have not yet been investigated.Study of hemimetabolous insect small RNAs could provide more insights into evolution and function of small RNAs in hemi- and holometabolous insects. The locust is an important, economically harmful hemimetabolous insect and its phase changes is an interesting phenomenon.Here, we used high-throughput sequencing to characterize and compare the small RNA transcriptomes of gregarious and solitary phases in locusts. We found abundant small RNAs and their different expression profiles in the two phases.

overall_design: Small RNAs were sequenced from gregarious and solitary phases of Locusta migratoria,respectively.

first: Shuang

last: Chen

first: Pengcheng

last: Yang

first: Le

last: Kang