title: Transcriptome-wide study of alternative splicing in nonsense-mediated mRNA decay-deficient mouse tissues

city: Copenhagen

first: Johannes

middle: Eichler

last: Waage

address: Ole Maaløesvej 5

department: Biotech Research and Innovation Center

country: Denmark

institute: University Of Copenhagen

summary: A major task in analyzing alternative splicing (AS) by RNA-Seq is to explore and quantify the differential representation of various splice classes and the protein-coding potential of the mapped reads. To this end, we have generated a streamlined bioinformatic pipeline available to the scientific community, which maps RNA-Seq reads to a complex combinatorial database of exon-exon junctions for the unique junction discovery, PTC detection and identification of splice isoforms. Hence, we have incorporated a splice isoform inference and PTC detection algorithm, which facilitates a highly accurate mapping and prediction of splice isoform junctions and nonsense-mediated mRNA decay (NMD)-susceptibility. We used this pipeline to investigate the complexity of the transcriptome and global role of AS-NMD in vivo in mammalian cells, by taking advantage of our Upf2 conditional knock-out mouse. Using tissue-specific Cre deleter strains, we have previously demonstrated in vivo that NMD acts to degrade many transcripts that results from AS. Also, ablating UPF2 (a core NMD factor) in the mouse leads to rapid mortality and collapse of most organs tested (Thoren et al., 2010; Weischenfeldt et al., 2008), suggesting an essential function for NMD in AS. To explore the effect of NMD on global splicing and to generate and validate an attractive bioinformatic pipeline for the scientific community to study AS and NMD, we chose to analyze two different mammalian organ systems with distinct phenotypes upon UPF2 ablation to test the robustness and dynamic range of the analysis. In one end of the spectrum, we chose to analyze the liver, wherein removal of UPF2 results in failure in liver metabolism and a high mortality rate (Thoren et al., 2010). To profile a less affected tissue, we chose to analyze bone marrow-derived macrophages (BMMs). These are macrophages that differentiate in vitro and Upf2 deleted BMMs are completely devoid of NMD activity but nevertheless show no morphological or functional phenotype compared to wild-type controls (Weischenfeldt et al., 2008). Thus, we were able to make a direct comparison between these two tissues to demonstrate the potency of our Isoform and PTC detection pipeline described below, which will allow the scientific community to analyze transcriptome data for global AS and NMD-susceptibility.

overall_design: Examination of UPF2 WT vs. KO in two different murine tissues (liver, bone marrow-derived macrophages).

first: Joachim

last: Weischenfeldt

first: Bo

last: Porse


title: Epigenetic alterations of a DNMT3B-mutant ICF patient at base-pair resolution

status: Public on Apr 26 2012

city: L'Hospitalet de Llobregat

first: Manel

last: Esteller

state: Barcelona

address: Hospital Duran i Reynals Av. Gran Via s/n km, 2.7

department: PEBC

country: Spain

institute: IDIBELL

type: Methylation profiling by high throughput sequencing

summary: The immunodeficiency, centromere instability and facial anomalies (ICF) syndrome is associated with mutation of the DNA methyl-transferase DNMT3B, resulting in a reduction of enzyme activity. Aberrant expression of immune system genes and hypomethylation of pericentromeric regions accompanied by chromosomal instability were determined as alterations driving the disease phenotype. However, so far only technologies capable of analyzing single loci were applied to determine epigenetic alterations in ICF patients. In the current study, we performed whole-genome bisulphite sequencing to assess alteration in DNA methylation at base-pair resolution.

overall_design: Whole-genome bisulphite sequencing was performed to assess alteration in DNA methylation of one ICF patient and one healthy control sample at base-pair resolution.

first: Holger

last: Heyn


title: Identification and functional characterization of Rca1, a transcription factor involved in both antifungal susceptibility and host response in Candida albicans

status: Public on May 02 2012

city: Lausanne

first: Dominique

last: Sanglard

address: Rue Bugnon 48

country: Switzerland

institute: University of Lausanne and University Hospital Center

type: Expression profiling by high throughput sequencing

summary: The identification of novel transcription factors associated to antifungal response may allow the discovery of fungal specific targets for new therapeutic strategies. A collection of 241 C. albicans transcriptional regulators mutants was screened for altered susceptibility to fluconazole, caspofungin, amphotericin B, and 5-fluorocytosine. Thirteen of these mutants not yet identified in their role in antifungal response were further investigated, and the function of one of them, mutant for orf19.6102 (RCA1) was characterized by transcriptome analysis. Strand specific RNA sequencing and phenotypic tests assigned Rca1 as the regulator of hyphal formation through the cAMP/PKA signaling pathway and the transcription factor Efg1, but also probably through its interaction with a transcriptional repressor, most likely Tup1. The mechanisms responsible for the high level of resistance to caspofungin and fluconazole observed resulting from Rca1 deletion were investigated. From our observations, we propose that caspofungin resistance was the consequence of the deregulation of cell wall gene expression, and that fluconazole resistance was linked to the modulation of the cAMP/PKA signaling pathway activity. In conclusion, our large-scale screening of a C. albicans transcription factor mutants collection allowed the identification of new effectors of the response to antifungals. The functional characterization of Rca1 assigned this TF and its downstream targets as promising candidates for the development of new therapeutic strategies, as Rca1 influences host sensing, hyphal development, and antifungal response.

overall_design: Total RNAs from RCA1 mutant and revertant isolates were extracted in duplicate from exponential growth-phase cultures in YEPD media. Differential expression was resolved by strand specific RNA sequencing.

first: Patrick

last: Vandeputte

first: Sylvain

last: Pradervand

first: Françoise

last: Ischer

first: Alix

middle: T

last: Coste

first: Sélène

last: Ferrari

first: Keith

last: Harshman


title: RNA-seq from ENCODE/HudsonAlpha

status: Public on Apr 27 2012

city: Stanford

first: ENCODE

last: DCC

state: CA

address: 300 Pasteur Dr

country: USA

institute: ENCODE DCC

summary:
This data was generated by ENCODE. If you have questions about the data, contact the submitting laboratory directly (Florencia Pauli mailto:fpauli@hudsonalpha.org). If you have questions about the Genome Browser track associated with this data, contact ENCODE (mailto:genome@soe.ucsc.edu).

This track is produced as part of the ENCODE Project. RNA-seq is a method for mapping and quantifying the transcriptome of any organism that has a genomic DNA sequence assembly (Mortazavi et al., 2008). Biological replicates of ENCODE cell lines were grown on separate culture plates, total RNA was purified and polyA selected two times. mRNA was then fragmented by magnesium-catalyzed hydrolysis, reverse transcribed to cDNA by random priming and amplified. The cDNA was sequenced on an Illumina Genome Analyzer (GAI or GAIIx).
The DNA sequences were aligned to the NCBI Build37 (hg19) version of the human genome using the sequence alignment programs ELAND (Illumina) or Bowtie (Langmead et al., 2009). The first 10 residues of sequencing have a weak characteristic nucleotide bias of unknown origin. This RNA-seq protocol does not specify the coding strand. As a result, there will be ambiguity at loci where both strands are transcribed.
This is the first NCBI Build37 (hg19) release of this track (Jan 2012). This release includes the 3 datasets (Jurkat, A549/DEX100nm, and A549/EtOH2pct) previously released on NCBI Build36 (hg18) and adds data for several more cell types and growth conditions in replicate. Four types of download files are available for each replicate including the Raw Data (fastq), Transcripts GencodeV7 (gtf), Raw Signal (bigwig), and Alignments (bam).

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:
Experimental Procedures
Cells were grown according to the approved ENCODE cell culture protocols (http://hgwdev.cse.ucsc.edu/ENCODE/protocols/cell) except for H1-hESC for which frozen cell pellets were purchased from Cellular Dynamics. Cells were lysed in RLT buffer (Qiagen RNEasy kit) and processed on RNEasy midi columns according to the manufacturer's protocol, with the inclusion of the "on-column" DNase digestion step to remove residual genomic DNA. mRNA was isolated from at least 10 ug of total RNA with oligo(dT) two times (Dynabeads mRNA PurificationgKit, Invitrogen). Alternatively, cells were lysed and mRNA was purified directly two times with oligo(dT) (Dynabeads mRNA DIRECT Kit, Invitrogen). 100 ng of mRNA was fragmented by magnesium-catalyzed hydrolysis and reverse transcribed to cDNA by random priming according to the protocol in Mortazavi et al. (2008). cDNA was prepared for sequencing on the Genome Analyzer flowcell according to the protocol for the ChIPSeq DNA genomic DNA kit (Illumina). The sequencing libraries were size-selected around 225 bp and amplified with 15 rounds of PCR.
Libraries were sequenced with an Illumina Genome Analyzer I or an Illumina Genome Analyzer IIx according to the manufacturer's recommendations. Single end reads of 36 nt in length were obtained.
Data Processing and Analysis
Fastq files were made from qseq files generated by the Illumina pipeline (Casava 1.7). The Raw Signal files (bigWig) were generated from bedgraph files and the score was calculated as the number of reads at that position divided by the total number of reads divided by one million.
Casava export files were aligned to the NCBI Build37 (hg19) version of the human genome with ELAND (Illumina), generating SAM files. Fastq files of experiments that were previously aligned to NCBI Build36 (hg18) were aligned to NCBI Build37 (hg19) using Bowtie (Langmead et al., 2009; parameters: -S -n 2 -k 11 -m 10 --best), also generating SAM files. SAM files were converted to BAM with SAMtools (Li et al., 2009).
Gene expression within Gencode.v7 (Harrow et al., 2006) gene models was estimated using Cufflinks v0.9.3 (Roberts et al., 2011). Estimates of transcript abundance were reported in Fragments Per Kilobase of exon per Million fragments mapped (FPKM). FPKM is calculated by dividing the total number of fragments that align to the gene model by the size of the spliced transcript (exons) in kilobases. This number is then divided by the total number of reads in millions for the experiment. FPKM is reported in the last column of the gtf (TranscriptGencV7) files.
Raw Data (fastq), Raw Signal (bigWig), Alignments (bam) and Transcript Gencode V7 (gtf) files are available from the Downloads (http://hgwdev.cse.ucsc.edu/cgi-bin/hgFileUi?g=wgEncodeHaibRnaSeq) page.

first: Richard

last: Myers

first: Florencia

last: Pauli


title: Genome-wide identification of non-coding RNAs targeted by XRN exoribonucleases in collaboration with FRY1 using RNA-Seq in Arabidopsis thaliana.

status: Public on May 03 2012

city: La Jolla

first: Joseph

middle: R

last: Ecker

address: 10010 North Torrey Pines Road

department: Genomic Analysis Laboratory

institute: HHMI-Salk-Institute

type: Non-coding RNA profiling by high throughput sequencing

summary: We report new insight of non-coding RNA degradation mediated by XRN exoribonucleases and FRY1 in Arabidopsis thaliana. We suggest that XRN3, in combination with FRY1, is required to prevent the accumulation of 3’ extensions that arise from thousands of mRNA and miRNA precursor transcripts.

overall_design: Examination of genome-wide transcriptomes of Arabidopsis genotypes such as fry1-6, xrn3-3, xrn2-1xrn3-3, xrn2-1xrn4-6 and xrn3-3xrn4-6 using directional RNA-Seq method.

first: Yukio

last: Kurihara

first: Robert

middle: J

last: Schmitz

last: Nery

first: Matthew

middle: D

last: Schultz

first: Emiko

last: Okubo-Kurihara


title: TMPRSS2-ERG, HDACs and EZH2 are involved in an AR-centric transcriptional circuitry that calibrates androgenic response for prostate cancer progression

city: Singapore

first: Kern Rei

last: Chng

address: 60 Biopolis Street #02-01 Genome

department: Cancer Biology and Pharmacology 3

country: Singapore

institute: GIS

type: Genome binding/occupancy profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Transcription factor AbVf19 induces hydrolytic enzyme genes during pathogenesis in Alternaria brassicicola

city: Honolulu

first: Yangrae

last: Cho

state: HI

address: 3190 Maile Way, St John 317

department: Plant and Environmental Protection Sciences

institute: University of Hawaii at Manoa

summary: Alternaria brassicicola is a successful saprophyte and necrotrophic pathogen with a broad host range.  It produces secondary metabolites that marginally affect virulence, in contrast to many A. alternata strains that produce secondary metabolites as host-specific pathogenicity factors.  Cell wall-degrading enzymes (CWDEs) have been considered important for pathogenesis, but no CWDEs have been identified as significant virulence factors in A. brassicicola.  In this study, we discovered mutants of a gene, AbVf19, which consistently produced smaller lesions than the wild type.  The mutants grew slower than the wild type on an axenic medium with pectin as a major carbon source.  Gene expression comparisons identified several hydrolytic enzyme-coding genes being down-regulated in the mutant during a late stage of infection.  These down-regulated genes comprised a small fraction of genes within each family.  Three of these genes had mutants that showed no or little change in virulence.  This suggested that each down regulated gene only made a small contribution to virulence, or that their functions were redundant.  This study demonstrated the existence and importance of a transcription factor that regulates a suite of genes that are probably important for decomposing and utilizing plant material during the late stage of plant infection.

overall_design: gene expression profile comparisons between wild type and a transcription factor mutant during host infection

first: Robin

last: Ohm


title: Reshaping of global gene expression networks and sex-biased gene expression by integration of a nascent gene

status: Public on May 01 2012

city: Cambridge

first: Sidi

last: Chen

state: MA

department: Biology

institute: MIT

type: Genome binding/occupancy profiling by genome tiling array

sample_organism:
Drosophila melanogaster
Drosophila simulans
Drosophila yakuba


title: Dynamic epigenetic enhancer signatures are predictive for key transcriptional regulators associated with cellular differentiation states

city: Regensburg

first: Michael

last: Rehli

address: F.-J.-Strauss-Allee 11

department: Internal Med III

country: Germany

institute: University Hospital Regensburg

summary: Cellular differentiation is orchestrated by lineage-specific transcription factors and associates with cell type-specific epigenetic signatures. Here, we utilized stage-specific, epigenetic "fingerprints" to deduce key transcriptional regulators of a cellular differentiation process. In the model of human macrophage differentiation, we globally mapped the distribution of epigenetic enhancer marks (histone H3 lysine 4 monomethylation, histone H3 lysine 27 acetylation, and the histone variant H2AZ) and show that cell type-specific epigenetic "fingerprints" correlate with specific, de novo derived motif signatures at all differentiation stages studied (hematopoietic progenitor cell, monocyte, macrophage). We validated the novel, de novo derived, macrophage-specific enhancer signature which included ETS, CEBP, bZIP, EGR, E-Box and NFkB motifs by ChIP-sequencing for a subset of motif corresponding transcription factors (PU.1, C/EBPbeta, and EGR2) which confirmed their predicted association with differentiation-associated epigenetic changes. This study highlights the power of genome-wide epigenetic profiling studies to reveal novel functional insights. It describes the dynamic enhancer landscape of human macrophage differentiation and provides a unique resource for macrophage biologists.

overall_design: ChIP-seq of 3 histone marks and 3 transcription factors in human blood monocytes and macrophages

first: Thu-Hang

last: Pham


title: Distinct STAT5 concentrations uniquely control establishment and differentiation of mammary epithelium during pregnancy

city: Cheonan

first: Keunsoo

last: Kang

state: Chungnam

address: Dandae-ro 119

department: Microbiology

country: South Korea

institute: Dankook University

summary: To dissect regulatory processes of cell proliferation and differentiation we generated mouse strains carrying any combination of the four Stat5 alleles, thus expressing STAT5 from 0 to 100%. RNA-Seq analyses revealed that different STAT5 levels activate specific genetic programs linked to cell proliferation and differentiation.

overall_design: We refer to wild-type mice and Stat5abfl/fl mice as AABB mice; Stat5abfl/fl;MMTV-Cre (with Stat5ab-deficient mammary epithelial cells) as Null mice; Stat5a-/- mice as BB mice; Stat5b-/- mice as AA mice; Stat5ab+/null mice as AB mice.

first: Yamaji

last: Daisuke

first: Kang

last: Keunsoo

first: Robinson

middle: W

last: Gertraud

first: Hennighausen

last: Lothar


title: Ultra short and progressive 4sU-tagging reveals key characteristics of RNA processing at nucleotide resolution

city: München

first: Caroline

last: Friedel

address: Amalienstr. 17

department: Institut für Informatik

institute: Ludwig-Maximilians-Universität München

summary: RNA synthesis and decay rates determine the steady-state levels of cellular RNAs. Metabolic tagging of newly transcribed RNA by 4-thiouridine (4sU) can reveal the relative contributions of RNA synthesis and decay rates. The kinetics of RNA processing, however, so far remained unresolved. Here, we show that ultra-short 4sU-tagging not only provides snap-shot pictures of eukaryotic gene expression but, when combined with progressive 4sU-tagging and RNA-seq, reveals global RNA processing kinetics at nucleotide resolution. Using this method, we identified classes of rapidly and slowly spliced/degraded introns. Interestingly, each class of splicing kinetics was characterized by a distinct association with intron length, gene length and splice site strength. For a large group of introns, we also observed long lasting retention in the primary transcript, but efficient secondary splicing or degradation at later time points. Finally, we show that processing of most, but not all small nucleolar (sno)RNA-containing introns is remarkably inefficient with the majority of introns being spliced and degraded rather than processed into mature snoRNAs. In summary, our study yields unparalleled insights into the kinetics of RNA processing and provides the tools to study molecular mechanisms of RNA processing and their contribution to the regulation of gene expression.

overall_design: 4sU-tagging was performed in human DG75 B-cells by adding 500 uM 4sU to cell culture medium for 5, 10, 15, 20 or 60 min. Following isolation of total cellular RNA, this was separated into nascent and untagged, pre-existing RNA.  Nascent RNA as well as total and untagged RNA from 60 min 4sU-tagging were subjected to SOLiD sequencing (SOLiD II) obtaining 35 nt reads.

first: Lukas

last: Windhager

first: Thomas

last: Bonfert

first: Kaspar

last: Burger

first: Zsolt

last: Ruzsics

first: Stefan

last: Krebs

first: Stefanie

last: Kaufmann

first: Georg

last: Malterer

first: Anne

last: L'Hernault

first: Markus

last: Schilhabel

last: Schreiber

first: Philip

last: Rosenstiel

first: Ralf

last: Zimmer

first: Dirk

last: Eick

middle: C

first: Lars

last: Dölken


title: [E-MTAB-935] GENCODE PCR-Seq Batch VII

status: Public on Apr 25 2012

city: Hinxton

first: ArrayExpress

last: EBI

state: Cambridgeshire

address: Wellcome Trust Genome Campus

country: United Kingdom

institute: European Bioinformatics Institute

summary:
As part of the ENCODE consortium the GENCODE project is producing a reference gene set through manual and automated gene prediction. Selected transcript models are verified experimentally by RT-PCR amplification followed by sequencing. This is batch VII, which targets annotations from Gencode 8 (March 2011).

ArrayExpress Release Date: 2012-02-01

Publication Title: GENCODE: producing a reference annotation for ENCODE
Publication Author List: Harrow J, Denoeud F, Frankish A, Reymond A, Chen CK, Chrast J, Lagarde J, Gilbert JG, Storey R, Swarbreck D, Rossier C, Ucla C, Hubbard T, Antonarakis SE, Guigo R

Person Roles: submitter
Person Last Name: Gonzalez
Person First Name: Jose
Person Mid Initials: M
Person Email: jmg@sanger.ac.uk
Person Phone: -498006
Person Address: Wellcome Trust Genome Campus, Hinxton, UK
Person Affiliation: Wellcome Trust Sanger Institute

Person Roles: investigator
Person Last Name: Hubbard
Person First Name: Tim
Person Mid Initials:
Person Email: th@sanger.ac.uk
Person Phone: -498055
Person Address: Wellcome Trust Genome Campus, Hinxton, UK
Person Affiliation: Wellcome Trust Sanger Institute

Person Roles: investigator
Person Last Name: Reymond
Person First Name: Alexandre
Person Mid Initials:
Person Email: Alexandre.Reymond@unil.ch
Person Phone:
Person Address: Lausanne, Switzerland
Person Affiliation: University of Lausanne

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:
Experimental Design: organism_part_comparison_design
Experimental Design: reference_design
Experimental Design: co-expression_design
Experimental Factor Name: ORGANISMPART
Experimental Factor Type: organism_part


title: The RDE-10/RDE-11 complex triggers RNAi induced mRNA degradation by association with target mRNA in C. elegans

city: Kansas City

first: Huan

last: Yang

state: MO

address: 1000 East 50th Street

institute: Stowers Institute for Medical Research

summary: The molecular mechanisms for target mRNA degradation in C. elegans undergoing RNA interference (RNAi) are not fully understood.  Using a combination of genetic, proteomic and biochemical approaches, we report a divergent RDE-10/RDE-11 complex that is required for RNAi in C. elegans.  The RDE-10/RDE-11 complex acts in parallel of nuclear RNAi.  Association of the complex with target mRNA is dependent on RDE-1 but not RRF-1, suggesting that target mRNA recognition depends on primary but not secondary siRNA.  Furthermore, RDE-11 is required for mRNA degradation subsequent to target engagement.  Deep sequencing reveals a 5-fold decrease in secondary siRNA abundance in rde-10 and rde-11 mutant animals, while primary siRNA and micro-RNA biogenesis is normal. Therefore, the RDE-10/RDE-11 complex is critical for amplifying the exogenous RNAi response. Our work uncovers an essential output of the RNAi pathway in C. elegans.

overall_design: 21-24nt small RNA were purifed from different C. elegans strain populations that underwent sel-1 RNAi

first: Jim

last: Vallandingham

first: Hua

last: Li

first: Ho

middle: Y

last: Mak


title: Genome-wide mapping of ligand-dependent progesterone receptor chromatin interactions in human breast cell lines using PR ChIP-seq

city: Westmead

first: J

middle: Dinny

last: Graham

state: NSW

address: 176 Hawkesbury Rd

department: Westmead Institute for Medical Research

country: Australia

institute: University of Sydney

summary: The effects of progesterone are pleiotropic, resulting in diverse outcomes in a range of target tissues. Progesterone signalling is mediated through the nuclear progesterone receptor (PR) and to identify whether the cell specificity of the PR transcriptome arises from distinct patterns of genomic interaction of PR, we have mapped the genomic binding sites for PR in breast cancer cells and minimally transformed breast cells. PR binding was correlated with transcriptional outcome in both cell lines.

overall_design: Three replicate PR ChIP samples and two replicate input DNA control samples from T-47D breast cancer cells and two replicate PR ChIP samples and two replicate input DNA control samples from AB32 transformed normal breast cells.

first: Christine

middle: L

last: Clarke


title: Non-overlapping progesterone receptor cistromes contribute to cell-specific transcriptional outcomes in breast cells


title: m6A mapping in mouse RNA (mouse liver and human brain)

city: Tel-Hashomer

first: Sharon

last: Moshitch-Moshkovitz

address: Laboratory wing

department: Cancer Research Center

country: Israel

institute: Sheba Medical Center

summary: We developed a novel approach, m6A-seq, for high-resolution mapping of the transcriptome-wide m6A landscape, based on antibody-mediated capture followed by massively parallel sequencing

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Identification of m6A modified sequences in mouse liver and human brain

first: Dan

last: Dominissini

first: Schraga

last: Schwartz

first: Mali

last: Salmon-Divon

first: Karen

last: Cesarkas

first: Gideon

last: Rechavi


title: TMPRSS2-ERG, HDACs and EZH2 are involved in an AR-centric transcriptional circuitry that calibrates androgenic response for prostate cancer progression (ChIP-Seq data)

first: Kern Rei

last: Chng

summary: Deregulation of the Androgen Receptor (AR) transcriptional network is a common hallmark in prostate cancers. To achieve its precise transcriptional role, AR needs to co-operate specifically with a plethora of cofactors. In prostate cancers, AR transcription collaborators are frequently aberrantly over-expressed, altering the AR signaling pathway to one that promotes oncogenesis. Recently, the prostate cancer recurrent fusion gene, ERG, was shown to promote tumor progression by acting as a repressor of AR signaling. However, the exact mechanics and the functional consequences associated with this crosstalk between ERG and AR still remains relatively unknown. Interestingly, through chromatin immunoprecipitation coupled with massively parallel sequencing, we discover that ERG and other commonly over-expressed transcriptional co-repressors (HDAC1, HDAC2, HDAC3 and EZH2) are wired into an AR-centric transcriptional network via a spectrum of distal enhancers and/or proximal promoters. We show that ERG represses several AR target genes involved in epithelial differentiation. Furthermore, we demonstrated that suppression of the androgen-induced gene, Vinculin, by ERG and histone deacetylases increases cancer cell invasiveness. From our results, we propose that ERG, histone deactelyases and the histone methyltransferase, EZH2, could impede epithelial differentiation and contribute to prostate cancer progression, in part through modulating the transcriptional output of AR.

overall_design: Genome-wide binding analysis of AR, ERG, HDAC1, HDAC2, HDAC3 and EZH2 in VCaP with and without DHT (dihydrotestosterone) stimulation using ChIP-Seq. 15 samples including 1 control (input).

first: Kern

first: Cheng

last: Chang

first: Si

middle: K

last: Tan

first: Chong

first: Shu

middle: Z

last: Hong

first: Edwin

last: Cheung


title: m6A mapping in human RNA (untreated)

summary: We developed a novel approach, m6A-seq, for high-resolution mapping of the transcriptome-wide m6A landscape, based on antibody-mediated capture followed by massively parallel sequencing.

overall_design: Identification of m6A modified sequences in HepG2 cells.


title: Critical Role of STAT5 Transcription Factor Tetramerization for Cytokine Responses and Normal Immune Function

city: Bethesda

first: Peng

state: MD

address: 9000 Rockville Pike

department: NHLBI

institute: NIH


title: m6A mapping in human RNA (with treatments)

overall_design: Identification of m6A modified sequences in HepG2 cells. HepG2 cells were incubated with either IFNg (200ng/ml) or HGF/SF (10 ng/ml) over night. Stress effects were tested in HepG2 cells by either 30 minutes incubation at 43ºC (heat shock) or UV irradiation of 0.04 J/cm2 followed by 4 hours of recovery in normal growing conditions prior to harvesting using Trypsin.


title: METTL3 KD in HepG2 cells

summary: To gain insight into possible processes that require m6A for their function, METTL3 was knocked down (KD) in HepG2 cells by siRNA transfections

overall_design: Differential expression analysis of METTL3 KD versus mock-transfected HepG2 cells, in 2 biological replicates


title: Topology of the human and mouse m6A RNA methylomes revealed by m6A-seq

platform_organism:
Schizosaccharomyces pombe
Saccharomyces cerevisiae
Homo sapiens
Mus musculus


title: Transcriptome analysis reveals strain-specific and conserved stemness genes in Schmidtea mediterranea

city: Farmington

first: Brenton

last: Graveley

state: CT

address: 400 Farmington Avenue

department: Genetics and Genome Sciences

institute: University of Connecticut Health Center

summary: The transcriptome of the planarian Schmidtea mediterranea is not well characterized.  We have used RNA-Seq to characterize the transcriptome in both sexual and asexual strains of S. mediterranea from both untreated and irradiated animals.  Moreover, we have performed RNA-Seq on RNA purified from FACS sorted neoblasts and differentiated cells.  Together these studies expand our understanding of the planarian transcriptome and have identified strain-specific, neoblast-specific, and conserved transcripts.

overall_design: RNA-Seq was performed on RNA isolated from untreated and irradiated S. mediterranea animals from both the sexual and asexual strains, on FACS purified X1 neoblasts, X2 neoblasts, and Xins differentiated cells.

first: Alissa

middle: M

last: Resch

first: Dasaradhi

last: Palakodeti

first: Yi-Chien

last: Lu


title: Small RNAs in Arabidopsis hyl1 and hyl1 suppressor mutants

city: University Park

first: Michael

last: Axtell

state: PA

address: 280 Mueller Lab

institute: Pennsylvania State University

summary: DCL1 and HYL1 are the core components of miRNA biogenesis machinery. hyl1-null mutants accumulate low levels of miRNAs and cause pleiotropic morphological phenotypes. Here, we reported that we identified 5 new alleles of DCL1 which are able to suppress the hyl1 morphological phenotypes and restore the miRNA accumulation level in hyl1 plants. These new alleles are located in the helicase and RNaseIIIa domains of DCL1, highlighting the critical functions of these domains. Biochemical analyses of these suppressors indicated that they process pri-miRNA more efficiently, with both higher Kcat and lower Km values. In addition, we investigated the functions of the DCL1 helicase domain. Our results indicated that the helicase domain is able to modulate the DCL1 processing activities. Based on these results, we proposed that HYL1 might exert its function through interaction with the DCL1 helicase domain.  Small RNA sequencing from hyl1 and select hyl1/dcl1 suppressor mutants confirms thats suppression of the hyl1 phenotype is due to an increase in the accumulation, but not the precision, of miRNAs in the suppressor mutants.

overall_design: Wild-type, hyl1-2, hyl1-2/dcl1-20, and hyl1-2/dcl1-21 plants were sampled from both rosette leaves and flowers.

first: Chenggang

last: Liu

first: Nina

middle: V

last: Fedoroff


title: Alternative splicing controlled by heterogeneous nuclear ribonucleoprotein L (hnRNP L) regulates development, proliferation and migration of thymic pre-T cells

status: Public on Apr 24 2012

city: Montreal

first: Brian

last: Wilhelm

address: 2950 Chemin Polytechnique

country: Canada

institute: University of Montreal

summary: The regulation of post-transcriptional modifications of pre-mRNA by alternative splicing is important for cellular function, development and immunity The receptor tyrosine phosphatase CD45, which is expressed on all hematopoietic cells, is known for its role in the development and activation of T cells. To investigate the role of hnRNP L further, we have generated conditional hnRNP L knockout mice carrying floxed alleles and the T-cell specific Lck-Cre recombinase transgene. The Lck-Cre transgene is active in DN3, DN4 as well as in DP and SP cells. We found that deletion of hnRNP L results in a decreased thymic cellularity caused by a partial block at the transition stage between DN4 and DP cells. In addition, hnRNP L-/- thymocytes express aberrant levels of the CD45RA splice isoform and show high levels of phosphorylated Lck at the activator tyrosine Y394 but lacking phosphorylation of the inhibitory tyrosine Y505. This is indicative of an increased basal Lck activation and correlated with a higher proliferation rate of DN4 cells in hnRNP L-/- mice. Deletion of hnRNP L also blocked egress of SP cells to peripheral lymphoid organs and the migration of SP thymocytes in response to the chemokines CCL21 and SDF-1a. Since we found that actin polymerization was also compromised in hnRNP L-/- SP cells, we propose that a defect in the signal transduction cascade downstream of the chemokine receptors CCR7 and CXCR4 caused by the absence of hnRNP L is responsible for this phenotype. Our results indicate that hnRNPL regulates pre-T cell development and migration by regulating CD45 pre-mRNA splicing and chemokine receptor signaling.

overall_design: RNA-seq from Thymocytes from WT mice compared to KO (hnRNP L) mice

first: Tarik

last: Moroy

first: Marie-Claude

last: Gaudreau


title: iCLIP analysis of EZH2-interacting RNAs

status: Public on Apr 23 2012

city: L'Hospitalet de Llobregat

first: Manel

last: Esteller

address: Hospital Duran i Reynals Av. Gran Via s/n km, 2.7

institute: IDIBELL

summary: RNAs directly interacting with EZH2 were isolated from human colorectal HCT116 cells using an in vivo crosslinking and immunoprecipitation strategy (iCLIP, König J et al, Nat Struct Mol Biol 2010) coupled to an ultrasequencing approach.

overall_design: RIP sequencing for 1 Human sample

first: Sonia

last: Guil


title: S2+DTT-4h smallRNA RNA-Seq

status: Public on Apr 19 2012

city: Toronto

first: DCC

last: modENCODE

state: Ontario

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

institute: Ontario Institute for Cancer Research

summary:
modENCODE_submission_3318
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: S2-HDR; Tissue: embryo-derived cell-line; Developmental Stage: late embryonic stage; Sex: Male;  EXPERIMENTAL FACTORS: Cell Line S2-HDR

first: Eric

last: Lai


title: Critical Role of STAT5 Transcription Factor Tetramerization for Cytokine Responses and Normal Immune Function (ChIP-Seq and RNA-Seq)

last: Li

summary: Cytokine-activated STAT proteins dimerize and bind to high-affinity motifs, and N-terminal domain-mediated oligomerization of dimers allows tetramer formation and binding to low-affinity tandem motifs, but the functions of dimers versus tetramers are unknown. We generated Stat5a and Stat5b double knock-in (DKI) N-domain mutant mice that form dimers but not tetramers, identified cytokine-regulated genes whose expression required STAT5 tetramers, and defined consensus motifs for dimers versus tetramers. Whereas Stat5- deficient mice exhibited perinatal lethality, DKI mice were viable, indicating that STAT5 dimers were sufficient for survival. Nevertheless, STAT5 DKI mice had fewer CD4+CD25+ T cells, NK cells, and CD8+ T cells, with impaired cytokine-induced proliferation and homeostatic proliferation of CD8+ T cells. DKI CD8+ T cell proliferation following viral infection was diminished and DKI Treg cells did not efficiently control colitis. Thus, tetramerization of STAT5 is dispensable for survival but is critical for cytokine responses and normal immune function.

overall_design: Genome-wide mapping of STAT5A,STAT5B binding in mouse WT and DKI T cells (cultured with or without IL-2 for 1 hr) was conducted. RNA-Seq is conducted in mouse CD8+ T cells (WT and DKI, non-treated or treated with IL-2/IL-15 for 4 hr, 24 hr, 48 hr and 72 hr)

first: Jian-Xin

last: Lin

first: Delong

first: Hyun

last: T Jin

first: Jianping

last: He

first: Mohammedr

last: Rasheed

first: Yrina

last: Rochman

first: Lu

last: Wang

first: Kairong

last: Cui

first: Chengyu

last: Kelsall

first: Rafi

last: Ahmed

first: Warren

last: Leonard


title: Lai-mbn2

summary:
modENCODE_submission_3741
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: Mbn2; Tissue: embryonic/larval hemocyte; Developmental Stage: late embryonic stage; Genotype: l(2)malignant blood neoplasm; Sex: Unknown;  EXPERIMENTAL FACTORS: Cell Line Mbn2


title: mRNA-seq of L1 larvae recovering from arrest

city: Durham

first: Colin

last: Maxwell

state: NC

address: 4314 FFSC

institute: Duke

summary: We used RNA-seq to sequence the transcriptome of worms arrested in the first larval stage after hatching without food and starved for 12 hours, then a timecourse of animals recovering from arrest after feeding. We collected samples after 12 hours of starvation and 1, 3 and 6 hours after feeding.

overall_design: Four time points (12 hr starvation and 1, 3, and 6 hr of recovery) with three biological replicates at times 0 and 6 and two biological replicates at times 1 and 3.

middle: S

first: Larry

last: Baugh


title: Lai-ML-DmD16c3/BS13

summary:
modENCODE_submission_3309
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: ML-DmD16-c3; Tissue: dorsal mesothoracic disc; Developmental Stage: third instar larval stage; Genotype: y v f mal; Sex: Female;  EXPERIMENTAL FACTORS: Cell Line ML-DmD16-c3


title: Lai-CMEW2

summary:
modENCODE_submission_3743
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: CME W2; Tissue: dorsal mesothoracic disc; Developmental Stage: third instar larval stage; Sex: Unknown;  EXPERIMENTAL FACTORS: Cell Line CME W2


title: Nucleosome Position by MNase-seq from ENCODE/Stanford/BYU

status: Public on Apr 20 2012

city: Stanford

first: ENCODE

last: DCC

address: 300 Pasteur Dr

institute: ENCODE DCC

summary:
This data was generated by ENCODE. If you have questions about the data, contact the submitting laboratory directly (Anshul Kundaje mailto:akundaje@stanford.edu). If you have questions about the Genome Browser track associated with this data, contact ENCODE (mailto:genome@soe.ucsc.edu).

Nucleosomes are part of the first level of chromatin packaging. They each consist of a histone heterooctamer around which DNA wraps 1.6 times. The histone heterooctomamer is made up of two copies of histones 2A, 2B, 3 and 5. The segment of DNA wrapped around the histones, the so-called "core" fragment, is 147 base pairs long. Neighboring nucleosomes are separated from one another by a stretch of DNA called the "linker," whose size varies depending on organism, cell type, and even chromatin activity. Certain chromatin remodeling factors govern accessibility of DNA to regulatory proteins by repositioning nucleosomes to reveal regulatory sites that would otherwise be occluded by a nucleosome.
In contrast to histone modifications such as methylation or acetylation, which are investigated by ChIP-seq, nucleosome positioning data are generated without immunoprecipitation (see Methods below). Instead, micrococcal nuclease is used to digest chromatin to apparent completion, the (well-defined and clearly visible) mononucleosomal core fragment fraction is isolated by gel purification, and one end is then sequenced. Mapping the sequence tag back to the genome reveals the precise position of one end of the core fragment that was protected by the nucleosome; the position of the other end can then simply be inferred by extending the read to a virtual length of 147 bases. Statistical analyses such as occupancy and positioning stringency can then be employed to analyze the local nucleosome landscape anywhere in the (mappable) genome or to infer global parameters of nucleosome organization.
In the context of the ENCODE project, nucleosome positioning data are particularly valuable for analysis of the relationship between transcription factor binding, histone modifications, and gene activity.
For a general primer on these types of data and analyses, refer to Valouev et al. (2008).

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:
To isolate mononucleosome core DNA fragments from the GM12878 and K562 ENCODE cell lines we followed the micrococcal nuclease (MNase) digestion and isolation protocol as described in Johnson et al. (2006), Valouev et al. (2008), and Valouev et al. (2011) with the following modifications. The precise concentrations of the two flash-frozen cell samples received from the Snyder Lab were not known so, per our standard procedure, we performed a series of digestions titrating the amount of MNase to determine the concentration of MNase for optimal digestion of each sample. Final concentrations of 25 U/µL and 50 U/µL of MNase were used to digest the GM12878 cells and K562 cells respectively at 20°C for 12 min. All other steps in the digestion and isolation protocol were as described.
Cells were grown according to the approved ENCODE cell culture protocols (http://hgwdev.cse.ucsc.edu/ENCODE/protocols/cell). K562 and GM12878 were each grown to ~2.5×108 cells. The cells were harvested, frozen and the nucleosome core isolation followed (Valouev et al. 2008).
The SOLiD reads were mapped in color-space with the probabilistic mapper, DNAnexus (https://dnanexus.com/). The DNAnexus mapper measures and propagates mapping uncertainty by including both quality values and mismatches in the alignment score calculation. The scores are then scaled across all possible mappings of the read to estimate the posterior probability for alignment to each genomic location. Reads corresponding to posterior probability of correct mapping > 0.9 were reported.
Nucleosome density signal maps (bedgraph and bigwig files) were generated by first shifting reads by 74 bp in the 5´ to 3´ direction and counting the total number of reads starting at each genomic coordinate on both strands. These counts are then smoothed using un-normalized kernel density smoothing with a triweight kernel. A bandwidth of 30 bp is used which is equivalent to a smoothing window of 60 bp. The smoothed counts at each position are then divided by the expected number of reads from an equivalent uniform distribution of reads in a ± 30 bp window around that position. If less than 25% of the positions in a ± 30 bp window around a genomic location are uniquely mappable or if the location is part of an assembly gap, the signal value at that position is considered unreliable and not recorded in the signal files. Hence, genomic coordinates that do not have any associated signal value should be considered missing or unreliable data. Genomic coordinates associated with a signal value of 0 are reliably mapable but do not have any signal in the dataset.

first: Michael

last: Snyder

first: Debasish

last: Raha

first: Steven

last: Johnson

first: Elliot

last: Winters

first: Arend

last: Sidow

first: Ziming

last: Weng

first: Cheryl

last: Smith

first: Philippe

last: Lacroute

first: Philipe

last: Cayting

first: Anshul

last: Kundaje


title: Lai–MLDmD8/BS37/BS38

first: DCC

last: modENCODE

summary:
modENCODE_submission_3513
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: ML-DmD8; Tissue: dorsal mesothoracic disc; Developmental Stage: third instar larval stage; Genotype: y v f mal;  EXPERIMENTAL FACTORS: Cell Line ML-DmD8

first: Eric

last: Lai


title: Epigenome and transcriptome of naive pluripotent mouse embryonic stem (ES) cells cultured in 2i serum-free medium

city: Nijmegen

first: Hendrik

last: Marks

address: Geert Grooteplein 26/28

department: Molecular Biology

country: Netherlands

institute: Radboud University Nijmegen, RIMLS

summary: This study describes the epigenetic and transcriptome profiling of mouse ES cells cultures in 2i serum-free medium. The proteins for which ChIP-Seq profiles were generated in these cells, as well as for cells grown in serum conditions,  include H3K4me3, H3K36me3, H3K27me3, H3K9me3, Suz12 and Ezh2. A sequence profile of genomic DNA is also included. Furthermore, we included RNA-Seq of ds_cDNA synthesized from double poly(A) selected mRNA and from rRNA depleted (strand specific) total RNA.

overall_design: ChIP-Seq profiling and RNA-Seq profiling on mouse naive ES cells

first: Roberta

last: Menafra

first: TÃ¼zer

last: Kalkan

first: Sergey

last: Denissov

first: Kenneth

last: Jones

first: Helmut

last: Hofemeister

first: Jennifer

last: Nichols

first: Andrea

last: Kranz

first: A

middle: F

last: Stewart

first: Austin

middle: G

last: Stunnenberg


title: Lai-S1/BS79

summary:
modENCODE_submission_3311
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: S1; Tissue: embryo-derived cell-line; Developmental Stage: late embryonic stage; Sex: Unknown;  EXPERIMENTAL FACTORS: Cell Line S1


title: RNA-seq from ENCODE/Stanford/Yale/USC/Harvard

first: ENCODE

last: DCC

summary:
This data was generated by ENCODE. If you have questions about the data, contact the submitting laboratory directly (Gerstein Lab mailto:datasubmission@gersteinlab.org). If you have questions about the Genome Browser track associated with this data, contact ENCODE (mailto:genome@soe.ucsc.edu).

The tracks show enrichment of RNA sequence tags generated by high throughput sequencing (RNA-seq) and mapped to the human genome. Double stranded cDNA was synthesized from polyadenylated RNA (polyA+) . PCR amplified, adapter ligated cDNA, 150-300nt long, was sequenced on an Illumina GA sequencer.
Where designated, cell lines received specific treatments prior to RNA isolation. As indicated, K562 cells were treated with either interferon-a or interferon-g for 30 minutes or 6 hours. These experiments were carried out in conjunction with ChIP-Seq experiments on the transcription factors STAT1 and STAT2 in order to examine the effects that inducers of a specific transcriptional response might have on gene expression and on transcription factor binding site discovery. K562 cells were treated with a-amanitin in order to examine the effects of RNA polymerase II inhibition on RNA polymerase III-mediated transcription.
This track shows expression data generated as confirmation of the SYDH TFBS (http://genome-preview.ucsc.edu/cgi-bin/hgTrackUi?db=hg19&g=wgEncodeSydhTfbs) tracks currently available on genome-preview.

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:
Cells were grown according to the approved ENCODE cell culture protocols (http://hgwdev.cse.ucsc.edu/ENCODE/protocols/cell). Total RNA was extracted using TRIzol reagents (15596-018, Life Tech), following the manufacturer's protocol. For polyA+ samples, polyadenylated RNA was purified using the MicroPoly(A) Purist kit (AM1919, Life Tech) and fragmented using RNA Fragmentation Reagent (AM8740, Life Tech). Illumina adapters were ligated to double stranded cDNA which was synthesized using reagents from Life Tech (11917-010).
PCR amplified adapter ligated cDNA (150-300 bp) was sequenced using Illumina GA. Sequence reads of 27-33nt long with 0-2 mismatches were mapped to the genome. The signal height corresponds to the number of overlapping fragments at each nucleotide position in the genome. Samples originally mapped to the hg18 version of the human genome were remapped to hg19 using the BWA aligner, version 0.5.7.

first: Michael

last: Snyder

first: Mark

last: Gerstein

first: Sherman

last: Weissman

first: Peggy

last: Farnham

first: Kevin

last: Struhl


title: Lai_OSCcell_V144_M031

status: Public on Apr 18 2012

first: DCC

last: modENCODE

summary:
modENCODE_submission_4788
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: OSC; Tissue: ovary stem cells; Developmental Stage: adult stage; Sex: Female;  EXPERIMENTAL FACTORS: Cell Line OSC

first: Eric

last: Lai


title: Mating of the stichotrichous ciliate Oxytricha trifallax induces production of a class of 27nt small RNAs derived from the parental macronucleus

city: Santa Cruz

first: Alan

last: Zahler

address: Sinsheimer Labs

department: MCD Biology

institute: UC Santa Cruz

summary: Here we report that soon after mating of Oxytricha trifallax, abundant 27nt small RNAs are produced that are not present prior to mating. We performed next generation sequencing of these 27nt RNAs from various times after mating. Using sequence comparisons between macronuclear and micronuclear versions of genes, we found that this 27nt RNA class derives from the parental macronucleus, not the developing macronucleus. These small RNAs are produced equally from both strands of macronuclear nanochromosomes, but in a highly non-uniform distribution along the length of the nanochromosome, and with a particular depletion in the 30 nt telomere-proximal positions. Unlike the Tetrahymena scanRNAs, the Oxytricha macronuclear-derived 27mers are not modified by 2'O-methylation at their 3' ends.

overall_design: Examination of small RNAs produced in Oxytricha trifallax during vegetative growth and at various timepoints during the mating process

first: Sol

last: Katzman


title: Lai_S1cell_V146

summary:
modENCODE_submission_4787
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


title: Lai_G2cell_V127

summary:
modENCODE_submission_4782
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: G2; Tissue: embryo-derived cell-line; Developmental Stage: late embryonic stage; Sex: Unknown;  EXPERIMENTAL FACTORS: Cell Line G2


title: Lai_CME_L1cell_V126

summary:
modENCODE_submission_4785
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: CME_L1; Tissue: ventral prothoracic disc; Developmental Stage: third instar larval stage; Sex: Unknown;  EXPERIMENTAL FACTORS: Cell Line CME_L1


title: Lai_fGS_OSScell_V091

summary:
modENCODE_submission_4780
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: fGS/OSS; Tissue: ovary; Developmental Stage: adult stage; Sex: Female;  EXPERIMENTAL FACTORS: Cell Line fGS/OSS


title: Altered Subcellular Localization of Transcription Factor TEAD4 Regulates First Mammalian Cell Lineage Commitment

first: Byunggil

last: Yoo

state: KS

address: 3901 Rainbow Blvd

institute: KUMC

summary: In the preimplantation mouse embryo TEAD4 is critical to establishing the trophectoderm (TE)-specific transcriptional program and segregating TE from the inner cell mass (ICM). However, TEAD4 is expressed both in the TE and the ICM. Thus, differential function of TEAD4 rather than expression itself regulates specification of the first two cell lineages. We used ChIP-seq to define genome-wide TEAD4 target genes and asked how transcription of TEAD4 target genes is specifically maintained in the TE. Our analyses revealed an evolutionarily conserved mechanism, in which lack of nuclear localization of TEAD4 impairs the TE-specific transcriptional program in inner blastomeres, thereby allowing their maturation towards the ICM lineage. Restoration of TEAD4 nuclear localization maintains the TE-specific transcriptional program in the inner blastomeres and prevents segregation of the TE and ICM lineages and blastocyst formation. We propose that altered subcellular localization of TEAD4 in blastomeres dictates first mammalian cell fate specification.

overall_design: ChIPseq profiles of TEAD4, IgG, Input in Mouse trophoblast stem cells using Illumina HiSeq 2000 and Illumina Genome Analyzer IIx

first: Pratik

last: Home

first: Sumedha

last: Gunewardena

first: Vincent

last: Schulz

first: Patrick

last: Gallagher

first: Soumen

last: Paul


title: Histone Modifications by ChIP-seq from ENCODE/Caltech

summary:
This data was generated by ENCODE. If you have questions about the data, contact the submitting laboratory directly (Barbara Wold mailto:woldb@caltech.edu, Georgi K. Marinov mailto:georgi@caltech.edu, Diane Trout mailto:diane@caltech.edu). If you have questions about the Genome Browser track associated with this data, contact ENCODE (mailto:genome@soe.ucsc.edu).

Our knowledge of the function of genomic DNA sequences comes from three basic approaches. Genetics uses changes in behavior or structure of a cell or organism in response to changes in DNA sequence to infer function of the altered sequence. Biochemical approaches monitor states of histone modification, binding of specific transcription factors, accessibility to DNases and other epigenetic features along genomic DNA. In general, these are associated with gene activity, but the precise relationships remain to be established. The third approach is evolutionary, using comparisons among homologous DNA sequences to find segments that are evolving more slowly or more rapidly than expected given the local rate of neutral change. These are inferred to be under negative or positive selection, respectively, and we interpret these as DNA sequences needed for a preserved (negative selection) or adaptive (positive selection) function.
The ENCODE project aims to discover all the DNA sequences associated with various epigenetic features, with the reasonable expectation that these will also be functional (best tested by genetic methods). However, it is not clear how to relate these results with those from evolutionary analyses. The mouse ENCODE project aims to make this connection explicitly and with a moderate breadth. Assays identical to those being used in the ENCODE project are performed in cell types in mouse that are similar or homologous to those studied in the human project. Thus, we will be able to discover which epigenetic features are conserved between mouse and human, and we can examine the extent to which these overlap with the DNA sequences under negative selection. The contribution of DNA that with a function preserved in mammals versus that with a function in only one species will be discovered. The results will have a significant impact on our understanding of the evolution of gene regulation.
Maps of Occupancy by Transcription Factors
Genome-wide occupancy maps of transcription factors (TFs) are generated by ChIP-seq. A ChIP-Seq experiment combines a chromatin immunoprecipitation (ChIP) experiment that enriches genomic DNA for the segments bound by specific proteins (the antigens recognized by the antibody) with high-throughput short read sequencing of the enriched DNA fragments (Wold & Myers, 2008). Proteins are crosslinked to DNA (usually with formaldehyde), chromatin is sheared and immunoprecipitated with the antibody of interest. The immunoprecipitated material is turned into a sequencing library and sequenced. The sequencing reads are then aligned to the genome. A control sample consisting of sonicated chromatin that has not been immunoprecipitated or immunoprecipitated with a non-specific immunoglobulin is also sequenced. The ChIP and the control datasets are analyzed with a variety of software packages to identify regions occupied by the target protein. The sequencing data, alignments and analysis files for these experiments are available for download.
In specific, the Ren lab examined RNA polymerase II (PolII), co-activator protein p300, the insulator protein CTCF, and two chromatin modification marks, H3K4me3 and H3K4me1, due to their demonstrated utilities in identifying promoters, enhancers and insulator elements (Barski et al., 2007; Blow et al., 2010; Heintzman et al., 2009; Kim et al., 2007; Kim et al., 2005a; Visel et al., 2009). Enrichment of H3K4me3 or PolII signals is a strong indicator of an active promoter, while the presence of p300 or H3K4me1 outside of promoter regions has been used as a mark for enhancers. CTCF binding sites are considered as a mark for potential insulator elements. For each transcription factor or chromatin mark in each tissue, ChIP-seq was carried out with at least two biological replicates. Each experiment produced 20-30 million monoclonal, uniquely mapped tags.
Our knowledge of the function of genomic DNA sequences comes from three basic approaches. Genetics uses changes in behavior or structure of a cell or organism in response to changes in DNA sequence to infer function of the altered sequence. Biochemical approaches monitor states of histone modification, binding of specific transcription factors, accessibility to DNases and other epigenetic features along genomic DNA. In general, these are associated with gene activity, but the precise relationships remain to be established. The third approach is evolutionary, using comparisons among homologous DNA sequences to find segments that are evolving more slowly or more rapidly than expected given the local rate of neutral change. These are inferred to be under negative or positive selection, respectively, and we interpret these as DNA sequences needed for a preserved (negative selection) or adaptive (positive selection) function.
The ENCODE project aims to discover all the DNA sequences associated with various epigenetic features, with the reasonable expectation that these will also be functional (best tested by genetic methods). However, it is not clear how to relate these results with those from evolutionary analyses. The mouse ENCODE project aims to make this connection explicitly and with a moderate breadth. Assays identical to those being used in the ENCODE project are performed in cell types in mouse that are similar or homologous to those studied in the human project. Thus we will be able to discover which epigenetic features are conserved between mouse and human, and we can examine the extent to which these overlap with the DNA sequences under negative selection. The contribution of DNA that with a function preserved in mammals versus that with a function in only one species will be discovered. The results will have a significant impact on our understanding of the evolution of gene regulation.
Maps of histone modifications
Levels of three histone modifications are being determined. H3K4me1 (monomethylation of lysine 4 of histone H3) is a mark for active chromatin and in the absence of H3K4me3, it is one indicator of an enhancer. H3K4me3 (trimethylation of lysine 4 of histone H3) is highly enriched at active promoters. One repressive (Polycomb) mark, H3K27me3, is associated with some silenced genes. Maps of genomic DNA in chromatin with these histone modifications are generated by ChIP-seq. This consists of two basic steps: chromatin immunoprecipitation (ChIP) is used to highly enrich genomic DNA for the segments bound by specific proteins (the antigens recognized by the antibodies) followed by massively parallel short read sequencing to tag the enriched DNA segments. Sequencing is done on the Illumina GAIIx and HiSeq. The sequence tags are mapped back to the mouse genome (Langmead et al. 2009), and a graph of the enrichment for histone modifications are displayed as the "Signal" track (essentially the counts of mapped reads per interval) and the deduced probable binding sites from the MACS program (Zhang et al. 2008) are shown in the "Peaks" track. Each experiment is associated with an input signal, which represents the control condition where immunoprecipitation with non-specific immunoglobulin was performed in the same cell type. The sequence reads, quality scores, and alignment coordinates from these experiments are available for download.
Our knowledge of the function of genomic DNA sequences comes from three basic approaches. Genetics uses changes in behavior or structure of a cell or organism in response to changes in DNA sequence to infer function of the altered sequence. Biochemical approaches monitor states of histone modification, binding of specific transcription factors, accessibility to DNases and other epigenetic features along genomic DNA. In general, these are associated with gene activity, but the precise relationships remain to be established. The third approach is evolutionary, using comparisons among homologous DNA sequences to find segments that are evolving more slowly or more rapidly than expected given the local rate of neutral change. These are inferred to be under negative or positive selection, respectively, and we interpret these as DNA sequences needed for a preserved (negative selection) or adaptive (positive selection) function.
The ENCODE project aims to discover all the DNA sequences associated with various epigenetic features, with the reasonable expectation that these will also be functional (best tested by genetic methods). However, it is not clear how to relate these results with those from evolutionary analyses. The mouse ENCODE project aims to make this connection explicitly and with a moderate breadth. Assays identical to those being used in the ENCODE project are performed in cell types in mouse that are similar or homologous to those studied in the human project. Thus we will be able to discover which epigenetic features are conserved between mouse and human, and we can examine the extent to which these overlap with the DNA sequences under negative selection. The contribution of DNA that with a function preserved in mammals versus that with a function in only one species will be discovered. The results will have a significant impact on our understanding of the evolution of gene regulation.
Maps of Occupancy by Transcription Factors
Maps of occupancy of genomic DNA by transcription factors (TFs) are determined by ChIP-seq. This consists of two basic steps: chromatin immunoprecipitation (ChIP) is used to highly enrich genomic DNA for the segments bound by specific proteins (the antigens recognized by the antibodies) followed by massively parallel short read sequencing to tag the enriched DNA segments. Sequencing is done on the Illumina GAIIx and HiSeq. The sequence tags are mapped back to the mouse genome (Langmead et al. 2009), and a graph of the enrichment for TF binding are displayed as the "Signal" track (essentially the counts of mapped reads per interval) and the deduced probable binding sites from the MACS program (Zhang et al. 2008) are shown in the "Peaks" track. Each experiment is associated with an input signal, which represents the control condition where immunoprecipitation with non-specific immunoglobulin was performed in the same cell type. The sequence reads, quality scores, and alignment coordinates from these experiments are available for download.
The sequence reads, quality scores, and alignment coordinates from these experiments are available for download.

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:
Cells were grown according to the approved ENCODE cell culture protocols (http://genome-test.cse.ucsc.edu/ENCODE/protocols/cell/mouse).
Chromatin immunoprecipitation followed published methods (Johnson & Mortazavi et al., 2007) with the exception of certain experiments for which glutaraldehyde was added to the crosslink reaction. Information on the antibodies used is available via the metadata for each subtrack. Libraries were constructed using the Illumina ChIP-seq Sample Preparation Kit or using a modified protocol that includes the addition of multiplexing tags to the fragments. DNA fragments were repaired to generate blunt ends and a single A nucleotide was added to each end. Double-stranded Illumina adaptors or Double-stranded Illumina adaptors with multiplexing tags were ligated to the fragments. Ligation products were amplified by 18 cycles of PCR, and the DNA between 150-250 bp was gel purified. Completed libraries were quantified with Quant-iT dsDNA HS Assay Kit. The DNA library was sequenced on the Illumina GAII and GAIIx sequencing systems, and more recently, for multiplexed libraries, several of them were pooled and sequenced on the HiSeq platform. Cluster generation, linearization, blocking and sequencing primer reagents were provided in the Illumina Cluster Amplification kits. Older libraries were generated using 2 rounds of PCR. Matched input samples were sequenced for each variation of fixation conditions and the number of PCR rounds. Reads of 32 bp, 36 bp or 50 bp length were generated.
Sequencing reads (fastq files) were assigned to the corresponding libraries based on the multiplexing tag for pooled libraries (all tags have been removed from reads in the fastq files available for download) or directly processed. Bowtie (Langmead et al., 2009) was used to map reads to the male or female version of the mouse genome (excluding the _random chromosomes in the assembly) depending on the cell line sex. The following parameters were used: "-v 2 -k 11 -m 10 -t --best --strata". Aligned reads were converted into rds files using the ERANGE package (Johnson & Mortazavi et al., 2007) and the findall.py program in ERANGE was used to identify enriched regions against the matching input sample. The following settings were used for point-source transcription factors: "--shift learn --ratio 3 --minimum 2 --listPeak --revbackground". For histone modifications, the settings were changed to "--notrim --nodirectionality --spacing 100 --ratio 3 --minimum 2 --listPeak --revbackground".
Cells were grown according to the approved ENCODE cell culture protocols (http://genome.ucsc.edu/ENCODE/protocols/cell/mouse).
RNA-Seq
RNA samples from tissues and primary cells were extracted from Trizol® according to protocol (Invitrogen). PolyA+ RNA was purified with the Dynabeads mRNA purification kit (Invitrogen). The mRNA libraries were prepared for strand-specific sequencing as described previously (Parkhomchuk et al., 2009).
Sequencing and Analysis
Samples were sequenced on Illumina Genome Analyzer II, Genome Analyzer IIx and HiSeq 2000 platforms for 36 cycles. Image analysis, base calling and alignment to the mouse genome version mm9 were performed using Illumina's RTA. Alignment to the mouse genome was performed using TopHat (Trapnell et al., 2009). Wig files were generated by TopHat and expression levels were calculated with Cufflinks (Trapnell et al., 2010).
Cells were grown according to the approved ENCODE cell culture protocols (http://genome.ucsc.edu/ENCODE/protocols/cell/mouse).
Enrichment and Library Preparation
Chromatin immunoprecipitation was performed according to Ren Lab ChIP Protocol (http://bioinformatics-renlab.ucsd.edu/RenLabChipProtocolV1.pdf).
Library construction was performed according to Ren Lab Library Protocol (http://bioinformatics-renlab.ucsd.edu/RenLabLibraryProtocolV1.pdf).
Sequencing and Analysis
Samples were sequenced on Illumina Genome Analyzer II, Genome Analyzer IIx and HiSeq 2000 platforms for 36 cycles. Image analysis, base calling and alignment to the mouse genome version mm9 were performed using Illumina's RTA and Genome Analyzer Pipeline software. Alignment to the mouse genome was performed using ELAND or Bowtie (Langmead et al., 2009) with a seed length of 25 and allowing up to two mismatches. Only the sequences that mapped to one location were used for further analysis. Of those sequences, clonal reads, defined as having the same start position on the same strand, were discarded. BED and wig files were created using custom perl scripts.
Cells were grown according to the approved ENCODE cell culture protocols.
The chromatin immunoprecipitation followed published methods (Welch et al. 2004). Information on antibodies used is available via the hyperlinks in the "Select subtracks" menu. Samples passing initial quality thresholds (enrichment and depletion for positive and negative controls - if available - by quantitative PCR of ChIP material) are processed for library construction for Illumina sequencing, using the ChIP-seq Sample Preparation Kit purchased from Illumina. Starting with a 10 ng sample of ChIP DNA, DNA fragments were repaired to generate blunt ends and a single A nucleotide was added to each end. Double-stranded Illumina adaptors were ligated to the fragments. Ligation products were amplified by 18 cycles of PCR, and the DNA between 250-350 bp was gel purified. Completed libraries were quantified with Quant-iT dsDNA HS Assay Kit. The DNA library was sequenced on the Illumina Genome Analyzer II sequencing system, and more recently on the HiSeq. Cluster generation, linearization, blocking and sequencing primer reagents were provided in the Illumina Cluster Amplification kits. All samples are being determined as biological replicates except time course samples. The data displayed are from the pooled reads for all replicates, but individual replicates are available by download.
The resulting 36-nucleotide sequence reads (fastq files) were moved to a data library in Galaxy, and the tools implemented in Galaxy were used for further processing via workflows (Blankenberg et al. 2010). The reads were mapped to the mouse genome (mm9 assembly) using the program bowtie (Langmead et al. 2009), and the files of mapped reads for the ChIP sample and from the "input" control (no antibody) were processed by MACs (Zhang et al. 2008) to call peaks for occupancy by transcription factors, using the parameters mfold=15, bandwidth=125. Because the signal for some histone modifications is not expected to be tightly localized (compared to a transcription factor), peak calling programs may not be appropriate. Thus in addition, we provide wiggle tracks with tag counts for every 10 bp segment. Per-replicate aligments and sequences are available for download at downloads page.
Cells were grown according to the approved ENCODE cell culture protocols (http://hgwdev.cse.ucsc.edu/ENCODE/protocols/cell/mouse).
The chromatin immunoprecipitation followed published methods (Welch et al. 2004). Information on antibodies used is available via the hyperlinks in the "Select subtracks" menu. Samples passing initial quality thresholds (enrichment and depletion for positive and negative controls - if available - by quantitative PCR of ChIP material) are processed for library construction for Illumina sequencing, using the ChIP-seq Sample Preparation Kit purchased from Illumina. Starting with a 10 ng sample of ChIP DNA, DNA fragments were repaired to generate blunt ends and a single A nucleotide was added to each end. Double-stranded Illumina adaptors were ligated to the fragments. Ligation products were amplified by 18 cycles of PCR, and the DNA between 250-350 bp was gel purified. Completed libraries were quantified with Quant-iT dsDNA HS Assay Kit. The DNA library was sequenced on the Illumina Genome Analyzer II sequencing system, and more recently on the HiSeq. Cluster generation, linearization, blocking and sequencing primer reagents were provided in the Illumina Cluster Amplification kits. All samples are being determined as biological replicates except time course samples. The data displayed are from the pooled reads for all replicates, but individual replicates are available by download.
The resulting 36-nucleotide sequence reads (fastq files) were moved to a data library in Galaxy, and the tools implemented in Galaxy were used for further processing via workflows (Blankenberg et al. 2010). The reads were mapped to the mouse genome (mm9 assembly) using the program bowtie (Langmead et al. 2009), and the files of mapped reads for the ChIP sample and from the "input" control (no antibody) were processed by MACs (Zhang et al. 2008) to call peaks for occupancy by transcription factors, using the parameters mfold=15, bandwidth=125. Per-replicate aligments and sequences are available for download at downloads page (http://hgdownload.cse.ucsc.edu/goldenPath/mm9/encodeDCC/wgEncodePsuTfbs/).
Cells were grown according to the approved ENCODE cell culture protocols (http://hgwdev.cse.ucsc.edu/ENCODE/protocols/cell/mouse). For details on the chromatin immunoprecipitation protocol used, see Euskirchen et. al., (2007), Rozowsky et. al. (2009) and Auerbach et. al. (2009).
DNA recovered from the precipitated chromatin was sequenced on the Illumina (Solexa) sequencing platform and mapped to the genome using the Eland alignment program. ChIP-seq data was scored based on sequence reads (length ~30 bps) that align uniquely to the human genome. From the mapped tags, a signal map of ChIP DNA fragments (average fragment length ~ 200 bp) was constructed where the signal height is the number of overlapping fragments at each nucleotide position in the genome. Reads were pooled from all submitted replicates to generate the Peak and Signal files. Per-replicate aligments and sequences are available for download at downloads page (http://hgdownload.cse.ucsc.edu/goldenPath/mm9/encodeDCC/wgEncodeSydhTfbs/).
For each 1 Mb segment of each chromosome, a peak height threshold was determined by requiring a false discovery rate <= 0.01 when comparing the number of peaks above said threshold to the number of peaks obtained from multiple simulations of a random null background with the same number of mapped reads (also accounting for the fraction of mapable bases for sequence tags in that 1 Mb segment). The number of mapped tags in a putative binding region is compared to the normalized (normalized by correlating tag counts in genomic 10 kb windows) number of mapped tags in the same region from an input DNA control. Using a binomial test, only regions that have a p-value = 0.01 are considered to be significantly enriched compared to the input DNA control.
Cells were grown according to the approved ENCODE cell culture protocols (http://hgwdev.cse.ucsc.edu/ENCODE/protocols/cell/mouse). Fresh tissues were harvested from mice and the nuclei prepared according to the tissue appropriate protocol (http://hgwdev.cse.ucsc.edu/ENCODE/protocols/cell/mouse). Digital DNaseI was performed by DNaseI digestion of intact nuclei, isolating DNaseI 'double-hit' fragments as described in Sabo et al. (2006), and direct sequencing of fragment ends (which correspond to in vivo DNaseI cleavage sites) using the Illumina IIx (and Illumina HiSeq by early 2011) platform (36 bp reads). Uniquely mapping high-quality reads were mapped to the genome using the bowtie aligner. DNaseI sensitivity is directly reflected in raw tag density, which is shown in the track as density of tags mapping within a 150 bp sliding window (at a 20 bp step across the genome). DNaseI sensitive zones (HotSpots) were identified using the HotSpot algorithm described in Sabo et al. (2004). 1.0% false discovery rate thresholds (FDR 0.01) were computed for each cell type by applying the HotSpot algorithm to an equivalent number of random uniquely mapping 36mers. DNaseI hypersensitive sites (DHSs or Peaks) were identified as signal peaks within FDR 1.0% hypersensitive zones using a peak-finding algorithm (I-max).
Cells were grown according to the approved ENCODE cell culture protocols (http://hgwdev.cse.ucsc.edu/ENCODE/protocols/cell/mouse). Fresh tissues were harvested from mice and the nuclei prepared according to the tissue appropriate protocol (http://hgwdev.cse.ucsc.edu/ENCODE/protocols/cell/mouse).
Reads were aligned to mm9 reference using ABI BioScope software version 1.2.1. Colorspace FASTQ format files were created using Heng Li's solid2fastq.pl script version 0.1.4, representing 0,1,2,3 color codes with the letters A,C,G,T respectively. Signal files were created from the BAM alignments using BEDTools.

first: Barbara

last: Wold

first: Millard

last: Jacobs

first: Muriel

first: Georgi

last: Marinov

first: Katherine

last: Fisher

first: Gordon

last: Kwan

first: Antony

last: Kirilusha

first: Ali

last: Mortazavi

first: Gilberto

last: DeSalvo

first: Brian

last: Williams

first: Lorianne

last: Schaeffer

first: Diane

last: Trout

first: Igor

last: Antoschechkin

first: Lu

last: Zhang

first: Gary

last: Schroth

first: Yin

last: Shen

first: Bing

last: Ren

first: Ross

last: Hardison

first: Mitchell

last: Weiss

first: Gerd

middle: A

last: Blobel

first: Stephan

last: Schuster

first: James

last: Taylor

first: Anton

last: Nekrutenko

first: Philip

last: Cayting

first: Richard

last: Sandstrom


title: Epigenomic enhancer profiling defines a signature of colon cancer

status: Public on Apr 16 2012

city: Cleveland Heights

first: Peter

last: Scacheri

state: OH

address: 10900 Euclid Ave; BRB 647

institute: Case Western Reserve University

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

overall_design: Refer to individual Series


title: Analysis of plant LTR-retrotransposons at the fine-scale family level reveals individual molecular patterns

city: Campinas

first: Renato

last: Vicentini

state: SP

address: Cidade Universitaria "Zeferino Vaz"

department: Centro de Biologia Molecular e Engenharia Genetica - CBMEG

country: Brazil

institute: Universidade Estadual de Campinas - UNICAMP

summary: Sugarcane is an important crop worldwide for sugar production and increasingly, as a renewable energy source. Modern cultivars have polyploid, large complex genomes, with highly unequal contributions from ancestral genomes. Long Terminal Repeat retrotransposons (LTR-RTs) are the single largest components of most plant genomes and can substantially impact the genome in many ways. It is therefore crucial to understand their contribution to the genome and transcriptome, however a detailed study of LTR-RTs in sugarcane has not been previously carried out. Sixty complete LTR-RT elements were classified into 35 families within four Copia and three Gypsy lineages. Structurally, within lineages elements were similar, between lineages there were large size differences. Four distinct patterns were observed in sRNA mapping, the most unusual of which was that of Ale1, with very large numbers of 24nt sRNAs in the coding region. The results presented support the conclusion that distinct small RNA-regulated pathways in sugarcane target the lineages of LTR-RT elements. Individual LTR-RT sugarcane families have distinct structures, and transcriptional and regulatory signatures. Our results indicate that in sugarcane individual LTR-RT families have distinct behaviors and can potentially impact the genome in diverse ways. For instance, these transposable elements may affect nearby genes by generating a diverse set of small RNA's that trigger gene silencing mechanisms.  There is also some evidence that ancestral genomes contribute significantly different element numbers from particular LTR-RT lineages to the modern sugarcane cultivar genome.

overall_design: Examination of small RNA populations in the sugarcane leaves that show matches against sugarcane LTR-RTs.

first: Douglas

last: Domingues

first: Guilherme

last: Cruz

first: Cushla

last: Metcalfe

first: Fabio

middle: T

last: Nogueira

first: Cristiane

last: Alves

first: Marie-Anne

last: Van Sluys


title: Lai_ML-DmD20c5/BS16/BS18

first: DCC

last: modENCODE

summary:
modENCODE_submission_3310
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: ML-DmD20-c5; Tissue: antenna disc-derived cell-line; Developmental Stage: third instar larval stage; Genotype: y v f mal; Sex: Male;  EXPERIMENTAL FACTORS: Cell Line ML-DmD20-c5

first: Eric

last: Lai


title: Sequence and expression analysis of human chromosome 20 gaps

status: Public on Apr 17 2012

city: Copenhagen

first: Sheroy

last: Minocherhomji

state: Copenhagen N

address: Blegdamsvej 3B, Building 18.1

department: Nordea Centre for Healthy Aging, Faculty Of Health Sciences Department of Cellular and Molecular Medicine

institute: University of Copenhagen

summary: The finished human genome-assemblies comprise several hundred un-sequenced euchromatic gaps, which may be rich in long polypurine/polypyrimidine stretches. Human chromosome 20 currently has three remaining un-sequenced gaps on its q-arm. All three gaps are within gene-dense regions, or overlap loci associated with human disorders, including one gap, which is at DLGAP4. In this study we sequenced, determined the complete sizes and assessed epigenetic landscapes of all three un-sequenced gaps on human chromosome 20 using a methodological approach involving Sanger sequencing, mate-pair paired-end high-throughput sequencing and chromatin and methylation analysis. We found histone H3K27me3 to be distributed across all three gaps in immortalized B-lymphocytes. We found five novel CpG islands in one gap to be highly hypermethylated in genomic DNA from both peripheral blood lymphocytes and human cerebellum. One of these CpG islands was differentially methylated and paternally hypermethylated. Furthermore, computational analyses predicted the presence of structured non-coding RNAs (ncRNAs) in all three chromosome 20 gaps. We verified expression for thirteen candidate ncRNAs, some of which showed tissue-specificity. Four ncRNAs expressed within the gap at DLGAP4 show elevated expression particularly in the human brain. Our data suggests that un-sequenced human genome gaps may comprise functional elements.

overall_design: Mate-pair paired end sequencing using genomic DNA from human translocation carriers having chromosomal rearrangments of chromosomes other than chromosome 20 and chromatin, DNA methylation analysis using human peripheral blood lymphocytes and/or human cerebellum tissue. Analysis done for three remaining human chromosome 20 un-sequenced gap regions.

first: Mads

last: Bak

first: Niels

last: Tommerup

first: Asli

last: Silahtaroglu


title: H1.4K34ac ChIP-sequencing in MCF-7 cells

status: Public on Apr 15 2012

city: Oxford

first: Kinga

last: Kamieniarz-Gdula

address: South Parks Road

department: Sir William Dunn School of Pathology

institute: University of Oxford

summary: The linker histone H1 is a key player in chromatin organization, yet our understanding of the regulation of H1 functions by posttranslational modifications is very limited. We provide here the first functional characterization of H1 acetylation. We show that H1.4K34 acetylation (H1.4K34ac) is mediated by GCN5 and is preferentially enriched at promoters of active genes, where it stimulates transcription by increasing H1 mobility and recruiting a general transcription factor. H1.4K34ac is dynamic during spermatogenesis and marks undifferentiated cells, such as induced pluripotent stem (iPS) cells and testicular germ cell tumors. We propose a model for H1.4K34ac as a novel regulator of chromatin function with a dual role in transcriptional activation.

overall_design: Examination of H1.4K34ac in the MCF-7 cell line by ChIP-Seq - two replicates of H1.4K34ac IP as two sequencing runs used to produce 1 BED file, one control IP sample.

last: Kamieniarz

first: Robert

last: Schneider


title: Genome-wide maps of polyadenylation sites in OPMD-model and control mice

city: Tel Aviv

first: Ran

last: Elkon

address: Ramat Aviv

department: Human Genetics

institute: Tel Aviv University

summary: We applied deep-sequencing based technique, 3'-Seq, to obtain comprehansive maps of poly-A sites in OPMD-model mice expressing trePABPN1(A17) mutant and control mice

overall_design: Examination of poly-A sites in control and OPMD-model mice (in two ages: 20 weeks and 12 months)

first: Mathias

last: Jenal

first: Reuven

last: Agami


title: PABPN1 role in regulation of alternative cleavage and polyadenylation


title: Epigenomic enhancer profiling defines a signature of colon cancer [ChIP-seq]

first: Peter

last: Scacheri

summary: Cancer is characterized by gene expression aberrations. Studies have largely focused on coding sequences and promoters, despite the fact that distal regulatory elements play a central role in controlling transcription patterns.  Here we utilize the histone mark H3K4me1 to analyze gain and loss of enhancer activity genome wide in primary colon cancer lines relative to normal colon crypts. We identified thousands of variant enhancer loci (VELs) that comprise a signature that is robustly predictive of the in vivo colon cancer transcriptome.  Furthermore, VELs are enriched in haplotype blocks containing colon cancer genetic risk variants, implicating these genomic regions in colon cancer pathogenesis.  We propose that reproducible changes in the epigenome at enhancer elements drive a unique transcriptional program to promote colon carcinogenesis.

overall_design: Examination of 3 histone modifications and global expression data in primary colon cancer cell lines and normal colon crypt controls

first: Batool

last: Akhtar-Zaidi

first: Alina

last: Saiakhova


title: Dynamic Transformations of Genome-wide Epigenetic Marking and Transcriptional Control Establish T Cell Identity [ChIP-Seq]

status: Public on Apr 14 2012

city: Pasadena

first: Jingli

last: Zhang

address: 1200 E California Blvd

institute: California Institute of Technology

summary: T cell development comprises a stepwise process of commitment from a multipotent precursor. To define molecular mechanisms controlling this progression, we probed five stages spanning the commitment process using deep sequencing RNA-seq and ChIP-seq methods to track genome-wide shifts in transcription, cohorts of active transcription factor genes, histone modifications at diverse classes of cis-regulatory elements, and binding patterns of GATA-3 and PU.1, transcription factors with complementary roles in T-cell development. The results locate potential promoter-distal cis-elements in play and reveal both activation sites and diverse mechanisms of repression that silence genes used in alternative lineages. Histone marking is dynamic and reversible, and while permissive marks anticipate, repressive marks often lag behind changes in transcription. In vivo binding of PU.1 and GATA-3 relative to epigenetic marking reveals distinctive, factor-specific rules for recruitment of these crucial transcription factors to different subsets of their potential sites, dependent on dose and developmental context.

overall_design: Genome-wide expression profiles, global distributions of three different histone modifications, and global occupancies of two transcription factors were examined in five developmentally related immature T populations. High throughput sequencing generated on average 9-30 million of mappable reads (single-read) for each ChIP-seq sample, and 10-15 million (single-read) for RNA-seq. Independent biological replicates were analyzed for individual populations. Terminology: while FLDN1_H3Ac_sample1.1 and FLDN1_H3Ac_sample1.2 are two lanes from the same sample, FLDN1_H3Ac_sample1.X and FLDN1_H3Ac_sample2 are from independent biological replicates. The sequence data of the input DNA from the same cell type was used as ChIP-seq background control.

first: Ali

last: Mortazavi

first: Brian

last: Williams

first: Barbara

last: Wold

first: Ellen

last: Rothenberg


title: Genome-wide maps of methylation state in cisplatin sensitive and resistant ovarian cancer cell line

status: Public on Apr 11 2012

city: HangZhou

first: Biaoyang

last: Lin

address: 866 Yu Hang Tang Road Zi Jin Gang Campus Zhejiang University.

department: ZCNI

country: China

institute: ZheJiang University

summary: We report the application of methyl-binding protein sequencing (MBD-seq) to global identify aberrant methylation between cisplatin sensitive and resistant ovarian cancer cell lines

overall_design: Detection of aberrant methylation states between cisplatin sensitive and resistant cell lines

first: Wei

last: Yu


title: Dynamic Transformations of Genome-wide Epigenetic Marking and Transcriptional Control Establish T Cell Identity


title: Analysis of ToxT regulon in Vibrio cholerae El Tor

status: Public on Apr 13 2012

city: Boston

first: Bryan

last: Davies

address: 200 Longwood Avenue

department: Microbiology and Immunobiology

institute: Harvard Medical School

summary: Using a combination of ChIP-seq and RNA-seq we show that ToxT regulates expression of the small RNA TarB. TarB modulates the expression level of the transcriptioanl repressor VC0177. VC0177 controls expression of several genes including VC0179.

overall_design: The function of the Vibrio 7th pandemic island-1 (VSP-1) in cholera pathogenesis has remained obscure. Utilizing ChIP-seq and RNA-seq to map the regulon of the master virulence regulator ToxT, we identify a TCP island encoded small RNA that reduces the expression of a previously unrecognized VSP-1 encoded transcription factor termed VspR. VspR modulates the expression of several VSP-1 genes including one that encodes a novel class of di-nucleotide cyclase (DncV), which preferentially synthesizes a previously undescribed hybrid cyclic AMP-GMP molecule.  We show that DncV is required for efficient intestinal colonization and down-regulates V. cholerae chemotaxis, a phenotype previously associated with hyperinfectivity. This pathway couples the actions of previously disparate genomic islands, defines VSP-1 as a pathogenicity island in V. cholerae and implicates its occurrence in 7th pandemic strains as a benefit for host adaptation through the production of a regulatory cyclic di-nucleotide.