title: Open Chromatin by DNaseI HS from ENCODE/OpenChrom(Duke University)

status: Public on Oct 14 2011

first: ENCODE

last: DCC

state: CA

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 (Terry Furey mailto:tsfurey@duke.edu). If you have questions about the Genome Browser track associated with this data, contact ENCODE (mailto:genome@soe.ucsc.edu).

These tracks display DNaseI hypersensitivity (HS) evidence as part of the four Open Chromatin track sets. DNaseI is an enzyme that has long been used to map general chromatin accessibility, and DNaseI "hypersensitivity" is a feature of active cis-regulatory sequences. The use of this method has led to the discovery of functional regulatory elements that include promoters, enhancers, silencers, insulators, locus control regions, and novel elements. DNaseI hypersensitivity signifies chromatin accessibility following binding of trans-acting factors in place of a canonical nucleosome.
Together with FAIRE and ChIP-seq experiments, these tracks display the locations of active regulatory elements identified as open chromatin in multiple cell types (http://hgwdev.cse.ucsc.edu/cgi-bin/hgEncodeVocab?type=cellType) from the Duke, UNC-Chapel Hill, UT-Austin, and EBI ENCODE group. Within this project, open chromatin was identified using two independent and complementary methods: these DNaseI HS assays and Formaldehyde-Assisted Isolation of Regulatory Elements (FAIRE), combined with chromatin immunoprecipitation (ChIP) for select regulatory factors. DNaseI HS and FAIRE provide assay cross-validation with commonly identified regions delineating the highest confidence areas of open chromatin. ChIP assays provide functional validation and preliminary annotation of a subset of open chromatin sites. Each method employed Illumina (formerly Solexa) sequencing by synthesis as the detection platform. The Tier 1 and Tier 2 cell types were additional verified by a second platform, high-resolution 1% ENCODE tiled microarrays supplied by NimbleGen.

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).
DNaseI hypersensitive sites were isolated using methods called DNase-seq or DNase-chip (Song and Crawford, 2010; Boyle et al., 2008a; Crawford et al., 2006). Briefly, cells were lysed with NP40, and intact nuclei were digested with optimal levels of DNaseI enzyme. DNaseI digested ends were captured from three different DNase concentrations, and material was sequenced using Illumina (Solexa) sequencing. DNase-seq data for Tier 1 and Tier 2 cell lines were verified by comparing multiple independent growths (replicates) and determining the reproducibility of the data. In general, cell lines were verified if 80% of the top 50,000 peaks in one replicate are detected in the top 100,000 peaks of a second replicate. For some cell types, additional verification was performed using similar material hybridized to NimbleGen Human ENCODE tiling arrays (1% of the genome) along with the input DNA as reference (DNase-chip). A more detailed protocol is available at http://hgwdev.cse.ucsc.edu/ENCODE/protocols/general/Duke_DNase_protocol.pdf.
The read length for sequences from DNase-seq are 20 bases long due to a MmeI cutting step of the approximately >50kb DNA fragments extracted after DNaseI digestion. Sequences from each experiment were aligned to the genome using BWA (Li et al., 2010) for the NCBI 36 (hg19) assembly. The command used for these alignments was > bwa aln -t 8 genome.fa s_1.sequence.txt.bfq > s_1.sequence.txt.sai Where genome.fa is the whole genome sequence and s_1.sequence.txt.bfq is one lane of sequences convert into the required bfq format.
Sequences from multiple lanes are combined for a single replicate using the bwa samse command, and converted in the sam/bam format using samtools. Only those that aligned to 4 or fewer locations were retained. Other sequences were also filtered based on their alignment to problematic regions (such as satellites and rRNA genes - see supplemental materials at http://hgdownload.cse.ucsc.edu/goldenPath/hg19/encodeDCC/wgEncodeOpenChromDnase/supplemental/). The mappings of these short reads to the genome are available for download at http://hgwdev.cse.ucsc.edu/cgi-bin/hgFileUi?g=wgEncodeOpenChromDnase.
The resulting digital signal was converted to a continuous wiggle track using F-Seq that employs Parzen kernel density estimation to create base pair scores (Boyle et al., 2008b). Input data has been generated for several cell lines. These are used directly to create a control/background model used for F-Seq when generating signal annotations for these cell lines. These models are meant to correct for sequencing biases, alignment artifacts, and copy number changes in these cell lines. Input data is not being generated directly for other cell lines. Instead, a general background model was derived from the available Input data sets. This should provide corrections for sequencing biases and alignment artifacts, but will not correct for cell type specific copy number changes. The exact command used for this step is > fseq -l 600 -v -f 0 -b <bff files> -p <iff files> aligments.bed where the (bff files) are the background files based on alignability, the (iff files) are the background files based on the Input experiments, and alignments.bed are a bed file of filtered sequence alignments.
Discrete DNaseI HS sites (peaks) were identified from DNase-seq F-seq density signal. Significant regions were determined by fitting the data to a gamma distribution to calculate p-values. Contiguous regions where p-values were below a 0.05/0.01 threshold were considered significant.
Data from the high-resolution 1% ENCODE tiled microarrays supplied by NimbleGen were normalized using the Tukey biweight normalization, and peaks were called using ChIPOTle (Buck, et al., 2005) at multiple levels of significance. Regions matched on size to these peaks that were devoid of any significant signal were also created as a null model. These data were used for additional verification of Tier 1 and Tier 2 cell lines by ROC analysis. Files containing this data can be found in the Downloads directory (http://hgwdev.cse.ucsc.edu/cgi-bin/hgFileUi?db=hg19&g=wgEncodeOpenChromDnase) labeled Validation view.
Release 1 (April 2011) of this track consists of a remapping of all previously released experiments to the human reference genome GRCh37/hg19 (these data were previously mapped to NCBI36/hg18; please see the Release Notes section of the hg18 Open Chromatin track (http://hgwdev.cse.ucsc.edu/cgi-bin/hgTrackUi?db=hg18&g=wgEncodeChromatinMap) for information on the NCBI36/hg18 releases of the data). There are 21 new DNaseI experiments in this release, on 19 new cell lines. New to this release is a reconfiguration of how this track is displayed in relation to other tracks from the Duke/UNC/UT-Austin/EBI group. A synthesis of open chromatin evidence from the three assay types was compiled for Tier 1 and 2 cell lines plus NHEK will also be added in this release and can be previewed in: Open Chromatin Synthesis (http://genome-preview.cse.ucsc.edu/cgi-bin/hgTrackUi?db=hg19&g=wgEncodeOpenChromSynth). Enhancer and Insulator Functional assays: A subset of DNase and FAIRE regions were cloned into functional tissue culture reporter assays to test for enhancer and insulator activity. Coordinates and results from these experiments can be found at http://hgdownload.cse.ucsc.edu/goldenPath/hg19/encodeDCC/wgEncodeOpenChromDnase/supplemental/.

first: Terry

last: Furey

first: Alan

last: Boyle

first: Lingyun

last: Song

first: Greg

last: Crawford

first: Paul

last: Giresi

first: Jason

last: Lieb

first: Zheng

last: Liu

first: Ryan

last: McDaniell

first: Bum-Kyu

last: Lee

last: Flicek

first: Damian

last: Keefe

first: Ewan

last: Birney

first: Stefan

last: Graf


title: Sequencing Apoptotic DNA Breakpoints

status: Public on Oct 15 2011

city: Singapore

first: Melissa

last: Fullwood

address: 8 College Road

country: Singapore

institute: A*STAR-Duke-NUS Neuroscience Research Partnership

type: Genome binding/occupancy profiling by high throughput sequencing

summary: DNA fragmentation is a well-recognized hallmark of apoptosis. However, the precise DNA sequences cleaved during apoptosis triggered by distinct mechanisms remain unclear. We used next-generation sequencing of DNA fragments generated in Actinomycin D-treated human HL-60 leukemic cells to generate a high-throughput, global map of apoptotic DNA breakpoints. These data highlighted that DNA breaks are non-random and show a significant association with active genes and open chromatin regions. We noted that transcription factor binding sites were also enriched within a fraction of the apoptotic breakpoints. Interestingly, extensive apoptotic cleavage was noted within genes that are frequently translocated in human cancers. We speculate that the non-random fragmentation of DNA during apoptosis may contribute to gene translocations and the development of human cancers.

overall_design: 3 files including 2 biological replicates (AHH001 and AHH002). Note that AHH002 is made up of 2 technical replicates (AHH002A and AHH002B)

middle: J

first: Joanne

first: Lifang

last: Lin

first: Guoliang

last: Li

first: Mikael

last: Huss

first: Patrick

last: Ng

first: Wing-Kin

last: Sung

first: Shirish

last: Shenolikar


title: Distribution of Drosophila insulator proteins after ecdysone treatment in Kc cells

city: Atlanta

first: Ashley

middle: M

last: Wood

state: GA

address: 1510 Clifton Rd. RRC 1071B

country: USA

institute: Emory University

summary: Here we examine changes in the distribution of Drosophila insulator proteins during the ecdysone response.  We performed ChIP-seq analysis in Kc cells at 0, 3, and 48 hours of ecdysone treatment with antibodies against CP190, Su(Hw), dCTCF, and BEAF-32B.

overall_design: Examination of 4 different insulator proteins at 3 time points of ecdysone treatment.

first: Victor

middle: G

last: Corces

first: Kevin

last: Van Bortle

first: Edward

last: Ramos


title: The Dynamic Architecture of Hox Gene Clusters

status: Public on Oct 13 2011

city: Lausanne

first: Marion

last: LELEU

address: EPFL-SV-ISREC-UPDUB- Station 19

department: School of Life Sciences

country: Switzerland

institute: EPFL

summary: The spatial and temporal control of Hox gene transcription is essential for patterning the vertebrate body axis. Although this process involves changes in histone posttranslational modifications, the existence of particular three-dimensional (3D) architectures remained to be assessed in vivo. Using high-resolution chromatin conformation capture methodology, we examined the spatial configuration of Hox clusters in embryonic mouse tissues where different Hox genes are active. When the cluster is transcriptionally inactive, Hox genes associate into a single 3D structure delimited from flanking regions. Once transcription starts, Hox clusters switch to a bimodal 3D organization where newly activated genes progressively cluster into a transcriptionally active compartment. This transition in spatial configurations coincides with the dynamics of chromatin marks, which label the progression of the gene clusters from a negative to a positive transcription status. This spatial compartmentalization may be key to process the collinear activation of these compact gene clusters.

overall_design: Examination of gene expression in 3 cell types. Examination of 2 different histone modifications in 2 cell types.

first: Daan

last: Noordermeer

last: Leleu

first: Denis

last: Duboule


title: The evolution of gene expression levels in mammalian organs

first: Henrik

last: Kaessmann

address: Génopode

department: Center For Integrative Genomics

institute: University of Lausanne

type: Expression profiling by high throughput sequencing

summary: Changes in gene expression are thought to underlie many of the phenotypic differences between species. However, large-scale analyses of gene expression evolution were until recently prevented by technological limitations. Here we report the sequencing of polyadenylated RNA from six organs across ten species that represent all major mammalian lineages (placentals, marsupials and monotremes) and birds (the evolutionary outgroup), with the goal of understanding the dynamics of mammalian transcriptome evolution. We show that the rate of gene expression evolution varies among organs, lineages and chromosomes, owing to differences in selective pressures: transcriptome change was slow in nervous tissues and rapid in testes, slower in rodents than in apes and monotremes, and rapid for the X chromosome right after its formation. Although gene expression evolution in mammals was strongly shaped by purifying selection, we identify numerous potentially selectively driven expression switches, which occurred at different rates across lineages and tissues and which probably contributed to the specific organ biology of various mammals. Our transcriptome data provide a valuable resource for functional and evolutionary analyses of mammalian genomes.

sample_organism:
Gallus gallus
Ornithorhynchus anatinus
Macaca mulatta
Gorilla gorilla
Pan paniscus
Pan troglodytes
Pongo pygmaeus
Homo sapiens
Mus musculus
Monodelphis domestica

overall_design:
To study mammalian transcriptome evolution at high resolution, we generated RNA-Seq data (∼3.2 billion Illumina Genome Analyser IIx reads of 76 base pairs) for the polyadenylated RNA fraction of brain (cerebral cortex or whole brain without cerebellum), cerebellum, heart, kidney, liver and testis (usually from one male and one female per somatic tissue and two males for testis) from nine mammalian species: placental mammals (great apes, including humans; rhesus macaque; mouse), marsupials (gray short-tailed opossum) and monotremes (platypus). Corresponding data (∼0.3 billion reads) were generated for a bird (red jungle fowl, a non-domesticated chicken) and used as an evolutionary outgroup.

Important note:
Fluorophore intensity files were processed with the Ibis base caller (version 1.11). As illustrated in Supplementary Note Table 1 and Supplementary Note Figure 1 of Brawand et al. (Nature 2011), we found that Ibis significantly increased the number of usable reads and drastically reduced the error rate. However, Ibis quality values are not comparable to those from Illumina values, which should be taken into account when using our data and interpreting their quality relative to data established using other base calling approaches.

first: David

last: Brawand

first: Magali

last: Soumillon

first: Anamaria

last: Necsulea

first: Philippe

last: Julien

first: Gábor

last: Csárdi

last: Harrigan

first: Manuela

last: Weier

first: Angélica

last: Liechti

first: Ayinuer

last: Aximu-Petri

first: Martin

last: Kircher

first: Frank

middle: W

last: Albert

first: Ulrich

last: Zeller

first: Philipp

last: Khaitovich

last: Grützner

first: Sven

last: Bergmann

first: Rasmus

last: Nielsen

first: Svante

last: Pääbo


title: Kc cells from Drosophila melanogaster during ecdysone treatment and CP190 knockdown

first: Ashley

last: Wood

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

overall_design: Refer to individual Series


title: Loss of Cardiac microRNA-Mediated Regulation Leads to Dilated Cardiomyopathy and Heart Failure.

status: Public on Oct 12 2011

city: Cambridge

first: Whitehead

last: Institute

state: MA

address: 9 Cambridge Center

department: Genome Technology Core

institute: Whitehead Institute

type: Non-coding RNA profiling by high throughput sequencing

summary:
RATIONALE:
Heart failure is a deadly and devastating disease that places immense costs on an aging society. To develop therapies aimed at rescuing the failing heart, it is important to understand the molecular mechanisms underlying cardiomyocyte structure and function.
OBJECTIVE:
microRNAs are important regulators of gene expression, and we sought to define the global contributions made by microRNAs toward maintaining cardiomyocyte integrity.
METHODS AND RESULTS:
First, we performed deep sequencing analysis to catalog the miRNA population in the adult heart. Second, we genetically deleted, in cardiac myocytes, an essential component of the machinery that is required to generate miRNAs. Deep sequencing of miRNAs from the heart revealed the enrichment of a small number of microRNAs with one, miR-1, accounting for 40% of all microRNAs. Cardiomyocyte-specific deletion of dgcr8, a gene required for microRNA biogenesis, revealed a fully penetrant phenotype that begins with left ventricular malfunction progressing to a dilated cardiomyopathy and premature lethality.
CONCLUSIONS:
These observations reveal a critical role for microRNAs in maintaining cardiac function in mature cardiomyocytes and raise the possibility that only a handful of microRNAs may ultimately be responsible for the dramatic cardiac phenotype seen in the absence of dgcr8.

overall_design: We report the miRNA composition of 6-8 week old murine heart. We performed deep sequencing analysis to catalog the miRNA population in the adult heart. 2 sample were sequenced.

first: Prakash

last: Rao

first: Harvey

last: Lodish


title: Transcriptome sequencing of naked mole rat, Heterocephalus glaber

city: shenzhen

first: zhiyong

last: huang

state: Guangdong

address: F5, Main Building, Beishan Industrial Zone, Beishan Road, Yantian District

country: China

institute: BGI-Shenzhen, Shenzhen 518083, China

summary: Deep sequencing of mRNA from naked mole rat

overall_design: Analysis of ploy(A)+ RNA of different specimens: brain, kidney, liver from new born , 4 years old , 20 years old and 4 years old hypoxia-exposed naked mole rat

first: Eun-Bae

last: Kim

first: Xiaodong

last: Fang

first: Alexey

middle: A

last: Fushan

first: Zhiyong

last: Huang

first: Alexei

middle: V

last: Lobanov

first: Lijuan

last: Han

first: Stefano

last: Marino

first: Xiaoqing

last: Sun

first: Anton

last: Turanov

first: Pengcheng

last: Yang

first: Sun-Hee

last: Yim

first: Xiang

last: Zhao

first: Marina

last: Kasaikina

first: Nina

last: Stoletzki

first: Chunfang

last: Peng

first: Paz

last: Polak

first: Zhiqiang

last: Xiong

first: Adam

last: Kiezun

first: Yabing

last: Zhu

first: Yuanxin

last: Chen

first: Gregory

last: Kryukov

first: Qiang

last: Zhang

first: Leon

last: Peshkin

first: Lan

first: Roderick

middle: T

last: Bronson

first: Bo

last: Wang

first: Changlei

first: Qiye

last: Li

first: Li

first: Wei

first: Shamil

middle: R

last: Sunyaev

first: Thomas

last: Park

first: Guojie

first: Jun

first: Vadim

middle: N

last: Gladyshev


title: Single-tube Linear DNA amplification (LinDA) for robust ChIP-seq

status: Public on Oct 11 2011

city: Illkirch; Strasbourg

first: Marco Antonio

last: Mendoza-Parra

address: 1, rue Laurent Fries

country: France

institute: IGBMC

summary: Genome-wide profiling of transcription factors based on massive parallel sequencing of immunoprecipitated chromatin (ChIP-seq) requires nanogram amounts of DNA. Here we describe a high-fidelity, single-tube linear DNA amplification method (LinDA) for ChIP-seq and reChIP-seq with picogram DNA amounts obtained from a few thousand cells. This amplification technology will facilitate global analyses of transcription-factor binding and chromatin with very small cell populations, such as stem or cancer-initiating cells.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: In total 5 samples were generated from the F9 teratocarcinoma cell system: mRXRa-RARg-LinDA;mRXRa_100xfold-diluted_LinDA; mRXRa(1) and two others previously described in GSE30538 (mRXRa(2): !Series_sample_id=GSM757796; mRARg : !Series_sample_id=GSM757803). In addition, 14 samples were generated from the H3396 breast cancer cell system. Those samples containing the label "LinDA" has been amplified following the Linear DNA amplification method developed by Pattabhiraman et al. ((2011),  Nature Methods 8, 565-67) prior library preparation for Solexa sequencing.

first: Pattabhiraman

last: Shankaranarayanan

first: Marco-Antonio

first: Mannu

last: Walia

first: Wouter

last: Van Gool

first: Ning

first: Luisa

last: Trindade

first: Hinrich

last: Gronemeyer


title: ENCODE Cold Spring Harbor Labs Long RNA-seq (hg18)

city: Stanford

first: ENCODE

last: DCC

summary:
This track depicts high throughput sequencing of long RNAs (>200 nt) from RNA samples from tissues or subcellular compartments from ENCODE cell lines. The overall goal of the ENCODE project is to identify and characterize all functional elements in the sequence of the human genome.
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.
Sample preparation and sequencing:
K562 and GM12878 total cell, total RNA: Standard Illumina Pair-end kit with the sole exception that a "tagged" random hexamer was used to prime the 1st strand synthesis: 5'-ACTGTAGGN6-3'. The addition of this tag is what permits us to make strand assignments for the reads. The sequence of the tag is reported in the 5' end of the read. Asymmetric PCR can place the tag on either the 1st or 2nd read depending on which strand it used as a template. Strand assignments are made by looking for the tag at the 5' end of either read 1 or read 2. Read 1 is physically linked to read 2. Therefore, if a tag is present on one end strand assignments are made for both ends. We noted during analysis that the tags are generally 5' truncated. We only "strand" reads that contain ACTGTAGG, CTGTAGG, TGTAGG, GTAGG. Between 63-68% of reads could be stranded in these libraries. It is possible to cull additional stranded reads that contain non-templated TAGG, AGG, GG, or G sequences at their 5' end. The peak in insert size distribution is between 200-250 nucleotides.
K562 cytosol, polyA+ RNA: Oligo-dT selected poly-A+ RNA was RiboMinus-treated according to the manufacturer's protocol (Invitrogen). The RNA was treated with tobacco alkaline pyrophosphatase to eliminate any 5' cap structures and hydrolyzed to ~200 bases via alkaline hydrolysis. The 3' end was repaired using calf intestinal alkaline phosphatase, and poly-A polymerase was used to catalyze the addition of Cs to the 3' end. The 5' end was phosphorylated using T4 PNK, and an RNA linker was ligated onto the 5' end. Reverse transcription was carried out using a poly-G oligo with a defined 5' extension. The inserts were then amplified using oligos targeting the 5' linker and poly-G extension. This cloning protocol generated stranded reads that were read from the 5' ends of the inserts. The library was sequenced on a Solexa platform for a total of 36 cycles; however, the reads underwent post-processing, resulting in trimming of their 3' ends. Consequently, the mapped read lengths are variable.
Analysis:
K562 and GM12878 total cell, total RNA: Tags were removed from the 5' ends of the reads in accordance to their lengths and strand assignments made. Subsequently, the reads were trimmed from their 3' ends to a final length of 50 nucleotides and were mapped using NexAlign, a program developed by Timo Lassman, RIKEN. We allowed up to 2 mismatches across the entire length and only report reads that mapped to a single/unique locus in the assembled hg18 genome.
K562 cytosol, polyA+ RNA: Reads were mapped to the human (hg18, March 2006) assembly using Nexalign, with only uniquely mapping (one loci), exactly matching (no mis-matches) aligned reads reported in the processed files, as follows: 1) Collect the read sequences from Illumina non-filtered output files. 2) Filter out all reads that contain undefined nucleotides ('N'). 3) Perform iterative alignment/C-tail chopping algorithm (below). On each alignment step, the reads are aligned to the genome with 100% identity. All reads that align to a single locus are withdrawn from the alignment pool and only the reads that could not be aligned continue to the next step. a) Align to the hg18 genome using Nexalign 1.3.3 (© Timo Lassmann) without chopping off any nucleotides. b) Chop off any C-blocks (until the first non-C) at the ends of the reads. c) Align to the genome -> remove and save those that align. d) Chop off any non-Cs until the next C. e) Chop off C-block until the next non-C. f) Align to the genome -> remove and save those that align. g) Repeat steps d, e, and f until the reads align to the genome, or chopping results in the reduction of the reads' lengths to below 16 (default), or there are no non-Cs left.

first: Tom

last: Gingeras

first: Carrie

last: Davis


title: Dissecting the retinoid-induced differentiation of F9 embryonal stem cells by integrative genomics


title: Transposon-based construction of strand-specific RNA-seq libraries

status: Public on Oct 10 2011

city: Huntsville

first: Jason

last: Gertz

state: AL

address: 601 Genome Way

institute: HudsonAlpha Institute

summary: We have developed two methods for efficiently consructing RNA-seq libraries using transposition.  Each method constructs high quality RNA-seq libraries when compared to standard approaches. One of the methods (Directional Tn-RNA-seq) maintains strand-of-origin information and exhibits strand specificity comparable to current approaches.

overall_design: RNA-seq libraries were constructed from ECC-1, a human cell line, and Universal Human Reference RNA using transposon-based and standard RNA-seq library construciton methods.

first: Richard

last: Myers


title: Chromatin Conformation Capture in two biological replicates of K562 cells to identify interactions between tRNA genes in human cells.

city: Santa Cruz

first: Jesse

last: Raab

address: 1156 High St

department: MCD Biology

institute: UC Santa Cruz

summary: tRNA genes are believed to form clusters in three-dimensional space in smaller eukaryotes (yeasts). In the course of investigating a cluster of tRNA genes for insulator activity in humans, we addressed whether these tRNA genes clustered with other tRNA genes. We used an anchor point to generate 4C material from a tRNA cluster located on chromosome 17 (hg18 7.962400 MB). We identify interactions occurring within the 300kb region of this point, but only extending in one direction and suggesting these interactions occur between this tRNA cluster and other tRNA gene clusters from this locus. We did not observed any specific interactions outside of this region or on other chromsomes, perhaps due to an low number of mappable reads from our samples.

first: Sol

last: Katzman

first: Rohinton

last: Kamakaka


title: Ubiquitous expression of long dsRNA in mice causes mainly RNAi effects in the oocyte

city: Prague 4

first: Radek

last: Malik

address: Videnska 1083

department: Dep. of Epigenetics Regulations

country: Czech Republic

institute: Institute of Molecular Genetics of the ASCR

summary: Double-stranded RNA (dsRNA) can enter different pathways in mammalian cells, such as the sequence-specific RNA interference (RNAi) pathway, the sequence-independent interferon response, and RNA editing by adenosine deaminases. To study routing of long dsRNA into different pathways in different tissues, we generated transgenic mice carrying an inverted repeat transcribed by a strong, ubiquitously active polII promoter. Here, we provide the first report of effects caused by ubiquitous long dsRNA expression in the adult mouse. Long dsRNA is poorly processed into siRNAs in somatic cells but readily induces the RNAi effects in the oocyte, suggesting that somatic cells are missing a component of RNAi that facilitates siRNA biogenesis. Expression of dsRNA is not sufficient to activate the interferon pathway and has a minimal effect on the transcriptome in somatic cells. The interferon response in somatic cells could be induced with high doses of dsRNA expression, suggesting that somatic cells can tolerate endogenous dsRNA expression to a large extent. Our data demonstrate that the association between long dsRNA and the interferon pathway in somatic cells cannot be generalized and that cells recognize other features of dsRNA molecules, which increase or reduce the likelihood activation of a particular dsRNA-induced pathway.

overall_design: Brain or kidney tissue (transgenic mice), or HEK293 or HeLa cell lines (human), expressing long dsRNA within the 3'-UTR of the EGFP reporter gene.

first: Petr

last: Svoboda

first: Jana

last: Nejepinska


title: modENCODE_White Lab: genome-wide ChIP data of CNC from WPP on Illumina Genome Analyzer.

status: Public on Oct 07 2011

city: Chicago

first: Kevin

middle: P.

last: White

state: IL

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

department: Institute for Genomics and Systems Biology

institute: University of Chicago

summary:
This is a dataset generated by the Drosophila Regulatory Elements modENCODE Project led by Kevin P. White at the University of Chicago. It contains genome-wide binding profile of the factor CNC from WPP generated by ChIP and analyzed on Illumina Genome Analyzer.

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:
A validated dataset is comprised of three biological replicates for ChIP-chip experiments and two replicates for ChIP-seq and meet the modENCODE quality standards. The control sample is the chromatin Input used for ChIP. Most factors binding profiles are generated by using specific antibodies for the protein of interest. However, some factors have been tagged by GFP in a transgenic line. In that case, ChIP is generated using an anti-GFP antibody.

This submission represents the ChIP-seq component of the study.

first: Nicolas

last: Negre

first: Nicholas

last: Bild

middle: P


title: Methylation specifies distinct estrogen-induced binding site repertoires of CBP to chromatin (ChIP-Seq)

first: Marco Antonio

last: Mendoza-Parra

summary: Multiple signaling pathways ultimately modulate the epigenetic information embedded in the chromatin of gene promoters by recruiting epigenetic enzymes. We found that, in estrogen-regulated gene programming, the acetyltransferase CREB-binding protein (CBP) is specifically and exclusively methylated by the coactivator-associated arginine methyltransferase (CARM1) in vivo. CARM1-dependent CBP methylation and p160 coactivators were required for estrogen-induced recruitment to chromatin targets. Notably, methylation increased the histone acetyltransferase (HAT) activity of CBP and stimulated its autoacetylation. Comparative genome-wide chromatin immunoprecipitation sequencing (ChIP-seq) studies revealed a variety of patterns by which p160, CBP, and methyl-CBP (meCBP) are recruited (or not) by estrogen to chromatin targets. Moreover, significant target gene-specific variation in the recruitment of (1) the p160 RAC3 protein, (2) the fraction of a given meCBP species within the total CBP, and (3) the relative recruitment of different meCBP species suggests the existence of a target gene-specific âfingerprintâ for coregulator recruitment. Crossing ChIP-seq and transcriptomics profiles revealed the existence of meCBP âhubsâ within the network of estrogen-regulated genes. Together, our data provide evidence for an unprecedented mechanism by which CARM1-dependent CBP methylation results in gene-selective association of estrogen-recruited meCBP species with different HAT activities and specifies distinct target gene hubs, thus diversifying estrogen receptor programming.

overall_design: Examination of estrogen-induced binding site in H3396 cells under Estrogen (E2) or Ethanol (EtOH) treatment:  CBP in E2; CBP in EtOH (two technical replicate); CBPR742m in E2 (two technical replicates); CBPR768m in E2; CBPR768m in EtOH; CBPR2151m in E2; CBPR2151m in EtOH; Estrogen receptor alpha (ERa) in E2; ERa in EtOH; Total Histone 3 and 4 acetylated in E2; H3K18ac in E2; H3K18ac in EtOH; Polymerase II (PolII) in E2; PolII in EtOH; RAC3 in E2; Input DNA sample in E2

first: Danilo

last: Ceschin

first: Mannu

last: Walia

first: Sandra

middle: S

last: Wenk

first: Carine

last: DuboÃ©

first: Claudine

last: Gaudon

first: Yu

last: Xiao

first: Lucas

last: Fauquier

first: Martial

last: Sankar

first: Laurence

last: Vandel


title: Methylation specifies distinct estrogen-induced binding site repertoires of CBP to chromatin


title: Lineage regulators direct BMP and Wnt pathways to cell-specific programs during differentiation and regeneration

city: Boston

first: Teresa

middle: Venezia

last: Bowman

address: One Blackfan Circle, 7th floor

institute: Children's Hospital Boston


title: Transcriptome sequencing of two macaques, crab-eating macaque and Indian rhesus macaque

first: zhiyong

last: huang

summary: Deep sequencing of mRNA from two macaques, crab-eating macaque and Indian rhesus macaque

sample_organism:
Macaca fascicularis
Macaca mulatta

platform_organism:
Macaca fascicularis
Macaca mulatta

overall_design: Analysis of ploy(A)+ RNA of different specimens:brain,ileum,kidney,liver,testes and white adipose for crab-eating macaque while brain,heart,,kidney,liver,quadriceps and testes for Indian rhesus macaque

first: Guojie

last: Zhang

first: Xiaodong

last: Fang

first: Jun

last: Wang


title: Identification of 67 histone marks and histone lysine crotonylation as a new type of histone modification

status: Public on Oct 06 2011

city: Cleveland

first: Fulai

last: Jin

state: OH - OHIO

address: 10900 Euclid Avenue

institute: Case Western Reserve University

summary: We report the identification of 67 previously undescribed histone modifications, increasing the current number of known histone marks by about 70%. We further investigated one of the marks, lysine crotonylation (Kcr), confirming that it represents an evolutionarily-conserved histone posttranslational modification. The unique structure and genomic localization of histone Kcr suggest that it is mechanistically and functionally different from histone lysine acetylation (Kac). Specifically, in both human somatic and mouse male germ cell genomes, histone Kcr marks either active promoters or potential enhancers. In male germinal cells immediately following meiosis, Kcr is enriched on sex chromosomes and specifically marks testis-specific genes, including a significant proportion of X-linked genes that escape sex chromosome inactivation in haploid cells. These results therefore dramatically extend the repertoire of histone PTM sites and designate Kcr as a specific mark of active sex chromosome-linked genes in postmeiotic male germ cells.

overall_design: 2 histone marks (pan-lysine acetylation and pan-lysine crotonylation) were studied in 1 human cell type and 2 mouse cell types using ChIP-Seq.  Input was sequenced for each cell type as a control. Pan-anti_Kac and pan-anti_Kcr antibodies were custom developed with PTM BioLab, Co., Ltd (Chicago, IL).

first: Jin

last: Fulai

first: Saadi

last: Khochbin

first: Bing

last: Ren

first: Yingming

last: Zhao


title: Genome-wide location analysis of WNT (Tcf7l2) and BMP (SMAD1) in human hematopoeitic cell lines co-occupied with lineage specific regulators (GATA1, GATA2, CEBPA)

summary: Lineage regulators direct BMP and Wnt pathways to cell-specific programs during differentiation and regeneration. This represents the human ChIP-seq portion of this dataset on leukemia cell lines.

overall_design: Human hematopoietic cells were cross-linked with formaldehyde for 20 min. DNA was enriched by chromatin immunoprecipitation (ChIP) and analyzed by Solexa sequencing. A sample of whole cell extract (WCE) was sequenced and used as the background to determine enrichment. ChIP was performed using an antibody against total Smad1 (Santa Cruz SC-7965), Tcf7l2 (Santa Cruz SC-8631),Gata1 (Santa Cruz SC-265),  Gata2 (Santa Cruz SC-9008), CEBPA (SC-9314) or H3K4me1 (Abcam ab8895).

first: E

last: Trompouki

first: T

first: L

last: Lawton

first: Z

last: Fan

first: D

last: Wu

first: A

last: DiBiase

first: C

last: Martin

first: J

last: Cech

middle: K

last: Sessa

middle: L

last: Leblanc

first: P

last: Li

last: Durand

last: Mosimann

first: G

middle: C

last: Heffner

middle: Q

last: Daley

first: R

middle: F

last: Paulson

last: Young

middle: I

last: Zon


title: Genome-wide location analysis of WNT (Tcf7l2) and BMP (SMAD1) in human hematopoeitic progenitors co-occupied with lineage specific regulators (GATA1, GATA2)

first: Teresa

last: Bowman

summary: Lineage regulators direct BMP and Wnt pathways to cell-specific programs during differentiation and regeneration. This represents the human CD34 ChIP-seq portion of this dataset.

overall_design: Human hematopoietic cells were cross-linked with formaldehyde for 20 min. DNA was enriched by chromatin immunoprecipitation (ChIP) and analyzed by Solexa sequencing. A sample of whole cell extract (WCE) was sequenced and used as the background to determine enrichment. ChIP was performed using an antibody against total Smad1 (Santa Cruz SC-7965), Tcf7l2 (Santa Cruz SC-8631),Gata1 (Santa Cruz SC-265),  Gata2 (Santa Cruz SC-9008) or CEBPA (SC-9314).

last: Trompouki

middle: M


title: Neuronal activity regulates hippocampal miRNA expression

city: Groningen

first: Eugene

last: Berezikov

address: Antonius Deusinglaan 1

department: European Research Institute for the Biology of Ageing

country: Netherlands

institute: University Medical Center Groningen

summary: Small RNA fractions from 6-8 week old C57BL/6 mouse hippocampus following electroconvulsive shock (ECS)

overall_design: Size selected RNA clones using Illumina v1.0 DGE small RNA kit, sequenced using Illumina

first: Stephen

last: Eacker

first: Ted

last: Dawson


title: Genome-wide  mapping  of  Sox6  binding  sites  in  mouse fetal myotubes

city: Davis

first: Chung-Il

last: An

address: One Shields Ave.

institute: University of California, Davis

summary: In  order  to  understand  how  Sox6  coordinately  regulates  the  transcription  of  multiple fiber type specific genes during muscle development, we have performed ChIP-seq analyses to identify Sox6 target genes as well as RNA polymerase II (Pol II) binding sites in mouse fetal myotubes.

overall_design: Examination of Sox6 and Pol II binding sites in mouse fetal primary myotubes

first: Nobuko

last: Hagiwara


title: CTCF promotes RNA pol II pausing and links DNA methylation to alternative splicing [RNA-Seq]

city: Frederick

first: Shalini

last: Oberdoerffer

state: MD

address: 1050 Boyles Street

institute: National Cancer Institute

summary: The goal of this study was to investigate the role of intragenic CTCF in alternative pre-mRNA splicing through a combined CTCF-ChIP-seq and RNA-seq approach. CTCF depletion led to decreased inclusion of weak upstream exons.

overall_design: CTCF ChIP-seq was performed in BJAB and BL41 B cell lines and normalized relative to Rabbit Ig control IP-seq reads. RNA-seq was performed in BJAB and BL41 cells transduced with shRNA against CTCF or RFP as a control, and in untransduced cells as well.

first: Sanjeev

last: Shukla

first: Rickard

last: Sandberg


title: Patterns and mechanisms of ancestral histone protein inheritance in budding yeast

city: Jerusalem

first: Assaf

last: Weiner

address: Givat Ram

department: Computer Science

country: Israel

institute: The Hebrew University of Jerusalem

summary: It is widely believed that chromatin, the nucleoprotein packaging state of eukaryotic genomes, can carry epigenetic information to propagate gene expression patterns in replicating cells. However, inheritance of genomic packaging status is subject to mechanistic challenges that do not confront the inheritance of genomic DNA sequence. Most notably, histone proteins must at least transiently dissociate from the maternal genome during replication, and it is unknown whether maternal proteins re-associate with daughter genomes near the sequence they originally occupied on the maternal genome. Here, we use a novel method for tracking old proteins to measure where histone proteins accumulate after 1, 3, or 6 generations of growth in yeast. To our surprise, ancestral histones accumulate near the 5’ end of long, relatively inactive genes. Using a mathematical model, we show that our results can be explained by the combined effects of histone replacement, histone movement along genes from 3’ to 5’, and histone spreading during replication. Our results show that old histones do move, but stay relatively close (~400 bp) to their original location, which places important constraints on how chromatin could potentially carry epigenetic information. Our findings also suggest that accumulation of ancestral histones can influence histone modification patterns.

overall_design: Solexa/Illumina sequencing of H3-HA and H3-T7 MNase-ChIP. Samples of HA and T7 libraries were taken after the tag swap (a) before release from arrest (0 generations), (b) at 1, 3 and 6 cell divisions or (c) after 5 hours of G2/M arrest. Gene deletion mutants for (d) cac1 (e) top1 and (f) H4-tail deletion were taken after 3 cell divisions.

first: Marta

last: Radman-Livaja

first: Kitty

last: Verzijlbergen

first: Tibor

last: van Welsem

first: Nir

last: Friedman

first: Oliver

middle: J

last: Rando

first: Fred

last: van Leeuwen


title: Genome-wide location analysis of  BMP (SMAD1) in mouse erythroid progenitors co-occupted with lineage specific regulators (GATA1, GATA2)

last: Bowman

summary: Lineage regulators direct BMP and Wnt pathways to cell-specific programs during differentiation and regeneration

overall_design: Mouse G1E or G1ER cells were cross-linked with formaldehyde for 20 min. DNA was enriched by chromatin immunoprecipitation (ChIP) and analyzed by Solexa sequencing. A sample of whole cell extract (WCE) was sequenced and used as the background to determine enrichment. ChIP was performed using an antibody against total Smad1 (Santa Cruz SC-7965), Gata1 (Santa Cruz SC-265), or Gata2 (Santa Cruz SC-9008).  This represents the Mouse ChIP-seq portion of this dataset.

first: E

first: T

first: L

last: Lawton

first: Z

last: Fan

first: D

last: Wu

first: A

last: DiBiase

first: C

last: Martin

first: J

last: Cech

last: Sessa

last: Leblanc

first: P

last: Li

last: Durand

last: Mosimann

first: G

last: Heffner

last: Daley

first: R

last: Paulson

middle: A

last: Young

last: Zon


title: CTCF promotes RNA pol II pausing and links DNA methylation to alternative splicing

first: Shalini

last: Oberdoerffer

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

overall_design: Refer to individual Series

first: Sanjeev

last: Shukla

first: Rickard

last: Sandberg


title: Dissecting the retinoid-induced differentiation of F9 embryonal stem cells by integrative genomics [ChIP-seq]

first: Marco Antonio

last: Mendoza-Parra

summary: Retinoic acid (RA) triggers physiological processes by activating heterodimeric transcription factors comprising retinoic acid (RARa,b,g) and retinoid X (RXRa,b,g) receptors. How a single signal induces highly complex temporally controlled networks that ultimately orchestrate physiological processes is unclear. Using an RA-inducible differentiation model we defined the temporal changes in the genome-wide binding patterns of RARg and RXRa and correlated them with transcription regulation. Unexpectedly, both receptors displayed a highly dynamic binding, with different RXRa heterodimers targeting identical loci. Comparison of RARg and RXRa co-binding at RA-regulated genes identified putative RXRa-RARg target genes that were validated with subtype-selective agonists. Gene regulatory decisions during differentiation were inferred from transcription factor target gene information and temporal gene expression.  This analysis revealed 6 distinct co-expression paths of which RXRa-RARg is associated with transcription activation, while Sox2 and Egr1 were predicted to regulate repression. Finally, RXRa-RARg regulatory networks were reconstructed through integration of functional co-citations. Our analysis provides a dynamic view of RA signalling during cell differentiation, reveals RA heterodimer dynamics and promiscuity, and predicts decisions that diversify the RA signal into distinct gene-regulatory programs.

overall_design: RXRa; RARg and RNA Polymerase II chromatin binding has been assessed at five time points; in addition an input control for F9 cells as well as a RXRa ChIP-seq assay from a rxra-/- ko strain has been performed.

first: Marco

first: Mannu

last: Walia

first: Martial

last: Sankar

first: Hinrich

last: Gronemeyer


title: The Estrogen receptor alpha cistrome defined by DamIP

status: Public on Nov 19 2011

city: Houston

first: Rui

last: Xiao

state: Texas

address: One Baylor Plaza

institute: Baylor College of Medicine

summary: We have developed a new method to study DNA-protein interaction in vivo called DamIP, which is based on fusing a protein of interest to a mutant form of DNA adenine methylatransferase (Dam) from E. coli. We showed previously that DamIP can efficiently identify in vivo binding sites of Dam-tethered human estrogen receptor alpha (hERα). In current study, we present the cistrome of hERα determined by DamIP and high throughput sequencing (DamIP-seq). The DamIP-seq defined hERα cistrome overlaps significantly with those determined by ChIP-chip or ChIP-seq, but identifies many new binding regions. As shown by conventional ChIP assay, many DamIP-seq unique hERα binding regions show relatively stable hERα binding, whereas DamIP-seq misses some regions with very transient hERα binding. The methyl-adenine modifications introduced by Dam are stable and do not decrease over 12 days. In summary, the current study provides both an alternate view of the hERα cistrome to further understand the mechanism of hERα mediated transcription, and a new tool to explore other transcriptional factors and cofactors.

overall_design: MCF7 cells were transfected with DamK9A or DamK9A-hERalpha. DamIP were performed from each sample and subjected to solexa sequencing

first: David

middle: D

last: Moore

first: Stephen

last: Ayers

first: Deqiang

last: Sun

first: Yuanxin

last: Xi


title: Targeting the hemangioblast with a novel cell type-specific enhancer

status: Public on Sep 30 2011

city: Oeiras

first: Ana

middle: Teresa

last: Tavares

address: Rua da Quinta Grande, 6

country: Portugal

institute: IGC

summary:
Hemangioblasts are known as the common precursors for primitive hematopoietic and endothelial lineages. Their existence has been supported mainly by the observation that both cell types develop in close proximity and by in vitro differentiation and genetic studies. However, more compelling evidence will arise from tracking their cell fates using a lineage-specific marker. We report the identification of a hemangioblast-specific enhancer (Hb) located in the cis-regulatory region of chick Cerberus gene (cCer) that is able to direct the expression of enhanced green fluorescent protein (eGFP) to the precursors of yolk sac blood and endothelial cells in electroporated chick embryos. Moreover, we present the Hb-eGFP reporter as a powerful live imaging tool for visualizing hemangioblast cell fate and blood island morphogenesis. We hereby introduce the Hb enhancer as a valuable resource for genetically targeting the hemangioblast population as well as for studying the dynamics of vascular and blood cell development.
We used microarray analysis of Hb-eGFP expressing cells to verify the expression of hemangioblast-specific genes in this cell population.

overall_design: In order to characterize the gene expression profile of the Hb-eGFP-positive population, we have isolated Hb-eGFP-positive cells and compared their expression profile with the control RFP-positive population. In brief, chicken embryos were electroporated at HH3 with Hb-eGFP and pCAGGS-RFP reporter constructs, observed under a fluorescence stereoscope and harvested at stage HH5-6 into three groups of four embryos each. Embryos were dissociated into single cell suspensions and the eGFP-positive and eGFP-negative/RFP-positive cell populations were sorted by Fluorescence Activated Cell Sorting (FACS). Cell populations were collected simultaneously into two different tubes, containing RNAlater (Ambion) and subsequently used for RNA extraction. Total RNA was isolated from triplicates of each population, evaluated for RNA integrity, reverse transcribed, amplified and hybridized against six Affymetrix Chicken Genome microarrays.

middle: T


title: Autophagy driven by a master regulator of hematopoiesis

city: MADISON

first: Emery

middle: H

last: Bresnick

state: Wisconsin

address: 1111 Highland Avenue

department: Department of Cell and Regenerative Biology

institute: University of Wisconsin School of Medicine and Public Health

summary: Developmental and homeostatic remodeling of cellular organelles is mediated by a complex process termed autophagy. The cohort of proteins that constitute the autophagy machinery function in a multistep biochemical pathway. Though components of the autophagy machinery are broadly expressed, autophagy can occur in specialized cellular contexts, and mechanisms underlying cell type-specific autophagy are poorly understood.  We demonstrate that the master regulator of hematopoiesis GATA-1 directly activates transcription of genes encoding the essential autophagy component Microtubule Associated Protein 1 Light Chain 3B (LC3B) and its homologs (MAP1LC3A, GABARAP, GABARAPL1, GATE-16). In addition, GATA-1 directly activates genes involved in the biogenesis/function of lysosomes, which mediate autophagic protein turnover. We demonstrate that GATA-1 utilizes the forkhead protein FoxO3 to activate select autophagy genes.  GATA-1-dependent LC3B induction is tightly coupled to accumulation of the active form of LC3B and autophagosomes, which mediate mitochondrial clearance as a critical step in erythropoiesis. These results illustrate a novel mechanism by which a master regulator of development establishes a genetic network to instigate cell type-specific autophagy.

overall_design: Genome-wide maps of GATA1 factor occupancy in primary human PBMC derived erythroblasts

first: Yoon-A

last: Kang

first: Rajendran

last: Sanalkumar

first: Henriette

last: O'Geen

first: Amelia

last: Linnemann

first: Chan-Jung

last: Chang

first: Eric

middle: E

last: Bouhassira

first: Peggy

last: Farnham

first: Sunduz

last: Keles


title: Transcription Factor Binding Sites by ChIP-seq from ENCODE/HAIB

status: Public on Sep 29 2011

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 (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).

The ChIP-Seq method was used to assay chromatin fragments bound by specific or general transcription factors as described below. DNA isolated by ChIP-Seq was size-selected (~225 bp) and sequenced. Short reads of 25-36 bp were mapped to the human reference genome, and enriched regions of high read density relative to a total input chromatin control reads were identified.
The sequence reads with quality scores (fastq files) and alignment coordinates (BAM files) 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://hgwdev.cse.ucsc.edu/ENCODE/protocols/cell). Cross-linked chromatin was immunoprecipitated with an antibody. The Protein:DNA crosslinks were then reversed and the DNA fragments were recovered and sequenced. Please see protocol notes below and go to http://hudsonalpha.org/myers-lab/protocols for the most current version of the protocol. Biological replicates from each experiment were completed.
Libraries were sequenced with an Illumina Genome Analyzer I or an Illumina Genome Analyzer IIx according to the manufacturer's recommendations. Sequence data produced by the Illumina data pipeline software were quality filtered and then mapped to NCBI Build37 (hg19) using the integrated Eland software; 32 nt of the sequence reads were used for alignment; up to two mismatches were tolerated; reads that mapped to multiple sites in the genome were discarded.
To identify likely binding sites, peak calling was applied to the aligned sequence data sets using Model-based Analysis of Chip-Seq MACS (Zhang Y, et al., 2008) (http://liulab.dfci.harvard.edu/MACS/00README.html). MACS models the shift size of ChIP-Seq tags empirically, and uses it to improve the spatial resolution of predicted binding sites. MACS also uses a dynamic Poisson distribution to capture local biases in the genome, allowing for more robust predictions (Zhang Y, et al., 2008).
Protocol Notes:
Several changes and improvements were made to the original ChIP-Seq protocol (Jonshon et al.,2008). The major differences between protocols are the number of cells and magnetic beads used for IP, the method of sonication used to fragment DNA, and the number of cycles of PCR used to amplify the sequencing library. The most current protocol used by the Myers lab can be found at http://hudsonalpha.org/myers-lab/protocols. The protocol field for each file denotes the version of the protocol used as being PCR1x, PCR2x or a version number (for examples, v041610.1).
The sequencing libraries labeled as PCR2x were made with two rounds of amplification (25 and 15 cycles) and those labeled as PCR1x were made with one 15-cycle round of amplification. These experiments were completed prior to January 2010 and were originally aligned to NCBI Build36 (hg18). They have been re-aligned to NCBI Build37 (hg19) with the Bowtie software (Langmead, et al., 2009) for this data release (http://bowtie-bio.sourceforge.net/index.shtml). The libraries labeled with a protocol version number were competed after January 2010 and were only aligned to NCBI Build37 (hg19). Please refer to the Myers Lab website (http://hudsonalpha.org/myers-lab/protocols) for details on each protocol version.
Verification:
The MACS (http://liulab.dfci.harvard.edu/MACS/00README.html) peak caller was used to call significant peaks on the individual replicates of a ChIP-Seq experiment. Afterwards, the irreproducible discovery rate (IDR) method, developed by Li et al. (submitted), was used to quantify the consistency between pairs of ranked peaks lists from replicates. The IDR methods uses a model that assumes that the ranked lists of peaks in a pair of replicates consist of two groups - a reproducible group and an irreproducible group. In general, the signals in the reproducible group are more consistent (i.e. with a larger rank correlation coefficient) and are ranked higher than the irreproducible group. The proportion of peaks that belong to the irreproducible component and the correlation of the reproducible component are estimated adaptively from the data. The model also provides an IDR score for each peak, which reflects the posterior probability of the peak belonging to the irreproducible group. The aligned reads were pooled from all replicates and the MACS peak caller was used to call significant peaks on the pooled data. Only datasets containing at least 100 peaks passing the IDR threshold are considered valid and submitted for release.

first: Richard

last: Myers

first: Flo

last: Pauli


title: Strand-specific transcriptome profiles of the oral pathogen Porphyromonas gingivalis using genomic tiling microarray and RNA sequencing

status: Public on Oct 01 2011

city: Cambridge

first: Tsute

last: Chen

address: 245 First Street

department: Microbiology

institute: The Forsyth Institute

summary: High-density tiling microarray and RNA sequencing technologies were used to analyze the transcriptome of Porphyromonas gingivalis. The compiled P. gingivalis transcriptome were based on total RNA samples isolated from three different laboratory culturing conditions and the strand-specific transcription profiles generated covered the entire genome including both protein coding and non-coding regions. The transcription profiles revealed various operon structures, 5' and 3' end untranslated regions (UTRs), differential expression patterns, and several not-yet annotated transcripts within intergenic and antisense regions. Further transcriptome analysis identified the majority of the genes to be expressed within operons and most 5' and 3' ends to be protruding UTRs of which several 3â UTRs were extended to overlap genes encoded on the opposite/antisense strand. Extensive antisense RNAs were identified opposite to most of insertion sequence (IS) elements. With the growing realization that non-coding RNAs play important biological functions, the comprehensive transcriptome profiles compiled in this study are of great value for the further understanding of gene regulation and virulence mechanism in this periodontal pathogen. The transcriptome profiles can be viewed at and downloaded from the 'Microbial Transcriptome Database' web site http://bioinformatics.forsyth.org/mtd.

overall_design: 3 total RNA samples from growing P. gingivalis strain W83 under different media conditions subjected to RNA-Seq.

first: Hedda

last: Høvik

first: Wen

last: Yu

first: Ingar

last: Olsen


title: High-throughput sequencing of WT and the yqhC mutant of Salmonella using Illumina Genome Analyzer II

city: Madison

first: Nicholas

last: Eakley

state: WI

address: 1675 Observatory Drive

department: Animal Sciences

institute: UW-Madison

summary: Total RNA was isolated from WT and the mutant and the ribosomal RNA was removed using Ambion Kit. Construction of cDNA library, addition of adapters and sequence read were performed.

sample_organism: Salmonella enterica subsp. enterica serovar Typhimurium

overall_design: 2 samples were used, WT Salmonella and its yqhC mutant

first: A

last: Fadl

first: N

first: S

last: Splinter

first: E

last: Cabot


title: Transcriptome Analysis of Human Supragingival Dental Plaque Microbiome from 19 Twin Pairs with and without Dental Caries

status: Public on Sep 26 2011

city: Rockville

first: Vasily

last: Sharov

address: 9704 Medical Center Drive

department: PFGRC

institute: JCVI

summary:
The study aims to assess gene expression in plaque samples collected from twin pairs that are both concordant and discordant with respect to dental Caries diagnosis.   File Naming Conventions are as follows:    Patient ID : 4 digit identifier    Diagnosis : Caries Negative(CN) or Caries Positive(CP)    Type of Twin: Monozygotic(MZ)or Dizygotic(DZ)    Pair to xxxx: 4 digit twin identifier maps to the Patient ID    E.g: 2126_CP_MZ_PairTo_2125_fastqc - 2126 is a caries positive patient and pairs to monozygotic twin pair 2125.
Plaque samples from twin pairs that are both concordant and discordant with respect to dental Caries diagnosis are enriched for bacterial messenger RNA to study the gene expression differences in the samples.

overall_design: RNA was extracted from RNAprotect (Qiagen, In c.) treated dental plaque scrapings from 38 patients. Amplified cDNA was created and rRNA sequence was removed by subtractive hybridization. Individual patient samples were run on a single lane of an Illumina Genome Analyzer.

first: Ana

last: Ong

first: Erik

last: Snesrud

first: Jia

last: Liu

first: John

last: Braisted

first: Walter

last: Bretz

first: Scott

middle: N

last: Peterson


title: de novo DNA Methylation Balances Hematopoietic Stem Cell Self-Renewal and Differentiation

first: deqiang

last: sun

state: TX

address: 2121 W Holcombe Blvd

institute: Texas A&M University

type: Methylation profiling by high throughput sequencing

summary: Cytosine methylation is an epigenetic mark usually associated with gene repression. Despite a requirement for de novo DNA methylation for differentiation of embryonic stem cells, its role in somatic stem cells is unknown. Using conditional ablation, we show that loss of either, or both, Dnmt3a or Dnmt3b, progressively impedes hematopoietic stem cell (HSC) differentiation during serial in vivo passage. Concomitantly, HSC self-renewal is immensely augmented in absence of either Dnmt3, particularly Dnmt3a. Dnmt3-KO HSCs show upregulation of HSC multipotency genes and downregulation of early differentiation factors, and the differentiated progeny of Dnmt3-KO HSCs exhibit hypomethylation and incomplete repression of HSC-specific genes. HSCs lacking Dnmt3a manifest hyper-methylation of CpG islands and hypo-methylation of genes which are highly correlated with human hematologic malignancies. These data establish that aberrant DNA methylation has direct pathologic consequences for somatic stem cell development, leading to inefficient differentiation and maintenance of a self-renewal program.

overall_design: Reduced representation bisulfite sequencing (MspI,~40-220bp size fraction) of secondarily-transplanted wild-type and Dnmt3a conditional knockout hematopoietic stem cells. We used microarrays to detail the global expression of genes in secondarily-transplanted control-HSCs and Dnmt3a-KO-HSCs.

first: Grant

last: Challen

first: Deqiang

last: Sun

first: Mira

last: Jeong

first: Jonathan

middle: S

last: Berg

first: Jaraslav

last: Jelinek

first: Christoph

last: Bock

first: Min

last: Luo

first: Aparna

last: Vasanthakuma

first: Hongcang

last: Gu

first: Yuanxin

last: Xi

first: Shoudan

last: Liang

first: Yue

last: Lu

first: Gretchen

last: Darlington

first: Alexander

last: Meissner

first: John-Pierre

last: Issa

first: Lucy

last: Godley

first: Wei

last: Li

first: Margaret

last: Goodel


title: 2-week-old Arabidopsis seedlings (Columbia ecotype)

status: Public on Sep 24 2011

city: Fort Collins

first: Mark

last: Rogers

state: CO

address: 1873 Campus Delivery

institute: Colorado State University

summary: Comparison of TopHat alignments and assessment of spurious splice junctions for 32nt and 76nt read lengths.  Total RNA from 2-week-old Arabidopsis thaliana (ecotype Columbia) seedlings grown on MS plates was isolated using RNeasy Plant Mini Kit from Qiagen. To remove any contaminating DNA, RNA was treated with DNAse.  Isolation of poly (A) mRNA and preparation of cDNA library were carried out using the Illumina TrueSeq RNA kit. Sequencing (72 cycle) was done on Illumina Genome Analyzer II.

overall_design: 2 replicates


title: A Comparative Profile of the MicroRNA Transcriptome in Immature and Mature Porcine Testes using Solexa Deep Sequencing

status: Public on Sep 23 2011

city: haerbin

first: Chuanjiang

last: Lian

address: maduan Street 427

institute: HVRI,CAAS

summary: MicroRNAs (miRNAs) are small non-coding regulatory RNAs that play key roles in many diverse biological processes such as spermatogenesis. However, no study has been performed on the miRNA transcriptome of developing porcine testes. Here, we employed Solexa deep sequencing technology to extend the repertoire of porcine testis miRNAs and extensively compare the expression patterns of the sexually immature and mature porcine testes. Solexa sequencing of two small RNA libraries derived from immature (30 days) and mature (180 days) pig testis samples yielded over 25 million high-quality reads. Overall, the two developmental stages had significantly different small RNA compositions. A custom data analysis pipeline identified 398 known and/or homologous conserved porcine miRNAs, 15 novel pig-specific miRNAs, and 56 novel candidate miRNAs. We further observed multiple mature miRNA variants (isomiRs) and identified a new bidirectional transcribed miRNA locus, ssc-mir-181a. One hundred twenty-two miRNAs were differentially expressed in the immature and mature testes, and 10 were validated using quantitative RT-PCR. Furthermore, GO and KEGG pathway analyses of the predicted miRNA targets further illustrate the likely roles for these differentially expressed miRNAs in spermatogenesis. This study is the first comparative profile of the miRNA transcriptome in immature and mature porcine testes using a deep sequencing approach, and it provides a useful resource for future studies on the role of miRNAs in spermatogenesis and male infertility treatment.

overall_design: microRNA profiling and discovery in two small RNA cDNA libraries derived from sexually immature (30-day) and mature (180-day) pig testes.

first: Zhihui

last: Zhao


title: Budding yeast mRNA poly(A) site mapping

status: Public on Sep 25 2011

city: Aurora

first: David

middle: L.

last: Bentley

department: Biochemistry and Mol. Genetics

institute: U. Colorado School of Medicine

summary:
The yeast mRNA export adaptor Yra1 binds the Pcf11 subunit of cleavage-polyadenylation factor CF1A linking export to 3'-end formation. We found a surprising consequence of this interaction is that Yra1 influences cleavage-polyadenylation. Yra1 competes with the CF1A subunit, Clp1, for binding to Pcf11, and excess Yra1 inhibits 3' processing in vitro. Release of Yra1 at the 3' ends of genes coincides with recruitment of Clp1, and depletion of Yra1 enhances Clp1 recruitment within some genes. These results suggest that CF1A is not necessarily recruited as a complete unit, but instead Clp1 can be incorporated co-transcriptionally in a process regulated by Yra1. Yra1 depletion causes widespread changes in poly(A) site choice particularly at sites where the efficiency element is divergently positioned. We propose that one way Yra1 modulates cleavage-polyadenylation is by influencing co-transcriptional assembly of the CF1A/B 3' processing factor.

Key Words: Yra1, cleavage-polyadenylation, mRNA export, Pcf11, Clp1, Sub2, alternative polyadenylation

overall_design:
mRNA poly (A) sites were mapped by sequencing 3' ends  in WT and  Yra1-depleted cells using a GAL1-YRA1 mutant.
RNA seq of mRNA 3' ends using Illumina platform.

first: Hyunmin

last: Kim


title: CTCF promotes RNA pol II pausing and links DNA methylation to alternative splicing [ChIP-Seq]

first: Shalini

last: Oberdoerffer

first: Sanjeev

last: Shukla

first: Rickard

last: Sandberg


title: Budding yeast mRNA export and processing factors

type: Genome binding/occupancy profiling by genome tiling array


title: Effect of downregulation of the high grain protein content (GPC) gene on the wheat transcriptome during monocarpic senescence

first: Dario

last: Cantu

address: One Shields Ave

department: Plant Sciences

institute: UC Davis

summary: When GPC transcription factor in hexaploid wheat is down-regulated by stable RNA interference (GPC RNAi), senescence is significantly delayed and grain protein content together with the overall nutrient partitioning to the grain is greatly reduced. mRNA-seq was used to catalogue the genes that are regulated by the GPC transcription factor during monocarpic senescence.

overall_design: cDNA was prepared from wild type bread wheat plants and GPC RNAi  plants 12 days after anthesis and sequenced by Illumina. Four biological replications per genotypes were sequenced. To determine gene expression levels, reads were aligned to a reference transcriptome generated by assemblying 454-reads obtained from the same biological material (454 assembled sequences: TSA project 59945 - accession numbers: HP608076-HP639668).

first: Jorge

last: Dubcovsky


title: Population of viral small RNAs expressed in common marmoset (Callithrix jacchus) T cells latently infected with Herpesvirus saimiri (strain A11)

status: Public on Sep 21 2011

city: Salt Lake City

first: Demian

last: Cazalla

state: UT

address: 15 N Medical Drive East, Rm 4100

department: Biochemistry

institute: University of Utah

summary: We report the discovery of six novel miRNAs expressed by Herpesvirus saimiri (strain A11). These miRNAs are generated by a non-canonical biogenesis pathway that does not require the Microprocessor complex.

overall_design: Examination of one sample prepared from common marmoset (Callithrix jacchus) T cells latently infected with Hespesvirus saimiri (strain A11).

first: DemiÃ¡n

first: Joan

last: Steitz


title: Accurate Identification of A-to-I RNA editing in human by transcriptome sequencing

city: Los Angeles

first: Gang

address: 611 Charles E. Young Drive Rm. 660

institute: UCLA

summary: RNA editing enhances the diversity of gene products at the post-transcriptional level. Approaches for genome-wide identification of RNA editing face two main challenges: separating true editing sites from false discoveries and accurate estimation of editing levels. We developed an approach to analyze transcriptome sequencing data (RNA-Seq) for global identification of RNA editing in cells for which whole-genome sequencing data are available. We applied the method to analyze RNA-Seq data of a human glioblastoma cell line, U87MG. Around 10,000 DNA-RNA differences were identified, the majority being putative A-to-I editing sites. These predicted A-to-I events were associated with a low false discovery rate (~5%). Moreover, the estimated editing levels from RNA-Seq correlated well with those based on traditional clonal sequencing. Our results further facilitated unbiased characterization of the sequence and evolutionary features flanking predicted A-to-I editing sites and discovery of a conserved RNA structural motif that may be functionally relevant to editing. Genes with predicted A-to-I editing were significantly enriched with those known to be involved in cancer, supporting the potential importance of cancer-specific RNA editing. A similar profile of DNA-RNA differences as in U87MG was predicted for another RNA-Seq data set obtained from primary breast cancer samples. Remarkably, significant overlap exists between the putative editing sites of the two transcriptomes despite their difference in cell type, cancer type and genomic backgrounds. Our approach enabled de novo identification of the RNA editome, which sets the stage for further mechanistic studies of this important step of post-transcriptional regulation.

overall_design: Examine mRNA expression in U87MG cells following ADAR1 or control siRNA knockdown

first: Jae-Hoon

last: Bahn

first: Jae-Hyung

last: Lee

first: Christopher

last: Greer

first: Guangdun

last: Peng

first: Xinshu

last: Xiao


title: Bisulfite sequencing of ChIP DNA on breast cancer cells (HCC1954)

status: Public on Sep 16 2011

city: Dallas

first: Gary

middle: Chung

last: Hon

address: 5323 Harry Hines Blvd.

department: OB/GYN

institute: UT Southwestern

summary: While genetic mutation is a hallmark of cancer, many cancers also acquire epigenetic alterations during tumorigenesis including aberrant DNA hypermethylation of tumor suppressors as well as changes in chromatin modifications as caused by genetic mutations of the chromatin-modifying machinery. However, the extent of epigenetic alterations in cancer cells has not been fully characterized. Here, we describe the first complete methylome maps at single nucleotide resolution of a low-passage breast cancer cell line and primary human mammary epithelial cells. We find widespread DNA hypomethylation in the cancer cell, primarily at partially methylated domains (PMDs) in normal breast cells. Unexpectedly, genes within these regions are largely silenced in cancer cells. The loss of DNA methylation in these regions is accompanied by formation of repressive chromatin, with a significant fraction displaying allelic DNA methylation where one allele is DNA methylated while the other allele is occupied by histone modifications H3K9me3 or H3K27me3. Our results show a mutually exclusive and complementary relationship between DNA methylation and H3K9me3 or H3K27me3. These results suggest that global DNA hypomethylation in breast cancer is tightly linked to the formation of repressive chromatin domains and gene silencing, thus identifying a potential epigenetic pathway for gene regulation in cancer cells and suggesting a possible new approach toward the development of cancer therapeutics.

overall_design: ChIP-methylC-Seq on H3K9me3, H3K27me3, and H3K36me3 in breast cancer HCC1954. 36 cycles of sequencing on Illumina platform.

first: Bing

last: Ren


title: Gene expression and H3K9ac genome-wide maps following HDAC inhibition in mouse ES cells

status: Public on Sep 17 2011

first: Eran

last: Meshorer

address: Edmond J. Safra campus

department: Genetics

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

overall_design: Refer to individual Series


title: Genome-wide location analysis in breast cancer cells (HCC1954)

overall_design: ChIP-Seq on H3K4me1, H3K4me3, H3K9me3, H3K27ac, H3K27me3, H3K36me3, and input on breast cancer HCC1954. 36 cycles of sequencing on Illumina platform


title: Gene expression analysis of breast cancer (HCC1954) and normal breast cells (HMEC)

overall_design: mRNA-Seq of polyA-selected RNA from breast cancer HCC1954 and normal breast HMEC. 36 cycles of sequencing on Illumina platform.


title: An Extended dsRBD with a Novel Zinc-Binding Motif Mediates Nuclear Retention of Fission Yeast Dicer

city: Basel

first: Dimos

last: Gaidatzis

address: Maulbeerstrasse 66

institute: Friedrich Miescher Institute

summary: Dicer proteins function in RNA interference (RNAi) pathways by generating small RNAs (sRNAs). Here we report the solution structure of the C-terminal domain of Schizosaccharomyces pombe Dicer (Dcr1). The structure reveals an unusual double-stranded RNA binding domain (dsRBD) fold embedding a novel zinc-binding motif that is conserved among dicers in yeast. Although the C-terminal domain of Dcr1 still binds nucleic acids, this property is dispensable for proper functioning of Dcr1. In contrast, disruption of zinc coordination renders Dcr1 mainly cytoplasmic and leads to remarkable changes in gene expression and loss of heterochromatin assembly. In summary, our results reveal novel insights into the mechanism of nuclear retention of Dcr1 and raise the possibility that this new class of dsRBDs might generally function in nucleo-cytoplasmic trafficking and not substrate binding. The C-terminal domain of Dcr1 constitutes a novel regulatory module that might represent a potential target for therapeutic intervention with fungal diseases.

overall_design: Small RNA libraries from total RNA isolations of wild-type, dcr1Delta, dcr1SHSS, and dcr1Deltaloop2 cells and subjected to high-throughput sequencing.

first: Pierre

last: Barraud

first: Stephan

last: Emmerth

first: Yukiko

last: Shimada

first: Hans-Rudolf

last: Hotz

first: FrÃ©dÃ©ric

last: Allain

first: Marc

last: Buhler


title: Integrated approaches reveal determinants of genome-wide binding and function of the transcription factor Pho4

status: Public on Sep 28 2011

city: Boston

first: Xu

last: Zhou

address: 300 Longwood Ave

department: Pediatrics

institute: Boston Children's Hospital

summary: Binding of transcription factors to DNA is a key regulatory step in the control of gene expression.  DNA sequences with high affinity for transcription factors occur more frequently in the genome than instances of genes bound or regulated by these factors.  Although several mechanisms have been identified that influence the specificity of transcriptional regulation, it is not known if these can explain the observed genome-wide pattern of binding or regulation for a given transcription factor.  We used genome-wide approaches to study how trans influences shape the binding and regulatory landscape of Pho4, a budding yeast transcription factor that activates gene expression in response to phosphate limitation.  We find that nucleosomes significantly restrict the sites to which Pho4 binds. At nucleosome-depleted sites, competition between Pho4 and another transcription factor, Cbf1, determines Pho4 occupancy, raising the threshold for transcriptional activation by Pho4 in phosphate replete conditions and preventing Pho4 activation of genes outside the phosphate regulon during phosphate starvation.  Pho4 binding is not sufficient for transcriptional activation - a cooperative interaction between the transcription factor Pho2 and Pho4 occurs specifically at genes that are activated.  Combining these experimental observations, we are able to globally predict Pho4 binding and its functionality.  Our study provides insights into the mechanisms of global control by sequence-specific transcription factors.

overall_design: ChIP-Seq experiments of Pho4, Pho2 and Cbf1 samples and paired-end nucleosome sequencing in no Pi conditions.

first: Erin

last: O'Shea


title: A role for Snf2 related nucleosome spacing enzymes in genome-wide nucleosome organization

city: Dundee

first: Triantafyllos

last: Gkikopoulos

state: Angus

address: Dow Street

department: College of Life Sciences

country: United Kingdom

institute: University of Dundee

summary: The positioning of nucleosomes within the coding regions of eukaryotic genes is aligned with respect to transcriptional start sites. This organization is likely to influence many genetic processes, requiring access to the underlying DNA. Here we show that the combined action of Isw1 and Chd1 nucleosome spacing enzymes is required to maintain this organization. In the absence of these enzymes regular positioning of the majority of nucleosomes is lost. Exceptions include the region upstream of the promoter, the +1 nucleosome and a subset of locations distributed throughout coding regions where other factors are likely to be involved. These observations indicated that ATP-dependent remodeling enzymes are responsible for directing the positioning of the majority of nucleosomes within the Saccharomyces cerevisiae genome.

overall_design:
Examination of nucleosome positioning in mutants of snf2-related enzymes
Other data used in this study are provided in GEO Series GSE31301 and GSE31833.

first: Pieta

last: Schofield

first: Vijender

last: Singh

first: Jane

last: Mellor

first: Michaela

last: Smolle

first: Geoffrey

last: Barton

first: Jerry

middle: L

last: Workman

first: Marina

last: Pinskaya

first: Tom

last: Owen-Hughes


title: MicroRNA expression during cell cycle arrest

status: Public on Sep 15 2011

first: Olivia

last: Rissland

address: 9 Cambridge Center

institute: Whitehead Institute for Biomedical Research

summary: The miR-16 family, which targets genes important for the G1-S transition, is a known modulator of the cell cycle, and members of this family are often deleted or down-regulated in many types of cancers. Here we report the reciprocal relationship - that of the cell cycle controlling the miR-16 family. Levels of this family increase rapidly as cells are arrested in G0. Conversely, as cells are released from G0 arrest, levels of the miR-16 family rapidly decrease. Such rapid changes are made possible by the unusual instabilities of several family members. The repression mediated by the miR-16 family is sensitive to these cell cycle changes, which suggests that the rapid up-regulation of the miR-16 family reinforces cell cycle arrest in G0. Upon cell cycle re-entry, the rapid decay of several members allows levels of the family to decrease, alleviating repression of target genes and allowing proper resumption of the cell cycle.

overall_design: Small RNAs were profiled by high-throughput sequencing either during synchronous release after serum starvation or during cell-cycle arrest by contact inhibition.

first: Sue-Jean

last: Hong

first: David

middle: P

last: Bartel


title: BCL6 is required for the initiation and maintenance of chronic myeloid leukemia

city: New York

first: Katerina

last: Hatzi

state: NY

address: 413 E 69th Street, BB-1462

department: Hematology/Oncology

institute: WCMC

summary: We identified the BCL6 protooncogene as a critical downstream effector of FoxO3A in self-renewal signaling of CML-initiating cells. BCL6 represses Arf and p53 in CML cells and is required for leukemia stem cell maintenance, colony formation and initiation of leukemia in transplant recipients. Importantly, peptide inhibition of BCL6 in human CML cells compromises colony formation and leukemia-initiation in xenotransplanted mouse recipients. These findings identify peptide-inhibition of BCL6 as a novel strategy to eradicate leukemia-initiating cells in CML.

overall_design: Identification of BCL6 binding sites in human CML cell line JURL-MK1

first: Christian

last: Hurtz

first: Markus

last: Müschen


title: An Alternative Splicing Switch Regulates Embryonic Stem Cell Pluripotency and Reprogramming [RNA-Seq]

first: Xinchen

last: Wang

address: 32 Vassar St, D514

institute: MIT

summary: Alternative splicing (AS) is a key process underlying the expansion of proteomic diversity and the regulation of gene expression. However, the contribution of AS to the control of embryonic stem cell (ESC) pluripotency is not well understood. Here, we identify an evolutionarily conserved ESC-specific AS event that changes the DNA binding preference of the forkhead family transcription factor FOXP1. We show that the ESC-specific isoform of FOXP1 stimulates the expression of transcription factor genes required for pluripotency including OCT4, NANOG, NR5A2 and GDF3, while concomitantly repressing genes required for ESC differentiation. Remarkably, this isoform also promotes the maintenance of ESC pluripotency and the efficient reprogramming of somatic cells to induced pluripotent stem cells. These results thus reveal that an AS switch plays a pivotal role in the regulation of pluripotency and functions by controlling critical ESC-specific transcriptional programs.

overall_design:
Exons 18 and 18b form a mutually exclusive splicing event. The FOXP1 (non-ES) isoform contains only exon 18 and not 18b, while the FOXP1-ES isoform contains only exon 18b and not 18.
To investigate whether FOXP1 and FOXP1-ES control different sets of genes, we performed knockdowns using custom siRNA pools targeting FOXP1 exons 18 or 18b in undifferentiated H9 cells, followed by RNA-Seq profiling.

first: Mathieu

last: Gabut

first: Payman

last: Samavarchi-Tehrani

first: Valentina

last: Slobodeniuc

first: Dave

last: O'Hanlon

first: Hoon-Ki

last: Sung

first: Manuel

last: Alvarez

first: Shaheynoor

last: Talukder

first: Qun

last: Pan

first: Knut

last: Woltjen

first: Esteban

middle: O

last: Mazzoni

first: Stephane

last: Nedelec

first: Hynek

last: Wichterle

first: Timothy

middle: R

last: Hughes

first: Peter

last: Zandstra

first: Andras

last: Nagy

first: Jeffrey

last: Wrana

first: Benjamin

last: Blencowe


title: An Alternative Splicing Switch Regulates Embryonic Stem Cell Pluripotency and Reprogramming [ChIP-Seq]

first: Xinchen

last: Wang

overall_design: To identify genes potentially directly regulated by FOXP1-ES and FOXP1 in H9 ESCs, we performed chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-Seq). Using an antibody capable of efficiently immunoprecipitating both isoforms, >3400 significant ChIP-Seq peaks were detected across the human genome. To assess if these peaks are sites of FOXP1 and FOXP1-ES occupancy, we determined whether they are significantly enriched in individual PBM-derived 8-mers that bind to either or both isoforms.

first: Mathieu

last: Gabut


title: The ERI-6/7 helicase acts at the first stage of an siRNA amplification pathway that targets recent gene duplications.

first: Taiowa

last: Montgomery

address: 185 Cambridge Street, CPZN-7250

department: Molecular Biology

institute: Massachusetts General Hospital

summary: To characterize the role of the ERI-6/7 helicase in endogenous small RNA pathways in C. elegans, small RNA populations from null alleles of eri-6 and eri-7, and from mutants of known endogenous RNAi pathway factors, eri-1 and ergo-1, were determined by deep sequencing, and compared to wild type.

overall_design: Small RNA analysis in wild type and eri-1, ergo-1, eri-6 and eri-7 mutant C. elegans strains.

first: Sylvia

last: Fischer


title: Regions of Focal DNA Hypermethylation and Long-Range Hypomethylation in Colorectal Cancer Coincide with Nuclear Lamina-Associated Domains

status: Public on Sep 27 2011

last: Berman

department: USC Epigenome Center

institute: University of Southern California

summary:
Extensive changes in DNA methylation are common in cancer and may contribute to oncogenesis through transcriptional silencing of tumor suppressor genes. Genome-scale studies have yielded important insights into these changes, but have focused on CpG islands or gene promoters. We used whole-genome bisulfite sequencing (Bisulfite-Seq) to comprehensively profile a primary human colorectal tumor and adjacent-normal colon tissue at single-basepair resolution. Regions of focal hypermethylation in the tumor were located primarily at CpG islands and were concentrated within regions of long-range (>100 kb) hypomethylation. These hypomethylated domains covered nearly half the genome and coincided with late replication and attachment to the nuclear lamina in human cell lines. The confluence of hypermethylation and hypomethylation within these domains was confirmed in 25 diverse colorectal tumors with matched adjacent tissue. We propose that widespread DNA methylation changes in cancer are linked to silencing programs orchestrated by the 3D organization of chromatin within the nucleus.
dbGAP study: phs000385

overall_design: Primary tissue samples sequenced using 76-bp bisulfite sequencing (WGBS or Methyl-seq) using the Illumina GAII platform. Two independent libraries were constructed for each sample, and these libraries were combined into a single data file for each sample.

first: D

last: Weisenberger

first: P

middle: W

last: Laird


title: An Alternative Splicing Switch Regulates Embryonic Stem Cell Pluripotency and Reprogramming

platform_organism:
Homo sapiens
synthetic construct


title: Dual Role of FoxA1 in Androgen Receptor Binding to Chromatin, Androgen Signaling and Prostate Cancer

status: Public on Sep 14 2011

city: Helsinki

first: Olli

middle: A.

last: Jänne

department: Inst. of Biomedicine/Physiology

country: Finland

institute: University of Helsinki

first: Biswajyoti

last: Sahu

first: Marko

last: Laakso

first: Kristian

last: Ovaska

first: Tuomas

last: Mirtti

first: Johan

last: Lundin

first: Antti

last: Rannikko

first: Anna

last: Sankila

first: Juha-Pekka

last: Turunen

first: Mikael

first: Juho

last: Konsti

first: Tiina

last: Vesterinen

first: Stig

last: Nordling

last: Kallioniemi

first: Sampsa

last: Hautaniemi


title: RNA-seq expression profiles during terminal erythropoiesis

first: Bill

last: Wong

institute: Whitehead Institute

summary: It is unclear how epigenetic changes regulate the induction of erythroid-specific genes during terminal erythropoiesis. Here we use global mRNA sequencing (mRNA-seq) and chromatin immunoprecipitation coupled to high-throughput sequencing (CHIP-seq) to investigate the changes that occur in mRNA levels, RNA Polymerase II (Pol II) occupancy and multiple post-translational histone modifications when erythroid progenitors differentiate into late erythroblasts. Among genes induced during this developmental transition, there was an increase in the occupancy of Pol II, the activation marks H3K4me2, H3K4me3, H3K9Ac and H4K16Ac, and the elongation methylation mark H3K79me2. In contrast, genes that were repressed during differentiation showed relative decreases in H3K79me2 levels yet had levels of Pol II binding and active histone marks similar to those in erythroid progenitors. We also found that relative changes in histone modification levels-in particular, H3K79me2 and H4K16ac-were most predictive of gene expression patterns. Our results suggest that in terminal erythropoiesis both promoter and elongation-associated marks contribute to the induction of erythroid genes, while gene repression is marked by changes in histone modifications mediating Pol II elongation. Our data maps the epigenetic landscape of terminal erythropoiesis and suggests that control of transcription elongation regulates gene expression during terminal erythroid differentiation.

overall_design: Mouse fetal liver cells are double-labeled for erythroid-specific TER119 and non erythroid-specific transferrin receptor (CD71) and then sorted by flow-cytometry. E14.5 fetal livers contain at least five distinct populations of cells (R1 through R5); as they progressively differentiate they gain TER119 and then gain and subsequently lose CD71.  CFU-E cells and proerythroblasts make up the R1 population; R2 consists of proerythroblasts and early basophilic erythroblasts; R3 includes early and late basophilic erythroblasts; R4 is mostly polychromatophilic and orthochromatophilic erythroblasts; and R5 is comprised of late orthochromatophilic erythroblasts and reticulocytes. We have sorted for R2-R5 cells for RNA-seq experiment.

first: B

first: A

last: Cheng


title: Gene induction and repression during terminal erythropoiesis are mediated by distinct epigenetic changes.


title: Genome-wide H3K9 acetylation state before and after VPA treatment

city: Jerusalem

first: Eran

last: Meshorer

institute: The Hebrew University of Jerusalem

summary: Genome-wide maps of the H3K9 acetylation state in embryonic stem cells (ESCs) before and after treatment with low levels of the histone deacetylase (HDAC) inhibitor valproic acid (VPA).

overall_design: ChIP-seq for 3 samples: untreated E14 cells, cells treated with VPA for 4 hrs and cells treated with VPA for 16 hrs. Unprecipitated DNA was used as the input control (Input).

first: Hadas

last: Hezroni


title: Epigenetic profiling of hematopoietic stem cells and leukemia stem cells

status: Public on Sep 13 2011

first: Amit

last: Sinha

address: 44 Binney St

department: Pediatric Oncology

institute: Dana-Farber Cancer Institute

summary: The histone 3 lysine 79 (H3K79) methyltransferase Dot1l has been implicated in the development of leukemias bearing translocations that involve the Mixed Lineage Leukemia (MLL) gene. We identified the MLL-fusion targets in a murine MLL-AF9 leukemia model, and  conducted epigenetic profiling for H3K79me2, H3K4me3, H3K27me3 and H3K36me3. Histone methylation patterns are highly abnormal on MLL-AF9 fusion target loci, defining a distinct epigenetic lesion involving H3K79. Conditional inactivation of Dot1l  leads to specific down-regulation of direct MLL-AF9 targets and an MLL-translocation associated gene expression signature, while global transcription levels remain largely unaffected. This correlated with a greater sensitivity of leukemic blasts towards loss of Dot1l compared to normal hematopoietic cells. Development of in vivo leukemia was absolutely dependent on Dot1l.

overall_design: Chromatin immunoprecipitation followed by Solexa sequencing for H3K4me3, H3K27me3, H3K36me3, H3K79me2 and biotinylated MLL-AF9 in HSC, GMP and LSC.

last: Zhu

first: Scott

last: Armstrong


title: Dual Role of FoxA1 in Androgen Receptor Binding to Chromatin, Androgen Signaling and Prostate Cancer [ChIP_seq, DHS_seq]

first: Olli

last: Jänne

summary: We report the dual role of FoxA1 in androgen receptor recruitment to the chromatin of androgen responsive prostate cancer cell line LNCaP-1F5 using ChIP-sequencing. Depletion of FoxA1 reprograms both androgen and glucocorticoid receptor recruitment and subsequent gene expression. The ChIP-seq has been performed using AR, FoxA1, GR, H3K4me2 antibodies. We have also mapped the DNaseI-hypersensitive sites (DHS) using deep sequencing.

overall_design: Examination of AR, FoxA1, GR, H3K4me2 binding sites and DHS sites in parental and FoxA1 depleted LNCaP-1F5 cells.

first: Biswajyoti

last: Sahu

first: Marko

last: Laakso

first: Kristian

last: Ovaska

first: Tuomas

last: Mirtti

last: JÃ¤nne


title: Global DNA Hypomethylation Coupled to Repressive Chromatin Domain Formation and Gene Silencing in Breast Cancer

first: Gary

last: Hon

summary:
While genetic mutation is a hallmark of cancer, many cancers also acquire epigenetic alterations during tumorigenesis including aberrant DNA hypermethylation of tumor suppressors as well as changes in chromatin modifications as caused by genetic mutations of the chromatin-modifying machinery. However, the extent of epigenetic alterations in cancer cells has not been fully characterized. Here, we describe the first complete methylome maps at single nucleotide resolution of a low-passage breast cancer cell line and primary human mammary epithelial cells. We find widespread DNA hypomethylation in the cancer cell, primarily at partially methylated domains (PMDs) in normal breast cells. Unexpectedly, genes within these regions are largely silenced in cancer cells. The loss of DNA methylation in these regions is accompanied by formation of repressive chromatin, with a significant fraction displaying allelic DNA methylation where one allele is DNA methylated while the other allele is occupied by histone modifications H3K9me3 or H3K27me3. Our results show a mutually exclusive and complementary relationship between DNA methylation and H3K9me3 or H3K27me3. These results suggest that global DNA hypomethylation in breast cancer is tightly linked to the formation of repressive chromatin domains and gene silencing, thus identifying a potential epigenetic pathway for gene regulation in cancer cells and suggesting a possible new approach toward the development of cancer therapeutics.

This SuperSeries is composed of the SubSeries listed below.

overall_design: Examination of histone modifications, DNA methylation, and gene expression in HCC1954 breast cancer cells and HMEC primary human mammary epithelial cells

middle: C

first: Bing

last: Ren


title: small RNA profiling in human cell line following Dicer silencing

city: Berlin

first: Marc

middle: Riemer

last: Friedlaender

state: Berlin

address: Robert-Rössle-Str. 10

country: Germany

institute: MDC, Berlin-Buch

summary: microRNAs (miRNAs) are a large class of small non-coding RNAs which post-transcriptionally regulate the expression of a large fraction of all animal genes and are important in a wide range of biological processes. Recent advances in high-throughput sequencing allow miRNA detection at unprecedented sensitivity, but the computational task of accurately identifying the miRNAs in the background of sequenced RNAs remains challenging. For this purpose we have designed miRDeep2, a substantially improved algorithm which identifies canonical and non-canonical miRNAs such as those derived from transposable elements and informs on high-confidence candidates that are detected in multiple independent samples. Analyzing data from seven animal species representing the major animal clades, miRDeep2 identified miRNAs with an accuracy of 98.6-99.9% and reported hundreds of novel miRNAs. To test the accuracy of miRDeep2, we knocked down the miRNA biogenesis pathway in a human cell line and sequenced small RNAs before and after. The vast majority of the >100 novel miRNAs expressed in this cell line were indeed specifically down-regulated, validating most miRDeep2 predictions.  Last, a new miRNA expression profiling routine, low time and memory usage and user-friendly interactive graphic output can make miRDeep2 useful to a wide range of researchers."

overall_design: high-throughput sequencing was used to profile small RNA expression in a human MCF-7 cell line before and after Dicer knock-down

last: Friedländer

first: Sebastian

last: Mackowiak

last: Li

first: Wei

last: Chen

first: Nikolaus

last: Rajewsky


title: Genome-wide DNA methylation mapping in breast cancer cells (HCC1954) and normal breast cells (HMEC)

overall_design: methylC-Seq on breast cancer HCC1954 and normal breast HMEC. 100 cycles of sequencing on Illumina platform.


title: RNA Capture-Seq resolves the deep complexity of the human transcriptome

status: Public on Sep 12 2011

city: Sydney

first: Tim

middle: R.

last: Mercer

state: New South Wales

address: 384 Victoria Street Darlinghurst NSW 2010

country: Australia

institute: Garvan

overall_design: Refer to individual Series


title: Directional DNA methylation changes and complex intermediate states accompany lineage specificity in the adult hematopoietic compartment

first: Andrew

middle: D

last: Smith

state: CA

address: 1050 Childs Way

department: Molecular and Computational Biology

summary: DNA methylation has been implicated as an epigenetic component of mechanisms that stabilize cell fate decisions. Here, we have characterized the methylomes of human female hematopoietic stem/progenitor cells (HSPCs) and mature cells from the myeloid and lymphoid lineages. Hypomethylated regions (HMRs) associated with lineage-specific genes were often methylated in the opposing lineage. In HSPCs, these sites tended to show intermediate, complex patterns that resolve to uniformity upon differentiation, by increased or decreased methylation. Promoter HMRs shared across diverse cell-types typically display a constitutive core that expands and contracts in a lineage-specific manner to fine-tune the expression of associated genes. Many newly identified intergenic HMRs, both constitutive and lineage-specific, were enriched for factor binding sites with an implied role in genome organization and regulation of gene expression, respectively. Overall, our studies represent an important reference dataset and provide insights into directional changes in DNA methylation as cells adopt terminal fates.

overall_design: Genomic DNA from purified human peripheral blood cells was isolated, bisulfite converted and sequenced using the Illumina GA2 paltform

first: Emily

last: Hodges

first: Antoine

last: Molaro

first: Camila

last: Dos Santos

first: Pramod

last: Thekkat

first: Qiang

last: Song

first: Philip

last: Uren

first: Jin

last: Park

first: Jason

last: Butler

first: Shahin

last: Rafii

first: W

last: McCombie

first: Gregory

middle: J

last: Hannon