title: Profiling of Argonaute (AGO) 2-associated sRNAs after infection of bacterial pathogen Pseudomonas syringae

city: Hsin-Chu

first: Wei-Chi

last: Wang

address: Room 112 Zhu-Ming Building, 75 Bo-Ai Street

institute: Institute of Bioinformatics and Systems Biology, NCTU

summary: Transgenic Arabidopsis plants (AGO2::HA:AGO2) were treated with either mock (10 mM MgCl2) or Pseudomonas syringae pv. tomato (Pst) expressing avrRpt2 (R2) at a concentration of 2 x 107 cfu/ml for 14 hours. sRNAs associated with AGO1 and AGO2 were co-immunoprecipitated using antibodies against either AGO1 (AGO1-IP), or HA (hemagglutinin) (AGO2-IP). As controls, we also gel-purified the 18-28 nt fraction of the total RNAs from  an AGO2 mutant (ago2-1). The co-immunoprecipitated or gel-purified RNAs were cloned and sequenced by Illumina deep sequencing.

overall_design: Examination of AGO-associated sRNAs in pathogen-treated or control plants

first: Hongwei

last: Zhao

first: Hailing

last: Jin


title: Global Analysis of the Immediate Transcriptional Effects of Estrogen Signaling Reveals a Rapid, Extensive, and Transient Response

status: Public on May 06 2011

city: Dallas

first: W. Lee

last: Kraus

state: TX

address: 5323 Harry Hines Blvd.

country: USA

institute: UT Southwestern Medical Center

type: Expression profiling by high throughput sequencing

summary:
We report the immediate effects of estrogen signaling on the transcriptome of breast cancer cells using Global Run-On and sequencing (GRO-seq). We found that estrogen signaling directly regulates a strikingly large fraction of the transcriptome in a rapid, robust, and unexpectedly transient manner. In addition to protein-coding genes, estrogen regulates the distribution and activity of all three RNA polymerases, and virtually every class of non-coding RNA that has been described to date. This data submission covers >95% of mapped reads comprising nearly all transcript classes described.  Reads mapping to intergenic and enhancer transcripts were removed from this data submission and will be reported separately (manuscripts in preparation).

Bed files are tab-separated text files in which columns represent: chrom, chromStart (5' End of the read), chromEnd (chromStart+1), name (unused always 'n'), score (the number of mismatches), and strand.  Note that because of the inclusion of reads mapping to the rRNA chromosome, bed files cannot be uploaded to the UCSC genome browser directly.  Instead, use the wiggle files (coming soon!) for this purpose.

overall_design: Using GRO-seq over a time course (0, 10, 40, 160 min) of estrogen signaling in ER-alpha positive MCF-7 human breast cancer cells.

first: Nasun

last: Hah

first: Charles

middle: G

last: Danko

first: Leighton

last: Core

first: Joshua

middle: J

last: Watterfall

first: Adam

last: Siepel

first: John

middle: T

last: Lis

first: W

middle: L


title: Genome-wide remodeling of the epigenetic landscape during myogenic differentiation

status: Public on May 08 2011

city: New York

first: Brian

last: Dynlacht

address: 550 1st avenue

institute: NYU Langone Medical Center

type: Genome binding/occupancy profiling by high throughput sequencing

summary: We have examined changes in the chromatin landscape during muscle differentiation by mapping the genome-wide location of ten key histone marks and transcription factors in mouse myoblasts and terminally differentiated myotubes, providing an exceptionally rich dataset that has enabled discovery of key epigenetic changes underlying myogenesis. Using this compendium, we focused on a well-known repressive mark, histone H3 lysine 27 trimethylation, and identified novel regulatory elements flanking the myogenin gene that function as a key differentiation-dependent switch during myogenesis. Next, we examined the role of Polycomb-mediated H3K27 methylation in gene repression by systematically ablating components of both PRC1 and PRC2 complexes. Surprisingly, we found mechanistic differences between transient and permanent repression of muscle differentiation and lineage commitment genes and observed that the loss of PRC1 and PRC2 components produced opposing differentiation defects. These phenotypes illustrate striking differences as compared to embryonic stem cell differentiation and suggest that PRC1 and PRC2 do not operate sequentially in muscle cells. Our studies of PRC1 occupancy also suggested a "fail-safe" mechanism, whereby PRC1/Bmi1 concentrates at genes specifying nonmuscle lineages, helping to retain H3K27me3 in the face of declining Ezh2-mediated methyltransferase activity in differentiated cells.

overall_design: Mapping of PoII, H3K4me1, H3K4me2, H3K4me3, H3K27me3, H3K36me3, H3K9Ac, H3K18Ac and H4K12Ac in growing myoblasts (MB) and fully differentiated myotubes (MT).

first: Patrik

last: Asp

first: Roy

last: Blum

first: Vasupradha

last: Vethantham

first: Fabio

last: Parisi

first: Mariann

last: Micsinai

first: Jemmie

last: Cheng

first: Christopher

last: Bowman

first: Yuval

last: Kluger

middle: D


title: seq-SDQ4663_RPC1_N2_Mxemb

status: Public on May 02 2011

city: Chapel Hill

first: Tristan

last: De Buysscher

state: NC

address: CB# 3280, Coker Hall

department: Biology

institute: University of North Carolina at Chapel Hill

summary:
ChIP-seq analysis of elements that specify nucleosome positioning and occupancy, control domains of gene expression, induce repression of the X chromosome, guide mitotic segregation and genome duplication, govern homolog pairing and recombination during meiosis, and organize chromosome positioning within the nucleus.

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: ChIP-seq against RPC-1 in L3 stage N2 worms.

first: Jason

last: Lieb

first: Kohta

last: Ikegami


title: DNA methylation analysis of prostate cancer cell lines and tissues using Next Generation Sequencing

status: Public on May 09 2011

city: Ann Arbor

first: Jung

last: Kim

state: MI

address: 1400 E. Medical Center Drive

institute: University of Michigan

type: Methylation profiling by high throughput sequencing

summary: Beginning with precursor lesions, aberrant DNA methylation marks the entire spectrum of prostate cancer progression.  We mapped the global DNA methylation patterns in prostate tissues (n=17) and cells (n=2) from fifty nanograms of genomic DNA using Methylplex-Next Generation Sequencing (M-NGS).    A Hidden Markov Model (HMM)-based algorithm previously used for Chip-Seq data analysis(http://www.sph.umich.edu/csg/qin/HPeak) was used to locate peaks from mapped reads obtained in each sequencing run. The total methylation events in intergenic/intronic regions between benign adjacent and cancer tissues were comparable.  While approximately 20% of all CpG islands (CGIs) (68,508) were methylated in tissues, promoter CGI methylation gradually increased from ~12.6% in benign samples to 19.3% and 21.8% in localized and metastatic cancer tissues.  We found distinct patterns in promoter methylation around transcription start sites, where methylation occurred directly on the CGIs, flanking regions and on CGI sparse promoters.  Among the 6,691 methylated promoters in prostate tissues, 2481 differentially methylated regions (DMRs) are cancer specific and several previously studied targets were among them.   A novel cancer specific DMR in WFDC2 promoter showed 77% methylation in cancer (17/22), 100% methylation in transformed prostate cell lines (6/6), none in the benign tissues (0/10) and normal PrEC cells.  Integration of LNCaP DNA methylation and H3K4me3 data suggested a role for DNA methylation in alternate transcription start site utilization.  While methylated promoters containing CGIs had mutually exclusive H3K4me3 modification, the histone mark was absent in CGI sparse promoters.   Finally, we observed difference in methylation of LINE-1 elements between transcription factor ERG positive and negative cancers.  The comprehensive methylome map presented here will further our understanding of epigenetic regulation of the prostate cancer genome.

overall_design: We mapped the global DNA methylation patterns in prostate tissues (n=17; data not available in GEO - being deposited in dbGaP for controlled access) and cells (n=2) from fifty nanograms of genomic DNA using Methylplex-Next Generation Sequencing (M-NGS).  For replicate analysis in cell lines, a total of 4 runs were completed for PrEC prostate normal cell line, and 5 runs were completed for LNCaP prostate cancer cell line. For tissue samples, 2 benign prostate samples were ran twice on illumina next generation sequencing platform to access overall repeatability of M-NGS.

middle: H

first: Saravana

middle: M

last: Dhanasekaran

first: Jindan

last: Yu

first: Daniel

last: Robinson

middle: R

last: Prensner

first: Ming

last: Hu

first: Steve

last: Qin

first: Arul

last: Chinnaiyan


title: seq-WA30534819_H3K4me3_N2_Mxemb

overall_design: ChIP-seq against H3K4me3 in L3 stage N2 worms.

first: Sevinc

last: Ercan


title: seq-SDQ4470_TAG315_N2_MXemb

overall_design: ChIP-seq against  TAG-315 in L3 stage N2 worms.


title: seq-WA30932379_H3K9ac_N2_L3

overall_design: ChIP-seq against H3K9ac in L3 stage N2 worms.

first: Julie

last: Ahringer

first: Isabel

last: Latorre


title: seq-WA30534819_H3K4me3_N2_L3


title: seq-SDQ4526_SFC1_N2_Mxemb

overall_design: ChIP-seq against SFC-1 in L3 stage N2 worms.


title: seq-SDQ2370_LIN53_N2_L3

overall_design: ChIP-seq against LIN-53 in L3 stage N2 worms.


title: seq-SDQ0820_EPC1_N2_L3_EGS

overall_design: ChIP-seq against EPC-1 in L3 stage N2 worms.


title: seq-SDQ3590_EFL1_N2_L3

overall_design: ChIP-seq against EFL-1 in L3 stage N2 worms.


title: seq-JL00002_SDC3_N2_L3

overall_design: ChIP-seq against SDC3 in L3 stage N2 worms.


title: The human TTAGGG Repeat Factors 1 and 2 bind to a subset of interstitial telomeric sequences and satellite repeats

status: Public on May 04 2011

city: Valbonne - Sophia-Antipolis

first: Kevin

last: Lebrigand

address: 660 route des lucioles

country: France

institute: IPMC/CNRS

summary: The study of the proteins that bind to telomeric DNA in mammals has provided a deep understanding of the mechanisms of chromosome-end protection. However, very little is known on the binding of these proteins to nontelomeric DNA sequences. The TTAGGG DNA repeat proteins 1 and 2 (TRF1 and TRF2) bind to mammalian telomeres as part of the shelterin complex and are essential for maintaining chromosome end stability. In this study, we combined chromatin immunoprecipitation with high-throughput sequencing to map at high sensitivity and resolution, the human chromosomal sites to which TRF1 and TRF2 bind. While most of the identified sequences correspond to telomeric regions, we showed that these two proteins also bind to extratelomeric sites. The vast majority of these extra-telomeric sites contains interstitial telomeric sequences (or ITSs). However we also identified non-ITS sites, which are also satellite DNA but the ones mainly constitutive of centromeric and pericentromeric regions. Interestingly, the TRF-binding sites are often located in the proximity of genes or within introns. We propose that, by binding to extratelomeric sequences, TRF1 and TRF2 couple the functional state of telomeres to the long-range organization of chromosomes and gene regulation networks.

overall_design: ChIP-SEQ experiment of transformed human fibroblast BJ cells with 3 antibodies (1 monoclonal anti-TRF1, 1 monoclonal anti-TRF2, 1 polyclonal anti-TRF2) and a negative control (proteinG without antibody used as the ChIP background)

first: Thomas

last: Simonet

first: Laure-Emmanuelle

last: Zaragosi

first: Claude

last: Philippe

first: Clémentine

last: Schouteden

first: Adeline

last: Augereau

first: Serge

last: Bauwens

first: Jing

last: Ye

first: Marco

last: Santagostino

first: Elena

last: Giulotto

first: Frédérique

last: Magdinier

first: Béatrice

last: Horard

first: Pascal

last: Barbry

first: Rainer

last: Waldmann

first: Eric

last: Gilson


title: seq-JA00011_LIN35_N2_L3

first: Tristan

last: De Buysscher

overall_design: ChIP-seq against LIN-35 in L3 stage N2 worms.

first: Julie

last: Ahringer


title: seq-LPAR109_H4tetraac_109_N2_L3

overall_design: ChIP-seq against H4tetraac in L3 stage N2 worms.

first: Isabel

last: Latorre


title: RNA-Seq anlalysis of prostate cancer cell lines using Next Generation Sequencing

first: Jung

last: Kim

summary: Beginning with precursor lesions, aberrant DNA methylation marks the entire spectrum of prostate cancer progression. We mapped the global DNA methylation patterns in selected prostate tissues and cell lines using Methylplex-Next Generation Sequencing (M-NGS). Hidden Markov Model based next generation sequence analysis identified ~68,000 methylated regions per sample. While global CpG Island (CGI) methylation was not differential between benign adjacent and cancer samples, overall promoter CGI methylation increased from ~12.6% in benign samples to 19.3% and 21.8% in localized and metastatic cancer tissues, respectively. We found distinct patterns of promoter methylation around transcription start sites, where methylation occurred not only on the CGIs, but also on flanking regions and CGI sparse promoters. Among the 6,691 methylated promoters in prostate tissues, 2481 differentially methylated regions (DMRs) are cancer specific, including numerous novel DMRs. A novel cancer specific DMR in WFDC2 promoter showed heavy methylation in cancer (17/22 tissues, 6/6 cell lines), but not in the benign tissues (0/10) and normal PrEC cells. Integration of LNCaP DNA methylation and H3K4me3 data suggested an epigenetic mechanism for alternate transcription start site utilization and these modifications segregated into distinct regions when present on the same promoter. Finally, we observed differences in repeat element methylation, particularly LINE-1, between ERG gene fusion positive and negative cancers. This comprehensive methylome map will further our understanding of epigenetic regulation in prostate cancer progression.

overall_design: Next generation Sequencing for Gene expression using the RNA-Seq methodology from LNCaP and PrEC cell lines

first: Mohan

last: Dhanasekaran

first: Arul

last: Chinaiyan


title: seq-JA00001_HTZ1_N2_L3

overall_design: ChIP-seq against HTZ-1 in L3 stage N2 worms.


title: seq-AB1791_H3_N2_L3

overall_design: ChIP-seq against H3 in L3 stage N2 worms.


title: seq-BH00001_LIN52_N2_L3

overall_design: ChIP-seq against LIN-52 in L3 stage N2 worms.


title: seq-BH00004_LIN54_N2_L3

overall_design: ChIP-seq against LIN54 in L3 stage N2 worms.


title: seq-BH00003_LIN37_N2_L3

overall_design: ChIP-seq against LIN-37 in L3 stage N2 worms.


title: seq-AR0144_H3_144_N2_L3


title: seq-SDQ3599_DPL1_N2_L3

overall_design: ChIP-seq against DPL-1 in L3 stage N2 worms.


title: Comparative transcriptomic analysis of prostate cancer and matched normal tissue using RNA-seq

city: Rochester

first: LIGUO

last: WANG

state: MN

address: 200 1st St SW

department: Division of Computational Biology

institute: Mayo Clinic

summary: We used RNA-seq to interrogate prostate cancer specific gene fusions, alternative splicings, somatic mutations and novel transcripts.

overall_design: We sequenced the transcriptome (polyA+) of 20 prostate cancer tumors and 10 matched normal tissues using Illumina GAII platform. Then we used bioinformatic approaches to identify prostate cancer specific aberrations which include gene fusion, alternative splicing, somatic mutation, etc.

first: Liguo

last: Wang

first: Kalpana

last: Kannan

first: Laising

last: Yen

first: Wei

last: Li


title: High-resolution analysis of parent-of-origin-allelic expression in Arabidopsis seeds

status: Public on Apr 30 2011

city: Vienna

first: Marc

last: Rehmsmeier

address: Dr. Bohr-Gasse 3

country: Austria

institute: GMI - Gregor Mendel Institute of Molecular Plant Biology GmbH

summary: We report the identification of parent-of-origin specific gene expression in Arabidopsis seeds. F1 hybrid seeds were generated by reciprocally crossing  Arabidopsis accessions Col-0 and Bur-0. Seeds were harvested at 4 DAP and total RNA was isolated and analyzed by deep sequencing.

overall_design: Overall identification of parent-of-origin-allelic expression by analysis of SNP distribution

first: Claudia

last: Köhler


title: Deep sequencing reveals distinct patterns of DNA methylation and transcript isoform regulation in prostate cancer

status: Public on May 03 2011

type: Methylation profiling by genome tiling array

summary:
Beginning with precursor lesions, aberrant DNA methylation marks the entire spectrum of prostate cancer progression.  We mapped the global DNA methylation patterns in selected prostate tissues and cell lines using Methylplex-Next Generation Sequencing (M-NGS).  Hidden Markov Model based next generation sequence analysis identified ~68,000 methylated regions per sample.  While global CpG Island (CGI) methylation was not differential between benign adjacent and cancer samples, overall promoter CGI methylation increased from ~12.6% in benign samples to 19.3% and 21.8% in localized and metastatic cancer tissues, respectively.  We found distinct patterns of promoter methylation around transcription start sites, where methylation occurred not only on the CGIs, but also on flanking regions and CGI sparse promoters.  Among the 6,691 methylated promoters in prostate tissues, 2481 differentially methylated regions (DMRs) are cancer specific, including numerous novel DMRs.   A novel cancer specific DMR in WFDC2 promoter showed heavy methylation in cancer (17/22 tissues, 6/6 cell lines), but not in the benign tissues (0/10) and normal PrEC cells.  Integration of LNCaP DNA methylation and H3K4me3 data suggested an epigenetic mechanism for alternate transcription start site utilization and these modifications segregated into distinct regions when present on the same promoter.  Finally, we observed differences in repeat element methylation, particularly LINE-1, between ERG gene fusion positive and negative cancers.  This comprehensive methylome map will further our understanding of epigenetic regulation in prostate cancer progression.

This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series.


title: ENCODE Genome Institute of Singapore DNA Paired-End Ditags

status: Public on Apr 27 2011

city: Stanford

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 (Yijun Ruan mailto:ruanyj@gis.a-star.edu.sg). If you have questions about the Genome Browser track associated with this data, contact ENCODE (mailto:genome@soe.ucsc.edu).

This track, produced as part of the ENCODE Project, contains deep sequencing DNase data that will be used to identify sites where regulatory factors bind to the genome (footprints).
Footprinting is a technique used to define the DNA sequences that interact with and bind DNA-binding proteins, such as transcription factors, zinc-finger proteins, hormone-receptor complexes, and other chromatin-modulating factors like CTCF. The technique depends upon the strength and tight nature of protein-DNA interactions. In their native chromatin state, DNA sequences that interact directly with DNA-binding proteins are relatively protected from DNA degrading endonucleases, while the exposed/unbound portions are readily degraded by such endonucleases. A massively parallel next-generation sequencing technique to define the DNase hypersensitive sites in the genome was adopted. Sequencing these next-generation-sequencing DNase samples to significantly higher depths of 300-fold or greater produces a base-pair level resolution of the DNase susceptibility maps of the native chromatin state. These base-pair resolution maps represent and are dependent upon the nature and the specificity of interaction of the DNA with the regulatory/modulatory proteins binding at specific loci in the genome; thus they represent the native chromatin state of the genome under investigation. The deep sequencing approach has been used to define the footprint landscape of the genome by identifying DNA motifs that interact with known or novel DNA binding proteins.
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. Digital DNaseI was performed by DNaseI digestion of intact nuclei, followed by isolating DNaseI 'double-hit' fragments as described in Sabo et al. (2006), and direct sequencing of fragment ends (which correspond to in vivo DNaseI cleavage sites) using the Solexa platform (27 bp reads). High-quality reads were mapped to the GRCh37/hg19 human genome using Bowtie 0.12.5 (Eland was used to map to NCBI36/hg18); only unique mappings were kept. DNaseI sensitivity is directly reflected in raw tag density (Signal), which is shown in the track as density of tags mapping within a 150 bp sliding window (at a 20 bp step across the genome). DNaseI hypersensitive zones (HotSpots) were identified using the HotSpot algorithm described in Sabo et al. (2004). False discovery rate thresholds of 1.0% (FDR 0.01) were computed for each cell type by applying the HotSpot algorithm to an equivalent number of random uniquely mapping 36-mers. DNaseI hypersensitive sites (DHSs or Peaks) were identified as signal peaks within 1.0% (FDR 0.01) hypersensitive zones using a peak-finding algorithm. Only DNase Solexa libraries from unique cell types producing the highest quality data, as defined by Percent Tags in Hotspots (PTIH ~40%) were designated for deep sequencing to a depth of over 200 million tags.

first: Xiaoan

last: Ruan

first: Atif

last: Shahab

first: Chialin

last: Wei

first: Yijun


title: Cytokine receptor modulation by interleukin-2 broadly regulates T helper cell lineage differentiation

status: Public on Apr 24 2011

city: Bethesda

first: Peng

state: MD

address: 9000 Rockville Pike

department: NHLBI

institute: NIH

summary: T helper (Th) cells control host defense to pathogens. IL 12R expression is required for Th1, IL-4Rα for Th2, and IL-6Rα/gp130 for Th17 differentiation to allow responsiveness to IL-12, IL-4, and IL-6, respectively. IL-2 via STAT5 controls Th2 differentiation by regulating the Th2 cytokine gene cluster and Il4ra expression. Here we show that IL-2 regulates Th1 differentiation, inducing STAT5-dependent IL-12Rβ2 and T-bet expression, with impaired human Th1 differentiation when IL-2 was blocked. Th1 differentiation was also impaired in mouse Il2-/- T cells but restored by IL-12Rβ2 expression. Consistent with IL-2’s inhibition of Th17 differentiation, IL-2 decreased Il6ra and Il6st/gp130 expression, and Il6st augmented Th17 differentiation even when IL-2 was present. Thus, IL-2 influences T-cell differentiation by modulating cytokine receptor expression to help specify/maintain differentiated states.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Genome-wide mapping of STAT1,STAT4,STAT5A,STAT5B binding to their target genes in Th1 or human CD4+ cells was conducted

last: Liao

first: Jian-Xin

last: Lin

first: Lu

first: Warren

last: Leonard


title: A Genome-Wide Characterization of New and Drought Stress Responsive MicroRNAs in Populus euphratica

status: Public on Apr 25 2011

city: Shenzhen

first: Bosheng

state: Guangdong

address: SUStech Huiyuan #1 406

department: MCDB

country: China

institute: SUSTC (current, may different with previous institution )

summary: We present an efficient method to genome-wide discover new and drought stress responsive miRNAs in P. euphratica. High throughput sequencing of P. euphratica leaves found 197 conserved miRNAs between P. euphratica and Populus trichocarpa. Meanwhile, 189 new miRNAs which belonged to 120 families were identified, a large increasing to the number of P. euphratica miRNAs. Target prediction and degradome sequencing verification of 22 new and 21 conserved miRNA targets showed these targets were involved in multiple biological processes, including transcription regulation and response to stimulus. Furthermore, comparison of high-throughput sequencing with miRNA microarray profiling data indicated that 104 miRNA sequences were up-regulated, while 27 were down-regulated under drought stress. This preliminary characterization based on our findings provided a framework for future analysis of miRNA genes and their roles in key traits of poplar as stress resistance plant breeding and environment protection usage.

overall_design: Examination of sRNA expression in 2 poplar leaf samples in drought and normal growth conditions.

first: Xinli

last: Xia


title: The Nucleosome Map of the Mammalian Liver

city: Philadelphia

first: Jonathan

last: Schug

state: Pennsylvania

address: 12-156 Smilow Ctr Trans Research

department: School of Medicine

institute: University of Pennsylvania

summary: We report the genome-wide map of nucleosome positions in the mouse liver, with emphasis on transcriptional start sites, CpG islands, Foxa2 binding sites, and their correlation with gene expression. Despite the heterogeneity of liver tissue, we could clearly discern the nucleosome pattern of the predominant liver cell, the hepatocyte. By analyzing nucleosome occupancy and the distributions of heterochromatin protein 1 (Hp1), CBP, and p300 in Foxa1/2-deficient livers we find, surprisingly, that the maintenance of nucleosome position and chromatin structure surrounding Foxa2 binding sites is independent of Foxa1/2.

overall_design: Examination of nucleosome map and Foxa2 binding in the mouse liver.

first: Zhaoyu

first: Klaus

middle: K

last: Kaestner


title: Comparative sequence analysis of plant small RNAs

status: Public on May 10 2011

city: Udine

first: Emanuele

last: De Paoli

state: UDINE

address: via delle Scienze 206

department: Agricultural, Food, Environmental and Animal Sciences

country: Italy

institute: University of Udine

type: Non-coding RNA profiling by high throughput sequencing

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

sample_organism:
Citrus sinensis
Chlamydomonas reinhardtii
Volvox carteri
Ginkgo biloba
Picea abies
Persea americana
Carica papaya
Cucurbita maxima
Populus trichocarpa
Capsicum annuum
Solanum lycopersicum
Nicotiana tabacum
Petunia x hybrida
Solanum tuberosum
Erythranthe guttata
Lactuca sativa
Hordeum vulgare
Setaria italica
Sorghum bicolor
Triticum aestivum
Zea mays
Musa acuminata
Amborella trichopoda
Marsilea quadrifolia
Zostera marina
Gossypium arboreum
Vitis vinifera
Silene latifolia
Panicum virgatum
Chara corallina
Cycas rumphii
Nuphar advena
Aristolochia fimbriata
Miscanthus x giganteus

overall_design: Refer to individual Series


title: Temporal response of DCs to LPS stimulation: 4sU_sequencing

city: Jerusalem

first: Michal

last: Rabani

address: Hebrew University, Edmond J. Safra Campus, Silberman building

country: Israel

institute: Hebrew University of Jerusalem

summary: Regulation of RNA levels is critical for the response to external stimuli and determined through the interplay between RNA production, processing and degradation. Despite the centrality of these processes, most global studies of RNA regulation do not distinguish their separate contributions and relatively little is known about how they are temporally integrated. Here, we combine metabolic labeling of RNA with advanced RNA quantification assays and computational modeling to estimate RNA transcription and degradation during the response of immune dendritic cells (DCs) to pathogens, a critical and tightly regulated step in innate immunity. We find that transcription regulation plays a major role in shaping most temporal changes in RNA levels, but that changes in degradation rate are important for shaping sharp ‘peaked’ responses. We find that transcription changes precede corresponding RNA changes by a small lag (15-30 min), which is shorter for induced than for repressed genes. Massively parallel sequencing of the entire RNA population – including non-polyadenylated transcripts – allows us to estimate RNA processing, and identify specific groups of transcripts, mostly cytokines and transcription factors, undergoing enhanced mRNA maturation. This suggests an additional role for splicing in regulating mRNA maturation. Our method provides a new quantitative approach to study key steps in the integrative process of RNA regulation.

overall_design:
Sequencing of 4sU-labeled RNA taken from a 7 samples time-series (one sample every 1 hour) during the response of DCs to LPS stimulation. 4-thiouridine was added 45 minutes prior to sample collection.
Data presented here for six timepoints: 0, 1, 3-6 hrs. 2hr timepoint not included.

first: Joshua

middle: Z

last: Levin


title: seq-BH00005_LIN9_N2_L3

first: Tristan

last: De Buysscher

overall_design: ChIP-seq against LIN-9 in L3 stage N2 worms.

first: Julie

last: Ahringer

first: Isabel

last: Latorre


title: Unamplified Cap Analysis of Gene Expression on a single molecule sequencer (HeliScopeCAGE)

status: Public on Apr 28 2011

city: Yokohama

first: Hideya

last: Kawaji

state: Kanagawa

address: 1-7-22 Suehiro-cho, Tsurumi-ku

department: Omics Science Center

country: Japan

institute: RIKEN

summary: We report the development of a simplified Cap Analysis of Gene Expression (CAGE) protocol adapted for single molecule sequencers which avoids second strand synthesis, ligation, digestion and PCR. HeliScopeCAGE directly sequences the 3’ end of cap trapped first strand cDNAs. As with previous versions of CAGE, we better define transcription start sites (TSS) than known models, identify novel regions of transcription and alternative promoters, and find two major classes of TSS signal, sharp peaks and broad regions. However using this protocol we observe reproducible evidence of regulation at the much finer level of individual TSS positions. The libraries are quantitative over 5 orders of magnitude and highly reproducible (pearson’s correlation coefficient of 0.987). We have also scaled down the sample requirement to 5ug total RNA for a standard HeliScopeCAGE library and 100ng for a low quantity version. When the same RNA was run as 5ug and 100ng versions, the 100ng was still able to detect expression for ~60% of the 13468 loci detected by a 5ug library using the same threshold, allowing comparative analysis of even rare cell populations. Testing the protocol for differential gene expression measurements on triplicate HeLa and THP-1 samples, we find the log fold change compared to Illumina microarray measurements is highly correlated (0.871). In addition, HeliScopeCAGE finds differential expression for 1000s more loci including those with probes on the array. Finally although the majority of tags are 5’ associated we also observe a low level of signal on exons which is useful for defining gene-structures.

overall_design: The same total RNAs (two pools, from HeLa and THP-1 cells) are profiled with microarray and HeliScopeCAGE. All replicates are technical replicates.

first: Mutsumi

last: Kanamori-Katayama

first: Masayoshi

last: Itoh

first: Timo

last: Lassmann

first: Shintaro

last: Katayama

first: Miki

last: Kojima

first: Nicolas

last: Bertin

first: Ai

last: Kaiho

first: Noriko

last: Ninomiya

first: Carsten

middle: O

last: Daub

first: Piero

last: Carninci

first: Alistair

last: Forrest

first: Yoshihide

last: Hayashizaki


title: Genome-wide Regulation of 5hmC, 5mC and Gene Expression by Tet1 Hydroxylase in Mouse Embryonic Stem Cells (ChIP-seq data)

status: Public on Apr 21 2011

city: Shanghai

first: Feizhen

last: Wu

state: Shanghai

address: Zhangheng Road 825, Pudong

department: Intelligent Medicine Institute

institute: Fudan Univ, Shanghai, China

summary: DNA methylation of C5-cytosine (5mC) in the mammalian genome is a key epigenetic event that is critical for various cellular processes. However, how the genome-wide 5mC pattern is dynamically regulated remains a fundamental question in epigenetic biology. The TET family of 5mC hydroxylases, which convert 5mC to 5-hydroxymethylcytosine (5hmC), have provided a new potential mechanism for the dynamic regulation of DNA methylation. The extent to which individual Tet family members contribute to the genome-wide 5mC and 5hmC patterns and associated gene network remains largely unknown. Here we report genome-wide mapping of Tet1 and 5hmC in mESCs and reveal a mechanism of action by which Tet1 controls 5hmC and 5mC levels in mESCs. In combination with microarray and mRNA-seq expression profiling, we identify a comprehensive yet intricate gene network influenced by Tet1. We propose a model whereby Tet1 controls DNA methylation both by binding to CpG-rich regions to prevent unwanted DNA methyltransferase activity, and by converting the existing 5mC to 5hmC through its enzymatic activity. This Tet1-mediated antagonism of CpG methylation imparts differential maintenance of DNA methylation status at Tet1 target loci, thereby providing a new regulatory mechanism for establishing the epigenetic landscape of mESCs, which ultimately contributes to mESC differentiation and the onset of embryonic development.

overall_design:
To determine the genome-wide distribution of Tet1 and 5hmC in mouse ES cells, as well as identify the gene transcription changes after Tet1 depletion.

GSM706669-GSM706671: We used GST pull-down followed by deep sequencing to map the DNA bound by the Tet1 CXXC domain in vitro. We made two mutants that have a single point mutation (Cys574 to Ala or Cys586 to Ala) in the core CXXC domain to ascertain the essential role of the CXXC domain in DNA binding by comparing the sequencing profile of DNA bound by wild type CXXC with the profiles of the CXXC mutants.

GSM706672-GSM706673: Tet1 ChIP-seq was performed to identify the genome-wide distribution of Tet1 in mouse ES cells.

GSM706674-GSM706679: We performed hydroxymethylated DNA immunoprecipitation (hMeDIP)-seq combined with a shRNA-mediated gene depletion strategy. To identify the loci specific 5hmC regulation by Tet1, we compared the 5hmC genome-wide distributions in control (Luc shRNA) and Tet1-depleted (Tet1 shRNA2863) mouse ES cells.

GSM706680-GSM706682: To identify the gene regulation network by Tet1, we compared the gene expression profiles of control (scramble shRNA) and Tet1-depleted (Tet1 shRNA 2863 and Tet1 shRNA 3387) mouse ES cells determined by mRNA-seq.

first: Yufei

last: Xu

first: Yujiang

last: Shi


title: Genome-wide Regulation of 5hmC, 5mC and Gene Expression by Tet1 Hydroxylase in Mouse Embryonic Stem Cells


title: Characterization of AGO1-/AGO4-associated smRNAs

status: Public on Apr 19 2011

first: Huan

last: Wang

state: New York

address: 1230 York Avenue

institute: The Rockefeller University

summary: To fully characterize smRNAs associated with AGO1 and AGO4, we developed a two-step protocol to purify AGO/smRNA complexes from flowers, leaves, roots and seedlings with enhanced purity, and sequenced the smRNAs by Illuminaâs technology. We identified some additional miRNAs, collateral miRNAs encoded in known miRNA precursors, phased smRNA clusters and nat-siRNAs. Organ specific sequencing provided digital expression profiles of all obtained smRNAs, especially miRNAs.

overall_design: We used extracts from Arabidopsis flowers, leaves and roots as well as ten-day old seedlings to purify smRNAs associated with AGO1 and AGO4 protein complexes using a two-step immunoprecipitation method.

first: Xiuren

last: Zhang

first: Jun

last: Liu

first: Takatoshi

last: Kiba

first: Jongchan

last: Woo

first: Tolulope

last: Ojo

first: Markus

last: Hafner

first: Thomas

last: Tuschl

first: Nam-Hai

last: Chua

first: Xiu-Jie


title: Genome-wide mapping of 5-hydroxymethylcytosine in embryonic stem cells

city: Montreal

first: William

middle: Abraham

last: Pastor

state: QC

address: 3655 Promenade Sir William Osler

department: Biochemistry

institute: McGill University

summary: 5hmC and TET proteins have been implicated in stem cell biology and cancer, but information on the genome-wide distribution of 5hmC is limited. Here we describe two novel and specific approaches to profile the genomic localisation of 5hmC.  The first approach, termed GLIB (GLucosylation, perIodate oxidation, Biotinylation) uses a combination of enzymatic and chemical steps to isolate DNA fragments containing as few as a single 5hmC.  The second approach involves conversion of 5hmC to cytosine 5-methylenesulphonate (CMS) by treatment of genomic DNA with sodium bisulphite, followed by immunoprecipitation of CMS-containing DNA with a specific antiserum to CMS. High-throughput sequencing of 5hmC-containing DNA from mouse embryonic stem (ES) cells showed strong enrichment within exons and near transcriptional start sites (TSS). 5hmC was especially enriched at the start sites of genes whose promoters bear dual histone 3 lysine 27 trimethylation (H3K27me3) and histone 3 lysine 4 trimethylation (H3K4me3) marks.  Our results indicate that 5hmC has a likely role in transcriptional regulation, and suggest a model in which 5hmC contributes to the “poised” chromatin signature found at developmentally-regulated genes in ES cells.

overall_design: Mapping of 5-hydroxymethylcytosine in ES cells by GLIB and anti-CMS methodologies

middle: A

first: Utz

last: Pape

first: Yun

last: Huang

first: Anjana

last: Rao


title: Using RNA-Seq for gene identification, polymorphism detection and transcript profiling in two alfalfa genotypes with divergent cell wall composition in stems

city: St Paul

first: Sam

last: Yang

address: 411 Upper Buford Circle

department: ARS

institute: USDA

type: Genome variation profiling by high throughput sequencing

summary: Alfalfa, [Medicago sativa (L.) sativa], a widely-grown perennial forage has potential for development as a cellulosic ethanol feedstock. The application of genomic approaches would advance development of alfalfa as a cellulosic feedstock. However, the genomics of alfalfa, a non-model species, is still in its infancy. The recent advent of RNA-Seq, a massively parallel sequencing method for transcriptome analysis, provides an opportunity to expand the identification of alfalfa genes and polymorphisms, and conduct in-depth transcript profiling. Cell walls in stems of alfalfa genotype 708 have higher cellulose and lower lignin concentrations compared to cell walls in stems of genotype 773. Using the Illumina GA-II platform, a total of 198,861,304 expression sequence tags (ESTs, 76 bp in length) were generated from cDNA libraries derived from elongating stem (ES) and post-elongation stem (PES) internodes of 708 and 773. These ESTs were de novo assembled into 132,153 unique sequences.  By combining the de novo assembled ESTs (132,153 sequences) with our previously identified EST sequences (341,984 sequences, unpublished data), and the ESTs available from GenBank (12,371 sequences), we built the first Alfalfa Gene Index (MSGI 1.0). MSGI 1.0 contains 124,025 unique sequences including 22,729 tentative consensus sequences (TCs), 22,315 singletons and 78,981 pseudo-singletons. We identified a total of 1, 294 simple sequence repeats (SSR) among the sequences in MSGI 1.0. In addition, a total of 10,826 single nucleotide polymorphisms (SNPs) were predicted between the two genotypes. Transcript profiling of stem internodes of genotypes 708 and 773 was conducted by quantifying the number of Illumina EST reads that were mapped to sequences in MSGI 1.0. We identified numerous candidate genes that may play a role in stem development as well as candidate genes that may contribute to the differences in cell wall composition in stems of the two genotypes. Our results demonstrate that RNA-Seq can be successfully used for gene identification, polymorphism detection and transcript profiling in alfalfa, a non-model, allogamous, autotetraploid species. The alfalfa gene index (MSGI 1.0) assembled in this study, and the SNPs, SSRs and candidate genes identified can be used to improve alfalfa as a cellulosic feedstock.

overall_design: Examination of 2 different tissue types at different developmental stages (Elongating vs. post-elongation stem internodes) in two alfalfa genotypes (708 and 773) with divergent cell wall composition in stems.

first: S

middle: S

first: Carroll

middle: P

last: Vance

first: John

middle: W

last: Gronwald


title: Genome-wide Regulation of 5hmC, 5mC and Gene Expression by Tet1 Hydroxylase in Mouse Embryonic Stem Cells (bisulfite sequencing data)

first: Feizhen

last: Wu

overall_design: To determine the genome-wide DNA methylation changes caused by Tet1 depletion in mouse ES cells. Tet1 protein was depleted by specific siRNA treatment. The DNA methylation levels in control and Tet1 siRNA-transfected ES cells were determined by targeted bisulfite sequencing.

first: Yufei

last: Xu

first: Yujiang

last: Shi


title: Cdx2 transcription factor binding in intestinal villus and gene expression profiling in Cdx mutant mice

status: Public on Apr 13 2011

city: Piscataway

first: Michael

last: Verzi

state: NJ

address: 145 Bevier Road

department: Genetics

institute: Rutgers, the State University of New Jersey

summary: We conditionally inactivated mouse Cdx2, a dominant regulator of intestinal development, and mapped its genome occupancy in adult intestinal villi. Although homeotic transformation, observed in Cdx2-null embryos, was absent in mutant adults, gene expression and cell morphology were vitally compromised. Lethality was accelerated in mice lacking both Cdx2 and its homolog Cdx1, with exaggeration of defects in crypt cell replication and enterocyte differentiation. Cdx2 occupancy correlated with hundreds of transcripts that fell but not with equal numbers that rose with Cdx loss, indicating a predominantly activating role at intestinal cis-regulatory regions. Integrated consideration of a mutant phenotype and cistrome hence reveals the continued and distinct requirement in adults of a master developmental regulator that activates tissue-specific genes.

overall_design: Cdx2 ChIP-seq in mouse villus, and gene expression data from Cdx1, Cdx2 and compound knockout mouse intestine

first: Ramesh

last: Shivdasani

first: X

last: Liu

first: Hyunjin

last: Shin


title: Tet1 and hydroxymethylcytosine in transcription and DNA methylation fidelity

city: Copenhagen

first: Kristine

last: Williams

address: Ole Maaløes vej 5

department: BRIC

country: Denmark

institute: University of Copenhagen


title: Tet1 and hydroxymethylcytosine in transcription and DNA methylation fidelity (ChIP/DIP-Seq data)

summary: We analyzed the genome-wide binding of Tet1 in control (shScr) and Tet1 knockdown (shTet1) mouse ES cells using two different Tet1 antibodies (Tet1-C and Tet1-N). Furthermore, we generated genome-wide mapping of hydroxymethyl cytosine (hmC) and methyl cytosine (mC). We find that hmC, in contrast to mC, is also found at transcription start sites (TSSs), and that there is a significant overlap between Tet1 binding and hmC positive regions. Surprisingly, our results also suggest, that Tet1 has a role in transcriptional repression. We showed that Tet1 associates with Sin3A co-repressor complex, and by performing ChIP-sequencing of Sin3A, we find co-localisation of Tet1 and Sin3a throughout the genome

overall_design: Examination of Tet1 and Sin3A binding as well as hmC and mC localization in mouse ES cells

first: Marianne

last: Pedersen


title: Wdr5 mediates self-renewal and reprogramming via the embryonic stem cell core transcriptional network

status: Public on Apr 08 2011

city: New York

first: Jonathan

last: Monk

state: NY

address: One Gustave L. Levy Place

department: Gene and Cell Medicine

institute: Mt. Sinai School of Medicine

summary: The embryonic stem (ES) cell transcriptional and epigenetic networks are critical for the maintenance of ES cell self-renewal. However, it remains unclear whether components of these networks functionally interact and if so, what factors mediate such interactions. Here we show that WD-repeat protein-5 (Wdr5), a core member of the mammalian Trithorax (trxG) complex, positively correlates with the undifferentiated state and is a novel regulator of ES cell self-renewal. We demonstrate that Wdr5, an ‘effector’ of H3K4 methylation, interacts with the pluripotency transcription factor Oct4. In concordance, genome-wide ChIP-Seq and transcriptome analyses demonstrate overlapping gene regulatory functions between Oct4 and Wdr5. We show that the Oct4-Sox2-Nanog circuitry cooperates with trxG for transcriptional activation of key self-renewal regulators. Furthermore, Wdr5 expression is required for the efficient formation of induced pluripotent stem (iPS) cells. Collectively, these findings implicate an integrated model of transcriptional and epigenetic control, mediated by select trxG members, for the maintenance of ES cell self-renewal and somatic cell reprogramming.

overall_design: 7 Samples

first: Yen-Sin

last: Ang

first: Su-Yi

last: Tsai

first: Dung-Fang

last: Lee

first: Jie

last: Su

first: Kajan

last: Ratnakumar

first: Yongchao

last: Ge

first: Henia

last: Darr

first: Betty

last: Chang

last: Rendl

first: Emily

last: Bernstein

first: Christoph

last: Schaniel

first: Ihor

last: Lemischka


title: Determinants of nucleosome organization in primary human cells

first: Anton

last: Valouev

address: 300 Pasteur Dr.

department: Pathology

institute: Stanford University

summary: Genome-wide maps of nucleosome organization in human CD4+ T-cells, CD8+ T-cells, granulocytes, and from in vitro reconstitution.

overall_design: 8 samples include: nucleosome-bound DNA isolated from primary human CD4+ T-cells, nucleosome bound-DNA isolated from primary human CD8+ T-cells, nucleosome-bound DNA isolated from primary human granulocytes, nucleosome-bound DNA isoalted from in vitro reconstitution of nucleosomes with human genomic DNA, control DNA generated by micrococcal nuclease treatment of human genomic DNA, RNA-seq from human CD4+ T-cells, RNA-seq from human CD8+ T-cells, RNA-seq from human Granulocytes.


title: Deep sequencing analysis of the developing mouse brain reveals a novel microRNA

status: Public on Apr 05 2011

city: UPM Serdang

first: King

middle: Hwa

last: Ling

state: Selangor Darul Ehsan

address: Faculty of Medicine and Health Sciences

department: Department of Obstetrics and Gynaecology

country: Malaysia

institute: Universiti Putra Malaysia

summary:
MicroRNAs (miRNAs) are small non-coding RNAs that can exert multilevel inhibition/repression at a post-transcriptional or protein synthesis level during disease or development. Characterisation of miRNAs in adult mammalian brains by deep sequencing has been reported previously. However, to date, no small RNA profiling of the developing brain has been undertaken using this method. We have performed deep sequencing and small RNA analysis of a developing (E15.5) mouse brain.

We identified the expression of 294 known miRNAs in the E15.5 developing mouse brain, which were mostly represented by let-7 family and other brain-specific miRNAs such as miR-9 and miR-124. We also discovered 4 putative 22-23nt miRNAs: mm_br_e15_1181, mm_br_e15_279920, mm_br_e15_96719 and mm_br_e15_294354 each with a 70-76nt predicted pre-miRNA. We validated the 4 putative miRNAs and further characterised one of them, mm_br_e15_1181, throughout embryogenesis. Mm_br_e15_1181 biogenesis was Dicer1-dependent and was expressed in E3.5 blastocysts and E7 whole embryos. Embryo-wide expression patterns were observed at E9.5 and E11.5 followed by a near complete loss of expression by E13.5, with expression restricted to a specialised layer of cells within the developing and early postnatal brain. Mm_br_e15_1181 was upregulated during neurodifferentiation of P19 teratocarcinoma cells. This novel miRNA has been identified as miR-3099.

We have generated and analysed the first deep sequencing dataset of small RNA sequences of the developing mouse brain. The analysis revealed a novel miRNA, miR-3099, with potential regulatory effects on early embryogenesis, and involvement in neuronal cell differentiation/function in the brain during late embryonic and early neonatal development.

overall_design: Deep sequencing analysis of small RNAs isolated from an E15.5 mouse brain.

first: Peter

last: Brautigan

first: Christopher

middle: N

last: Hahn

first: Tasman

last: Daish

first: John

last: Rayner

first: Pike-See

last: Cheah

first: Joy

last: Raison

first: Sandra

last: Piltz

first: Jeffrey

last: Mann

first: Deidre

last: Mattiske

first: Paul

middle: Q

last: Thomas

first: David

last: Adelson

first: Hamish

last: Scott


title: Expression data from Arabidopsis mature siliques

city: Freiburg im Breisgau

first: Annaick

last: Carles

address: Habsburgerstrasse 49

country: Germany

institute: University of Freiburg

summary: Analysis of the transcriptomes of nearly ripe siliques (18-19 DAP) of the rdo2-1, rdo3 and hub1-2 (rdo4) mutants in comparison with wild-type Ler, using Affymetrix GeneChip Arabidopsis ATH1 Genome Array.

overall_design: The experiment includes rdo2, rdo3, and rdo4 as well the Ler genotypes with three biological replicates for each. Therefore in total there are 12 arrays

first: Yongxiu

last: Liu

first: Yong

last: Li

first: Bruno

last: Huettel

first: Wim

middle: J

last: Soppe


title: Comprehensive Long Span Paired-End-Tag Mapping Reveals Characteristic Patterns of Structural Variations in Epithelial Cancer Genomes

status: Public on Apr 06 2011

city: Cologne

first: Axel

last: HILLMER

address: Kerpener Str. 62

department: Institute of Pathology

institute: University of Cologne

summary: Somatic genome rearrangements are thought to play important roles in cancer development. We optimized a long span paired-end-tag (PET) sequencing approach using 10 Kb genomic DNA inserts to study human genome structural variations (SVs). The use of 10 Kb insert size allows the identification of breakpoints within repetitive or homology containing regions of a few Kb in size and results in a higher physical coverage compared to small insert libraries with the same sequencing effort. We have applied this approach to comprehensively characterize the SVs of 15 cancer and 2 non-cancer genomes and used a filtering approach to strongly enrich for somatic SVs in the cancer genomes. Our analyses revealed that most inversions, deletions, and insertions are germline SVs, whereas tandem duplications, unpaired inversions, inter-chromosomal translocations, and complex rearrangements are overrepresented among somatic rearrangements in cancer genomes. We demonstrate that the quantitative and connective nature of DNA-PET data is precise in delineating the genealogy of complex rearrangement events, we observe signatures which are compatible with breakage-fusion-bridge cycles, and discover that large duplications are among the initial rearrangements that trigger genome instability for extensive amplification in epithelial cancers.

overall_design: Structural variations of 15 human cancer samples and 2 human normal samples were identified by long span paired-end sequencing

first: Yijun

last: Ruan

last: Hillmer


title: Genome-wide identification of micro-ribonucleic acids associated with human endometrial receptivity in natural and stimulated cycles by deep sequencing

status: Public on Apr 01 2011

city: Shantou

first: Zeng-Ming

last: Yang

address: 263 Daxue Road

institute: Shantou University

summary: In this study, we took advantage of deep sequencing approaches to investigate the miRNA expression profiles of human endometrium on days LH+2 and LH+7 in natural cycles, and compare them with those on days hCG+4 and hCG+7 in stimulated cycles during IVF treatment.

overall_design: Examination of miRNA expression in human endometrium during natural and stimulated cycles

first: Zengming

first: Jilong

first: Aiguo

last: Sha


title: Genome wide maps of Dmc1 in testis of Hop2 null mice.

status: Public on Apr 04 2011

first: Ivan

last: Gregoretti

address: 5 Memorial Dr, Building 5, Room 205.

department: National Institute of Diabetes and Digestive and Kidney Diseases

institute: National Institutes of Health

summary: We report the application of ChIP-seq targeted at the meiosis-specific protein DMC1 to reveal the genome-wide distribution of initiation of meiotic recombination. The mouse model here employed is Hop2-/- because it is unable to repair the DNA double-stranded breaks and therefore the DMC1 signal is more persistent. We also provide the resulting dataset of ChIP-seq targeted at RAD51 which is not meiosis specific but is also targeted at initiation of recombination loci in meiotic tissue. In addition, we report DMC1 ChIP-seq on wild type mouse pup testis. Finally, we present ChIP-seq targeted at H3K4me3 in testis and liver tissues.

overall_design: Lysates from mice testis cells were sonicated, clarified and DNA protein complexes were isolated with anti Dmc1 or anti Rad51 or H3K4Me3 antibodies. DNA purified from proteins and libraries were prepared according to Illumina's instructions accompanying the genomic DNA Sample preparing kit part number 1000181. Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenowpolymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (3’ to 5’ exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 18 cycles and library fragments of 150~250 bp were isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. To prepare Micrococcal Nuclease-digested fractions of chromatin DNA, homogenized testis cells were treated with MN and pelleted. The supernatant containing the digested fraction was separated and the DNA was purified from proteins. The library was prepared the same way as described above and all were sequenced on the Genome Analyzer following the manufacturer's protocols.

first: Fatima

last: Smagulova

middle: V

first: Kevin

last: Brick

first: Pavel

last: Khil

first: Rafael

last: Camerini-Otero

first: Galina

last: Petukhova


title: Genome-wide analysis of transcription factor E2F1 mutant proteins reveals that N- and C-terminal protein interaction domains do not participate in targeting E2F1 to the human genome.

city: Palo Alto

first: Philip

last: Cayting

address: 1501 S California Ave

summary:
Previous studies of E2F family members have suggested that protein-protein interactions may be the mechanism by which E2Fs are recruited to specific genomic regions. We have addressed this hypothesis on a genome-wide scale using ChIP-seq analysis of MCF7 cell lines that express tagged wildtype and mutant E2F1 proteins.  First, we performed ChIP-seq for tagged wt E2F1.  Then, we analyzed E2F1 proteins that lacked the N terminal SP1 and cyclin A binding domains, the C terminal transactivation and pocket protein binding domains, and the internal marked box domain. Surprisingly, we found that the ChIP-seq patterns of the mutant proteins were identical to that of wt E2F1.  However, mutation of the DNA binding domain abrogated all E2F1 binding to the genome.  These results suggested that the interaction between the E2F1 DNA binding domain and a consensus motif may be the primary determinant of E2F1 recruitment.  To address this possibility, we analyzed the in vivo binding sites for the in vitro-derived consensus E2F1 motif (TTTSSCGC) and also performed de novo motif analysis. We found that only 12% of the ChIP-seq peaks contained the TTTSSCGC motif. De novo motif analysis indicated that most of the in vivo sites lacked the 5â half of the in vitro derived consensus, having instead the in vivo consensus of CGCGC. In summary, our findings do not provide support for the model that protein-protein interactions are involved in recruiting E2F1 to the genome, but rather suggest that recognition of a motif found at most human promoters is the critical determinant.

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: 9 total ChIP-seq datasets; four different HA-ER-E2F1 mutants, and one HA ER E2F1 wild type dataset done in duplicate, from 5 different stable cell lines derived from MCF7 cells;  Three Input replicates from 2 different stable cell lines derived from MCF7 cells; HA ER E2F1 wild type duplicate dataset from MCF7 stable cells; 1 HA ER E2F1 DBDmut replicate from MCF7 stable cells cells, 1 HA ER E2F1ÎC replicate from MCF7 stable cells cells, 1 HA ER E2F1ÎN/C replicate from MCF7 stable cells cells, 1 HA ER E2F1ÎMB replicate from MCF7 stable cells cells, 1 HA ER E2F1ÎMB replicate from MCF7 stable cells cells, 3 Input replicates from MCF7 stable cells cells.

first: Alina

last: Cao

first: Peggy

last: Farnham


title: Genome-Wide Analysis of Somatic Copy Number Alterations and Gene Expression in Metastatic Melanoma

status: Public on Apr 07 2011

city: Lausanne

first: Armand

last: Valsesia

address: Bâtiment Génopode

country: Switzerland

institute: Ludwig Institute for Cancer Research

type: Genome variation profiling by genome tiling array

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

overall_design: Refer to individual Series

first: Brian

last: Stevenson


title: Metabolic Transcription Factors Mediate Conserved Functions Largely via Species-Specific Binding Regions

city: RTP

first: Logan

last: Everett

state: North Carolina

address: 109 T.W. Alexander Dr

department: Office of Research and Development

institute: U.S. Environmental Protection Agency

summary: The winged helix protein FOXA2 and the nuclear receptor PPARg are highly conserved, regionally-expressed transcription factors that regulate networks of genes controlling complex metabolic functions.  Cistrome analysis for FOXA2 in mouse liver and PPARg in mouse adipocytes has previously produced consensus binding sites that are nearly identical to those used by the factors in human cells.  Despite this conservation of the canonical binding motif, we report here that the great majority of specific binding regions for FOXA2 in human liver and for PPARg in human adipocytes are not in the orthologous locations to the mouse genome.  Nevertheless, gene-centric analysis reveals strong shared transcription factor occupancy near genes in tissue-specific metabolic pathways that are functionally conserved across species.  Genes with only species-specific binding sites fail to show enrichment for these pathways.  Thus, the biological functions of transcription factors that control specific metabolic functions are highly shared across species.

overall_design: Two TFs, FOXA2 and PPARg, were studied for genome-wide conservation of binding between mouse and human in specific tissues/cell-types (liver for FOXA2, adipocytes for PPARg).  The number of replicates for each TF was chosen to obtain a comparable number of reads between the TFs and species.  Human FOXA2 ChIP-seq was performed on two biological replicates of human liver samples, in three technical replicates each. Input DNA was also collected and sequenced from both biological samples.   Mouse FOXA2 ChIP-seq was performed on four biological replicates of mouse liver samples.  The ChIP and sequencing were repeated on two of these biological replicates to create technical replicates for additional sequence reads.  Input DNA was sequenced from three additional mouse livers.  Human PPARg ChIP-seq was performed on a human adipocyte cell-line (SGBS) differentiated in two replicate cultures.  Input DNA was also collected and sequenced from one culture.  Mouse PPARg ChIP-seq was performed on 3T3-L1 cells differentiated into adipocytes in culture in a single replicate, and this sequence data was pooled with existing data previously generated by the same lab, already available in GEO (GSE21314).  A standard pool of input DNA sample sequence from multiple mouse tissue was used for analyzing the Mouse PPARg ChIP-seq data.

first: Geetu

last: Tuteja

first: Raymond

middle: E

last: Soccio

first: Mitchell

last: Lazar

first: Klaus

last: Kaestner


title: Identification of Transcription Factor PHA-4::GFP Binding Regions in L2

status: Public on Mar 31 2011

city: Toronto

first: DCC

last: modENCODE

state: Ontario

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

country: Canada

institute: Ontario Institute for Cancer Research

summary:
modENCODE_submission_3158
This submission comes from a modENCODE project of Michael Snyder. For full list of modENCODE projects, see http://www.genome.gov/26524648
Project Goal: We are identifying the DNA binding sites for 300 transcription factors in C. elegans. Each transcription factor gene is tagged with the same GFP fusion protein, permitting validation of the gene's correct spatio-temporal expression pattern in transgenic animals.  Chromatin immunoprecipitation on each strain is peformed using an anti-GFP antibody, and any bound DNA is deep-sequenced using Solexa GA2 technology.
For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: EXPERIMENT TYPE: CHIP-seq. BIOLOGICAL SOURCE: Strain: OP37 (official name : OP37 genotype : unc-119(ed3) III; wgIs37 [unc-119(+) pha-4::TY1::EGFP::3xFLAG] outcross : 3 mutagen : Bombard tags : GFP::3xFlag description : This strain's transgene was constructed by Mihail Sarov at the Max Planck Institute for Cell Biology in Tubiginen using Tony Hyman's recombineering pipeline.  The resulting plasmid was used for biolistic transformation of an unc-119(ed3) strain.  The PHA-4::EGFP fusion protein is expressed in the correct pha-4 spatio-temporal expression pattern.  This strain was used for ChIP-seq experiments to map the in vivo binding sites for the PHA-4 transcription factor. made_by : R. Waterston and S. Kim ); Developmental Stage: L2; Genotype: unc-119(ed3) III; wgIs37 [unc-119(+) pha-4::TY1::EGFP::3xFLAG]; Sex: Hermaphrodite;  EXPERIMENTAL FACTORS: Developmental Stage L2; Target gene pha-4; Strain OP37 (official name : OP37 genotype : unc-119(ed3) III; wgIs37 [unc-119(+) pha-4::TY1::EGFP::3xFLAG] outcross : 3 mutagen : Bombard tags : GFP::3xFlag description : This strain's transgene was constructed by Mihail Sarov at the Max Planck Institute for Cell Biology in Tubiginen using Tony Hyman's recombineering pipeline.  The resulting plasmid was used for biolistic transformation of an unc-119(ed3) strain.  The PHA-4::EGFP fusion protein is expressed in the correct pha-4 spatio-temporal expression pattern.  This strain was used for ChIP-seq experiments to map the in vivo binding sites for the PHA-4 transcription factor. made_by : R. Waterston and S. Kim ); temp (temperature) 20 degree celsius

first: Mei

last: Zhong

first: Mike

last: Snyder

first: Cindie

last: Slightam

first: Stuart

last: Kim

first: John

last: Murray

first: Robert

last: Waterston

first: Mark

last: Gerstein

first: Wei

last: Niu

first: Judi

last: Janette

first: Debasish

last: Raha

first: Ashish

last: Agarwal

first: Valerie

last: Reinke

first: Mihail

last: Sarov

first: Anthony

last: Hyman


title: High-throughput pyrosequencing of endogenous small RNAs in O. Europaea

city: Murcia

first: Livia

last: Donaire

state: Murcia

address: Campus de Espinardo

department: Plant Pathology

country: Spain

institute: Centro de Edafología y Biología Aplicada del Segura

summary: Small RNAs of 20 to 25 nucleotides in length maintain genome integrity and control gene expression in a multitude of developmental and physiological processes. Despite RNA silencing has been primarily studied in model plants, the advent of high-throughput sequencing technologies has enabled profiling of the small RNA component of more than 40 plant species. Here, use deep sequencing and molecular methods to report the first inventory of small RNAs in olive (Olea europaea). Small RNAs of 24 nts dominate the small RNA transcriptome and atypically accumulate to levels never seen in other plant species, suggesting an active role of heterochromatin silencing in the maintenance and integrity of its large genome. By contrast, small RNAs of 20 to 22 nts were poorly represented in the population at levels lower than those found in most plant species tested. A total of 14 known miRNA families were identified in two libraries prepared from growing and dormant lateral buds. We found that some known miRNAs showed tissue- and/or developmental-specific expression. Also, seven novel, olive-specific miRNA candidates were found in our sequenced set of which 1 were supported by their star strands. Potential precursors for these miRNA candidates with intramolecular folding capacities were found in the olive EST database. Target mRNAs of conserved miRNAs and new olive-specific miRNA were computationally predicted among the olive EST collection and experimentally validated through endonucleolytic cleavage assays.

overall_design: Two samples analyzed: growing (juvenile) and dormant (adult) lateral buds from the progeny of a genetic cross between the Picual and Arbequina olive varieties

first: Laia

last: Pedrola

first: Raúl

last: de la Rosa

first: César

last: LLave


title: Nucleosome map of the IMR90 fibroblast cell line

city: Los Angeles

first: Benjamin

last: Berman

department: USC Epigenome Center

institute: University of Southern California

summary: We produced a map of nucleosome positions in IMR90 by sequencing the ends of MNase-digested chromatin fragments.

overall_design: IMR90 cells were grown in culture, about 1E6 cells were isolated and digested using micrococcal nuclease (MNase). Mononucleosomes were gel-selected and fragment ends were sequenced using the Illumina GAIIx sequencing platform.

first: Theresa

last: Kelly

first: Peter

last: Jones

last: Laird


title: Identification of Transcription Factor EGL-5::GFP Binding Regions in L3

first: DCC

last: modENCODE

summary:
modENCODE_submission_3159
This submission comes from a modENCODE project of Michael Snyder. For full list of modENCODE projects, see http://www.genome.gov/26524648
Project Goal: We are identifying the DNA binding sites for 300 transcription factors in C. elegans. Each transcription factor gene is tagged with the same GFP fusion protein, permitting validation of the gene's correct spatio-temporal expression pattern in transgenic animals.  Chromatin immunoprecipitation on each strain is peformed using an anti-GFP antibody, and any bound DNA is deep-sequenced using Solexa GA2 technology.
For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: EXPERIMENT TYPE: CHIP-seq. BIOLOGICAL SOURCE: Strain: OP54 (official name : OP54 genotype : unc-119(ed3) III; wgIs54 [unc-119(+) egl-5::TY1::EGFP::3xFLAG] outcross : 3 mutagen : Bombard tags : GFP::3xFlag description : This strain's transgene was constructed by Mihail Sarov at the Max Planck Institute for Cell Biology in Tubiginen using Tony Hyman's recombineering pipeline.  The resulting plasmid was used for biolistic transformation of an unc-119(ed3) strain.  The EGL-5::EGFP fusion protein is expressed in the correct egl-5 spatio-temporal expression pattern.  This strain was used for ChIP-seq experiments to map the in vivo binding sites for the EGL-5 transcription factor. made_by : R. Waterston and S. Kim ); Developmental Stage: L3; Genotype: unc-119(ed3) III; wgIs54 [unc-119(+) egl-5::TY1::EGFP::3xFLAG]; Sex: Hermaphrodite;  EXPERIMENTAL FACTORS: Developmental Stage L3; Target gene egl-5; Strain OP54 (official name : OP54 genotype : unc-119(ed3) III; wgIs54 [unc-119(+) egl-5::TY1::EGFP::3xFLAG] outcross : 3 mutagen : Bombard tags : GFP::3xFlag description : This strain's transgene was constructed by Mihail Sarov at the Max Planck Institute for Cell Biology in Tubiginen using Tony Hyman's recombineering pipeline.  The resulting plasmid was used for biolistic transformation of an unc-119(ed3) strain.  The EGL-5::EGFP fusion protein is expressed in the correct egl-5 spatio-temporal expression pattern.  This strain was used for ChIP-seq experiments to map the in vivo binding sites for the EGL-5 transcription factor. made_by : R. Waterston and S. Kim ); temp (temperature) 20 degree celsius

first: Mei

last: Zhong

first: Mike

last: Snyder

first: Cindie

last: Slightam

first: Stuart

last: Kim

first: John

last: Murray

last: Waterston


title: Genome-wide analysis reveals unique regulation of transcription of Th2-specific genes by GATA3

city: Chiba

first: Atsushi

last: Onodera

address: Chuo-ku Inohana 1-8-1

department: Immunology

institute: Chiba University

summary: Differentiation of naive CD4 T cells into type 2 helper (Th2) cells is accompanied by chromatin remodeling and increased expression of a set of Th2-specific genes including those encoding Th2 cytokines.  IL-4-mediated STAT6 activation induces high levels of transcription of GATA3, a master regulator of Th2 cell differentiation, and enforced expression of GATA3 induces Th2 cytokine expression.  However, it remains unclear whether the expression of other Th2-specific genes is induced directly by GATA3.  A genome-wide unbiased ChIP-seq analysis revealed that GATA3 bound to 1,279 genes selectively in Th2 cells, and 101 genes in both Th1 and Th2 cells.  Simultaneously, we identified 26 highly Th2-specific STAT6-dependent inducible genes by a DNA microarray analysis-based three-step selection processes, and among them 17 genes showed GATA3 binding.  We assessed dependency on GATA3 for the transcription of these 26 Th2-specific genes, and 10 genes showed increased transcription in a GATA3-dependent manner while 16 genes showed no significant responses.  The transcription of the 16 GATA3-nonresponding genes was clearly increased by the introduction of an active form of STAT6, STAT6VT.  Therefore, although GATA3 has been recognized as a master regulator of Th2 cell differentiation, many Th2-specific genes are not regulated by GATA3 itself but in collaboration with STAT6.

overall_design:
Th1 and Th2  subsets are profiled for mRNA expression
Examination of GATA3-binding and 3 different histone modifications in Th1 and Th2 cells.

first: S

last: Horiuchi

first: A

first: H

last: Hosokawa

first: Y

last: Watanabe

first: T

last: Tanaka

last: Sugano

last: Suzuki

last: Nakayama


title: A modified bacterial one-hybrid system yields improved quantitative models of transcription factor specificity

city: Worcester

first: Scot

last: Wolfe

state: MA

address: 364 Plantation Street, LRB 619

department: MCCB

institute: UMass Medical School

summary: We examine the use of high-throughput sequencing on binding sites recovered using a bacterial one-hybrid (B1H) system and find that improved models of transcription factor (TF) binding specificity can be obtained compared to standard methods of sequencing a small subset of the selected clones. We can obtain even more accurate binding models using a modified version of B1H selection method with constrained variation (CV-B1H). However, achieving these improved models using CV-B1H data required the development of a new method of analysis - GRaMS (Growth Rate Modeling of Specificity) - that estimates bacterial growth rates as a function of the quality of the recognition sequence.  We benchmark these different methods of motif discovery using Zif268, a well characterized C2H2 zinc finger transcription factor on both a 28bp randomized library for the standard B1H method and on 6bp randomized library for the CV-B1H method for which forty-five different experimental conditions were tested: 5 time points and three different IPTG and 3-AT concentrations.  We find that GRaMS analysis is robust to the different experimental parameters whereas other analysis methods give widely varying results depending on the conditions of the experiment. Finally, we demonstrate that the CV-B1H assay can be performed in liquid media, which produces recognition models that are similar in quality to sequences recovered from selection on solid media.

overall_design: All selections were performed using the bait transcription factor Zif268.  45 different B1H selections were performed on plates and the selected sites were multiplexed and sequenced in a single lane.  A single selection was performed in liquid media and sequenced along with other multiplexed sets of sites in a separate lane.  For the experiments performed on plates, the of the selections assays was varied (4, 8, 12, 18, 24 hours).  The stringency of the selections was also varied by changing the concentration of IPTG (0, 10, 50 uM) and 3-AT (0.5, 1, 2 mM).  The initial randomized library used for all selections was also sequenced in a single lane.  The randomized portion of each transcription factor binding site is 6bp long and is immediately 5' of the constant sequence, GCGG, which is the consensus binding site for finger 1 of Zif268.  Upstream of the randomized binding site is a 4bp randomized region referred to as the key region.  The purpose of the key region is to detect possible PCR 'jackpotting.'  The single liquid media experiment was run for 4 hours using 50uM IPTG and 5mM 3-AT.  Four selections were performed using a prey library with a 28bp long randomized region and not fixed consensus site region. These selections were performed at either 36 or 48 hours using 10uM IPTG and 0, 2, or 5 mM 3-AT.

first: Gary

middle: D

last: Stormo


title: Regulating RNA Polymerase Pausing and Elongation in Embryonic Stem Cell Transcription

city: Bethesda

first: Josh

last: Waterfall

address: 37 Convent Dr Room 6138

institute: NIH

summary: Transitions between pluripotent stem cells and differentiated cells are executed by key transcription regulators. Comparative measurements of RNA polymerase distribution over the genome’s primary transcription units in different cell states can identify the genes and steps in the transcription cycle that are regulated during such transitions. To identify the complete transcriptional profiles of RNA polymerases with high sensitivity and resolution, as well as the critical regulated steps upon which regulatory factors act, we used genome-wide, nuclear run-on (GRO-seq) to map the density and orientation of transcriptionally-engaged RNA polymerases in mouse embryonic stem cells (ESCs) and embryonic fibroblasts (MEFs).  In both cell types, progression of a promoter-proximal, paused RNA polymerase II (Pol II) into productive elongation is a rate-limiting step in transcription of ~40% of mRNA-encoding genes. Importantly, quantitative comparisons between cell types reveal that transcription is controlled frequently at paused Pol II’s entry into elongation. Furthermore, “bivalent” ESC genes (exhibiting both active and repressive histone modifications) bound by Polycomb Group Complexes PRC 1 and PRC2 show dramatically reduced levels of paused Pol II at promoters relative to an average gene. In contrast, bivalent promoters bound by only PRC2 allow Pol II pausing, but it is confined to extremely 5’ proximal regions. Altogether, these findings identify rate-limiting targets for transcription regulation during cell differentiation.

overall_design:
Mapping engaged RNA polymerase density in two cell types by sequencing run-on transcripts.

SUPPLEMENTARY FILES:
All fastq files have sanger-fastq format q values.
Alignments were generated with eland and the mm9 mouse genome assembly.  Reads aligning to regions annotated as similar to rRNA by RepeatMasker were then removed.
Wiggle files are in units of RPKM (reads per kilobase per million aligned reads) and are broken up by cell type and chromosome to aid in uploading to UCSC.  Each file furthermore contains two tracks - one for each strand.  As in the published paper, plus strand RPKM densities are in red with positive values and minus strand RPKM densities are in blue with negative values.

first: Irene

middle: M

last: Min

first: Joshua

middle: T

last: Lis


title: Dual functions of Tet1 in transcriptional regulation in mouse embryonic stem cells (ChIP-Seq)

status: Public on Mar 30 2011

city: Boston

first: Hao

last: Wu

state: Massachusetts

address: 149G Warren Alpert Building, 200 Longwood Avenue

institute: Harvard Medical School/HHMI

summary: Epigenetic modification of the mammalian genome by DNA methylation (5-methylcytosine) has a profound impact on chromatin structure, gene expression and maintenance of cellular identity. Recent demonstration that members of the Ten-eleven translocation (Tet) family proteins can convert 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) raised the possibility that Tet proteins are capable of establishing a distinct epigenetic state. We have recently demonstrated that Tet1 is specifically expressed in murine embryonic stem (ES) cells and is required for ES cell self-renewal and maintenance. Using chromatin immunoprecipitation coupled with high-throughput DNA sequencing (ChIP-seq), here we show that Tet1 is preferentially bound to CpG-rich sequences at promoters of both transcriptionally active and Polycomb-repressed genes. Despite a general increase in levels of DNA methylation at Tet1 binding-sites, Tet1 depletion does not lead to down-regulation of all the Tet1 targets. Interestingly, while Tet1-mediated promoter hypomethylation is required for maintaining the expression of a group of transcriptionally active genes, it is also required for repression of Polycomb-targeted developmental regulators. Tet1 contributes to silencing of this group of genes by facilitating recruitment of PRC2 to CpG-rich gene promoters. Thus, our study not only establishes a role for Tet1 in modulating DNA methylation levels at CpG-rich promoters, but also reveals a dual function of Tet1 in promoting transcription of pluripotency factors as well as participating in the repression of Polycomb-targeted developmental regulators.

overall_design: To determine the genome-wide distribution of Tet1 in mouse ES cells, we have performed ChIP-seq experiments using Tet1 antibodies in control knockdown (KD) and Tet1 KD ES cells.

first: Ana

last: D'Alessio

first: Shinsuke

last: Ito

first: Kai

last: Xia

first: Zhibin

last: Wang

first: Keji

last: Zhao

first: Yi

last: Sun

first: Kairong

last: Cui

last: Zhang


title: Two-step RNA sequencing identifies a tissue-specific landscape of sense/antisense transcription

status: Public on Mar 29 2011

city: Kiel

first: Robert

last: Haesler

address: Schittenhelmstr. 12

department: IKMB / 1st Dept. of Medicine

institute: University Clinic of Schleswig Holstein

summary: We report the complexity of the polyadenylated transcriptome of the mouse intestinal tract assessed by two step RNA sequencing.

overall_design: Screening and verification of non-annotated transcriptional units by different RNA-Seq protocols.

first: Ulrich

last: Klostermeier

first: Philip

last: Rosenstiel


title: High-throughput sequencing of mRNA from oocyte, 1 cell, 16/32 cells, 128/256 cells, 3.5hpf and 5.3hpf zebrafish embryos (Wild type; AB line)

city: Singapore

first: Justin

middle: Jeyakani

last: Joseph Gnanakkan

state: Singapore

address: 60, Biopolis street

department: Computational and Systems Biology

country: Singapore

institute: Genome Institute of Singapore

summary:
mRNA seq based approach to determine the transcriptome dynamics during early development
To study the mechanisms regulating this developmental event in zebrafish, we applied RNA deep sequencing technology and generated comprehensive transcriptome profiles of 6 developmental stages from oocyte to early gastrulation. We determined the expression levels of maternal and zygotic transcripts and clustered them based on expression pattern. We identified a large number of novel transcribed regions in un-annotated regions of the genome, as well as splice variants with an estimated frequency of 40-75% during early zebrafish embryogenesis. Our data constitute a useful resource for developmental studies, gene discovery, and genome annotation.

overall_design: RNA was extracted from pooled embryos of desired stages and one RNA seq library was generated for each sample.  Totally 6 stages were selected: Maternal, 1cell, 16/32 cells, 128/256 cells, 3.5hpf and 5.3hpf

first: Sinnakaruppan

last: Mathavan


title: Repertoire of porcine microRNAs in adult ovary and testis by deep sequencing.

city: Ya'an

first: Mingzhou

last: Li

state: Sichuan

address: No.46  Xinkang Road

department: College of Animal Science and Technology

institute: Sichuan Agricultural University

summary: Despite increasing efforts in miRNAs identification across various species and diverse tissue types, little is known about porcine gonad-specific miRNAs. Although the well-known importance of pig in agriculture, as well as a model for human biology, the miRNA catalog of pig has been largely undefined. Identification and preliminary characterization of gonad-specific miRNAs would be a prerequisite for a thorough understanding of their roles in regulating folliculogenesis and spermatogenesis. In the present study, we get insight into miRNA transcriptome in adult porcine ovary and testis using deep sequencing technology, and to elucidate their characteristic organ- and gender-specific profiles, genomic context and emphasize the features of X-linked miRNAs.

overall_design: Two small RNA libraries from adult porcine ovary and testis tissues were sequenced.

first: Haosi

last: Chen

first: Xuewei

first: Xiaolian

last: Gao


title: A genome-scale shRNA resource for transgenic RNAi in Drosophila

status: Public on Mar 26 2011

city: Cold Spring Harbor

first: Benjamin

last: Czech

address: 1 Bungtown Rd

institute: Cold Spring Harbor Laboratory

summary: Existing transgenic RNAi resources in Drosophila utilize the expression of long double stranded hairpin RNAs and are powerful tools for the identification and characterization of genes involved in various processes. However, they are ineffective to knockdown genes during oogenesis, an important model system for the study of numerous fundamental questions in biology. Here, we show that short hairpin RNAs (shRNAs), modeled on an endogenous microRNA, are extremely effective at silencing gene expression during oogenesis. We also describe our progress towards building a genome-wide shRNA resource.

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

first: Jian-Quan

last: Ni

first: Rui

last: Zhou

first: Lu-Ping

last: Liu

first: Laura

last: Holderbaum

first: Donghui

last: Yang-Zhou

first: Hye-Seok

last: Shim

first: Dominik

last: Handler

first: Phillip

last: Karpowicz

first: Richard

last: Binari

first: Matthew

last: Booker

first: Julius

last: Brennecke

first: Perkins

last: Lizabeth

first: Gregory

last: Hannon

first: Norbert

last: Perrimon


title: Dual functions of Tet1 in transcriptional regulation in mouse embryonic stem cells

first: Hao

last: Wu

type: Genome binding/occupancy profiling by genome tiling array


title: The histone methyltransferase SETDB1 is recurrently amplified in melanoma and accelerates its onset

status: Public on Mar 25 2011

city: Cambridge

last: Young

address: 9 Cambridge Center

institute: Whitehead Institute for Biomedical Research

summary: The most common mutation in human melanoma, BRAF(V600E), activates the serine/threonine kinase BRAF and causes excessive activity in the mitogen-activated protein kinase pathway. BRAF(V600E) mutations are also present in benign melanocytic naevi, highlighting the importance of additional genetic alterations in the genesis of malignant tumours. Such changes include recurrent copy number variations that result in the amplification of oncogenes. For certain amplifications, the large number of genes in the interval has precluded an understanding of the cooperating oncogenic events. Here we have used a zebrafish melanoma model to test genes in a recurrently amplified region of chromosome 1 for the ability to cooperate with BRAF(V600E) and accelerate melanoma. SETDB1, an enzyme that methylates histone H3 on lysine 9 (H3K9), was found to accelerate melanoma formation significantly in zebrafish. Chromatin immunoprecipitation coupled with massively parallel DNA sequencing and gene expression analyses uncovered genes, including HOX genes, that are transcriptionally dysregulated in response to increased levels of SETDB1. Our studies establish SETDB1 as an oncogene in melanoma and underscore the role of chromatin factors in regulating tumorigenesis.

overall_design: ChIP was performed from short-term cultures of WM262 and WM451Lu cells.

first: Steve

last: Bilodeau


title: The pan-genome of the dominant human gut-associated archaeon, Methanobrevibacter smithii

city: St Louis

first: Federico

last: Rey

state: MO

address: 4444 Forest Park Blvd

institute: Washington University

summary:
The human gut microbiota harbors methanogens represented by the dominant archaeon, Methanobrevibacter smithii, a polyphyletic group of acetogens, and sulfate-reducing bacteria.  Defining their roles in the H2-economy of the gut has potential therapeutic importance for modulating the efficiency of fermentation of dietary components. We quantified methanogens in fecal samples from 40 healthy adult female monozygotic(MZ) and 28 dizygotic(DZ) twin pairs, analyzed bacterial 16S rRNA datasets generated from their fecal samples to identify taxa that co-occur with methanogens, sequenced the genomes of 20 M. smithii strains isolated from families of MZ and DZ twins, and performed RNA-Seq of a subset of strains to identify their responses to varied formate concentrations. The concordance rate for methanogen carriage was significantly higher for MZ versus DZ twin pairs.  Co-occurrence analysis revealed 22 bacterial species-level taxa positively correlated with methanogens: all but two were members of the Clostridiales, with several being, or related to, known hydrogen-producing and -consuming bacteria. The M. smithii pan-genome contains 987 genes conserved in all strains, and 1860 variably represented genes. Strains from MZ and DZ twin pairs had a similar degree of shared genes and SNPs, and were significantly more similar than strains isolated from mothers or members of other families. The 101 adhesin-like proteins(ALPs) in the pan-genome (45±6/strain) exhibit strain-specific differences in expression and responsiveness to formate. We hypothesize that M. smithii strains use their different repertoires of ALPs to create diversity in their metabolic niches, by allowing them to establish syntrophic relationships with bacterial partners with differing metabolic capabilities and patterns of co-occurrence
These strains were isolated from human feces, but they are in pure culture now. All the information about each species is associated with the genome accession number

sample_organism:
Methanobrevibacter smithii
Methanobrevibacter smithii ATCC 35061

overall_design:
Fecal samples from 40 healthy adult female monozygotic(MZ) and 28 dizygotic(DZ) twin pairs, analyzed bacterial 16S rRNA datasets generated from their fecal samples to identify taxa that co-occur with methanogens, sequenced the genomes of 20 M. smithii strains isolated from families of MZ and DZ twins, and performed RNA-Seq of a subset of strains to identify their responses to varied formate concentrations.
Strains of Methanobrevibacter smithii were grown in vitro (modified MBC media) to mid-log phase, at 37°C in serum bottles pressurized with 80% hydrogen, 20% CO2 gasses at 30psi. Cells were harvested by centrifugation, and DNA was isolated by phenol-chloroform and ethanol precipitation.

first: Elizabeth

last: Hansen


title: Metatranscriptomics provides insight into the physiologies of the digestive tract symbionts within the medicinal leech, Hirudo verbana

city: Storrs

first: Joerg

last: Graf

state: CT

address: 91 North Eagleville Road

institute: University of Connectiuct

summary: Here, we report the use of Illumina RNA-Seq for investigating the physiology of the digestive-tract microbiome within the medicinal leech, Hirudo verbana.  About 12 million cDNA reads were mapped against the genomes of the two dominant members of this simple microbiome.  Results suggested that the most abundant, yet uncultured Rikenella-like bacterium forages host mucin glycans and ferments the carbohydrates to acetate that is secreted into the environment.  The second dominant symbiont, Aeromonas veronii, appears to utilize the acetate secreted by Rikenella as a carbon and energy source, possibly linking the physiologies of the dominant symbionts.  This study demonstrates how RNA-seq can be used to reveal the physiology of a naturally occurring microbiome.

sample_organism:
Hirudo verbana
gut metagenome

platform_organism:
Hirudo verbana
gut metagenome

overall_design: Examination of the physiology of the leech gut microbiome 42 hrs after ingestion of a blood meal

first: Lindsey

last: Bomar


title: The androgen receptor coordinates biosynthesis and proliferation in prostate cancer

status: Public on Mar 24 2011

first: Suraj

last: Menon

address: Li Ka Shing Center, Robinson Way

country: United Kingdom

institute: Cambridge Research Institute, Cancer Research UK

summary: The androgen receptor (AR) is the major therapeutic target in prostate cancer, although the important targets of the AR have remained obscure.  Here we report a detailed genomic profile of AR signalling and find that the AR directly regulates glycolysis, anabolic metabolism and cell cycle regulators in prostate cancer.  This coordinated transcriptional program promotes cancer cell proliferation and enhances the macromolecular synthesis needed to produce daughter cells.  Clinical gene expression profiles and mechanistic studies highlight the importance of CAMKK2, an AR target which regulates both cell proliferation and metabolism. Thus our genomics study identifies a direct link between AR signalling and aerobic glycolysis (the Warburg effect), providing a new perspective on the oncogenic function of the AR in prostate cancer.

overall_design: 8 Samples: Chromatin IP using AR and PolII in stimulated and unstimulated LNCaP and VCaP cells.

first: Massie

last: Charlie

first: Stark

last: Rory


title: Identification of Transcription Factor ALR-1::GFP Binding Regions in L2

first: DCC

last: modENCODE

summary:
modENCODE_submission_3156
This submission comes from a modENCODE project of Michael Snyder. For full list of modENCODE projects, see http://www.genome.gov/26524648
Project Goal: We are identifying the DNA binding sites for 300 transcription factors in C. elegans. Each transcription factor gene is tagged with the same GFP fusion protein, permitting validation of the gene's correct spatio-temporal expression pattern in transgenic animals.  Chromatin immunoprecipitation on each strain is peformed using an anti-GFP antibody, and any bound DNA is deep-sequenced using Solexa GA2 technology.
For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: EXPERIMENT TYPE: CHIP-seq. BIOLOGICAL SOURCE: Strain: OP200 (official name : OP200 genotype : unc119(ed3);wgIs200(alr-1::TY1 EGFP FLAG C;unc119) outcross : 3 mutagen : Bombard tags : GFP::3xFlag description : his strain's transgene was constructed by Mihail Sarov at the Max Planck Institute for Cell Biology in Tubiginen using Tony Hyman's recombineering pipeline.  The resulting plasmid was used for biolistic transformation of an unc-119(ed3) strain.  The ALR-1::EGFP fusion protein is expressed in the correct alr-1 spatio-temporal expression pattern.  This strain was used for ChIP-seq experiments to map the in vivo binding sites for the ALR-1 transcription factor. made_by : R. Waterston ); Developmental Stage: L2; Genotype: unc119(ed3);wgIs200(alr-1::TY1 EGFP FLAG C;unc119); Sex: Hermaphrodite;  EXPERIMENTAL FACTORS: Developmental Stage L2; Target gene alr-1; Strain OP200 (official name : OP200 genotype : unc119(ed3);wgIs200(alr-1::TY1 EGFP FLAG C;unc119) outcross : 3 mutagen : Bombard tags : GFP::3xFlag description : his strain's transgene was constructed by Mihail Sarov at the Max Planck Institute for Cell Biology in Tubiginen using Tony Hyman's recombineering pipeline.  The resulting plasmid was used for biolistic transformation of an unc-119(ed3) strain.  The ALR-1::EGFP fusion protein is expressed in the correct alr-1 spatio-temporal expression pattern.  This strain was used for ChIP-seq experiments to map the in vivo binding sites for the ALR-1 transcription factor. made_by : R. Waterston ); temp (temperature) 20 degree celsius

first: Mei

last: Zhong

first: Mike

last: Snyder

first: Cindie

last: Slightam

first: Stuart

last: Kim

first: John

last: Murray

first: Robert

last: Waterston

first: Mark

last: Gerstein

first: Wei

last: Niu

first: Judi

last: Janette

first: Debasish

last: Raha

first: Ashish

last: Agarwal

first: Valerie

last: Reinke

first: Mihail

last: Sarov

first: Anthony

last: Hyman


title: CNV CME W1 Cl.8+ Cells

status: Public on Mar 23 2011

summary:
modENCODE_submission_3201
This submission comes from a modENCODE project of David MacAlpine. For full list of modENCODE projects, see http://www.genome.gov/26524648
Project Goal: Analysis of copy number variation across modENCODE Drosophila cell line by high throughput sequencing.  We observe numerous copy number variations for the different cell lines, with each cell line having a distinct ploidy signature.
For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design:
EXPERIMENT TYPE: deep sequencing analysis. BIOLOGICAL SOURCE: Cell Line: CME-W1-Cl.8+; Tissue: dorsal mesothoracic disc; Developmental Stage: third instar larval stage; Sex: Male;  EXPERIMENTAL FACTORS: Cell Line CME-W1-Cl.8+
In this experiment, there is no ChIP step. Simply separate the polytene dna from the tissue (cell line) and seq them, if more reads (than other regions) then they are the CNV regions. Two independent samples were merged to generate the single wiggle file.

first: David

last: MacAlpine

first: Sara

last: Powell

first: Matthew

last: Eaton


title: Expression data from Pseudomonas putida KT2440

city: Hannover

first: Sarah

last: Frank

address: Carl-Neuberg-Str. 1

department: Clinic for Paediatric Pneumology and Neonatology

institute: Medical School Hannover

summary:
In this study we exploited next-generation Illumina sequencing technology (Wang et al., 2009) to refine the current annotation of the KT2440 genome. Transcriptome sequencing data were queried for yet undescribed small RNAs and ORFs and employed to validate predicted operons and gene coordinates. Expression profiles were measured at 10°C and 30°C to cover the physiologic temperature profile of this mesophilic bacterium. Moreover we compared the sensitivity and specificity of transcriptome data by taking the same RNA preparations for cDNA sequencing and hybridization of Progenika and Affymetrix microarrays.

This SuperSeries is composed of the SubSeries listed below.

overall_design:
P. putida KT2440 (strain DSM6125) (Bagdasarian et al., 1981) was obtained from the German Resource Centre for Biological Material (DSMZ, (Braunschweig, Germany). Bacterial cultures were inoculated from a frozen stock culture of P. putida wild type, and incubated at 30°C for 8 hours at 250 rpm in LB medium. An aliquot of 0.2 mL was added to 20 mL M9 medium (Na2HPO4 33.9 g/L, KH2PO4 15.0 g/L, NaCl 2.5 g/L, NH4Cl 5.0 g/L, MgSO4 2 mM, CaCl2 0.1 mM, FeSO4 x 7 H2O 0.01 mM, pH 6.8) supplemented with 15 mM succinate as sole carbon source in a 100 mL flask and incubated overnight at 30°C. For RNA extraction, bacteria were grown in a 1.5 L batch culture (M9 + 15 mM succinate) using the BioFlo 110 Fermenter (New Brunswick Scientific Co., Edison, NJ) to ensure constant pH, aeration and agitation. When cultures reached mid-exponential phase (OD600 ~ 0.8) the temperature was decreased from 30°C to 10°C. Three samples, each for parallel RNA extraction, were subsequently taken immediately before temperature downshift (30°C) and two hours after the media had been cooled to 10°C.
The gene expression profiles of P. putida KT2440 at 30°C and 10°C were analyzed using three different transcriptome platforms: RNA-Seq (Illumina cDNA sequencing), Affymetrix microarrays and Progenika oligonucleotide microarrays.
For a detailed description of sample preparation (RNA, cDNA , labelling, hybridization etc.) refer to individual Series.