title: RNA-seq reveals that Air1 and Air2 control TRAMP substrate specificity for nuclear RNA surveillance

status: Public on Nov 14 2012

city: Rochester

first: J. Scott

last: Butler

state: NY

address: 601 Elmwood Ave Box 672

department: Department of Microbiology and Immunology

institute: University of Rochester Medical Center

summary: The Saccharomyces cerevisiae TRAMP4 and TRAMP5 complexes, which consist of the poly(A) polymerase Trf4 or Trf5, respectively, the zinc knuckle proteins Air1 or Air2, and the RNA helicase Mtr4, play a critical role in nuclear RNA surveillance. Although it is known to enhance the nuclease activity of the exosome, relatively little is known about the exact mechanism and specificity of TRAMP. To better define the specificities of the TRAMP complexes, we used phenotypic analysis and RNA deep-sequencing technology to measure differences in global RNA polyadenylation in air mutants, revealing specific requirements for each Air protein in the regulation of the levels of non-coding and coding RNAs. These findings reveal differential functions for Air proteins in eukaryotic RNA metabolism and indicate that they control the substrate specificity of the RNA exosome.

overall_design: Poly(A)+ RNA from WT, rrp6-Δ, air1-Δ rrp6-Δ and air2-Δ rrp6-Δ was sequenced using ABI SOLiD platform, in duplicate.

first: David

last: Mathews


title: Chromatin occupancy of key transcriptional regulators in P493-6 cells with various levels of c-Myc expression

city: Cambridge

first: Richard

middle: A

last: Young

state: MA

address: 9 Cambridge Center

country: USA

institute: Whitehead Institute for Biomedical Research

type: Genome binding/occupancy profiling by high throughput sequencing

summary: P493-6 cells are immortalized human peripheral B cells that carry a conditional, tetracycline-regulated myc gene. We present ChIP-seq analysis of key transcritional regulators in P493-6 cells expressing various levels of c-Myc: 0hr (low c-Myc levels), 1hr (intermediate c-Myc levels), 24hr (very high c-Myc levels) and No Tet (steady-state c-Myc levels).

overall_design: Brd4, c-Myc, Max, Med1, RNAPII, and the chromatin modification H3K27Ac were profiled in P493-6 cells

first: Rahl

last: Peter

first: Jakob

last: Loven


title: Bromodomain-Containing-Protein 4 (BRD4) Regulates RNA Polymerase II Serine 2 Phosphorylation in Human CD4+ T Cells

city: Singapore

first: Keh-chuang

last: Chin

address: Biopolis, 8A Biomedical Grove, #03-06 Immunos

department: Laboratory of Gene Regulation

country: Singapore

institute: Singapore Immunology Network

type: Expression profiling by high throughput sequencing

summary: Transcriptional elongation by RNA polymerase II (Pol II) is regulated by positive transcription elongation factor b (P-TEFb) in association with Bromodomain-containing protein 4 (BRD4). We used genome-wide chromatin immunoprecipitation sequencing in primary human CD4+ T cells to reveal that BRD4 co-localizes with Ser2-phosphorylated Pol II (Pol II Ser2) at both enhancers and promoters of active genes. Disruption of bromodomain:histone acetylation interactions by JQ1, a small-molecule bromodomain inhibitor, resulted in decreased BRD4 binding, reduced Pol II Ser2, and reduced expression of lineage-specific genes in primary human CD4+ T cells. A large number of JQ1-disrupted BRD4 binding regions exhibited di-acetylated H4 (lysine-5 and -8) and H3K27 acetylation (H3K27ac), which correlated with the presence of histone acetyltransferases and deacetylases. Genes associated with BRD4/H3K27ac co-occupancy exhibited significantly higher activity than those associated with H3K27ac or BRD4 binding alone. Comparison of BRD4 binding in T cells and in human embryonic stem cells revealed that enhancer BRD4 binding sites were predominantly lineage-specific. Our findings suggest that BRD4-driven Pol II phosphorylation at serine 2 plays an important role in regulating lineage-specific gene transcription in human CD4+ T cells.

overall_design: Examination of BRD4, total Pol II, serine 2 phosphorylated Pol II and serine 5 phosphorylated Pol II binding in CD4+ T cells (with and without JQ1 treatment) and BRD4 binding in human embryonic  stems cell; PolyA RNA expression in CD4+ T cells( with and without JQ1 treatment) using RNA-seq


title: Identification of candidate enhancer elements in unstimulated and stimulated dendritic cells and fibroblasts

status: Public on Nov 19 2012

city: Nice

first: Dominic

last: van Essen

address: 28 Avenue Valombrose

country: France

institute: Institute for Research on Cancer & Aging, Nice (IRCAN)

summary: To identify candidate enhancer elements we analyzed the distribution of two histone modifications associated with enhancers - H3K4me1 and H3K27ac - and one histone modification associated with active transcription - H4 acetylation.

overall_design: ChIP-seq for H3K4me1, H3K27ac and H4ac, and input DNA controls, from 2 cell types (DCs & fibroblasts) under 2 conditions (unstimulated & stimulated)

first: Yina

last: Zhu

first: Simona

last: Saccani


title: Regulatory impact of RNA secondary structure across the Arabidopsis thaliana transcriptome

status: Public on Nov 15 2012

city: Los Angeles

first: Fan

last: Li

state: CA

address: 675 Charles E Young Drive South

department: Pediatrics

institute: University of California Los Angeles

summary: The secondary structure of an RNA molecule plays an integral role in its maturation, regulation, processing, and functionality. However, the global influence of this feature on plant gene expression is still for the most part unclear. Here, we use a high-throughput, sequencing-based, structure-mapping approach in conjunction with transcriptome-wide sequencing of polyA+-selected (RNA-seq), small (smRNA-seq), and ribosome-bound (ribo-seq) RNA populations to investigate the impact of RNA secondary structure on gene expression regulation in Arabidopsis. From this analysis, we find that highly unpaired and paired RNAs are strongly correlated with euchromatic and heterochromatic epigenetic histone modifications, respectively, providing further evidence that secondary structure is necessary for RNA-mediated posttranscriptional regulatory pathways. Additionally, we uncover key structural patterns across protein-coding transcripts that indicate RNA folding demarcates regions of protein translation and likely affects microRNA-mediated regulation of mRNAs in this model plant. We also reveal that RNA folding is significantly anti-correlated with overall transcript abundance, which is likely due to the increased propensity of highly structured mRNAs to be degraded and/or processed into smRNAs. Finally, we find that secondary structure affects mRNA translation, suggesting that this feature regulates plant gene expression at multiple levels. Overall, our findings provide the first global assessment of RNA folding and its significant regulatory effects in a plant transcriptome.

overall_design: Single-stranded RNA sequencing (ssRNA-seq) and ribosome-bound RNA sequencing (ribo-seq) in immature buds. A single replicate of each library.

first: Brian

middle: D

last: Gregory


title: Genome wide analysis of FOXO3 transcription regulation through RNA pol II profiling

city: Utrecht

first: Michal

last: Mokry

address: Lundlaan 6

country: Netherlands

institute: Wilhelmina Children's Hospital, University Medical Center Utrecht

summary:
FOXO transcription factors are key players in diverse cellular processes affecting tumorigenesis, stem cell maintenance and lifespan. To gain insight into mechanisms of FOXO regulated target gene expression, we studied genome-wide effects of FOXO3 activation. Profiling RNA polymerase II changes shows FOXO3 regulates gene expression through transcription initiation. Correlative analysis of FOXO3 and RNA polymerase II ChIP-seq profiles demonstrates FOXO3 to act as a transcriptional activator. Furthermore, this analysis reveals a significant part of FOXO3 gene regulation proceeds through enhancer regions. FOXO3 binds to pre-existing enhancers and further activates these enhancers as shown by changes in histone acetylation and RNA polymerase II recruitment. In addition, FOXO3-mediated enhancer activation correlates with regulation of adjacent genes and pre-existence of chromatin loops between FOXO3 bound enhancers and target genes. Combined, our data elucidate how FOXOs regulate gene transcription and provide insight into mechanisms by which FOXOs can induce different gene expression programs depending on chromatin architecture.
Paper with published gene expression: PMID 22139133.

overall_design: Examination of FOXO3 binding and changes in RNAPII occupancy (0, 4, 24 hours after induction) in colorectal cell line

first: Astrid

last: Eijkelenboom

first: Elzo

last: de Wit

first: Paulien

last: Polderman

first: Almut

last: Schulze

first: Wouter

last: de Laat

first: Edwin

last: Cuppen

first: Boudewijn

middle: M

last: Burgering


title: Polycomb PHF19 binds H3K36me3 and recruits PRC2 and demethylase NO66 to embryonic stem cell genes during differentiation

status: Public on Nov 17 2012

city: Dublin

first: Adrian

middle: P.

last: Bracken

state: Dublin

address: Trinity College, Dublin

department: Genetics

country: Ireland

institute: Trinity College, Dublin

summary: The Polycomb group proteins are repressive chromatin modifiers with essential roles in metazoan development, cellular differentiation and cell fate maintenance. How Polycomb proteins access active chromatin in order to confer transcriptional silencing during lineage transitions remains unclear. Here we show that the Polycomb Repressive Complex 2 (PRC2) component PHF19 binds the active chromatin mark H3K36me3 via its tudor domain. PHF19 associates with the H3K36me3 demethylase NO66, and is required to recruit the PRC2 complex and NO66 to stem cells genes during differentiation, leading to PRC2 mediated H3K27 tri-methylation, loss of H3K36me3 and transcriptional silencing. We propose a model whereby PHF19 functions during ES cell differentiation to transiently bind the H3K36me3 mark via its tudor domain, forming essential contact points that allow recruitment of PRC2 and H3K36me3 demethylase activity to active gene loci during their transition to a Polycomb-repressed state.

overall_design: Examination of PHF19 genome-wide binding in mouse embryonic stem cells

first: Gerard

middle: L

last: Brien

middle: P


title: K562 polyA RNA-Seq

first: Nataly

middle: Moran

last: Cabili

address: 7  Cambridge Center

institute: Broad Institute ;  Harvard University

summary: RNA-Seq reads and TopHat (Trapnell et al. Bioinformatics 2009) alignments of K562 cell-line transcriptome.  These were used to validate the expression of short peptides idenitified by Mass-Spectrometry in K562 cells.

overall_design: K562 polyA+ RNA (Batch 1) and total RNA (batch 2) was purchased from Ambion. We used oligo (dT)-selected polyA+ RNA to construct libraries for RNA-Seq.We then profiled the transcriptome of polyadenylated mRNA-Seq using Illumina sequencing platforms. We then used the sequenced reads to reconstruct the transcriptome using the Cufflinks de-novo assembler (Trapnell et al. Nat.Bio.Tech. 2010). Recent computational and ribosome profiling analyses suggest that many short open reading frames (sORFs) in eukaryotic genomes are translated. However, evidence that these sORFs produce stable polypeptides is lacking. Here we develop a strategy to discover and validate novel sORF-encoded polypeptides (SEPs) in human cells. In total, we detect 117 SEPs, 114 of which are novel, varying in length from 15 to 149 amino acids. Of these, 10 SEPs (0.5%) are derived from long intergenic non-coding RNAs (lincRNAs). We also observe the presence of polycistronic genes and the widespread use of non-AUG start codons, which is a phenomenon historically thought to be rare in the mammalian genome. Quantitative measurements reveal that SEPs can be found at concentrations between ~10-2000 copies per cell, which is within the range of typical cellular proteins. We confirm the translation of a number of these SEPs through heterologous expression of their encoding cDNAs. We also discover that several SEPs possess properties characteristic of functional proteins. These results demonstrate that human sORFs produce numerous stable polypeptides, revealing that the human proteome is larger and more diverse than previously appreciated.

first: Moran

middle: N

first: Joshua

middle: Z

last: Levin

first: Andrew

last: Mitchell

first: Sarah

last: Slavoff

first: Adam

last: Schwaid

first: John

last: Rinn

first: Alan

last: Saghatelian


title: Silencing of Germline-Expressed Genes by DNA Elimination in Somatic Cells

status: Public on Nov 13 2012

city: Aurora

first: Jianbin

last: Wang

state: CO

address: 12801 East 17th Ave

department: Biochemistry and Molecular Genetics

institute: University of Colorado Denver

summary: Chromatin diminution is the programmed elimination of specific DNA sequences during development. It occurs in diverse species, but the function(s) of diminution and the specificity of sequence loss remain largely unknown. Diminution in the nematode Ascaris suum occurs during early embryonic cleavages and leads to the loss of germline genome sequences and the formation of a distinct genome in somatic cells. We found that ∼43 Mb (∼13%) of genome sequence is eliminated in A. suum somatic cells, including ∼12.7 Mb of unique sequence. The eliminated sequences and location of the DNA breaks are the same in all somatic lineages from a single individual and between different individuals. At least 685 genes are eliminated. These genes are preferentially expressed in the germline and during early embryogenesis. We propose that diminution is a mechanism of germline gene regulation that specifically removes a large number of genes involved in gametogenesis and early embryogenesis.

overall_design: To profile the tissue expression of all A. suum genes, we used 8 different developmental stages/tissues of A. suum, including testis, ovary, embryo, larvae, intestine, muscle, male carcass and female carcass. In this submission there are 10 samples: 3 for early embryos (24hr, 46hr and 64hr) and the other 7 for testis, ovary, larvae, intestine, muscle, male carcass and female carcass. For each sample, 200 ug of total RNA was used for poly(A) selection and 200 ng of poly(A)+ RNA was used to make the cDNA libraries using standard Illumina protocol.

first: Richard

middle: E

last: Davis


title: High-throughput sequencing of small RNAs (25-70 nt) from Drosophila wild-type and Dnmt2 mutants

status: Public on Nov 12 2012

city: Vienna

first: Schaefer

last: Matthias

state: AT

address: Schwarzspanierstrasse 17-I

department: Cell and Developmental Biology

country: Austria

institute: Medical University Vienna, Center for Anatomy & Cell Biology

type: Non-coding RNA profiling by high throughput sequencing

summary: Dnmt2 genes are highly conserved tRNA methyltransferases with biological roles in cellular stress responses. The absence of obvious mutant phenotypes under laboratory conditions suggested a function for Dnmt2 under non-physiological conditions. Indeed, Dnmt2 has recently been implicated in various aspects of the cellular stress response  and the tRNA methyltransferase activity of Dnmt2 has been shown to interfere with stress-induced fragmentation of various tRNAs. We used adult animals and small RNA sequencing during a heat stress experiment to determine the tRNA fragment abundance and identities in wild-type and Dnmt2 mutant somatic tissues. Dnmt2 mutants produced tRNA fragments with different identities when compared to wild-type controls, indicating the accumulation of non-physiological tRNA-derived molecules in tissues without Dnmt2.

overall_design: 6 samples examined: heterozygous and Dnmt2 mutant under control, heat shock and recovery conditions

first: Zeljko

last: Durdevic

first: Matthias

last: Schaefer


title: High-throughput 16S rRNA amplicon pyrosequencing of contaminated aquifer sediment

status: Public on Nov 11 2012

city: Neuherberg

first: Giovanni

last: Pilloni

address: Ingolstädter Landstr. 1

country: Germany

institute: Helmholtz-Centre Munich

summary: Contaminated aquifer (Dusseldorf-Flinger, Germany) templates extracted from 5 sediment depths ranging between 6.4 and 8.4 m below ground and over 3 years of sampling were amplified for amplicon pyrosequencing using the primers Ba27f (5’-aga gtt tga tcm tgg ctc ag-3’) and Ba519r (5’- tat tac cgc ggc kgc tg-3’), extended as amplicon fusion primers with respective primer A or B adapters, key sequence and multiplex identifiers (MID) as recommended by 454/Roche. Amplicons were purified and pooled as specified by the manufacturer. Emulsion PCR (emPCR), purification of DNA-enriched beads and sequencing run were performed following protocols and using a 2nd generation pyrosequencer (454 GS FLX Titanium, Roche) as recommended by the developer. Quality filtering of the pyrosequencing reads was performed using the automatic amplicon pipeline of the GS Run Processor (Roche), with a slight modification concerning the valley filter (vfScanAllFlows false instead of TiOnly) to extract the sequences. Demultiplexed raw reads were furhter trimmed for quality and lenght (>250 bp).

overall_design: 15 samples examined in total from important plume zones of the aquifer sampled in Feb. 2006, Sep. 2008 and Jun. 2009 (5 every year of sampling).

first: Tillmann

last: Lueders

first: Marion

last: Engel


title: Nascent-Seq Reveals Novel Features of Mouse Circadian Transcriptional Regulation [StrandSpe_NascentSeq]

status: Public on Nov 10 2012

city: College Station

first: Jerome

middle: S

last: Menet

state: TX

address: 301 Old Main Drive, Building 1530, ILSB

department: Dept of Biology

institute: Texas A&M University

summary: Over the past decade, genome-wide assays have underscored the broad sweep of circadian gene expression. A substantial fraction of the transcriptome undergoes oscillations in many organisms and tissues, which governs the many biochemical, physiological and behavioral functions under circadian control.  Based predominantly on the transcription feedback loops important for core circadian timekeeping, it is commonly assumed that this widespread mRNA cycling reflects circadian transcriptional cycling. To address this issue, we directly measured dynamic changes in mouse liver transcription using Nascent-Seq. Many genes are rhythmically transcribed over the 24h day, which include precursors of several non-coding RNAs as well as the expected set of core clock genes. Surprisingly however, nascent RNA rhythms overlap poorly with mRNA abundance rhythms assayed by RNA-seq. This is because most mouse liver genes with rhythmic mRNA expression manifest poor transcriptional rhythms, indicating a prominent role of post-transcriptional regulation in setting mRNA cycling amplitude. To gain further insight into circadian transcriptional regulation, we also characterized the rhythmic transcription of liver genes targeted by the transcription factors CLOCK and BMAL1; they directly target other core clock genes and sit at the top of the molecular circadian clock hierarchy in mammals. CLK:BMAL1 rhythmically bind at the same discrete phase of the circadian cycle to all target genes, which not surprisingly have a much higher percentage of rhythmic transcription than the genome as a whole. However, there is a surprisingly heterogeneous set of cycling transcription phases of direct target genes, which even include core clock genes. This indicates a disconnect between rhythmic DNA binding and the peak of transcription, which is likely due to other transcription factors that collaborate with CLK:BMAL1. In summary, the application of Nascent-Seq to a mammalian tissue provides surprising insights into the rhythmic control of gene expression and should have broad applications beyond the analysis of circadian rhythms.

overall_design: Mouse liver nascent RNA profile over 6 time points of the 24h light:dark cycle, in duplicate, sequenced using Ilumina GAII (Nascent-Seq); Mouse liver mRNA profile over 6 time points of the 24h light:dark cycle, in duplicate, sequenced using Ilumina HiSeq2000 (RNA-Seq); CLK and BMAL1 DNA binding profile in the mouse liver at ZT8, sequenced along an Input sample using GAII (ChIP-Seq); Mouse liver strand-specific nascent RNA profile over 6 time points of the 24h light:dark cycle, in duplicate, sequenced using Ilumina HiSeq2000 (Strand-specific Nascent-Seq);

first: Joe

last: Rodriguez

first: Michael

last: Rosbash


title: Nascent-Seq Reveals Novel Features of Mouse Circadian Transcriptional Regulation [ChIP-seq]

overall_design:
CLK and BMAL1 DNA binding profile in the mouse liver at ZT8, sequenced along an Input sample using GAII (ChIP-Seq)
Supplementary file ChIPSeq_Mouse_Liver_Processed_data_Table1.txt represents annotated CLK and BMAL1 peaks.


title: Characterization of the EZH2-MMSET histone methyltransferase regulatory axis in cancer

city: Atlanta

first: Meng

last: Zhao

state: GA

address: 1518 Clifton Rd, #369

department: BIOS

institute: Emory University

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

overall_design: Refer to individual Series

first: Irfan

last: Asangani


title: Dual Binding of Chromomethylase Domains to H3K9me2-containing Nucleosomes Mediates DNA Methylation in Plants

city: Dallas

first: Hume

last: Stroud

address: 5323 Harry Hines Blvd

institute: UT Southwestern

summary: DNA methylation and histone modification exert epigenetic control over gene expression. CHG methylation by CHROMOMETHYLASE3 (CMT3) depends on histone H3K9 dimethylation (H3K9me2), but the mechanism underlying this relationship is poorly understood. Here, we report multiple lines of evidence that CMT3 interacts with H3K9me2-containing nucleosomes. CMT3 genome locations nearly perfectly correlated with H3K9me2 and CMT3 stably associated with H3K9me2-containing nucleosomes. Crystal structures of maize CMT3 homologue, ZMET2, in complex with H3K9me2 peptides, showed that ZMET2 binds H3K9me2 via both BAH and chromo domains. The structures reveal an aromatic cage within both BAH and chromo domains as interaction interfaces that capture H3K9me2. Mutations that abolish either interaction disrupt CMT3 binding to nucleosomes, and show a complete loss of CMT3 activity in vivo. Our study establishes dual recognition of H3K9me2 marks by BAH and chromo domains, and reveals a novel mechanism of interplay between DNA methylation and histone modification.

overall_design: Investigation of genome-wide occupancy of CMT3 by ChIP-seq

first: Xuehua

last: Zhong


title: Transcriptome-wide miR-155 binding map reveals widespread non-canonical microRNA targeting [CLIP-seq data]

status: Public on Nov 09 2012

city: Chicago

first: Aly

middle: Azeem

last: Khan

state: IL

address: 6045 S. Kenwood Ave.

institute: University of Chicago

summary:
microRNAs (miRNAs) are essential components of gene regulation, but identification of miRNA targets remains a major challenge.  Most target prediction and discovery relies on perfect complementarity of the miRNA seed to the 3’ untranslated region (UTR).  However, it is unclear to what extent miRNAs target sites without seed matches.  Here, we performed a transcriptome-wide identification of the endogenous targets of a single miRNA—miR-155—in a genetically controlled manner.  We found that approximately forty percent of miR-155-dependent Argonaute binding occurs at sites without perfect seed matches.  The majority of these non-canonical sites feature extensive complementarity to the miRNA seed with one mismatch.  These non-canonical sites confer regulation of gene expression albeit less potently than canonical sites.  Thus, non-canonical miRNA binding sites are widespread, often contain seed-like motifs, and can regulate gene expression, generating a continuum of targeting and regulation.
Argonaute (AGO) HITS-CLIP Libraries generated from wild type and miR-155 knockout activated T cells.

overall_design: AGO HITS-CLIP libraries were generated from activated wild type and miR-155 knockout T cells with two different 3' linkers.  Libraries were generated and sequenced with an 11nt index read that contained both a 5nt multiplexing index and a 6nt degenerate barcode.  Files have been demultiplexed and the 6nt degenerate barcode has been appended as the first 6 nucleotides of the read.

first: Gabriel

middle: B

last: Loeb

first: David

last: Canner

first: Joseph

last: Hiatt

first: Jay

last: Shendure

first: Robert

last: Darnell

first: Christina

last: Leslie

first: Alexander

middle: Y

last: Rudensky


title: Digital gene expression (DGE) of CMV-infected tobacco leaves

city: Beijing

first: Jie

last: Lu

address: Shuangqiaozhonglu

country: China

institute: CAIQ

summary:
Here we performed a transcriptomic study on complete symptom development process of CMV-infected Nicotiana tabacum using Solexa/Illumina's high-throughput digital gene expression (DGE) system. 12 DGE libraries (from six virus-infected samples and six corresponding mock-inoculated samples) were constructed, and the gene expression variations between the virus-infected sample and the mock-inoculated sample in each symptom stage were compared. Thousands of differentially expressed genes were obtained by the comparison, and KEGG pathway analysis of these genes suggested that many biological processes (such as phtosynthesis, pigment metabolism and plant-pathogen interaction) were related to the symptom development. The authenticity of the DGE data was further confirmed by analyzing real-time RT-PCR using several random-selected genes.
We sequenced a cDNA library constructed from mixture of total RNA from six virus-infected samples and six mock-inoculated samples to get gene information for tobacco leaves in different symptom stages, including vein clearing, mosaic, severe chlorosis, partial recovery, total recovery and re-mosaic, and 95,916 Unigenes were  obtained

overall_design:
Tobacco leaves were inoculated by CMV-infected leaf homogenate and healthy leaf homogenate, respectivley. Six time points with different symptom stage were selected, and one virus-infect sample and one mock-inoculated sample were collected at each time. In order to average out variation of different plants, five leaves from five different plants were mixed to prepare every RNA sample. Twelve individual tag libraries of samples (six infected samples and six mock-inoculated samples) were constructed in parallel. For the gene expression analysis, the twelve samples were grouped into six groups, and each group contained a virus-infected sample and a mock-inoculated sample collected at the same time. In each group, the DGE data of virus-infected sample were compared to that of mock-inoculated sample to obtain the gene expression variations.
Illumina sequencing of transcripts from virus-infected and mock-inoculated samples to get gene information for tobacco leaves in different symptom stages. In order to get more gene information, the systemically infected leaves and mock-inoculated leaves were harvested at six time points and five leaves from five different plants were collected at each time point. The RNA-Seq analysis provided gene information for mock-inoculated and virus-infected tobacco leaves in different symptom stages.

first: Zhixin

last: Du

first: Yu

last: Pang

first: Lingna

last: Chen

first: Xiaohua

last: Meng

first: Jun

last: Kong

first: Shuifang

last: Zhu


title: Nascent-Seq Reveals Novel Features of Mouse Circadian Transcriptional Regulation

first: Jerome

last: Menet


title: DNA methylation changes are a late event in Acute Promyelocytic Leukemia and coincide with loss of transcription factor binding

status: Public on Nov 08 2012

city: Heidelberg

first: Christian

last: Rohde

address: Im Neuenheimer Feld 410

institute: Heidelberg University

type: Methylation profiling by high throughput sequencing

type: Methylation profiling by genome tiling array

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus


title: DNA methylation changes are a late event in Acute Promyelocytic Leukemia and coincide with loss of transcription factor binding (sequencing)

summary: The origin of aberrant DNA methylation in cancer remains largely unknown. In this study, we elucidated the DNA methylome in primary Acute Promyelocytic Leukemia (APL) and the role of PML-RARa in establishing these patterns. APL patients showed increased genome-wide DNA methylation with higher variability than healthy CD34+ cells, promyelocytes and remission bone marrow. A core set of differentially methylated regions in APL was identified.  Age at diagnosis, Sanz score and Flt3-mutation status characterized methylation subtypes. Transcription factor binding sites, e.g. c-myc binding sites were associated with low methylation. SUZ12 and REST binding sites identified in embryonic stem cells were, however, preferentially DNA hypermethylated in APL. Unexpectedly, PML-RARa binding sites were also protected from aberrant DNA methylation in APL.  In line, myeloid cells from pre-leukemic PML-RARa knock-in mice did not show altered DNA methylation and expression of PML-RARa in hematopoietic progenitor cells prevented differentiation without affecting DNA methylation. ATRA treatment of APL blasts did also not result in DNA methylation changes. These results suggest that aberrant DNA methylation is associated with leukemia phenotype but not required for PML-RARa-mediated initiation of leukemogenesis.

overall_design: We used Reduced Representation Bisulfite Sequencing (RRBS) to determine the genome-wide methylation signature of 18 primary APL patient samples. We then compared the APL methylation signature with methylation patterns found in CD34+ progenitor cells (n=4), promyelocytes (n=4) and remission bone marrow samples (n=8). Differentially methylated regions found in all three comparisons (APL vs. all three control specimens) were then further analyzed for genomic localization, variability and association with clinical parameters. Finally, the relationship between differentially methylated regions in APL and specific transcription factor binding sites was analyzed. For this purpose, ChiP-Sequencing of SUZ12 and REST was performed in primary APL patient blasts. To further determine the contribution of the leukemogenic transcription factor PML-RARa to methylation in APL, we also performed RRBS in pre-leukemic PML-RARa knock-in mice and hematopoetic progenitor cells retrovirally transduced with PML-RARa.

first: Till

last: Schoofs

first: Carsten

last: Müller-Tidow


title: Conserved DNA methylation patterns in healthy blood cells and extensive changes in leukemia measured by a new quantitative technique.

city: Philadelphia

first: Jaroslav

last: Jelinek

state: PA

address: 3307 North Broad St

department: Fels Institute

institute: Temple University

summary: We describe a simple method, Digital Restriction Enzyme Analysis of Methylation (DREAM), based on sequential DNA digestion with a pair of methylation-blocked and methylation-tolerant neoschizomeric restriction enzymes SmaI/XmaI followed by end repair and ultra-deep sequencing. DREAM provides information on 160,000 unique CpG sites of which 39,000 are in CpG islands, and 33,000 are at transcription start sites (-1 kb to +1 kb) of 13,139 RefSeq genes. We compared DNA methylation values in white blood cells from 4 healthy individuals and found them to be remarkably uniform. Interindividual differences >30% were observed only at 227 of 28,331 (0.8%) of autosomal CCCGGG sites covered by 100+ sequencing reads. Similarly, differences at only 59 sites were observed between the cord and adult blood. Conserved methylation patterns in healthy blood cells contrasted with extensive changes affecting 18-40% of CpG sites in leukemia. The method is cost effective, quantitative (r2=0.93 when compared to bisulfite pyrosequencing), reproducible (r2=0.997), and can detect differences >25% with false positive rate <0.001. Accurate analysis of changes in DNA methylation will be useful in quantifying epigenetic effects of environment and nutrition, correlating developmental epigenetic variation with phenotypes, understanding epigenetics of cancer and chronic diseases, measuring the effects of drugs on DNA methylation or deriving new biological insights into mammalian genomes.

overall_design: Digital restriction enzyme analysis of methylation (DREAM) was performed to determine the DNA methylation profiles of healthy white blood cells from cord blood and adult blood, acute myeloid leukemia bone marrow, and two leukemia cell lines (HEL and K562). In this approach, genomic DNA is sequentially cut at CCCGGG sites with the methylation-sensitive enzyme SmaI (blunt ends) and its methylation-tolerant neoschizomer XmaI (5'CCGG overhangs), creating different end sequences that represent methylation status of the CCCGGG sites. These end sequences are analyzed by next-generation sequencing, and thereafter the methylation status at individual CCCGGG sites across the genome can be determined.

first: Shoudan

last: Liang

first: Yue

first: Rong

last: He

first: Louis

last: Ramagli

first: Elizabeth

middle: J

last: Shpall

first: Marcos

middle: R

last: Estecio

first: Jean

last: Issa


title: Integrative analysis of the zinc finger transcription factor Lame duck in the Drosophila myogenic gene regulatory network

city: Bethesda

first: Brian

last: Busser

state: Md

address: 10 Center Drive

institute: NHLBI/NIH

summary: Contemporary high throughput technologies permit the rapid identification of transcription factor (TF) target genes on a genome-wide scale, yet the functional significance of TFs requires knowledge of target gene expression patterns, cooperating TFs and cis-regulatory element (CRE) structures.  Here we investigated the myogenic regulatory network downstream of the Drosophila zinc finger TF Lame duck (Lmd) by combining both previously published and newly performed genomic data sets, including chromatin immunoprecipitation sequencing (ChIP-seq), genome-wide mRNA profiling, cell-specific expression patterns of putative transcriptional targets, analysis of histone mark signatures, studies of TF co-occupancy by additional mesodermal regulators, TF binding site determination using protein binding microarrays (PBMs), and machine learning of candidate CRE motif compositions.  Our findings suggest that Lmd orchestrates an extensive myogenic regulatory network, a conclusion supported by the identification of Lmd-dependent genes, histone signatures of Lmd-bound genomic regions, and the relationship of these features to cell-specific gene expression patterns.  The heterogeneous co-occupancy of Lmd-bound regions with additional mesodermal regulators revealed that different transcriptional inputs are used to mediate similar myogenic gene expression patterns. Machine learning further demonstrated diverse combinatorial motif patterns within tissue-specific Lmd-bound regions.  PBM analysis established the complete spectrum of Lmd DNA binding specificities, and site-directed mutagenesis of Lmd and additional, newly discovered motifs in known enhancers demonstrated the critical role of these TF binding sites in supporting full enhancer activity.  Collectively, these findings provide new insights into the transcriptional codes regulating muscle gene expression, and offer a generalizable approach for similar studies in other systems.

overall_design: Examination of Lmd occupancy to genomic DNA from sorted mesodermal cells

middle: W

first: Di

last: Huang

first: Kevin

last: Rogacki

last: Lane

first: Leila

last: Shokri

first: Ting

last: Ni

first: Caitlin

last: Gamble

first: Stephen

last: Gisselbrecht

first: Martha

last: Bulyk

first: Ivan

last: Ovcharenko

first: Alan

last: Michelson


title: β-catenin regulates FSHβ induction by GnRH: next generation RNA-Sequencing identifies Brms1L as a mediator of beta-catenin regulation of FSHβ gene expression

city: New York

first: Maria

last: Chikina

address: Annenberg 14-88 One Gustave Levy Pl

institute: Mount Sinai School of Medicine

summary: The regulation of gonadotropin synthesis by GnRH (Gonadotropin-releasing hormone) plays an essential role in the neuroendocrine control of reproduction. The known signaling mechanisms involved in gonadotropin synthesis have been expanding.  For example, involvement of β-catenin in LHβ induction by GnRH has been discovered. We examined the role of β-catenin in FSHβ gene expression in LβT2 gonadotrope cells. GnRH caused a sustained increase in nuclear β-catenin levels, which was significantly reduced by JNK inhibition. siRNA-mediated knockdown of β-catenin mRNA demonstrated that induction of FSHβ mRNA by GnRH depended on β-catenin and that regulation of FSHβ by β-catenin occurred independently of the JNK-c-jun pathway. β-catenin depletion had no impact on FSHβ mRNA stability. In LβT2 cells transfected with FSHβ promoter luciferase fusion constructs, GnRH responsiveness was conferred by the proximal promoter (-944/-1), and was markedly decreased by β-catenin knockdown. However, none of the TCF/LEF binding sites in that region were required for promoter activation by GnRH. Chromatin immunoprecipitation further corroborated the absence of direct interaction between β-catenin and the 1.8 kb FSHβ promoter.  To elucidate the mechanism for the β-catenin effect, we analyzed ~1 billion reads of next generation RNA sequencing β-catenin knockdown assays and selected the nuclear cofactor Brms1L as one candidate for further study. Subsequent experiments confirmed that Brms1L mRNA expression was decreased by β-catenin knockdown as well as by JNK inhibition. Furthermore, knockdown of Brms1L significantly attenuated GnRH-induced FSHβ expression. Thus, our findings indicate that the expression of Brms1L depends on β-catenin activity and contributes to FSHβ induction by GnRH.

overall_design: A total of 24 samples were analyzed, namely 6 different experimental conditions, each comprised of 4 replicates. Control samples are included in the analysis. For the RNA-Seq assay, samples were multiplexed in the form of 3 samples per lanes, resulting in a total of 8 lanes.

first: Qian

last: Wang

first: Elena

last: Zaslavsky

first: Hanna

last: Pincas

first: Stuart

middle: C

last: Sealfon


title: Genome-wide comparative ChIP-seq data of CTCF and RNA-seq data in Drosophila white prepupa on Illumina Genome Analyzer

last: White

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

department: Institute for Genomics and Systems Biology

summary:
This is a dataset which comprises the following two different kinds of genomic data in Drosophila species:

First, triplicate ChIP-seq data of CTCF (CCCTC binding factor) binding profiles in each of the four closely related Drosophila species : Drosophila melanogaster, Drosophila simulans, Drosophila yakuba and Drosophila pseudoobscura at white pre pupa stage;
Second, triplicate RNA-seq data of white pre pupa whole animals of three Drosophila species: Drosophila melanogaster, Drosophila simulans and Drosophila yakub. The binding site/region/peaks are called using a modified method of QuEST( please see details in our related publication). The sequence read counts and RPKM values are calculated following the method in Mortazavi et al 2008 Nature Methods paper.

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

sample_organism:
Drosophila melanogaster
Drosophila pseudoobscura
Drosophila simulans
Drosophila yakuba

overall_design: Examination of CTCF binding in 4 Drosophila species and their correlation with gene expression levels in the same development stages

first: Xiaochun


title: Rice whole transcriptome differences at two temperature levels surveyed by RNA-Seq

status: Public on Nov 07 2012

first: Dong

address: No.1 West Beichen Road,Chaoyang District

department: Institute of Genetics and Developmental Biology

institute: Chinese Academy of Sciences and National Center for Plant Gene Research

summary: We report that TOG1 coordinates global changes of gene expression upon high temperature to support rice thermotolerant growth. We performed gene ontology (GO) enrichment analysis of the differentially expressed genes in rice seedlings grown at 25 ℃ and 30 ℃ and found that when temperature increases, wild-type dramatically adjusts the expression level of hundreds of genes in GO terms polysaccharide metabolic process, carbohydrate metabolic process, glucan metabolic process, carbon fixation, response to stimulus and organic substance metabolic process required for growth. However, in a tog1 mutant, the differentially expressed genes in response to the temperature increase significantly enriched in GO terms response to stimulus, response to oxidative stress and response to stress, whereas the coordination of primary metabolic processes and carbon fixation were largely impaired or undetectable.

overall_design: Differentially expressed mRNA profiles of 22-day-old wild type (WT) and tog1 rice upon a temperature difference were generated by deep sequencing

first: Yongbiao

last: Xue


title: A model system for assessing the ability of tag sequencing to detect genes specific for malignant B-cells

city: Aalborg

middle: Bro

last: Kloster

address: Sdr. Skovvej 15

department: Department of Haematology

country: Denmark

institute: Aalborg Hospital

summary:
The purpose of this study was to develop a quantification method that can be used to assess the ability of tag-seq to detect malignant B-cell transcripts.
The data support that tumour cell concentration is an important variable with fundamental impact on gene expression pattern.

overall_design: We analysed eight serial dilutions of the malignant B-cell line, OCI-Ly8, into the embryonic kidney cell line, HEK293, by tag-sequencing. No technical replicates were performed.

first: Anders

last: Bilgrau

last: Rodrigo-Domingo

first: Kim

last: Bergkvist

first: Alexander

last: Schmitz

first: Mads

last: Sønderkær

first: Julie

last: Bødker

first: Steffen

last: Falgreen

first: Mette

last: Nyegaard

first: Hans

last: Johnsen

first: Kåre

last: Nielsen

first: Karen

last: Dybkaer

first: Martin

last: Bøgsted


title: Genome wide binding sites of BAP1, HCF1 and OGT

city: South San Francisco

first: Jinfeng

last: Liu

address: 1 DNA Way, MS93

department: Bioinformatics and Computational Biology

institute: Genentech Inc.

summary: We report the genome wide binding sites of BAP1, HCF1 and OGT in bone marrow derived macrophages. De-ubiquitinating enzyme BAP1 is mutated in a hereditary cancer syndrome with increased risk of mesothelioma and uveal melanoma. Somatic BAP1 mutations occur in various malignancies. We show that mouse Bap1 gene deletion is lethal during embryogenesis, but systemic or hematopoietic-restricted deletion in adults recapitulates features of human myelodysplastic syndrome (MDS). Knockin mice expressing BAP1 with a 3xFlag tag revealed that BAP1 interacts with host cell factor–1 (HCF-1), O-linked N-acetylglucosamine transferase (OGT), and the polycomb group proteins ASXL1 and ASXL2 in vivo. OGT and HCF-1 levels were decreased by Bap1 deletion, indicating a critical role for BAP1 in stabilizing these epigenetic regulators. Human ASXL1 is mutated frequently in chronic myelomonocytic leukemia (CMML) so an ASXL/BAP1 complex may suppress CMML. A BAP1 catalytic mutation found in a MDS patient implies that BAP1 loss of function has similar consequences in mice and humans.

overall_design: For BAP1, bone marrow derived macrophages were used differentiated from bone marrow cells of BAP1-3X Flag Tagged KI mice we generated. For OGT and HCF1, bone marrow derived macrophages were used from BAP1 WT mice.

first: Anwesha

last: Dey


title: A model system for assessing the ability of exon microarray and tag sequencing to detect genes specific for malignant B-cells

first: Maria


title: Spark: A navigational paradigm for genomic data exploration

status: Public on Nov 06 2012

city: Davis

first: Henriette

last: O'Geen

address: 1 Shields Avenue

institute: UC Davis

summary: Biologists possess the detailed knowledge critical for extracting biological insight from genome-wide data resources and yet they are increasingly faced with non-trivial computational analysis challenges posed by genome-scale methodologies. To lower this computational barrier, particularly in the early data exploration phases, we have developed an interactive pattern discovery and visualization prototype, Spark, designed with epigenomic data in mind. Here we demonstrate Spark’s ability to reveal both known and novel epigenetic signatures using genome-wide histone modification, DNA methylation, and transcription factor data from human embryonic stem cells.

overall_design: Human embryonic stem cells (hESCs) were obtained from Cellular Dynamics as part of a large batch of cells prepared for the ENCODE Consortium and the RoadMap Epigenome Consortium. Cell growth and crosslinking conditions can be found at http://www.genome.ucsc.edu/ENCODE/cellTypes.html. ChIP-seq experiments for the histone modifications have been described previously (Harris et al. 2010). The YY1ChIP assay was performed using 5x107 cells following the protocol provided at http://farnham.genomecenter.ucdavis.edu/pdf/FarnhamLabChIP%20Protocol.pdf except that StaphA cells were blocked only with BSA before use and the pre-clearing step was omitted. ChIP and Input samples (10% of the amount of chromatin used per ChIP) were purified using the QIAquick PCR purification kit (QIAGEN) according to manufacturer’s instructions and purified eluates were dissolved in 50uls of water. ChIP libraries were created and sequenced as described previously (Harris et al. 2010)  by the DNA Technologies Core Facility at the University of California-Davis (http://genomecenter.ucdavis.edu/dna_technologies/). Illumina read sequences were aligned to the reference human genome (hg18) using BWA (Li and Durbin 2009). FindPeaks 4.0.15 (Fejes et al. 2008) was subsequently used to detect enrichment peaks at an FDR of 0.01.

first: Cydney

first: Peggy

last: Farnham


title: Whole transcriptome analysis of a reversible neurodegenerative process in Drosophila reveals potential neuroprotective genes

city: Derio

first: Rosa

last: Barrio

address: Bizkaia Technology Park

country: Spain

institute: CIC bioGUNE

summary: Background. Neurodegenerative diseases are progressive and irreversible and they can be originated by mutations in specific genes. Spalt-like genes (Sall) encode transcription factors expressed in the central nervous system. In humans, SALL mutations are associated with hereditary syndromes characterized by mental retardation, sensorineural deafness and motoneuron problems, among others. Drosophila sall mutants exhibit severe neurodegeneration of the Central Nervous System, which surprisingly reverts later in development, suggesting a potential to recover from neurodegeneration. We hypothesize that this recovery is mediated by a reorganization of the transcriptome counteracting SALL lost. To identify genes associated to neurodegeneration and neuroprotection, we used mRNA-Seq to compare the transcriptome of Drosophila sall mutant and wild type embryos from neurodegeneration and reversal stages. Results. Neurodegeneration stage is temporally associated with transcriptional changes in 220 genes, of which only 5% were already described as relevant for neurodegeneration. Genes related to the groups of Redox, Lifespan/Aging and Mitochondrial diseases are significantly represented at this stage. By contrast, neurodegeneration reversal stage is associated with significant changes in 480 genes, including 424 not previously associated with neuroprotection. Immune response and Salt stress are the most represented groups at this stage. We validated our result by comparison with data from modENCODE project and by Quantitative PCR. Conclusions. We identify new genes associated to neurodegeneration and neuroprotection by using an mRNA-Seq approach. The strong homology between Drosophila and human genes raises the possibility to unveil novel genes involved in neurodegeneration and neuroprotection also in humans. Key words: mRNA-Seq, spalt, SALL, Neurodegeneration, Neuroprotection, Drosophila

overall_design: A total of 14 samples, belonging to 6 stage/genotype groups, have been sequenced (Ho16 (3 replicates), He16 (2 replicates), WT16 (2 replicates), Ho17 (3 replicates), He17 (2 replicates) and WT17 (2 replicates)) following mRNAseq protocol (Illumina Inc.) at GAII.

first: María-José

last: Ferreiro

first: Naiara

last: Rodríguez-Ezpeleta

first: Coralia

last: Pérez

first: Michael

last: Hackenberg

first: Ana-María

last: Aransay

first: Rafael

last: Cantera


title: Mediator promotes CENP-A incorporation at fission yeast centromeres

city: Gothenburg

first: Claes

middle: M

last: Gustafsson

address: Medcinaregatan 9A

department: Department of Medical biochemistry and cell biology

country: Sweden

institute: Gothenburg University

type: Expression profiling by genome tiling array


title: Chromatin proteins linked to dosage compensation in Drosophila captured by ChIP-mass spectrometry

city: Boston

first: Peter

last: Kharchenko

address: 10 Shattuck St.

department: DBMI

institute: Harvard Medical School

summary: We report the usage of ChIP-mass spectrometry in identifying proteins and histone modifications involved in Drosophila dosage compensation. We identified a chromatin targeting factor, CG4747, that is involved in recognition of H3K36me3 and robust recruitment of the Drosophila MSL complex to its correct targets on the male X chromosome.

overall_design: ChIP-seq with PAP antibody of Drosophila larvae expressing C-terminally TAP-tagged CG4747.

first: Charlotte

last: Wang

first: Mitzi

last: Kuroda


title: Degradome analysis in Phalaenopsis aphrodite subsp. formosana

city: Tainan

first: Feng-Ming

last: An

address: No. 59, Silaya Blvd., Sinshi Dist.

department: Biotechnology Center in Southern Taiwan

country: Taiwan

institute: Academia Sinica

summary: We performed one degradome sequencing for identification and characterization of novel microRNAs in Phalaenopsis aphrodite subsp. formosana. Plant tissues of leaves, stalk, and flower buds frozen in liquid nitrogen were ground to fine powder using a mortar and pestle, respectively. Total RNA extraction was freshly prepared using Trizol (Invitrogen, Carlsbad, CA, USA) following the manufacturer's instructions. About 50 μg of leaves with or without low temperature treatment, stalks and flower buds were taken separately and then mixed. The degradome library was constructed by the Genomics BioSci & Tech. Company following the manufactoring protocol. In brief, 200 μg of total RNA was passed though polyA column using the Oligotex kit (Qiagen). The 5'-RNA adapter containing a Mme I recognition site was ligated by T4 RNA ligase and further reverse transcribed to amplify the templates. After digesting with Mme I then ligated to a 3'-double DNA adapter, the products were amplified by additional PCR cycles and gel-purified for Illumina sequencing. The files contain the raw data of degradome sequencing.

overall_design: Examination of one degradome of mixed tissues of Phalaenopsis orchid

first: Ming-Tsair

last: Chan


title: Histone H3K4 Demethylation is Negatively Regulated by Histone H3 Acetylation in S. cerevisiae

status: Public on Nov 05 2012

city: Vancouver

first: Benjamin

last: Martin

state: BC

address: 2350 Health Sciences Mall

department: Biochemistry

country: Canada

institute: UBC

summary: Histone H3 lysine 4 tri-methylation (H3K4me3) is a hallmark of transcription initiation, but how H3K4me3 is demethylated during gene repression is poorly understood.  Jhd2, a JmjC domain protein, was recently identified as the major H3K4me3 histone demethylase (HDM) in S. cerevisiae.  While JHD2 is required for removal of methylation upon gene repression, deletion of JHD2 does not result in increased levels of H3K4me3 in bulk histones, indicating that this HDM is unable to demethylate histones during steady state conditions.  In this study, we showed that this was due to the negative regulation of Jhd2 activity by histone H3 lysine 14 acetylation, which co-localizes with H3K4me3 across the yeast genome.  We demonstrated that loss of the histone H3-specific acetyltransferases (HATs) resulted in genome-wide-depletion of H3K4me3, and this was not due to a transcription defect.  Moreover, H3K4me3 levels were reestablished in HAT mutants following loss of JHD2, which suggested that H3-specific HATs and Jhd2 served opposing functions in regulating H3K4me3 levels.  We revealed the molecular basis for this suppression by demonstrating that histone H3K14 acetylation negatively regulated Jhd2 demethylase activity on an acetylated peptide in vitro.  These results revealed the existence of a general mechanism for removal of H3K4me3 following gene repression.

overall_design: Examination of H3K4me3 in WT, ada2sas3, ada2sas3jhd2, and jhd2 strains.

first: Julie

last: Brind'Amour

first: LeAnn

last: Howe


title: Histone acetylation regulates intracellular pH

status: Public on Nov 04 2012

city: Los Angeles

first: Siavash

middle: K

last: Kurdistani

address: 615 Charles E Young Dr South

department: Biological Chemistry

institute: UCLA

summary: Differences in global levels of histone acetylation occur in normal and cancer cells, although the reason cells regulate these levels has remained unclear. Here we demonstrate a role for histone acetylation in regulating intracellular pH (pHi). As pHi decreases, histones are globally deacetylated by histone deacetylases (HDACs) and the released acetate anions are co-exported with protons out of the cell by monocarboxylate transporters (MCTs), preventing further reductions in pHi. Conversely, global histone acetylation increases at more alkaline pHi, such as when resting cells are induced to proliferate. Inhibition of HDACs or MCTs decreases acetate export and lowers pHi, particularly compromising pHi maintenance in acidic environments. Global deacetylation at low pH is reflected at a genomic level by decreased abundance and extensive redistribution of acetylation at promoters and intergenic regions. Thus acetylation of chromatin functions as a rheostat to regulate pHi with important implications for therapeutic use of HDAC inhibitors.

overall_design: To investigate the redistribution of H4K16ac throughout the genome upon treatment at pH 6.5

first: Matthew

middle: A

last: McBrian


title: Influence of p38 MAPK (PMK-1) on the heat stress response of C. elegans

city: Münster

first: Rüdiger

last: Paul

address: Schlossplatz 8

department: Institut für Zoophysiologie

summary: PMK-1 is involved in the heat stress response of C. elegans, translocates to the nucleus upon heat exposure and influences the expression of chaperone genes, proteasomal subunits and protein-biosynthesis related genes.

overall_design: Differential Gene expression of WT and pmk-1 deletion mutant (KU25) after 5 hours at 35°C

first: Ansgar

last: Mertenskötter


title: Post-transcriptional regulation of cell-cell interaction protein-encoding transcripts by Zfs1p in S. pombe (RNA-Seq)

city: RTP

first: NIEHS

last: Microarray Core

state: NC

address: 111 T.W. Alexander Drive

department: DIR

institute: NIEHS

summary: Members of the tristetraprolin (TTP) family of CCCH tandem zinc finger proteins can bind directly to AU-rich elements in mRNAs and promote transcript deadenylation and decay.  The yeast Schizosaccharomyces pombe expresses a single TTP family member, Zfs1p, that has been linked to the mating response pathway and septum formation.  We showed previously that Zfs1p can bind to and promote the destabilization of AU-rich element-containing transcripts.  In this study, we identified additional target transcripts by comparing transcript levels in wild type and zfs1 mutant yeast, using deep sequencing and microarray approaches.  We also used direct RNA sequencing to determine the locations of the polyA tails in both wild type and mutant strains, and to confirm the presence of potential Zfs1p target sequences within the mRNA.  These studies identified a set of transcripts containing potential Zfs1p binding sites that accumulated significantly in the zfs1 mutants; a subset of these turned over more slowly in the zfs1 mutant strain, and bound directly to Zfs1p in co-immunoprecipitations.  One apparent direct target encodes the transcription factor Cbf12p, which is known to increase cell-cell adhesion and flocculation when over-expressed.  Studies of zfs1 and cbf12 double mutants demonstrated that the increased flocculation seen in zfs1 mutants is due, at least in part, to a direct effect on the turnover of cbf12 mRNA, leading in turn to changes in the levels of its transcriptionally regulated genes.  These data suggest that Zfs1p can both directly and indirectly regulate the levels of transcripts involved in cell-cell adhesion in this species.

overall_design: Strains were grown in YES media to approximately OD600=1.0 and total RNA was harvested from two biological replicates each of WT and zfs1 deletion.

first: Melissa

middle: L

last: Wells

first: Perry

last: Blackshear

first: Weichun

last: Huang

first: Leping

last: Li

first: Kevin

last: Gerrish

first: David

last: Fargo

first: Fatih

last: Ozsolak


title: Transcriptome-wide miR-155 binding map reveals widespread non-canonical microRNA targeting

city: Chicago

first: Aly

last: Khan

address: 6045 S. Kenwood Ave.

institute: University of Chicago

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

overall_design: Refer to individual Series


title: In vivo NCL-targeting affects breast cancer aggressiveness through miRNA regulation

status: Public on Nov 02 2012

city: Bellinzona

first: Luciano

last: Cascione

state: Ticino

address: Via Chiesa 5

department: Lymphoma & Genomics Research Program

country: Switzerland

institute: IOR


title: The Oscillating miRNA 959-964 cluster impacts Drosophila feeding time and other circadian outputs.

city: Waltham

first: Stefan

last: Pescatore

address: 415 South Street

department: Biology

institute: Brandeis University / HHMI

first: Sadanand

last: Vodala

first: Joseph

last: Rodriguez

first: Marita

last: Buescher

first: Ya-Wen

last: Chen

first: Ruifen

last: Weng

first: Stephen

last: Cohen

first: Michael

last: Rosbash


title: Integrative Epigenomic Analysis Identifies Biomarkers and Therapeutic Targets in Adult B-Acute Lymphoblastic Leukemia [ChIP-seq]

status: Public on Nov 27 2012

city: San Francisco

first: Huimin

last: Geng

address: 513 Parnassus Ave., MSB HSW 1201

department: Department of Laboratory Medicine

institute: UCSF

summary:
Integrative epigenomic analysis identifies biomarkers and therapeutic targets in adult B-acute lymphoblastic leukemia.

We performed DNA methylation (HELP array) and gene expression profiling in 215 samples of adult B-lineage acute lymphoblastic leukemia (ALL) and 12 normal preB samples. Adult B-lineage acute lymphoblastic leukemia (B-ALL) is an aggressive disease with <40% long-term survival. Genetic alterations such as BCR/ABL, E2A/PBX1 and MLL rearrangement (tMLL) define distinct B-ALL subtypes, which are associated with poor clinical outcome. It has been shown that these B-ALL subtypes have distinct expression profiles. However, the role of the epigenome in shaping these expression profiles and how the aberrant epigenetic gene regulation contributes to the biological and clinical features of those ALL subtypes is largely unknown. To address this question, we performed genome-wide DNA methylation and gene expression profiling on a large cohort of 215 well-characterized adult B-ALL specimens from the ECOG E2993 phase III clinical trial and a cohort of normal precursor B (preB) cells from 12 healthy bone marrows. The integrative analysis of these profiles led to the identification of key gene networks deregulated at the epigenetic and transcriptional levels within each subtype. In BCR/ABL, we identified a network centered on IL2RA(CD25), which is itself hypomethylated and overexpressed in most BCR/ABL B-ALL and confers poor clinical outcomes.  In the tMLL subtype, we uncovered aberrant epigenetic and transcriptional activities that include hypomethylation and upregulation of FLT3 and BCL6. After showing that MLL/AF4 fusion protein binds to these genes as well as other hypomethylated and overexpressed genes in tMLL ALL cells, we showed that a specific BCL6 inhibitor, RI-BPI, kills tumor cells in both tMLL ALL cell lines and patient samples. BCL6 inhibition may therefore represent a novel therapeutic strategy for B-ALL patients with MLL translocations.

RUNX1 is a key target gene in MLL-AF4 leukemias and contributes to gene activation by interacting with the AF4-MLL complex.

The Mixed Lineage Leukemia 1 protein (MLL1) is an important epigenetic protein that is required for the maintenance of gene activation during development, but is also mutated in a subset of aggressive human leukemias. The most common leukemogenic MLL1 mutations are chromosome translocations that fuse MLL1 in-frame to produce novel fusion proteins. Different MLL1 fusion proteins cause unique leukemias even when they are expressed in the same cell type, suggesting that they function through unique molecular mechanisms. We used ChIP-seq in MLL-AF4 patient cell lines to identify target genes that are involved in leukemogenesis.

overall_design: ChIP-seq using MLLN, AF4, H3K4me3 and H3K79me2 antibodies in RS4;11 cells.

first: Thomas

last: Milne

first: Ari

last: Melnick

first: Robert

middle: G

last: Roeder

first: C

middle: D

last: Allis


title: Genome-wide binding of WDR5 and GRHL3

city: Irvine

first: Amelia

middle: Soto

last: Hopkin

address: 250 Sprague Hall

department: Biological Chemistry & Medicine

institute: University of California Irvine

summary: The antagonistic actions of Polycomb and Trithorax are responsible for proper cell fate determination in mammalian tissues. In the epidermis, a self-renewing epithelium, previous work has shown that release from Polycomb repression only partially explains differentiation gene activation. We now show that Trithorax is also a key regulator of epidermal differentiation, not only through activation of genes repressed by Polycomb in progenitor cells, but also through activation of genes independent of regulation by Polycomb. The differentiation associated transcription factor GRHL3/GET1 recruits the ubiquitously expressed Trithorax complex to a subset of differentiation genes.

overall_design: Examination of WDR5 and GRHL3 binding in human differentited primary keratinocytes (NHEK). High calcium medium was added to NHEK cells at 50% confluency to induce differentiation. Cells were collected for ChIP 24 hours after addition of high calcium medium. ChIP with Wdr5 and Grhl3 antibodies, and an input control were sequenced.

first: Bogi

last: Andersen


title: Activating transcription factor 3 regulates immune and metabolic homeostasis

city: Cologne

first: Mirka

last: Uhlirova

address: Zülpicher Str. 47a

department: Institute For Genetics

institute: CECAD, University of Cologne

summary: Integration of metabolic, stress and immune responses plays a fundamental role during animal development to maintain energy homeostasis while ensuring growth and proper developmental timing. Perturbation of metabolic and immune signaling circuits has detrimental consequences to animal development including growth retardation, organ malfunction and emergence of the metabolic syndrome. Here, we demonstrate that the Drosophila basic region-leucine zipper (bZIP) protein, Activating transcription factor 3 (Atf3), safeguards a balance of metabolic and immune system responses during fly development. Loss of Atf3 function results in lethality during late-larval and pupal stages. Atf3-deficient larvae exhibit phenotypes resembling the metabolic syndrome in mammals. Excessive accumulation of lipids in the larval fat body and gut is accompanied by altered expression of genes involved in lipid metabolism. Moreover, the fat body of atf3 mutants becomes infiltrated by hemocytes. The major pro-inflammatory pathways signaling through JNK and Imd are hyperactivated in atf3 mutants, causing ectopic expression of antimicrobial peptide genes. Suppression of the immune response, achieved by reducing the gene dose of the transcription factors FOXO or NF-kappaB/Relish, significantly improves lipid metabolism and normalizes gene expression profile of atf3 mutants. In addition, heterozygosity of relish partially rescues lethality of the atf3 mutants. Our data thus identify Atf3 as an essential player that links metabolic and immune system homeostasis during animal development.

overall_design:
Examination of mRNA levels from four genotypes of male, 3rd instar Drosophila melanogaster larvae.  mRNA levels from four genotypes relative to y w control were determined using two biological replicates per genotype.
Genome build: BDGP R5/dm3, April 2006

first: Jan

last: Rynes

first: Colin

last: Donohoe

first: Peter

last: Frommolt

first: Susanne

last: Brodesser

first: Marek

last: Jindra


title: CDX2 ChIP-seq

first: Keyan

last: Salari

address: Gray/Bigelow Rm 1102, 55 Fruit Street

institute: Massachusetts General Hospital

summary: Somatic DNA alteration underlies tumor development and progression, and gives rise to tumors with diverse genetic contexts. Here, we identify in a collection of 29 colorectal cancer cell lines and 226 primary colorectal tumors recurrent amplification of chromosome 13, an alteration highly restricted to colorectal-derived cancers. A minimal region of amplification on 13q12.2 pinpoints caudal type homeobox transcription factor CDX2, a master regulator of anterior-posterior patterning, midgut development, and intestinal epithelial cell differentiation and maintenance. In contrast to its described role as a colorectal tumor suppressor, we show that in the context of genomic amplification, CDX2 is required for proliferation and anchorage-independent growth of colorectal cancer cells. By genome-wide expression and location analysis, we reveal that CDX2 directly promotes expression of Wnt pathway genes. Further results suggest that CDX2 induces expression of intestinal differentiation markers and modulates Î²-catenin transcriptional activity. These data characterize CDX2 as a novel lineage-survival oncogene deregulated in colorectal cancer.

overall_design: ChIP-seq analysis of CDX2 binding sites in COLO320 cells

first: Jonathan

last: Pollack


title: Rho and NusG suppress pervasive antisense transcription in Escherichia coli

status: Public on Nov 01 2012

city: Madison

first: Jason

middle: Matthew

last: Peters

state: WI

address: 1550 Linden Drive

institute: UW-Madison

type: Genome binding/occupancy profiling by genome tiling array

platform_organism: Escherichia coli str. K-12 substr. MG1655

last: Landick


title: Genome-wide mapping of FOXM1 binding sites

city: Manchester

first: Xi

last: Chen

address: Michael Smith Building, Dover Street

department: Faculty of Life Sciences

country: United Kingdom

institute: The University of Manchester

summary: There are nearly fifty forkhead (FOX) transcription factors encoded in the human genome and due to sharing a common DNA binding domain, they are all thought to bind to similar DNA sequences. It is therefore unclear how these transcription factors are targeted to specific chromatin regions to elicit specific biological effects. Here we used ChIP-seq to investigate the genome-wide chromatin binding mechanisms used by the forkhead transcription factor FOXM1. In keeping with its previous association with cell cycle control, we demonstrate that FOXM1 binds and regulates a group of genes which are mainly involved in controlling late cell cycle events in the G2 and M phases. However, rather than being recruited through canonical RYAAAYA forkhead binding motifs, FOXM1 binding is directed via CHR elements. FOXM1 binds these elements through protein-protein interactions with the MMB transcriptional activator complex. Thus, we have uncovered a novel and unexpected mode of chromatin binding of a FOX  transcription factor which allows it to specifically control cell cycle-dependent gene expression.

overall_design: Examination of FOXM1 binding sites in G2/M U2OS cells

first: Andrew

last: Sharrocks


title: Colorectal Cancer Cells


title: Differential transcript isoform usage pre- and post-zygotic genome activation in zebrafish

city: Oslo

first: Håvard

last: Aanes

state: Oslo

address: Ullevålsveien 72

country: Norway

institute: Norwegian School of Veterinary Science

summary: Zebrafish embryos are transcriptional silent until activation of the zygotic genome during the 10th cell cycle. Onset of transcription is followed by cellular and morphological changes involving cell speciation and gastrulation. Previous genome-wide surveys of transcriptional changes only assessed gene expression levels; however, recent studies have shown the necessity to map isoform-specific transcriptional changes. Here we perform isoform discovery and quantification on transcriptome sequences from before and after zebrafish zygotic genome activation (ZGA). We identify novel isoforms and isoform switches during ZGA for genes related to cell adhesion, pluripotency and DNA methylation. Isoform switching events include alternative splicing and changes in transcriptional start sites and in 3’ untranslated regions. New isoforms are identified even for well-characterized genes such as pou5f1, sall4 and dnmt1. Genes involved in cell-cell interactions such as f11r and magi1 display isoform switches with alterations of coding sequences. We also detect over 1000 transcripts that acquire a longer 3’ terminal exon when transcribed zygotically relative to the maternal transcript counterparts. ChIP-seq data mapped onto skipped exon events reveals a correlation between histone H3K36trimethylation peaks and the skipped exons, suggesting epigenetic marks being part of alternative splicing regulation. The novel isoforms and isoform switches reported here include regulators of the transcriptional, cellular and morphological changes taking place around ZGA. Our data display an array of isoform-related functional changes and represent a valuable resource complementary to existing early embryo transcriptomes.

overall_design: Examination H3K36me3 in zebrafish whole embryos at the Post-MBT stage

first: Philippe

last: Collas


title: Mechanistic and structural insight into the functional dichotomy between interleukin-2 and interleukin-15

city: Bethesda

first: Peng

last: Li

state: MD

address: 9000 Rockville Pike

department: NHLBI

institute: NIH

summary: Interleukin-15 (IL-15) and IL-2 possess distinct immunological functions despite both signaling through IL-2Rβ and the common cytokine receptor γ-chain, γc, We find that in the IL-15/IL-15Rα/IL-2Rβ/γc quaternary complex structure, IL-15 heterodimerizes IL-2Rβ and γc identically to the IL-2/IL-2Rα/IL-2Rβ/γc complex, despite differing receptor-binding chemistries. IL-15Rα dramatically increases the affinity of IL-15 for IL-2Rβ, and this allostery is required for IL-15 trans-signaling versus IL-2 cis-signaling.  Consistent with the identical IL-2Rβ/γc dimer geometry, IL-2 and IL-15 exhibited similar signaling properties in lymphocytes, with any differences resulting from disparate receptor affinities. Thus, IL-15 and IL-2 induce similar signals, and the cytokine-specificity of IL-2Rα versus IL-15Rα determines cellular responsiveness.  These results provide important new insights for specific development of IL-15- versus IL-2-based immunotherapeutics.

overall_design: RNA-Seq is conducted in mouse CD8+ T cells, not treated or treated with IL2 or IL15 for indicated concentrations (1nM or 500nM) and times (4hr or 24hr).

first: Aaron

last: Ring

first: Jian-Xin

last: Lin

first: Dan

last: Feng

first: Suman

last: Mitra

first: Mathias

last: Rickert

first: Gregory

last: Bowman

first: Vijay

last: Pande

first: Ignacio

last: Moraga

first: Rosanne

last: Spolski

first: Engin

last: Özkan

first: Warren

last: Leonard

last: Garcia


title: The AHL- and BDSF-dependent quorum sensing systems control specific and overlapping sets of genes in Burkholderia cenocepacia H111

city: Zürich

first: Gabriella

last: Pessi

address: Winterthurerstrasse 190

institute: University of Zürich

summary: Quorum sensing in Burkholderia cenocepacia H111 involves two signalling systems that depend on different signal molecules, namely N-acyl homoserine lactones (AHLs) and the diffusible signal factor cis-2-dodecenoic acid (BDSF). Previous studies have shown that AHLs and BDSF control similar phenotypic traits, including biofilm formation, proteolytic activity and pathogenicity. In this study we mapped the BDSF stimulon by RNA-Seq and shotgun proteomics analysis. We demonstrate that a set of the identified BDSF-regulated genes or proteins are also controlled by AHLs, suggesting that the two regulons partially overlap. The detailed analysis of two mutually regulated operons, one encoding three lectins and the other one encoding the large surface protein BapA and its type I secretion machinery, revealed that both AHLs and BDSF are required for full expression, suggesting that the two signalling systems operate in parallel. In accordance with this, we show that both AHLs and BDSF are required for biofilm formation and protease production.

overall_design: Burkholderia cenocepacia BDSF stimulon was determined in complex media using Illumina RNA-Seq

first: Nadine

last: Schmid

first: Claudio

last: Aguilar

first: Yinyue

last: Deng

first: Aurelien

last: Carlier

first: Alexander

last: Grunau

first: Ulrich

last: Omasits

first: Lian-Hui

last: Zhang

first: Christian

middle: H

last: Ahrens

first: Leo

last: Eberl


title: The Oscillating miRNA 959-964 cluster impacts Drosophila feeding time and other circadian outputs [miRNA-seq].

first: Stefan

last: Pescatore

summary: Using high throughput sequencing of Drosophila head RNA, a small set of miRNAs that undergo robust circadian oscillations in levels were discovered. We concentrated on a cluster of six miRNAs, mir-959-964, all of which peak at about ZT12 or lights-off. The data indicate that the cluster pri-miRNA is transcribed under bona fide circadian transcriptional control and that all 6 mature miRNAs have short half-lives, a requirement for oscillating. Manipulation of food intake dramatically affects the levels and timing of cluster miRNA transcription with no more than minor effects on the core circadian oscillator. This indicates that the central clock regulates feeding, which in turn regulates proper levels and cycling of the cluster miRNAs. Viable Gal4 knock-in as well as cluster knock-out and over-expression strains were used to localize cluster miRNA expression as well as explore their functions. The adult head fat body is a major site of expression, and feeding behavior, innate immunity, metabolism, and perhaps stress responses are under cluster miRNA regulation. The feeding behavior results indicate that there is a feedback circuit between feeding time and cluster miRNA function as well as a surprising role of post-transcriptional regulation in these behaviors and physiology.

overall_design: Six samples of small RNA libraries (RNA size 19 to 29 nucleotides long) were prepared from Drosophila heads, each collected at one circadian time point during a light-dark cycle (ZT0, ZT4, ZT8, ZT12, ZT16, ZT20).

first: Sadanand

last: Vodala

first: Joseph

last: Rodriguez

first: Marita

last: Buescher

first: Ya-Wen

last: Chen

first: Ruifen

last: Weng

first: Stephen

last: Cohen

first: Michael

last: Rosbash


title: small RNA sequencing of the M.truncatula rdr6.1 mutant

city: Castanet Tolosan

first: Erika

last: Sallet

address: 24 Chemin de Borde Rouge, CS 52627

department: Plant Health and the Environment

institute: INRA/CNRS

summary: The rdr6.1 missense allele was found by TILLING in EMS mutagenized plants  of Medicago truncatula. To analyse the impact of this point mutation on small RNA populations, we sequenced small RNA from two sibling lines carrying the mutation and the corresponding wild-type plants.

overall_design: libraries from two sibling lines carrying the rdr6.1 mutation and the corresponding wild-type plants

first: Christine

last: Lelandais-Brière

first: Pilar

last: Bustos

first: Martin

last: Crespi

first: Jérome

last: Gouzy


title: Acute Stress Induced Increase in Histone H3 Lys 9 Trimetylation in Rat Hippocampus

city: Boston

first: Richard

middle: G.

last: Hunter

state: Massachusetts

address: 100 William Morrissey Blvd.

department: Psychology

institute: University of Massachusetts Bo

summary: We utilized ChIP Seq to examine the the genomic localization of the increase in H3K9 trimethylation we had previously observed in the hippocampus as a consequence of acute restraint stress in rats (Hunter et. al PNAS 2009).

overall_design: Rats were restrained for 30 minutes, allowed to recover for 1 hour and then sacrificed and their hippocampi extracted and processed for H3K9me3 ChIP using the Cell Signalling H3K9me3 antibody.

first: Donald

last: Pfaff

first: Bruce

middle: S

last: McEwen


title: Transcriptome profiling of wild type primordial germ cells (PGC) by RNAseq

status: Public on Oct 31 2012

city: La Jolla

first: Kun

last: Zhang

address: 9500 Gilman Dr Mailcode: 0412

department: Bioengineering

institute: University of California, San Diego

summary:
Purpose: To identify molecular pathways underlying epigenetic reprogramming in early germ cell precursors, we examined global gene expression of wild type primordial germ cells using mRNA sequencing.
Methods: Given the limited number of PGCs collected from E9.5 to E13.5 (ranging from 300 to 5000), we used a low-input RNA sequencing method, Smart-Seq. RNA libraries were pooled and sequenced by Illumina Hiseq.
Results: We generated >22 million uniquely mapped reads per sample and identified >10 thousand transcripts per genotype (RPKM>0.1). Hierarchical clustering and correlation analysis on gene expression indicates samples were clearly separated according to their genotypes with Spearman correlation coefficient of 0.98/0.99 within biological replicates. Compared with E9.5 PGCs, 479 genes were significantly up-regulated and 248 genes were down-regulated in E11.5 PGCs. When compared with E11.5 PGCs, male E13.5 PGCs had 362 up-regulated, and 239 down-regulated genes, whereas female E13.5 PGCs had 1163 up-regulated and 333 down-regulated genes. Overall, the number of up-regulated genes was greater than that of the down-regulated genes in every comparison, suggesting that gene expression is generally activated during PGC reprogramming.

overall_design: mRNA profiles of primordial germ cells derived from developmental embryos stages (E9.5, E11.5 and E13.5) were generated by deep sequencing, in  duplicates (E9.5 and E11.5) or triplicates (E13.5f and E13.5m), using Illumina Hiseq.


title: Mediator promotes CENP-A incorporation at fission yeast centromeres [ChIP-seq]

city: Gothenburg

first: Claes

last: Gustafsson

address: Medcinaregatan 9A

department: Department of Medical biochemistry and cell biology

institute: Gothenburg University

summary: At Schizosaccharomyces pombe centromeres, heterochromatin formation is required for de novo incorporation of the histone H3 variant CENP-A/Cnp1, which in turn directs kinetochore assembly and ultimately chromosome segregation during mitosis. Noncoding RNAs (ncRNAs) transcribed by RNA polymerase II (Pol II) directs heterochromatin formation via the RNAi machinery, but also through RNAiindependent RNA processing factors. Control of centromeric ncRNA transcription is therefore a key factor for proper centromere function. We here use transcriptional profiling, gene inactivation experiments, and chromatin immunoprecipitation analyses to demonstrate that the Mediator complex directs ncRNA transcription and regulates centromeric heterochromatin formation in fission yeast. Mediator co-localizes with Pol II at centromeres and loss of the Mediator subunit Med20 causes a dramatic increase in pericentromeric transcription and desilencing of the core centromere. As a consequence, heterochromatin formation is impaired both via the RNAi dependent and independent pathways, resulting in loss of CENP-A/Cnp1 from the core centromere, defect kinetochore function, and a severe chromosome segregation defect. Interestingly, the increased centromeric transcription observed in med20Δ appears to directly block CENP-A/Cnp1 incorporation and inhibition of Pol II transcription can suppress the observed phenotypes. Our data thus identify Mediator as a crucial regulator of ncRNA transcription at fission yeast centromeres and add another crucial layer of regulation to centromere function.

overall_design: 3 samples examined: wild type chromatin incubated with beads as the non antibody control, wild type chromatin incubated with RNA Polymerase II CTD domain antibody and Protein G beads,  and TAP-Med7 cells chromatin incubated with IgG  beads.

first: Jonas

middle: O

last: Carlsten

first: Xuefeng

last: Zhu

first: Marcela

last: Davila Lopez


title: The expression analyses of metafemales compared with normal females

city: Columbia

first: Lin

last: Sun

state: MO

address: 311 Tucker Hall

department: Biological Sciences

institute: University of Missouri

summary: Comapare the global expression of metafemales and normal femalems

overall_design: Collected the metafemales, females and males and made RNA-seq

first: James

last: Birchler


title: Natural variation in processability of individual Arabidopsis thaliana microRNAs

city: Tuebingen

first: Felix

last: Ott

address: Spemannstr. 35-39

institute: Max Planck Institute for Developmental Biology

summary: Like protein coding genes, loci that produce microRNAs (miRNAs) are generally considered to be under purifying selection, consistent with miRNA polymorphisms being able to cause disease. Nevertheless, it has been hypothesized that variation in miRNA genes may contribute to phenotypic diversity. Here we demonstrate that a naturally occurring polymorphism in the MIR164A gene interacts epistatically with an unlinked locus to affect leaf shape and shoot architecture in Arabidopsis thaliana. A single-base pair substitution in the miRNA complementary sequence alters the stability of the miRNA:miRNA* duplex. It thereby interferes with processing of the precursor and greatly reduces miRNA accumulation. We demonstrate that this is not a rare exception, but that natural strains of Arabidopsis thaliana harbor dozens of similar polymorphisms that affect processing of a wide range of miRNA precursors. Our results suggest that natural variation in miRNA processability due to cis mutations is a common contributor to phenotypic variation in plants.

overall_design: sRNA sequencing transgenic A. thaliana

first: Marco

last: Todesco

first: Sureshkumar

last: Balasubramanian

first: Jun

last: Cao

first: Sridevi

last: Sureshkumar

first: Korbinian

last: Schneeberger

first: Rhonda

last: Meyer

first: Thomas

last: Altmann

first: Detlef

last: Weigel


title: Efficient direct reprogramming of c-Kit− mature amniotic cells into endothelial cells by ETS factors and TGFβ suppression

status: Public on Oct 30 2012

city: New York

first: Olivier

last: Elemento

state: NY

address: 1305 York Avenue

institute: WEILL MEDICAL COLLEGE OF CORNELL UNIV

summary: ETS transcription factors ETV2, FLI1 and ERG1 specify pluripotent stem cells into endothelial cells (PSC-ECs). However, these PSC-ECs are unstable and often drift towards non-vascular cell fates. We show that human mid-gestation c-Kit- lineage-committed amniotic cells (ACs) can be reprogrammed into induced vascular endothelial cells (rAC-VECs). Transient ETV2 expression in ACs generated immature iVECs, while co-expression with FLI1/ERG1 endowed rAC-VECs with a vascular repertoire and morphology matching mature ECs. Brief TGFb-inhibition functionalizes VEGFR2 signaling, augmenting specification of ACs into rAC-VECs. Genome-wide transcriptional analyses showed that rAC-VECs are similar to adult ECs in which vascular-specific genes are expressed and non-vascular genes are silenced. Functionally, rAC-VECs form stable vasculature in Matrigel plugs and regenerating livers. Thus, short-term ETV2 expression and TGFb-inhibition along with constitutive ERG1/FLI1 co-expression reprogram mature ACs into generic rAC-VECs with clinical-scale expansion potential. Public banking of HLA-typed rAC-VECs would establish a vascular inventory for treatment of genetically diverse disorders.

overall_design: Transcriptome sequencing of clonal and non-clonal rAC-VECs, HUVECs, LSECs, CD34+/Lin-, BMS

first: Michael

last: Ginsberg

first: Shahin

last: Rafii


title: Correlated alterations in genome organization, histone methylation, and DNA-lamina interactions in Hutchinson-Gilford progeria syndrome

city: Worcester

first: Rachel

middle: Patton

last: McCord

state: MA

address: 364 Plantation St. LRB 570M

department: Program in Gene Function and Expression

institute: University of Massachusetts Medical School

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

overall_design: Refer to individual Series


title: Cleavage Factor Im as a key regulator of 3’ UTR length

city: Basel

first: Andreas

middle: R

last: Gruber

address: Klingelbergstrasse 50-70

department: Biozentrum

institute: University of Basel

summary: In eukaryotes, the 3' ends of RNA polymerase II-generated transcripts are made in the majority of cases by site-specific endonucleolytic cleavage, followed by the addition of a poly(A) tail. By alternative polyadenylation, a gene can give rise to multiple mRNA isoforms that differ in the length of their 3' UTRs and hence in their susceptibility to post-transcriptional regulatory factors such as microRNAs. A series of recently conducted high-throughput studies of poly(A) site usage revealed an extensive tissue-specific control of 3’ UTR length and drastic changes in 3’ UTR length of mRNAs upon induction of proliferation in resting cells. To understand the dynamics of polyadenylation site usage, we recently identified binding sites of the major pre-mRNA 3’ end processing factors - cleavage and polyadenylation specificity factor (CPSF), cleavage stimulation factor (CstF), and cleavage factor Im (CF Im) - and mapped cleaved polyadenylation sites in HEK293 cells. Our present study extends previous findings on the role of CF Im in alternative polyadenylation and reveals that subunits of the CF Im complex generally control 3’ UTR length. More specifically, we demonstrate that the  loss-of-function of CF Im68 and CF Im25 but not of CF Im59 leads to a transcriptome-wide increase of the use of proximal polyadenylation sites.

overall_design: 3' ends of transcripts were profiled by high-throughput sequencing in HEK 293 cells under normal conditions, and in HEK 293 cells depleted of 3' end processing factors CF Im25, CF Im59, and CF Im68.

first: Georges

last: Martin

first: Walter

last: Keller

first: Mihaela

last: Zavolan


title: Correlated alterations in genome organization, histone methylation, and DNA-lamina interactions in Hutchinson-Gilford progeria syndrome (ChIP-seq)

summary: Hutchinson-Gilford progeria syndrome (HGPS) is a premature aging disease that is frequently caused by a de novo point mutation at position 1824 in LMNA. This mutation activates a cryptic splice donor site in exon 11, and leads to an in-frame deletion within the prelamin A mRNA and the production of a dominant negative lamin A protein, known as progerin. Here we show that HGPS cells experience genome-wide alterations in patterns of H3K27me3 deposition, changes in the associations of genomic loci with nuclear lamin A/C, and, at late passages, genome-wide loss of spatial compartmentalization of active and inactive chromatin domains that characterizes chromosome folding in normal cells. We further demonstrate that the H3K27me3 changes associate with gene expression alterations in HGPS cells. Our results support a model that the accumulation of progerin in the nuclear lamina leads to altered H3K27me3 marks in heterochromatin, possibly through the down-regulation of EZH2, and disrupts heterochromatin-lamina interactions. These changes may then lead to the genomic disorganization and changes in transcriptional regulation we observe in HGPS fibroblasts.

overall_design: ChIP-Seq was performed for H3K27me3 and lamin A/C in the human genome in three primary fibroblast cell lines: an HGPS patient (HGPS), normal cells from the father of the HGPS patient (Father), and age-matched normal cells (Age Control). Two biological replicates were performed.

middle: P

first: Ashley

last: Nazario-Toole

first: Haoyue

last: Zhang

first: Peter

last: Chines

first: Ye

last: Zhan

last: Erdos

first: Francis

last: Collins

first: Job

last: Dekker

first: Kan


title: Hog1 bypasses stress-mediated down-regulation of transcription by PolII redistribution and chromatin remodeling

city: Barcelona

first: Francesc

last: Posas

address: Dr. Aiguader

institute: UPF

summary: Cells are subjected to dramatic changes on gene expression upon environmental changes. Stress causes a general down-regulation of gene expression together with the induction of a set of stress-responsive genes. Genome wide localisation studies showed major changes on Pol II localisation towards stress-responsive genes in contrast to housekeeping genes. Pol II relocalisation requires of the Hog1 SAPK, which also associates at stress-responsive loci. Chromatin structure was not significantly altered upon stress except for those genes that displayed Hog1 association. Together, Hog1 serves to bypass the general down-regulation of gene expression by targeting RNA Pol II machinery and inducing chromatin remodeling at stress-responsive loci.

overall_design: Hog1 and Pol II ChIP-Seq and Mnase-Seq experiments in both strains WT and Hog1 mutant, for 2 conditions: Exposed and not exposed to osmostress

first: Nadal-Ribelles

last: Mariona

first: Posas

last: Francesc


title: Correlated alterations in genome organization, histone methylation, and DNA-lamina interactions in Hutchinson-Gilford progeria syndrome (Hi-C)

overall_design: Hi-C was performed on three primary fibroblast cell lines: an HGPS patient (passage 17 and 19; HGPS), normal cells from the father of the HGPS patient (passage 18; Father), and age-matched normal cells (passage 20; Age Control).

first: Michael

last: Cao


title: Comparison of gene expression profiles in olfactory epithelium from Atf5+/+ and Atf5-/- mice

middle: R.

last: Green

address: 364 Plantation Street

department: Molecular, Cell and Cancer Biology

summary:
We have found that the expression pattern of Atf5 is highly restricted to the olfactory system, strongly suggesting that Atf5 is an olfactory sensory neuron-specific transcription factor. To test this possiblity, we compared
gene expression profiles in olfactory epithelium (septa and turbinates) from Atf5+/+ and Atf5-/- mice.

overall_design: Olfactory septa and turbinates were dissected from Atf5+/+ and Atf5-/- P0 pups from different litters (n=3 for each genotype) and pooled into a single sample. Three independent samples were prepared for each genotype.

first: Shu-Zong

last: Wang


title: Rho and NusG suppress pervasive antisense transcription in Escherichia coli [RNA-seq].

first: Jason

last: Peters

department: Biochemistry

summary: Despite the prevalence of antisense transcripts in bacterial transcriptomes, little is known about how their synthesis is controlled. We report that a major function of the Escherichia coli termination factor Rho and its co-factor NusG is suppression of ubiquitous antisense transcription genome-wide. Rho binds C-rich unstructured nascent RNA (high C/G ratio) prior to its ATP-dependent dissociation of transcription complexes. NusG is required for efficient termination at minority subsets (~20%) of both antisense and sense Rho-dependent terminators with lower C/G ratio sequences. In contrast, a widely studied nusA deletion proposed to compromise Rho-dependent termination had no effect on antisense or sense Rho-dependent terminators in vivo. Global co-localization of the nucleoid-associated protein H-NS with Rho-dependent terminators and genetic interactions between hns and rho suggest that H-NS aids Rho in suppression of antisense transcription. The combined actions of Rho, NusG, and H-NS appear to be analogous to the Sen1-Nrd1-Nab3 and nucleosome systems that suppress antisense transcription in eukaryotes.

overall_design: RNA-seq experiments were performed on ribosome-depleted RNA from cells treated with 20 ug/ml bicyclomycin or untreated cells. The series contains 4 datasets.

first: Robert

last: Landick


title: rem-1 analysis in C. elegans

city: Groningen

first: Eugene

last: Berezikov

address: Antonius Deusinglaan 1

department: European Research Institute for the Biology of Ageing

institute: University Medical Center Groningen

summary: small RNA libraries from total RNA isolated from young adult animals

overall_design: Wild-type and rem-1 mutant animals were used for RNA isolation. Regular libraries were made using adaptor ligations at both ends. In addition, librraies were made from oxidised and TAP treated RNA.

first: Rene

middle: F

last: Ketting


title: Analysis of soybean DNA methylomes shows CHH hypermethylation enhances gene expression in cotyledons of developing seeds [methylome]

city: beijing

first: Jinsong

address: No.1 West Beichen Road,Chaoyang District

institute: IGDB

summary: Cytosine methylation is an important mechanism for dynamical regulation of gene expression and transposon mobility during plant developmental processes. Recently, the variation of DNA methylation has been described between wild type and DNA methylation-related mutants in Arabidopsis thaliana. However, the elaborate representation of soybean DNA methylomes remains lacking. Here, we described the epigenome maps of soybean root, stem, leaf, and cotyledon of developing seed at a single-base resolution. We confirmed the transcription start sites of genes using high-throughput sequencing and reported the DNA methylation patterns in gene and transposon regions. The correlation between gene expression and DNA methylation was revealed through transcriptome sequencing. We found CHH methylation may function in promotion of gene expression and ten cotyledon-preferred genes were identified CHH hypermethylated in cotyledon. Small RNA library sequencing showed that DNA methylation was enhanced by small RNAs not by strand-specific way, and the variation of DNA methylation between the organs was highly related with expression of small RNAs.

overall_design: methylomes of roots, stems, leaves, and cotyledons of developing seeds

first: Qing-Xin

last: Song

first: Jin-Song


title: Methylome Analysis of FF HPV+ and HPV- HNSCC samples

city: London

first: Gareth

last: Wilson

address: 23 Huntley Street

institute: UCL Cancer Institute

summary: The analysis of the 6 methylomes by MeDIP-Seq comprise 3 HPV+ HNSCC samples and 3 HPV- HNSCC samples

overall_design: Analysis of the 6 methylomes, comprising 3 HPV+ HNSCC samples and 3 HPV- HNSCC samples

first: Matthias

last: Lechner

first: Tim

last: Fenton

first: James

last: West

first: Andrew

last: Feber

first: Christina

last: Thirlwell

first: Harpreet

last: Dibra

first: Amrita

last: Jay

first: Lee

last: Butcher

first: Fiona

last: Gratrix

first: Nirali

last: Patel

last: Vaz

first: Paul

last: O'Flynn

first: Nicholas

last: Kalavrezos

middle: E

last: Teschendorff

first: Chris

last: Boshoff

first: Stephan

last: Beck


title: Regulation of Puf2 Expression is Critical for Plasmodium Sporozoite Development and Maintenance of Sporozoite Transmissibility

status: Public on Oct 27 2012

city: Seattle

first: Wonjong

last: Moon

state: WA

address: 307 Westlake Avenue North, Suite 500

institute: Seattle BioMed

summary: Malaria’s cycle of infection requires parasite transmission between a mosquito vector and a vertebrate host. Plasmodium regulates transmission by translationally repressing specific mRNAs until their products are needed. We demonstrate that the Plasmodium yoelii Pumilio-FBF family member Puf2 allows the sporozoite to retain its infectivity in the mosquito salivary glands while awaiting transmission. Puf2 mediates this critical feature solely through its RNA-Binding Domain (RBD) likely by protecting and silencing specific mRNAs. Puf2 storage granules are distinct from stress granules and P-bodies and dissolve rapidly after infection of hepatocytes, likely releasing the protected and silenced transcripts for ‘just-in-time’ translation by early exoerythrocytic forms (EEFs). Further corroborating this model, pypuf2- sporozoites have no apparent defect in host infection early after invading the salivary glands, but become progressively noninfectious and subsequently prematurely transform into EEFs during prolonged salivary gland residence. In contrast, the premature overexpression of Puf2 in oocysts causes striking deregulation of sporozoite maturation, resulting in fewer oocyst sporozoites that are non-infectious and unable to colonize the salivary glands. Maintenance of Puf2 expression in liver stage parasites produces no phenotype, suggesting that a window of permissive expression exists.  Finally, by conducting the first comparative RNAseq analysis of Plasmodium sporozoites, we have uncovered that Puf2 may play a role in both the protection of specific transcripts as well as RNA turnover via the CCR4/Not complex. These findings uncover requirements for maintaining a window of opportunity for the malaria parasite to accommodate the unpredictable moment of transmission from mosquito to vertebrate host.

overall_design: Wild-type (Py17XNL) and pypuf2 -salivary gland sporozoites

first: Scott

last: Lindner

first: Sebastian

last: Mikolajczak

first: Ashley

middle: M

last: Vaughan

first: Brad

last: Joyce

first: William

middle: J

last: Sullivan

first: Stefan

last: Kappe