title: Differential gene expression in hypothalamus of non-recombinant subcongenic-derived F2 mice fed macronutrient selection diets

city: Baton Rouge

first: Brenda

last: Richards

state: LA

address: 6400 Perkins Rd.

department: Basic Science

institute: Pennington Biomedical Research Center

summary: The specific genes influencing the quantitative variation in macronutrient preference and food intake are virtually unknown.  We refined a previously identified mouse chromosome 17 (MMU17) region harboring quantitative trait loci (QTL) with large effects on preferential macronutrient intake-carbohydrate (Mnic1), total kilcalories (Kcal2), and total food volume (Tfv1) using interval-specific congenic strains.  These loci were isolated in the [C57BL/6J.CAST/EiJ-17.1-(D17Mit19-D17Mit50); B6.CAST-17.1] strain, developed by introgressing a ~40.1 Mb CAST MMU17 region into recipient B6 genome.  In a diet selection paradigm (carbohydrate/protein vs. fat/protein), these B6.CAST-17.1 sub-congenic mice eat 30% more calories from the carbohydrate-rich diet, ~10% more total calories, and ~9% more total food volume per body weight.  In the current study, this carbohydrate-preferring B6.CAST-17.1 subcongenic strain was crossed with the fat-preferring inbred B6 strain to generate a subcongenic-derived F2 mapping population; genotypes were determined using a high-density, custom SNP panel.  The main outcome of this study is that genetic linkage analysis greatly reduced the 95% confidence interval (CI) for Mnic1 (encompassing Kcal2 and Tfv1) from 40.1 to 29.5 Mb and more precisely established the QTL boundaries.  Specifically, the genetic architecture for Mnic1 (preferential carbohydrate intake) does not follow the same pattern as that for co-localized Kcal2/Tfv1 (total kcal and food volume, respectively), suggesting the presence of separate quantitative trait genes for these food intake traits.  No genetic linkage for self-selected fat intake was detected, underscoring the carbohydrate-specific effects of this MMU17 locus.  The Mnic1/Kcal2/Tfv1 QTL was further de-limited to a ~19.1 Mb interval, based on the absence of macronutrient diet selection phenotypes in subcongenic HQ17IIa mice that possess CAST MMU17 donor segment on a C57BL/6Jhg/hg background.  A second key finding is the separation of two energy balance QTLs:  Mnic1/Kcal2/Tfv1 for food intake and a newly discovered locus regulating short term body weight gain.  The genes Decr2, Ppard and Agapt1 in the critical QTL interval were identified and prioritized using a combination of genome sequence analysis, and tag-based transcriptome sequencing to measure hypothalamic gene expression in non-recombinant F2 controls, possessing cast/cast and b6/b6 genotypes across the sub-congenic segment.

overall_design: Global gene expression profiles in the hypothalamus were compared between non-recombinant subcongenic-derived F2 mice, possessing a genotype of cast/cast (n=12) or b6/b6 (n=12) across the Chr 17 subcongenic segment and b6/b6 across the rest of the genome.  RNA was isolated from individual mice of each genotype that were selected for study.  Specifically, twelve mice of each genotype that displayed the most divergent phenotypic values for self-selected carbohydrate and total kcal intake were chosen for gene expression analysis, i.e., cast/cast mice with the highest and b6/b6 mice with the lowest kcal intake from carbohydrate, or the 25% tails of the distribution.  Tissues were harvested ~48 h after re-initiation of the carbohydrate/protein vs. fat/protein diets, following an extended wash-out period on chow diet after completion of the 10 d macronutrient selection test.

first: Rodrigo

last: Gularte-Merida

first: Lisa

middle: M

last: DiCarlo

first: Ginger

last: Robertson

first: Jacob

last: Simon

first: William

middle: D

last: Johnson

first: Claudia

last: Kappen

first: Juan

last: Medrano

middle: K


title: A novel phenotypic drug screen identifies potent inhibitors of mycobacterial virulence protein secretion

status: Public on May 15 2015

city: Bern

first: Andrej

last: Benjak

address: Murtenstrasse 24-28

department: Department for BioMedical Research

country: Switzerland

institute: University of Bern

type: Expression profiling by high throughput sequencing

summary: Mycobacterium tuberculosis (Mtb) depends on protein secretion systems for intracellular survival and virulence. The major virulence determinant and ESX-1 dependent effector protein EsxA causes tissue damage and necrosis which promotes spread and dissemination of the bacterium. We developed a fibroblast survival assay (FSA) that exploits this phenotype by selecting for molecules that protect host cells from Mtb-induced lysis. Hit compounds identified in this high-throughput screen blocked the secretion of EsxA thus promoting phagosome maturation which led to substantially reduced bacterial burden in activated macrophages. Target identification studies performed on these drugs led to discovery of a benzothiophene containing histidine kinase inhibitor and a benzyloxybenzylidine hydrazine compound affecting mycobacterial metal ion homeostasis which enabled us to reveal zinc stress as a signal for EsxA secretion. Collectively, this novel drug screening approach can help to tackle the mounting problem of antibiotic-resistant mycobacteria by largely extending the target spectrum of small molecule libraries.

overall_design: RNA-Seq: M. tuberculosis H37Rv (GC1237) cultures grown in vitro to exponential phase and treated with 5 µM of compound or DMSO

first: Jan

last: Rybniker

last: Sala

first: Stewart

middle: T

last: Cole


title: Brg1 activates enhancer repertoires to establish B cell identity and modulate cell growth

city: Freiburg

last: Bossen

address: Breisacher Strasse 115

department: Center for Chronic Immunodeficiency

country: Germany

institute: University Hospital Freiburg (D)

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Early B cell development is orchestrated by the combined activities of the transcriptional regulators E2A, EBF1, Foxo1 and Ikaros. However, how the genome-wide binding patterns of these regulators are modulated during B-lineage development remains to be determined. Here, we found that in lymphoid progenitors the chromatin remodeler Brg1 specified the B cell fate. In committed pro-B cells Brg1 regulated Igh locus contraction and controlled c-Myc expression to modulate the expression of genes that regulate ribosome biogenesis. In committed pro-B cells Brg1 also suppressed a pre-B lineage-specific pattern of gene expression. Finally, we found that Brg1 acted mechanistically to establish B cell fate and modulate cell growth by facilitating access of lineage-specific transcription factors to poised enhancer repertoires.

overall_design: 8 ATAC-Seq samples from sorted ALP and BLP (duplicates, control and Brg1-deleted), 4 ATAC-Seq samples from cultured pro-B cells (duplicates, control and Brg1-deleted), 2 Ikaros ChIP-seq samples (performed in Rag1-/- pro-B cells and in E2A-/- pre-pro-B cells), 1 Brg1 ChIP-seq sample and accompanying Input sample (both in Rag1-/- pro-B cells), 4 RNA-Seq samples from cultured pro-B cells (duplicates, control and Brg1-deleted), 6 RNA-Seq samples from cultured Rag1-/- pro-B cells (triplicates, control and Brg1-knock down).

first: Caroline

middle: S

last: Murre

first: Aaron

middle: N

last: Chang

first: Robert

last: Mansson

first: Hans-Reimer

last: Rodewald

first: Cornelis


title: Argonaute 2 binds directly to tRNA genes and promotes gene repression in cis

status: Public on May 17 2015

city: Birmingham

first: keith

middle: e

last: giles

state: AL

address: 1825 University Blvd/ SHEL 706

department: Biochemistry & Molecular Genetics/ Stem Cell Institute

country: USA

institute: University of Alabama at Birmingham

summary: To further our understanding of the RNAi machinery within the human nucleus, we analyzed the chromatin and RNA binding of Argonaute 2 (AGO2) within human cancer cell lines. Our data indicated that AGO2 binds directly to nascent tRNA and 5S rRNA, and to the genomic loci from which these RNAs are transcribed, in a small RNA- and DICER-independent manner. AGO2 chromatin binding was not observed at non-TFIIIC-dependent RNA polymerase (Pol) III genes or at extra-TFIIIC (ETC) sites, indicating that the interaction is specific for TFIIIC-dependent Pol III genes. A genome-wide analysis indicated that loss of AGO2 caused a global increase in the mRNA expression level among genes that flank AGO2-bound tRNA genes. This effect was shown to be distinct from that of the disruption of DICER, DROSHA, or CTCF. We propose that AGO2 binding to tRNA genes has a novel and important regulatory role in human cells.

overall_design: ChIP-seq for AGO2 was performed from K562 cells using 2 commercially available monoclonal antibodies (mAbs) and 5 replicates.  Replicates 1-3 were performed with Millipore 04-642, and replicates 4 and 5 were performed with Abcam 57113.  RNA-seq was carried out on 2 replicates of shMock and 2 replicates of shAGO2.  Lentiviral vectors GIPZ Lentiviral #RHS4531-EG271611.

first: J

middle: L

last: Woolnough

first: K

middle: E

last: Giles


title: Regulation of histone methylation and gene expression reprogramming in rice inflorescence meristem

status: Public on May 14 2015

city: Wuhan

first: shaoli

last: zhou

state: Hubei

address: shizishan street

department: National Key Laboratory of Crop Genetic Improvement

country: China

institute: Huazhong Agricultural University

summary: Rice inflorescence meristem (IM) development is critical for panicle size and grain production. The transition from SAM to IM is an important development stage in rice. Here we report the application of ChIP-seq technology for profiling of H3K4me3 and H3K27me3 modification in SAM, IM, SDG711 RNAi IM and jmj703 IM which affect panicle size, respectively,  and besides, RNA-seq technology for detecting gene expression diversity betwwen SAM, IM and SDG711 RNAi IM. We found that genome-wide H3K27me3 modification vary between SAM and IM, and thouands of genes lost H3K27me3 in SDG711 and jmj703. Also, we found H3K27me3/H3K4me3 ratio was an important factor for the differential expression of many genes during the transition. Differential expressed genes were found between SAM and IM, 711RNAi and WT, repectively. Many important genes involved in  IM activity were repressed by SDG711-mediated H3K27me3 and some of which’s expressions were mis-regulated in SDG711 RNAi.

overall_design: Genome-wide mapping of two histone modifications (H3K4me3 and H3K27me3) in SAM, IM, SDG711 RNAi IM and jmj703 IM, and mRNA transcripts in SAM, IM, SDG711 RNAi IM.

first: X

last: Liu

first: S

last: Zhou

first: D


title: The zebrafish embryo as an alternative model for the evaluation of liver toxicity using histopathology and expression profiling

city: Bitlhoven

first: Marja

last: Driessen

state: Utrecht

address: Antonie van Leeuwenhoeklaan 9

department: Laboratory for Health Protection Research

country: Netherlands

institute: National Institute of Public Health and the Environment (RIVM)

summary: The zebrafish is as a powerful vertebrate model system for modeling human disease including liver pathology. In ZFE, hepatic responses can be expected after exposure to hepatotoxicants, because hepatocytes are present from 36-hpf and at 72-hpf the liver is fully functioning. These characteristics make the whole ZFE an attractive alternative model for compound-induced hepatotoxicity screening. Therefore, the main objective of this study is to further strengthen the applicability of whole ZFE as an alternative model for hepatotoxicity testing, with a special focus on the ability to identify gene expression responses in whole ZFE that are suggestive of hepatotoxicity. Deep sequence technology is applied to assess whether hepatotoxicity-specific transcripts that are identified in livers of hepatotoxicant treated adult zebrafish can be detected in whole ZFE as well.

overall_design: Whole zebrafish embryo and adult zebrafish are exposed for 48 hours to human hepatotoxicants. Afterwards, the adult liver is dissected and snap frozen untill processed for RNA isolation. The whole zebrafish embryos are pooled, one sample consists of 15 embryos, and are also snap frozen untill processed for RNA isolation. For every sample RNA is isolated according to the protocol of de Jong et al (2009). and then equal amounts of every sample is pooled. The two pools (one for the adult zebrafish liver, the other for the whole zebrafish embryo) are sequenced.

first: Anne

last: Kienhuis

first: Jeroen

middle: A

last: Pennings

first: Tessa

last: Pronk

first: Evert-Jan

last: van den Brandhof

first: Marianne

last: Roodbergen

first: Herman

middle: P

last: Spaink

first: Bob

last: van de Water

first: Leo

last: van der Ven


title: Genome-wide analysis of H3.3 dissociation in mouse embryonic stem cells

city: Bethesda

first: Misook

last: Ha

state: MD

address: NIH

institute: National Heart Lung Blood Institute

summary: Using Tet-inhibited expression of epitope-tagged H3.3 combined with ChIP-Seq we undertook genome-wide measurements of H3.3 dissociation rates across the ESC genome and examined the relationship between H3.3-nucleosome turnover and ESC-specific transcription factors, chromatin modifiers and epigenetic marks.

overall_design: To measure dissociation rates of H3.3, we utilized a TET-repressible ESC line, ES[MC1R(20)], with the expression cassette integrated at the ROSA26 locus. We transfected MC1R ESCs with HA/FLAG-tagged H3.3 controlled by tetracycline response elements. For ‘TET-OFF’ experiments, ESCs were cultured over several passages (weeks) on feeder cells in the absence of DOX and were subsequently passaged onto feeder-free plates prior to the inhibition of HA-H3.3 expression. To repress HA/FLAG-H3.3 expression we treated cells with 2 μg/ml doxycycline hyclate before crosslinking with formaldehyde at various time points. Measurement of H3.3-HA enrichment over time-course was performed using two replicates of ChIP-seq experiments. As a control, input DNA sequencing was performed for every time point

first: Daniel

last: Kraushaar

first: Keji

last: Zhao


title: Identification and characterization of cold-responsive microRNAs in tea plant (Camellia sinensis) and their targets using high-throughput sequencing and degradome analysis [miRNA-seq]

city: Nanjing

first: Yue

last: Zhang

state: Jiangsu

address: Weigang No.1

department: Tea Science Research Institute

institute: Nanjing Agricultural University

type: Non-coding RNA profiling by high throughput sequencing

summary: We discovered 31 up-regulated miRNAs and 43 down-regulated miRNAs in ‘Yingshuang’, and 46 up-regulated miRNA and 45 down-regulated miRNAs in ‘Baiye 1’ in response to cold stress, respectively. A total of 763 related target genes were detected by degradome sequencing. The RLM-5’RACE procedure was successfully used to map the cleavage sites in six target genes of C. sinensis. These findings reveal important information about the regulatory mechanism of miRNAs in C. sinensis, and promote the understanding of miRNA functions during the cold response. The miRNA genotype-specific expression model might explain the distinct cold sensitivities between tea lines.

overall_design: Solexa sequencing technology was used for high-throughput sequencing of the small RNA library from the cold treatment of tea leaves. To align the sequencing data with known plant miRNAs, we characterized 106 conserved C. sinensis miRNAs. In addition, 215 potential candidate miRNAs were found, among, which 98 candidates with star sequences were chosen as novel miRNAs. Both congruously and differentially regulated miRNAs were obtained, and cultivar-specific miRNAs were identified by microarray-based hybridization in response to cold stress. The results were also confirmed by quantitative real-time polymerase chain reaction. To confirm the targets of miRNAs, two degradome libraries from two treatments were constructed. According to degradome sequencing, 455 and 591 genes were identified as cleavage targets of miRNAs from cold treatments and control libraries, respectively, and 283 targets were present in both libraries. Functional analysis of these miRNA targets indicated their involvement in important activities, such as development, regulation of transcription, and stress response.


title: Transcriptomic analysis reveals novel long non-coding RNAs critical for vertebrate development [RNA-Seq]

city: La Jolla

first: Christopher

last: Benner

state: California

address: 9500 Gilman Dr. MC 0640

department: Medicine

institute: University of California, San Diego (UCSD)

summary: Long non-coding RNAs (lncRNAs) have emerged as critical regulators of gene expression and chromatin modifications, with important functions in development and disease. Here we sought to identify and functionally characterize lncRNAs critical for vascular vertebrate development with significant conservation across species. Genome-wide transcriptomic analyses during human vascular lineage specification enabled the identification of three conserved novel lncRNAs: TERMINATOR, ALIEN and PUNISHER that are specifically expressed in pluripotent stem cells, mesoderm and endothelial cells, respectively. Gene expression profiling, alongside RNA immunoprecipitation coupled to mass spectrometry, revealed a wide range of new molecular networks and protein interactors related to post-transcriptional modifications for all three lncRNAs. Functional experiments in zebrafish and murine embryos, as well as differentiating human cells, confirmed a developmental-stage specific role for each lncRNA during vertebrate development. The identification and functional characterization of these three novel non-coding provide a comprehensive transcriptomic roadmap and shed new light on the molecular mechanisms underlying human vascular development.

overall_design: Time course RNA-Seq analysis H1 ESCs differentiated into endothelial cells

last: Kurian

first: Aitor

last: Aguirre

first: Juan

middle: C

last: Izpisua-Belmonte


title: Molecular consequences of Dlx3 deletion in mouse enamel organ as determined by RNASeq analysis

first: Hong-wei

last: Sun

address: 9000 Rockville Pile

department: Office of Science & Technology

institute: NIAMS

summary:
DLX3 is a homeodomain transcription factor involved in ameloblast differentiation and enamel formation. Mutations in DLX3 in human lead to defects in enamel.
However, the downstream targets of Dlx3 transcriptional activity in the enamel organ have not been identified yet.
In this study, we compared the transcriptome of enamel organs where Dlx3 has been deleted to control tissues.

overall_design: Total RNA was extracted from enamel organs (mandibular incisors) from Dlx3-WT (N=4) and Dlx3-K14cKO (N=4) mice at P10. cDNA libraries were generated using NEBnext and NuGen kits. Sequencing was performed on the HiSeq2000.

first: Olivier

last: Duverger

first: Takahiro

last: Ohara

first: Maria

last: Morasso


title: Understanding strategy of nitrate and urea assimilation in a Chinese strain of Aureococcus anophagefferens through RNA-seq analysis

city: Guangzhou

first: Hongpo

last: Dong

address: West of Huangpu Road 601

department: Research Center for Harmful Algae and Marine Biology

institute: Jinan University

summary:
Purpose: RNA-seq technology was used to profile the transcriptome of a Chinese strain of A. anophagefferens that was grown on urea, nitrate, and a mixture of urea and nitrate, and that was under N-replete, limited and recovery conditions to understand the molecular mechanisms that underlie nitrate and urea utilization.
Methods: mRNA profiles of Aureococcus anophagefferens that was grown on urea, nitrate, and a mixture of urea and nitrate, and that was under N-replete, limited and recovery conditions were generated by deep sequencing using Illumina HiSeq 2000. The sequence reads that passed quality filters were analyzed by aligning to reference genome and transcript using SOAPaligner/SOAP2 (version 2.21).
Results: Deconvolution and filtering of raw reads yielded a mean of 6,938,798 reads (range: 6,539,842 to 7,242,083 reads) per individual RNA-seq library (Table 1). Subsequent alignment of the clean reads to the A. anophagefferens reference genome yielded a mean of 5,852,408 reads (84.3%) for each sample that mapped to at least one location in the A. anophagefferens genome. However, of these mapped reads, only 47.4 to 50.9% of the total reads were mapped to the reference transcript for each sample (Table 2).  In A. anophagefferens grown on three different N sources, 9148 to 9526 genes were detected for each sample. A comparison of gene expression among the three N sources was performed. 322 differentially expressed genes were detected between nitrate-grown and urea-grown cells, 237 between nitrate-grown and mixture N-grown cells and 29 between urea-grown and mixture N-grown cells. Fewer differentially expressed genes between urea-grown and mixture N-grown cells were identified, which further suggested the preferred utilization of cells for urea in media with mixture N. For nitrogen-limited and recovery experiments, 707 genes were up-regulated significantly, and 766 were down-regulated significantly in N-depleted cells relative to N-replete cells. N-depleted cells exhibited a broad transcriptional response to nitrogen re-addition, with 681 genes up-regulated and 874 genes down-regulated in urea recovery cells, and 312 genes up-regulated and 688 genes down-regulated in nitrate recovery cells.
Conclusions:  We noted that transcripts upregulated by nitrate and N-limitation included those encoding proteins involved in amino acid, nucleotide and aminosugar transport, degradation of amides and cyanates, and nitrate assimilation pathway. The data suggest that A. anophagefferens possesses an ability to utilize a variety of dissolved organic nitrogen. Moreover, transcripts for synthesis of proteins, glutamate-derived amino acids, spermines and sterols were upregulated by urea. Transcripts encoding key enzymes that are involved in the ornithine-urea and TCA cycles were differentially regulated by urea and nitrogen concentration, which suggests that the OUC may be linked to the TCA cycle and involved in reallocation of intracellular carbon and nitrogen. These genes regulated by urea may be crucial for the rapid proliferation of A. anophagefferens when urea is provided as the N source.

overall_design: Seven mRNA samples for Aureococcus anophagefferens were sequencd using Illumina HiSeqTM 2000, including three samples that was grown on urea, nitrate, and a mixture of urea and nitrate, and four samples that was under N-replete, limited and recovery conditions

first: Songhui

last: Lu


title: Shear Stress and NOTCH1 Regulate Calcification Related Genes in Human Aortic Valve Endothelium (RNA-Seq)

status: Public on May 16 2015

city: San Francisco

first: Deepak

last: Srivastava

state: CA

address: 1650 Owens St.

department: Gladstone Institute of Cardiovascular Disease

institute: J. David Gladstone Institutes

summary: We are interested in the role of NOTCH1 and Shear Stress in Aortic Valve Endothelium.

overall_design: Primary human aortic valve endothelium was subjected to 4 conditions in vitro. 1) Control siRNA, No shear stress. 2) NOTCH1 siRNA, No shear stress. 3) Control siRNA, 15 dynes/cm2 shear stress. 4) NOTCH1 siRNA, 15 dynes/cm2 shear stress. Triplicates of each condition were pooled for library perp and sequencing

first: Mark

last: White


title: Proteins that bind regulatory regions identified by histone modification chromatin immunoprecipitations and mass spectrometry

city: Rotterdam

first: Raymond

last: Poot

address: Wytemaweg 80

department: Cell Biology

institute: Erasmus MC

summary: The locations of transcriptional enhancers and promoters were recently mapped in many mammalian cell types. Proteins  that bind those regulatory regions can determine cell identity but have not been systematically identified. Here we purify native enhancers, promoters or heterochromatin from embryonic stem cells by chromatin immunoprecipitations (ChIP) for characteristic histone modifications and identify associated proteins using mass spectrometry (MS). 239 factors are identified and predicted to bind enhancers or promoters with different levels of activity, or heterochromatin. Published genome-wide data indicate a high accuracy of location prediction by ChIP-MS. A quarter of the identified factors are important for pluripotency and includes Oct4, Esrrb, Klf5, Mycn and Dppa2, factors that drive reprogramming to pluripotent stem cells. We determined the genome-wide binding sites of Dppa2 and find that Dppa2 operates outside the classical pluripotency network. Our ChIP-MS method provides a detailed read-out of the transcriptional landscape representative of the investigated cell type.

overall_design: Examination of Dppa2 binding in V5-Dppa2 CGR8 mouse ESCs. H3K4me1, H3K4me3, H3K27ac and H3K9me3 ChIP-seq experiments were conducted using the modified ChIP protocol that was adapted for Mass spectrometry.

first: Johannes

middle: H

last: Brandsma

first: Erik

last: Engelen

first: Christel

last: Kockx

first: Zehila

last: Ozgür

first: Wilfred

middle: J

last: van IJcken


title: Bile diversion to the distal small intestine has comparable metabolic benefits to bariatric surgery

status: Public on May 13 2015

city: Nashville

first: Yan

last: Guo

state: TN

address: 639 Preston Research Bld

institute: Vanderbilt Cancer Center

summary: Roux-en-Y gastric bypass (RYGB) is highly effective in reversing obesity and associated diabetes. Recent observations in humans suggest a contributing role of increased circulating bile acids in mediating such effects. Here we use a diet-induced obesity mouse model and compared metabolic remission when bile flow was diverted through a gallbladder anastomosis to jejunum, ileum or duodenum (sham control). We found that only bile diversion to the ileum results in physiologic changes similar to RYGB including sustained improvements in weight, glucose tolerance and hepatic steatosis despite differential effects on hepatic gene expression.  Circulating free fatty acids and triglycerides decrease while bile acids increase, particularly conjugated tauro-b-muricholic acid, an FXR antagonist. Activity of the hepatic FXR/FGF15 axis was reduced and associated with altered gut microbiota. Thus bile diversion, independent of surgical rearrangement of the gastrointestinal tract, imparts significant weight loss accompanied by improved glucose and lipid homeostasis that are hallmarks of RYGB.

overall_design: Total RNA from n = 5 DIO, n = 4 GB-IL, n = 5 RYGB mice livers was extracted of total RNA and submitted fro RNAseq

first: Charles

last: RFlynn

first: Vance

last: Albaugh

first: Steven

last: Cai

first: Joyce

last: Cheung-Flynn

first: Phillip

last: Williams

first: Robert

last: Brucker

first: Seth

middle: R

last: Bordenstein

first: David

last: Wasserman

first: Naji

last: Abumrad


title: Transcriptomic analysis reveals novel long non-coding RNAs critical for vertebrate development

first: Christopher

last: Benner

type: Methylation profiling by genome tiling array

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

overall_design: Refer to individual Series


title: Genome-wide activities of Polycomb complexes

city: Boston

first: Hun-Goo

last: Lee

state: MA

address: 185 Cambridge St

department: Molecular Biology

institute: Mass General Hospital

type: Genome binding/occupancy profiling by genome tiling array


title: mCasz1_conditional knockout

city: Raleigh

first: Nirav

last: Amin

state: NC

address: 1060 William Moore Drive

department: Molecular Biomedical Sciences

institute: North Carolina State University

summary: The overall goal of our studies is to elucidate the cellular and molecular mechanism by which the transcription factor Casz1 functions in murine heart development. We established that Casz1 is expressed in myocardial progenitor cells beginning at E7.5 and in differentiated cardiomyocytes throughout development. We generated conditional Casz1 knockout mice to show that ablation of CASZ1 in Nkx2.5-positive cardiomyocytes leads to cardiac hypoplasia, ventricular septal defects and lethality by E13.5. To identify the pathways and networks by which Casz1 regulates cardiomyocyte development, we used RNA-Seq and identified genes involved in cell proliferation are upregulated in Casz1 mutants compared to wild-type littermates. We conclude that Casz1 is essential for cardiac development and has a pivotal role in regulating part of the cardiac transcriptional program.

overall_design: 3 biological replicates of the two genotypes (Nkx2-5+/+,Casz1f/+ and Nkx2-5Cre/+,Casz1f/f) were used for RNA-seq to determine genes that are differentially expressed in the murine heart when Casz1 is mutated. Nkx2-5+/+,Casz1f/+  were used as wild-type controls for comparison.

first: Kerry

last: Dorr

first: Frank

last: Conlon


title: Genome-wide analysis of epigenetic marks in E(z) temperature sensitive cells [ChIP-seq]

summary: PcG protein complex PRC2 is a methyltransferase specific for histone H3 lysine27, and H3K27me3 is essential for stable transcription silencing. Less well known but quantitatively much more important is the genome-wide role of PRC2 that dimethylates ~70% of total H3K27. Here we show that H3K27me2 occurs in inverse proportion to transcriptional activity in genes and intergenic regions and its loss results in global transcriptional derepression proportionally greatest in previously silent or weakly transcribed regions. H3K27me2 levels are controlled by opposing roaming activities of PRC2 and the H3K27 demethylase dUTX. Unexpectedly, we find an equally pervasive distribution of histone H2A ubiquitylated at lysine 118 (H2AK118ub), attributed to the RING1 subunit of PRC1-type complexes.

overall_design: Examination of global changes in five epigenetic marks when E(z) is inactivated in E(z) temperature-sensitive cells at 25°C and 31°C. Genome-wide binding of Polycomb was also examined to classify canonical PcG target regions.

middle: G

first: Amanda

last: Simcox

first: Yuri

middle: B

last: Schwartz

first: Vincenzo

last: Pirrotta


title: Genome-wide analysis of histone marks in E(z) temperature sensitive cells [ChIP-seq]

overall_design: Examination of global changes in five epigenetic marks when E(z) is inactivated in E(z) temperature-sensitive cells at 25°C and 31°C.


title: Reprogrammed myeloid cell transcriptomes in NSCLC

city: New York

first: Dingcheng

last: Gao

state: NY

address: 1300 York Ave

department: CT Surgery

institute: Weill Cornell Medical College

summary: Lung cancer is the leading cause of cancer related mortality worldwide, with non-small cell lung cancer (NSCLC) as the most prevalent form.  Despite advances in treatment options including minimally invasive surgery, CT-guided radiation, novel chemotherapeutic regimens, and targeted therapeutics, prognosis remains dismal. Therefore, further molecular analysis of NSCLC is necessary to identify novel molecular targets that impact prognosis and the design of new-targeted therapies. In recent years, tumor “activated/reprogrammed” stromal cells that promote carcinogenesis have emerged as potential therapeutic targets. However, the contribution of stromal cells to NSCLC is poorly understood. Here, we show increased numbers of bone marrow (BM)-derived hematopoietic cells in the tumor parenchyma of NSCLC patients compared with matched adjacent non-neoplastic lung tissue. By sorting specific cellular fractions from lung cancer patients, we compared the transcriptomes of intratumoral myeloid compartments within the tumor bed with their counterparts within adjacent non-neoplastic tissue from NSCLC patients. The RNA sequencing of specific myeloid compartments (immature monocytic myeloid cells and polymorphonuclear neutrophils) identified differentially regulated genes and mRNA isoforms, which were inconspicuous in whole tumor analysis. Genes encoding secreted factors, including osteopontin (OPN), chemokine (C-C motif) ligand 7 (CCL7) and thrombospondin 1 (TSP1) were identified, which enhanced tumorigenic properties of lung cancer cells indicative of their potential as targets for therapy. This study demonstrates that analysis of homogeneous stromal populations isolated directly from fresh clinical specimens can detect important stromal genes of therapeutic value.

overall_design: We sorted pure populations of the immature monocytic myeloid cells (IMMCs), neutrophils (Neu), and epithelial cells (Epi) from tumors and adjacent lung tissues of stage I-III lung adenocarcinoma patients. RNA samples (totally 17 samples) were sequenced: from tumor IMMC (n=3), Neu (n=2), Epi (n=2); from adjacent lung IMMC (n=3), Neu (n=4), Epi (n=3).

first: Anna

last: Durrans

first: Vivek

last: Mittal


title: Identification and characterization of conserved microRNAs in pepper by high-throughput sequencing

city: SEOUL

first: Chanseok

last: Shin

address: Gwanak-gu Daehak-dong

country: South Korea

institute: Seoul National University

summary: High-throughput sequencing was employed to identify conserved microRNAs in pepper.

overall_design: 1 sample examined: flower and root in pepper

first: Dong-Gyu

last: Hwang

first: June-Hyun

last: Park

first: Jae-Yun

last: Lim


title: Comparative haploid genetic screens reveal divergent pathways in the biogenesis and trafficking of glycophosphatidylinositol (GPI)-anchored proteins

city: Boulder

first: Eric

middle: Michael

last: Davis

state: Colorado

address: MCDB 347 UCB, Gold A345C

department: MCDB

institute: University of Colorado, Boulder

summary: Comparative haploid genetic screens to annotate common and specialized genes required for the biogenesis of individual GPI anchored proteins. SEC62 and SEC63 were required for proper PrP targeting, and were dispensable for CD59. CD59 however, required a GPI side chain modification for maturation, in addition to the aspartyl intramembrane cleaving protease: SPPL3.

overall_design: Examination of GPI trafficking requirements for two GPI anchored proteins using unbiased haploid genetic screens.

middle: M

first: Jingshi

last: Shen


title: A Werner syndrome stem cell model unveils heterochromatin alterations as a driver of human aging

city: Beijing

first: Jingyi

last: Li

address: Yiheyuan Road, No.5, Haidian District

institute: Peking University

type: Methylation profiling by high throughput sequencing

summary: Our data throw light upon the effect of WRN deficiency on gene expression and epigenomic modification, which indicates aging-associated changes from both genomic and epigenomic level.

overall_design: It was compared between WRN+/+ and WRN-/- in hESCs and hMSCs that the gene expression landscapes and epigenetic modifications(H3K4me3, H3K27me3, H3K9me3 and  5-methylcytosine).

first: Weiqi

last: Zhang

first: Xiaomeng

last: Liu

first: Guanghui

first: Fuchou

last: Tang


title: Nuclear DICKKOPF-1 as a biomarker of chemoresistance and poor clinical outcome in colorectal cancer

city: Oviedo

first: Agustin

middle: F

last: Fernandez

email: affernandez@hca.es, agusff@gmail.com

state: Asturias

address: Avenida de Roma s/n

department: IUOPA

country: Spain

institute: Oviedo University-HUCA

summary: Sporadic colorectal cancer (CRC) insurgence and progression depend on the activation of Wnt/β-catenin signaling. Dickkopf (DKK)-1 is an extracellular inhibitor of Wnt/β-catenin signaling that also has undefined β-catenin-independent actions. Here we report for the first time that a proportion of DKK-1 locates within the nucleus of healthy small intestine and colon mucosa, and of CRC cells at specific chromatin sites of active transcription. Moreover, we show that DKK-1 regulates several cancer-related genes including the cancer stem cell marker aldehyde dehydrogenase 1A1 (ALDH1A1) and Ral-binding protein 1-associated Eps domain-containing 2 (REPS2), which are involved in detoxification of chemotherapeutic agents. Nuclear DKK-1 expression is lost along CRC progression; however, it remains high in a subset (15%) of CRC patients (n = 699) and associates with decreased progression-free survival (PFS) after chemotherapy administration and overall survival (OS) [adjusted HR, 1.65; 95% confidence interval (CI), 1.23-2.21; P = 0.002)]. Overexpression of ALDH1A1 and REPS2 associates with nuclear DKK-1 expression in tumors and correlates with decreased OS (P = 0.001 and 0.014) and PFS. In summary, our findings demonstrate a novel location of DKK-1 within the cell nucleus and support a role of nuclear DKK-1 as a predictive biomarker of chemoresistance in colorectal cancer.

overall_design: Analysis of DKK-1 location and associated roles in human colon cancer cells and  crypts of small and large bowel.

first: Oscar

last: Aguilera

first: Jose

last: González-Sancho

first: Sandra

last: Zazo

first: Raúl

last: Rincón

first: Olga

last: Tapia

first: Francesc

last: Canals

first: Beatriz

last: Morte

first: Calvanese

last: Vincenzo

last: Orgaz

first: Nuria

last: Niell

first: Susana

last: Aguilar

last: Freije

first: Osvaldo

last: Graña

first: David

last: Pisano

first: Aurea

last: Borrero

first: Javier

last: Martínez-Useros

first: Benilde

last: Jimenez

first: Mario

last: Fraga

first: Jesús

last: García-Foncillas

first: Carlos

last: López-Otin

first: Miguel

last: Lafarga

first: Federico

last: Rojo

first: Alberto

last: Muñoz


title: Sox9 controls self-renewal of oncogene targeted cells and links tumor initiation and invasion

city: Brussels

first: Jean-Christophe

last: Larsimont

state: Brussels

address: 808, route de Lennik

department: Faculté de médecine

country: Belgium

institute: Université Libre de Bruxelles (ULB)


title: Sox9 controls self-renewal of oncogene targeted cells and links tumor initiation and invasion [ChIP-Seq]

summary: Sox9 is a transcription factor expressed in most solid tumors. However, the molecular mechanisms underlying Sox9 function during tumorigenesis remain unclear. Here, using a genetic mouse model of basal cell carcinoma (BCC), the most frequent cancer in human, we show that Sox9 is expressed from the earliest step of tumor formation in a Wnt/β-catenin dependent manner. Deletion of Sox9 together with the constitutive activation of Hedgehog (HH) signaling completely prevents BCC formation and leads to a progressive loss of oncogene expressing cells. Transcriptional profiling of oncogene expressing cells with Sox9 deletion, combined with in vivo ChIP-sequencing uncovers a cancer-specific gene network regulated by Sox9 that promotes stemness, extracellular matrix (ECM) deposition and cytoskeleton remodeling while repressing epidermal differentiation. Our study identifies the molecular mechanisms regulated by Sox9 that links tumor initiation and invasion.

overall_design: Sox9 ChIP-seq analysis in K14CreER SmoM2 cells.

first: Matthieu

last: Defrance

first: Cédric

last: Blanpain


title: Genome-wide analysis of whole transcriptome in E(z) temperature sensitive cells [RNA-Seq]

first: Hun-Goo

last: Lee

overall_design: Examination of global changes in transcription genome-wide when E(z) is inactivated by monitoring total RNA from E(z) temperature-sensitive cells at  25°C and 31°C in duplicate

first: Amanda

last: Simcox

first: Yuri

last: Schwartz

first: Vincenzo

last: Pirrotta


title: SRSF2 mutations impair hematopoiesis and alter exon recognition

status: Public on May 12 2015

city: Seattle

first: Janine

middle: O

last: Ilagan

state: WA

address: 1100 Fairview Ave N

institute: Fred Hutchinson Cancer Research Center

summary: Mutations within genes encoding spliceosomal proteins are the most common class of mutations in patients with myelodysplastic syndromes, yet it is currently not well understood how these mutations impact hematopoiesis or RNA splicing. Here we report that mutations affecting the splicing factor SRSF2 alter its normal RNA recognition activity, resulting in impaired hematopoietic differentiation and myelodysplasia. Commonly occurring SRSF2 mutations impaired wildtype SRSF2’s normal RNA-binding avidity and preference for specific exonic splicing enhancer RNA motifs. Integration of murine and human transcriptome data identified recurrent mis-splicing of key transcriptional regulators in the presence of mutant SRSF2, including promotion of a highly conserved “poison” exon of EZH2 that results in nonsense-mediated decay and contributes to impaired hematopoiesis. These data provide a mechanistic basis for the enrichment of specific mutations in spliceosomal proteins in myelodysplasia, and suggest that altered RNA recognition activity is a novel mechanism of leukemogenesis.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: mRNA profiles of murine model and K562 cells expressing SRSF2 WT, mutants and knockdown of SRSF2 in TF-1 cells generated by deep sequencing.

first: Eunhee

last: Kim

first: Yang

last: Liang

first: Gerrit

last: Daubner

first: Stanley

first: Aravind

last: Ramakrishnan

first: Yue

last: Li

first: Young

last: Chung

first: Jean-Baptiste

last: Micol

first: Michele

last: Murphy

first: Hana

last: Cho

first: Min-Kyung

first: Ahmad

last: Zebari

first: Silvia

last: Buonamici

first: Peter

last: Smith

first: Joachim

last: Deeg

first: Camille

last: Lobry

first: Iannis

last: Aifantis

first: Yorgo

last: Modis

first: Frederic

last: Allain

first: Stephanie

last: Halene

first: Robert

middle: K

last: Bradley

first: Omar

last: Abdel-Wahab


title: The ChIP-Sequence data of STAT92E in Drosophila adult intestines

first: Wanshi

last: Cai

address: 1 Beichen West Road, Chaoyang District

department: INSTITUTE OF ZOOLOGY

institute: CHINESE ACADEMY OF SCIENCES

summary: JAK/STAT pathway plays important roles in controlling Drosophila intestinal homeostasis and regulating the ISC proliferation and differentiation. However,the downstream targets of its transcription factor-STAT92E remain largely unknown.To further identify the regualtory mechanisms of the JAK/STAT pathway in controlling intestinal homeostasis,we performed the ChIP-Seq assay with mouse raised STAT92E antibody using JAK/STAT signaling highly activated adult intestines.Through the ChIP assay, we have identified over 1000 significant peaks (p<0.01) around the putative targets.The well-characterized JAK/STAT downstream targets including Domeless,Socs36E,STAT92E and chinmo were identified in our ChIP assay,indicating that our experiment is workable to identify novel JAK/STAT downstream targets in adult intestines.This work will provide insights into our understanding of regulatory mechanisms of JAK/STAT signaling during Drosophila intestinal development.

overall_design: Identify the ChIP peaks of STAT92E antibody using JAK/STAT signaling highly actived Drosophila adult intestines, compared with input libaray as the control

first: Yan

last: Zhang

first: Wenyan

last: Ren

first: Xinhua

last: Lin


title: Transcriptome analysis of insecticide resistant housefly strain: insights about SNPs and regulatory elements in differentially expressed cytochrome P450 genes

city: Slagelse

first: Khalid

last: Mahmood

address: Forsøgsvej 1

department: Agro-ecology

country: Denmark

institute: Aarhus University

summary: The 791spin is the spinosad-selected strain derived from 791a, a laboratory strain derived from a multi-resistant field-collected sample of houseflies. The 791a strain proved highly resistant to pyrethroids and some anticholinesterases and showed some resistance to the chitin synthesis-disrupting larvicides.In order to understand the evolution of insecticide resistance, de novo assembly of a spinosad resistant housefly strain 791spin using 454 technology was initiated

overall_design: Transcriptome analysis of insecticide resistant housefly strain compared to susceptible strain

first: Dorte

last: Højland

first: Michael

last: Kristensen


title: Identification and characterization of cold responsive microRNAs in tea plant (Camellia sinensis) based on high-throughput sequencing and their targets using degradome analysis

first: Yue

address: Weigang No.1

institute: Nanjing Agricultural University

platform_organism:
Embryophyta
Camellia sinensis


title: Transcriptome analysis of Th2 CD4+ T cells differentiated from wild-type and NLRP3KO mice

city: Dijon

first: Romain

last: Boidot

address: 1, rue Pr Marion

country: France

institute: CGFL

summary: We report that the Th2 differentiation program is altered in absence of NRLP3 protein

overall_design: Examination of the expression profile of Th2 CD4+ T cells after 3 and 6 days of differentiation

first: Mélanie

last: Bruchard

first: Frédérique

last: Végran

first: Lionel

last: Apetoh

first: François

last: Ghiringhelli


title: Next Generation Sequencing (NGS) comparison of two MVT1 cells subpopulations, CD24- cells and CD24+ cells

city: Oakland

first: Ravi

last: Sachidanandam

address: 2945 Webster St

institute: Girihlet Inc.

summary: The goal of this study is to compare the transcriptome of the 2 MVT1 subpopulations in order to identify new genes and pathways that stands beyond the CD24+ aggressive phenotype

overall_design: mRNA profiles of CD24- and CD24+ cells were generated by deep sequencing, in triplicate, using Illumina HiSeq 2500

first: Ran

last: Rostoker

first: Anitha

last: Jayaprakash


title: Transcriptome data of interacting Salmonella Typhimurium and human host cells

city: Frankfurt

first: Fabian

last: Afonso-Grunz

address: Max-von-Laue-Str. 9

institute: University of Frankfurt

summary: The poly(A)+ and poly(A)− fractions of interacting and non-interacting cells were used for distinct library preparation of interacting and non-interacting prokaryotic pathogen and eukaryotic host cells by deepSuperSAGE. Sequencing was performed with the Illumina HiSeq 2000 platform, and one point of time post infection (early interaction) was additionally prepared by Massive Analysis of cDNA Ends (MACE) as alternative tag-based library preparation method.

sample_organism:
Homo sapiens
Salmonella enterica

platform_organism:
Homo sapiens
Salmonella enterica

overall_design: 10 deepSuperSAGE and 2 MACE libraries. Please consult the publication mentioned in the following for more details.

first: Klaus

last: Hoffmeier

first: Sören

last: Müller

first: Alexander

last: Westermann

first: Björn

last: Rotter

first: Jörg

last: Vogel

first: Peter

last: Winter

first: Günter

last: Kahl


title: RNA sequencing of common myeloid progenitors following short term induction of mutant U2AF1(S34F) expression in transgenic mice

status: Public on May 11 2015

first: Matt

last: Walter

address: 4940 parkview

department: oncology

institute: washington university in st.louis school of medicine

summary: Mutations in the splicing factor gene U2AF1 have been found in the bone marrow of patients with myelodysplastic syndrome and acute myeloid leukemia, as well as in other cancers.  To study the effects of mutant U2AF1(S34F) expression on hematopoiesis and pre-mRNA splicing in hematopoietic cells, we generated two inducible transgenic mouse lines expressing either mutant U2AF1(S34F) or U2AF1(wildtype, WT) as control.  We performed strand-specific transcriptome sequencing on bone marrow common myeloid progenitor cells from U2AF1(S34F) and U2AF1(WT)-expressing mice to examine the pre-mRNA splicing changes associated with expression of mutant U2AF1(S34F).

overall_design: Donor-derived common myeloid progenitor cells from mice transplanted with U2AF1(S34F)/rtTA or U2AF1(WT)/rtTA bone marrow were sorted by flow cytometry, and RNA was extracted for transcriptome analysis.  Ribosomal RNA was depleted prior to strand-specific RNA sequencing (TruSeq stranded library production, followed by 2 x 100bp paired-end sequencing performed on the HiSeq2000 platform from Illumina).  Three samples were sequenced per genotype (n=3), and each sample was composed of bone marrow from 4-5 mice pooled.  Reads were aligned to the mouse mm9 reference genome using TopHat (version 2.0.8), and we performed all subsequent analyses in R.

first: Cara

last: Shirai

first: Matthew


title: Identification of therapeutic targets for glioblastoma by network analysis

city: la jolla

first: vipul

last: bhargava

address: 9500 gilman drive

institute: ucsd

summary: In this study, we confirmed that transformed dedifferentiated astrocytes and neurons acquired a stem/progenitor cell state, although they still retained gene expression memory from their parental cell-type. Transcriptional network analysis on transformed cells revealed up-regulation of genes involved in three signaling pathways: Wnt signaling, cell cycle and focal adhesion with the gene Spp1, also known as osteopontin (OPN) serving as a key node connecting these three pathways. Inhibition of OPN blocked the formation of aggregated neurospheres, affected the proliferative capacity of transformed cell-types and reduced the expression levels of neural stem cell markers. Specific inhibition of OPN in murine glioma tumors prolonged mice survival. We conclude that OPN is an important player in dedifferentiation of cells during tumor formation, hence its inhibition can be a therapeutic target for glioblastoma.

overall_design: Cortical neurons and astrocytes were derived from 11 days old SynapsinI-Cre and GFAP-Cre mice, respectively. The cells were cultured in their respective media to maintain their identity. These cells were then transduced with HRas-shp53 lentivirus with a transduction efficiency of >90%. The transduced neurons and astrocytes were later switched to neural stem cell media devoid of serum and supplemented with FGF-2 (NSC media). Within one week, these cells became proliferative and aggregated to form free-floating neurospheres. These cells, hereinafter referred to as NSynR53 and AGR53, respectively, were later harvested and mRNA collected for sequencing library generation using DP-seq. To assess the regression of these cells to an undifferentiated state along the differentiation axis, enriched populations of mESC and NSC were also grown in vitro and mRNA obtained from these cells were subjected to sequencing library preparation.

first: Vipul

last: Bhargava

first: Shankar

last: Subramaniam


title: CRISPR Display: A modular method for locus-specific targeting of long noncoding RNAs and synthetic RNA devices in vivo

city: Kansas City

first: Scott

middle: T

last: Younger

state: MO

address: 2401 Gillham Road

institute: Children's Mercy Kansas City

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

overall_design: Refer to individual Series


title: CRISPR Display: A modular method for locus-specific targeting of long noncoding RNAs and synthetic RNA devices in vivo [RNA-Seq]

summary: Noncoding RNAs (ncRNAs) comprise an important class of natural regulators that mediate a vast array of biological processes, including the modulation of chromatin architecture. Moreover, artificial ncRNAs have revealed that the functional capabilities of RNA are extremely broad. To further investigate and harness these capabilities, we developed CRISPR-Display ("CRISP-Disp"), a targeted localization strategy that uses Cas9 to deploy large RNA cargos to specific DNA loci. We demonstrate that exogenous RNA domains can be functionally appended onto the CRISPR scaffold at multiple insertion points, allowing the construction of Cas9 complexes with RNAs nearing one kilobase in length, with structured RNAs, protein-binding cassettes, artificial aptamers and pools of random sequences. CRISP-Disp also allows the simultaneous multiplexing of disparate functions at multiple targets. We anticipate that this technology will provide a powerful method with which to ectopically localize functional RNAs and ribonuceloprotein complexes at specified genomic loci.

overall_design: Whole cell poly(A) selected RNA seq, from HEK293FT cells bearing lentivirally-integrated Gaussia and Cypridina luciferase reporter loci.  Cells were transiently transfected with dCas9~VP64 alone, or with dCas9~VP and one of several modified sgRNAs,each targeting the Gaussia reporter.

first: David

last: Shechner

first: Ezgi

last: Hacisuleyman

first: John

last: Rinn


title: CRISPR Display: A modular method for locus-specific targeting of long noncoding RNAs and synthetic RNA devices in vivo [RIP-Seq]

overall_design: RNA Immunoprecipitation (RIP) against FLAG-tagged Cas9 protein, coexpressed with a large pool of CRISPR RNAs bearing random internal insertions


title: Context- and cell-type specific function of miR155-Socs1 interaction in immune regulation

status: Public on May 10 2015

first: Yuheng

last: Lu

address: 417 East 68th Street

institute: Memorial Sloan Kettering Cancer Center

summary: The microRNA (miRNA) dependent regulation of gene expression confers robustness to cellular phenotypes and controls responses to extracellular stimuli. Although a single miRNA can regulate expression of hundreds of target genes, it is unclear whether any of its distinct biological functions can be due to the regulation of a single target. To explore in vivo the function of a single miRNA-mRNA interaction, we mutated the 3' UTR of a major miR-155 target SOCS1 to specifically disrupt its regulation by miR-155.  We found that under physiologic conditions and during autoimmune inflammation or viral infection some immunological functions of miR-155 were fully or largely attributable to the regulation of SOCS1, whereas others could be accounted only partially or not at all by this interaction. Our data suggest that the role of a single miRNA-mRNA interaction is cell type- and biological context-dependent.

overall_design: Naïve WT, SOCS1KI and miR-155KO OVA-specific OT-1 TCR transgenic CD8+ T cells (1x10e4 per mouse) were adoptively transferred into CD45.1+ wt mice prior to infection with MCMV-OVA. WT, SOCS1KI and miR-155KO NK cells (2x10e5 per mouse) were adoptively transferred into CD45.1+ Klra8KO (Ly49H-deficient) mice prior to infection with MCMV. On d4 post infection, CD45.2+ CD44+ CD8+ OT-1 and CD45.2+ Ly49H+ NK1.1+ CD3- NK cells were FACS-sorted (BD FACS ARIA2). Each condition has 3 sequencing replicates.

middle: Y

last: Rudensky


title: Chromatin interaction analysis of transcription factors and NLRP3 in CD4+ T cells differentiated from wild-type and NLRP3KO mice

first: Romain

last: Boidot

summary: We report that the NLRP3 protein is able to interact with chromatin during Th2 differentiation

overall_design: Examination of the interaction of NLRP3 and transcription factors during early stages of Th2 CD4+ T cell differentiation

first: Mélanie

last: Bruchard

first: Frédérique

last: Végran

first: François

last: Ghiringhelli


title: Coordinated epigenetic remodelling occurs during early breast carcinogenesis

status: Public on May 09 2015

city: Darlinghurst

first: Warwick

middle: James

last: Locke

state: NSW

address: 384 Victoria St

department: Cancer Deparment

country: Australia

institute: The Garvan Institute of Medical Research

type: Genome binding/occupancy profiling by array


title: Investigation about fibroblasts of different origins in culture

status: Public on May 08 2015

city: Fribourg

first: Rüegg

last: Curzio

address: Albert-Gockel 1

department: Département de Médecine 3

institute: Université de Fribourg

summary: The goal of this study was to determine if fibroblasts from different origin (skin, colon, tumors) were keeping their characteristic while extracted and cultured ex vivo for several passages. HUVEC was used as a control, being cells from a different background. Surprisingly, fibroblasts from different origins are losing their independant characteristic to cluster in a similar way after 5-6 passages in culture in vitro, showing an activated status.

overall_design: Fibroblasts were extracted from human skin, colon normal stroma and colon tumor stroma. HUVECs were extracted from human samples at the same time. All cells, each group from 3 different patients, were grown on plastic for 5 passages and mRNA was extracted to perform RNASeq analysis.

first: Curzio

last: Rüegg

first: Sarah

last: Knuchel

first: Pascale

last: Anderle


title: Coordinated epigenetic remodelling occurs during early breast carcinogenesis [ChIP-seq & MBD-Seq]

summary: Dysregulation of the epigenome is a common event in malignancy. However, deciphering the earliest cancer associated epigenetic events remains a challenge. Cancer epigenome studies to date have primarily utilised cancer cell lines or clinical samples, where it is difficult to identify the initial epigenetic lesions from those that occur over time. Here, we analysed the epigenome of normal Human Mammary Epithelial Cells (HMEC) and a matched variant cell population (vHMEC) that has escaped senescence and undergone partial carcinogenic transformation. Using this model system we sought to identify the earliest epigenetic changes that potentially occur during carcinogenesis. First we show that the transcriptome of vHMEC resembles that of basal-like breast cancer. Moreover, in vHMEC there is significant deregulation of MYC, p53, EZH2/polycomb, the Aryl Hydrocarbon Receptor (AHR) and miRNAs-143, 145, 199a and 519a at the transcriptional level. Second, we find that vHMEC exhibit genome-wide changes in DNA methylation affecting key cancer-associated pathways. Hypermethylation predominately impacted gene promoters (particularly those targeted by AHR and TP53) and polycomb associated loci, whereas hypomethylation frequently affected enhancers. Next we show that long range epigenetic deregulation occurred in vHMEC involving concordant change in chromatin modification and gene expression across ~0.5-1Mb regions. Finally, we demonstrate that the DNA methylation changes we observe in vHMECs, occur in basal-like breast cancer (notably FOXA1 hypermethylation).. Overall our results suggest that the first steps of carcinogenesis are associated with a co-ordinated deregulation of DNA methylation and chromatin modification spanning a range of genomic loci potentially targeted by key transcription factors and a corresponding deregulation of transcriptional networks.

overall_design: We sought to study the chromatin modification profile of human mammary epithelial cells (HMEC) and a matched isogenic variant population (vHMEC) utilising ChIP-seq. ChIP was performed against H3K27ac, H3K36me3 and H3K27me3 for a HMEC and vHMEC timpoint in one donor. H3K4me3 CHIP was performed in two donors, which were treated as biological replicates.


title: Chemical profiling of the genome with anti-cancer drugs defines target specificities

city: Palo Alto

first: Baoxu

last: Pang

state: CALIFORNIA

address: 3165 Porter Drive

department: Genetics

institute: Stanford University

summary: Many anti-cancer drugs induce DNA breaks to eliminate tumor cells. The anthracycline topoisomerase II inhibitors can also evict histones. We performed a genome-wide high-resolution mapping of chemotherapeutic effects of various topoisomerase I and II inhibitors. We show that different drugs target different types of chromatin for induction of DNA damage and histone eviction. Topoisomerase inhibitors topotecan and etoposide similarly target transcriptionally active chromatin for DNA damage. Daunorubicin induces DNA breaks and evicts histones in active chromatin, thus quenching local DNA damage response. The analog aclarubicin evicts histones in H3K27me3-marked heterochromatin. These results can guide rational treatment decisions regarding these genome manipulating anti-cancer drugs.

overall_design: FAIRE-seq and g-H2AX ChIP-seq were performed on K562 cells after drug exposure

first: Johann

last: de Jong

first: Xiaohang

last: Qiao

first: Arno

last: Velds

first: Ron

last: Kerkhoven

first: Lodewyk

last: Wessels

first: Jacques

last: Neefjes


title: Extrachromosomal DNA (microDNA) in Human, Mouse and Chicken

city: Charlottesville,

first: Anindya

last: Dutta

email: ad8q@virginia.edu, pk7z@virginia.edu

state: VA

address: Room no. 1232, Jordan Hall

department: MD-BIOC BC/Mole Genetics~School of medicine

institute: University of Virginia

type: Genome variation profiling by high throughput sequencing

summary: MicroDNAs are <400-base long extrachromosomal circles found in mammalian cells.  Tens of thousands of microDNAs were found in all tissue types, including sperm.  MicroDNAs arose preferentially from areas with high gene density, high GC content, high exon density, promoters with activating chromatin modifications and in sperm from the 5'UTR of full-length LINE-1 elements, but were depleted from lamin-associated heterochromatin. Furthermore, analysis of microDNAs from a set of human cancer cell lines revealed lineage-specific patterns of microDNA origins. A survey of microDNAs from chicken cell lines defective in various DNA repair proteins revealed that homologous recombination repair and non-homologous end joining repair pathways are not required for microDNA production. A deletion of the MSH3 protein, involved in DNA mismatch repair, resulted in a significant decrease in microDNA abundance specifically from non-CpG areas of the genome. Thus microDNAs arise as part of normal cellular physiology, either from DNA breaks associated with RNA metabolism or from replication slippage followed by mismatch repair.

sample_organism:
Gallus gallus
Homo sapiens
Mus musculus

platform_organism:
Gallus gallus
Homo sapiens
Mus musculus

overall_design: Circular DNA profiling by high throughput sequencing. Five human, ten mouse and nine chicken samples are analyzed

first: Pankaj

last: Kumar

first: Laura

middle: W

last: Dillon

first: Yoshiyuki

last: Shibata


title: Evaluation of RNA amplification and RNA-Seq library preparation protocols for spermatozoa RNA profiling

city: Detroit

first: Stephen

last: Krawetz

state: MI

address: 275 E. Hancock

department: Center for Molecular Medicine and Genetics

institute: Wayne State University School of Medicine

summary: RNA-Seq technique was applied to investigate the effects of four cDNA amplification kits and two RNA-Seq library preparation kits to the deep sequencing results at different perspectives.

overall_design: The same set of semen samples were applied to investigate the qualitative and quantitative effect of four cDNA amplification methods and two RNA-Seq library preparation methods on sperm transcript profiling.

first: Shihong

last: Mao

first: Edward

last: Sendler

first: Robert

last: Goodrich

first: Russ

last: Hauser

middle: A


title: CTCF and Smad signaling proteins and response to DPP

first: Kevin

last: Van Bortle

address: 3165 Porter Dr

summary: ChIP-seq profiles for CTCF and TGF-beta signaling factors before and after treatment with DPP

overall_design: Examination of transcription factor dynamics during signaling

first: K

first: N

last: Takenaka

first: V

last: Corces


title: 3' RNA-seq of flies over-expressing cabut protein or RNAi against cabut mRNA

city: Waltham

first: Sebastian

last: Kadener

address: 415 South Street

institute: Brandeis University

summary:
CbtOE (Tim-gal4; UAS-cbtFLAG), Tim-gal4 (control for CbtOE), cbtRNAi (Tim-gal4-UAS-Dcr2-UAS-cbtIR-cbtE1) and Tim-gal4;UAS-Dcr2 (control for CbtRNAi) flies.
Flies were entrained in LD (light: dark) condition for 3-4 days and harvested at six time points: ZT3, ZT7, ZT11, ZT15, ZT19, ZT23
Fly heads  were collected, RNA was extracted and  RNA-seq libraries were prepared as previously described (Engreitz et al., 2013)

overall_design: Three  samples of cbtRNAi and three samples of their controls.  Two samples of cbtOE with two samples of their controls.

first: Ville

last: Hietakangas


title: A potential role for snoRNA in PKR activation during metabolic stress

city: Salt Lake City

first: Osama

last: Youssef

state: UT

address: 15 N medical Dr

institute: University of Utah

summary: PKR (Protein Kinase RNA-activated) is activated by metabolic stress, such as that associated with obesity, and this activation depends on its RNA-binding domain. Here we investigated whether endogenous RNA triggers PKR activation in response to lipid exposure. Our results indicate that snoRNAs (small nucleolar RNAs) associate with PKR during metabolic stress. Our high-throughput sequence shows that snoRNAs are enriched in PKR_WT samples after palmitic acid treatment compared to mock-treated samples.

overall_design: MEF cells reconstituted with wild type PKR (PKR_WT) or PKR with a mutation in each dsRBM (PKR_RM) were mock- or palmitate-treated. RNAs associated with PKR were immunoprecipitated. IPed RNAs were extracted, cloned and sequenced. We performed differential expression to obtain RNAs enriched in PKRWT after palmitate treatment. We sequenced three different datasets of samples (A, B and C). Each A and B dataset contains three biological replicates. Dataset C contains a single sample.

first: Sarah

last: Safran

first: Takahisa

last: Nakamura

first: David

last: Nix

first: Gökhan

middle: S

last: Hotamisligil

first: Brenda

middle: L

last: Bass


title: Disruptions of Topological Chromatin Domains Causes Pathogenic Rewiring of Gene-Enhancer Interactions [4C-Seq-mouse]

city: Berlin

first: Darío

last: Lupiáñez

address: Ihnestrasse 73

institute: Max Planck Institute for Molecular Genetics

summary: Mammalian genomes are organized into megabase-scale topologically associated domains (TADs) that have been proposed to represent large regulatory units. Here we demonstrate that disruption of TADs can cause rewiring of long-range regulatory architecture and result in pathogenic phenotypes. We show that distinct human limb malformations are caused by deletions, inversions, or duplications altering the structure of the TAD-spanning WNT6/IHH/EPHA4/PAX3 locus. Using CRISPR/Cas genome editing, we generated mice with corresponding rearrangements. Both in mouse limb tissue and patient-derived fibroblasts, disease-relevant structural changes cause ectopic interactions between promoters and non-coding DNA, and a cluster of limb enhancers normally associated with Epha4 is misplaced relative to TAD boundaries and drives ectopic limb expression of another gene in the locus. Our results demonstrate the functional importance of TADs for orchestrating gene expression via genome architecture and indicate criteria for predicting the pathogenicity of human structural variants, particularly in non-coding regions of the human genome.

overall_design: Circular Chromosome Conformation Capture (4C seq) at the Wnt6/Ihh/Epha4/Pax3 locus in developing distal limbs of mutants and WT animals at E11.5

first: Stefan

last: Mundlos


title: Disruptions of Topological Chromatin Domains Causes Pathogenic Rewiring of Gene-Enhancer Interactions


title: Disruptions of Topological Chromatin Domains Causes Pathogenic Rewiring of Gene-Enhancer Interactions [4C-Seq-human]

overall_design: Circular Chromosome Conformation Capture (4C seq) at the WNT6/IHH/EPHA4/PAX3 locus in human adult fibroblasts (HAF) of adult patients and controls


title: Spalax galili and Spalax carmeli Placenta Transcriptomes

city: Tampa

first: Derek

last: Wildman

state: FL

address: 3720 Spectrum Blvd

institute: University of South Florida

summary:
Placenta transcriptome from two species of Spalax.
Associated with the article "Genome-wide adaptive complexes to underground stresses in blind mole rats Spalax."

sample_organism:
Spalax carmeli
Nannospalax galili

platform_organism:
Spalax carmeli
Nannospalax galili

overall_design: Transcriptome or Gene expression

first: William

middle: E

last: Gundling Jr


title: YBX1 HITS-CLIP

city: San Francisco

first: Hani

last: Goodarzi

department: Biochemistry and Biophysics

institute: UCSF

summary: In order to identify YBX1 binding sites on endogenous RNA, we performed HITS-CLIP on endogenous YBX1

overall_design: We used a previously published method to perform HITS-CLIP on endogenous YBX1 (Licatalosi D, et al. 2008, Nature 456:464-U22)

first: H

first: X

last: Liu

first: S

last: Zhang

first: L

last: Fish

last: Tavazoie


title: Disruptions of Topological Chromatin Domains Causes Pathogenic Rewiring of Gene-Enhancer Interactions [RNA-Seq]

overall_design: RNA-seq profile of developing distal limbs of mutants and WT animals at E11.5


title: YBX1 small-RNA HITS-CLIP

summary: In order to identify YBX1 binding sites on tRNA fragments, we performed small-RNA HITS-CLIP on endogenous YBX1

overall_design: We used a previously published method to perform HITS-CLIP on endogenous YBX1 (Chi SW, et al. 2009, Nature 460:479)


title: Dosage compensation can buffer copy-number variation in wild yeast

city: Madison

first: Audrey

last: Gasch

state: WI

address: 425-G Henry Mall

institute: University of Wisconsin - Madison

summary: We show that aneuploidy is common in wild isolates of yeast, which are inherently tolerant to chromosome amplification and down-regulate expression at 40% of amplified genes.  To dissect the mechanism of this dosage response, we generated isogenic strain panels in which diploid cells carried either two, three, or four copies of the affected chromosomes.  Using a mixture of linear regression (MLR) model to classify genes, we find that expression is actively down regulated in proportion to increased gene copy at up to 30% of genes. Genes subject to dosage control are under higher expression constraint – but show elevated rates of gene amplification – in wild populations, suggesting that dosage compensation buffers copy number variation (CNV) at toxic genes

overall_design: RNA-seq and transcriptome analysis of S. cerevisiae natural isolates having aneuploidy. Technical triplicate was performed for isogenic diploid strains having 2, 3 and 4 copies of a given chromosome (strain panels), while technical duplicate or singulate was performed on all other aneuploids.

first: James

last: Hose


title: Transcript 5'-end mapping was used to identify transcriptional start sites and RNA processing sites genome-wide in Mycobacterium tuberculosis

city: Worcester

first: Scarlet

last: Shell

address: 60 Prescott St

department: Biology and Biotechnology

institute: Worcester Polytechnic Institute

summary: Most eukaryotic mRNAs are enzymatically processed before they are translated, but less is known about mRNA processing in prokaryotes.  While bacterial rRNAs and tRNAs are known to be highly processed, only a handful of bacterial mRNAs have been shown to be processed by ribonucleases, typically altering transcript stability. We hypothesized that mRNA processing might be more widespread in prokaryotes and constitute a previously underappreciated layer of post-transcriptional regulation. We therefore adapted RNA-seq methodologies to map transcript 5’ ends across the Mycobacterium tuberculosis transcriptome, distinguishing between transcriptional start sites (TSSs) and RNA processing sites.  We report for the first time genome-wide mRNA endonucleolytic cleavage patterns in a prokaryote, and find that mRNA processing is widespread in M. tuberculosis. This has physiologic consequences, as mRNA cleavage plays a regulatory function in the ESX-1 locus by producing a transcript fragment encoding the secreted proteins EsxB and EsxA that is significantly more stable than the parent transcript, leading to greater steady-state transcript abundance. At least 20 other loci display similar patterns of processing-associated differences in transcript levels in M. tuberculosis. Mycobacteria therefore use mRNA processing to effect differential abundance of sub-sections of polycistronic transcripts, which may provide additional layers of regulation. Moreover, our results demonstrate that stable, processed mRNAs are a ubiquitous feature in this prokaryotic transcriptome, suggesting that mRNA processing may represent a major post-transcriptional regulatory mechanism.

overall_design: RNA from two biological replicate cultures was analyzed by RNA-seq for expression analysis and 5'-end-mapping for identification of transcriptional start sites and RNA processing sites. An additional set of three biological replicate cultures were analyzed by RNA-seq for expression analysis using a different library construction method.

last: Fortune

first: Michael

last: Chase

first: Thomas

last: Loerger


title: 3' RNA-seq of fly expressing cabut RNAi after exposure to different levels of sucrose

first: Sebastian

last: Kadener

summary:
Control (+/cbtE1-UAS-cbt RNAi) or cabut RNAi flies (Tim-gal4, UAS-cbt RNAi) were starved for 16 hours and then exposed to food containing different concentrations of sucrose: 0, 25, 50  and 100 %  for 18 hours.
Fly heads  were collected, RNA was extracted and  RNA-seq libraries were prepared as previously described (Engreitz et al., 2013)

overall_design: For each sucrose  concentration, two samples of cabut RNAi flies and one sample of control flies were sequenced.

first: Ville

last: Hietakangas


title: Frequent and Transient Acquisition of Pluripotency During Somatic Cell Trans-Differentiation with iPSCs Reprogramming Factors (WGBS)

status: Public on May 07 2015

city: Rehovot

first: Noa

last: Novershtern

address: Weizmann Institute

department: Molecular Genetics

country: Israel

institute: Weizmann Institute of Science

summary: Recent reports have proposed a new paradigm for obtaining mature somatic cell types from fibroblasts without going through a pluripotent state, by briefly expressing canonical iPSC reprogramming factors Oct4, Sox2, Klf4 and c-Myc (abbreviated as OSKM), in cells expanded in lineage differentiation promoting conditions. Here we apply genetic lineage tracing for endogenous Nanog, Oct4 and X chromosome reactivation during OSKM induced trans-differentiation, as these molecular events mark final stages for acquisition of induced pluripotency. Remarkably, the vast majority of reprogrammed cardiomyocytes or neural stem cells derived from mouse fibroblasts via OSKM mediated trans-differentiation were attained after transient acquisition of pluripotency, and followed by rapid differentiation. Our findings underscore a molecular and functional coupling between inducing pluripotency and obtaining “trans-differentiated” somatic cells via OSKM induction, and have implications on defining molecular trajectories assumed during different cell reprogramming methods.

overall_design: WGBS (Whole-Genome-Bisulfite-sequencing) were measured during conversion of mouse embryonic fibroblasts to neural stem cells using OSKM trans-differentiation method, as well as in mouse emrbyonic fibroblasts, and mouse ESCs.

first: Itay

last: Maza

first: Elad

last: Chomsky

first: Jacob

last: Hanna


title: Small RNA Solexa sequencing of chickpea (Cicer arietinum)

city: Kunming

first: Yun

last: Zheng

state: Yunnan

address: 452 Fengyuan Road

institute: Yunnan Agricultural University

summary: The total RNA were extracted from pooled tissues of leaves and flowers from several plants of chickpea (Cicer arietinum) using TRIzol reagent (Invitrogen) according to the manufacturer's instructions. Then small RNAs ranging in 18–30 nucleotides were size fractionated electrophoretically, isolated from the gel, ligated with the 5′ and 3′ RNA adapters. The ligated product was reverse transcribed and subsequently amplified using 10–12 PCR cycles. The purified PCR product was sequenced using Illumina Genome Analyzer II. The qualified reads were used to predict microRNAs and phased small interfering RNAs from chickpea.

overall_design: Identification of microRNAs and phased small inferfering RNAs in chickpea (Cicer arietinum) by analyzing small RNA sequencing profiles of leaves and flowers using Illumina GAII.

first: Ramanjulu

last: Sunkar


title: Allele-specific binding of ZFP57 in the regulation of imprinted and mono-allelic expression

city: Cambridge

first: Felix

last: Krueger

address: Granta Park

department: Bioinformatics

country: United Kingdom

institute: Altos Labs

summary: Selective maintenance of genomic methylation imprints during pre-implantation development is required for parental origin-specific expression of imprinted genes. The Kruppel-like zinc finger protein ZFP57 acts as a factor necessary for maintaining the DNA methylation memory at multiple imprinting control regions (ICRs) in early mouse embryos and ES cells. Maternal-zygotic deletion of ZFP57 in mice presents a highly penetrant phenotype with no animals surviving to birth. In addition, several cases of human transient neonatal diabetes (TND) are associated with somatic mutations in ZFP57 coding sequence. Here we comprehensively map sequence-specific ZFP57 binding sites in an allele-specific manner using hybrid ES cell lines from reciprocal crosses between C57BL/6J and Cast/EiJ mice assigning allele specificity to approximately two thirds of all binding sites. While half of these are biallelic and include ERV targets, the rest show mono-allelic binding based either on parental-origin or on genetic background of the allele. Parental-origin allele-specific binding was methylation-dependent and mapped only to imprinted DMRs established in the germline (gDMRs). No binding was evident at secondary somatically-derived DMRs. ZFP57-bound gDMRs can predict imprinted gene expression and we identify new imprinted genes, including the Fkbp6 gene with a critical function in mouse male germ cell development. Genetic-background specific sequence differences also influence ZFP57 binding. We show that genetic variation that disrupts the consensus binding motif and its methylation is associated with mono-allelic expression of neighbouring genes. The work described here uncovers further roles for ZFP57 mediated regulation of genomic imprinting and identifies a novel mechanism for genetically determined mono-allelic gene expression.

overall_design: Input and Zfp57 CHiP-Seq profiles of hybrid Black6/Cast ES cells were generated by sequencing using the Illumina GAIIx platform.

first: Ruslan

last: Strogantsev

first: Kazuki

last: Yamazawa

first: Hui

last: Shi

first: Poppy

last: Gould

first: Megan

last: Goldman-Roberts

first: Kirsten

last: McEwan

first: Bowen

last: Sun

first: Roger

last: Pederson

first: Anne

middle: C

last: Ferguson-Smith


title: Hepatic transcriptional signatures of exposure to oestradiol (E2) and the herbicide linuron in mature male brown trout

city: Swansea

first: Tamsyn

last: Uren Webster

address: Wallace Building

institute: Swansea University

summary: These studies aimed to investigate the hepatic transcriptional response of brown trout to the natural estrogen, E2, and the herbicide linuron. We exposed mature male brown trout to three concentrations of each chemical for 4 days and sequenced the hepatic transcriptome of 3 individuals per treatment group in order to determine the global mechanisms of toxicity of these environmental contaminants. We assembled the brown trout transcriptome using a de novo approach. Subsequent differential expression analysis identified a total of 2113 differentially-regulated transcripts in the group exposed to the highest E2 treatment concentration, and 822 differentially-regulated transcripts across all linuron treatments. For E2, differentially-expressed transcripts included those encoding known oestrogen-responsive genes, while regulated processes included those associated with vitellogenesis including lipid metabolism, cellular proliferation and ribosome biogenesis. For linuron, there was a striking down-regulation of transcripts encoding the majority of the enzymes involved in the cholesterol biosynthesis pathway, and also a considerable induction of transcripts involved in cellular stress response including Cyp1a.

overall_design: Fish were exposed to 3 concentrations of E2 (measured concentrations were 1.9, 18.1 and 34.4 ng/L), 3 concentrations of linuron (measured concentrations were 1.7, 15.3 and 225.9 µg/L) and water controls for 4 days. Liver mRNA from 3 replicate individuals per treatment was sequenced in an Illumina HiSeq 2500 platform. Two control groups (n=6 fish in total) were included. Using a de novo approach, we assembled the hepatic transcriptome for brown trout. Sequence reads were re-mapped to the assembled transcriptome using Bowtie2 and transcript expression profiling was conducted using EdgeR. ERCC spike controls were added to all individual samples, allowing for the assessment of the reproducibility and dynamic range for transcript expression quantification in our experiments.

first: Eduarda

last: Santos


title: Global Gene Expression analysis of CUTLL1 cell lines after treatment with Perhexiline

city: New York

first: Alberto

last: Ambesi-Impiombato

address: 1130 St. Nicholas Avenue

department: ICG

institute: Columbia University

summary: We identify perhexiline, a small molecule inhibitor of mitochondrial carnitine palmitoyltransferase-1, as a HES1-signature antagonist drug with robust antileukemic activity against NOTCH1 induced leukemias in vitro and in vivo.

overall_design: RNA-Seq from CUTLL1 cell lines treated with Perhexiline or vehicle for 3 days

first: Stephanie

last: Schnell

first: Luyao

last: Xu

first: Yue

last: Qin

first: Ryoichiro

last: Kageyama

first: Adolfo

last: Ferrando


title: Multiplex Single Cell Profiling of Chromatin Accessibility by Combinatorial Cellular Indexing [ATAC-seq]

city: Tucson

first: Darren

middle: A.

last: Cusanovich

state: AZ

address: 1230 N Cherry Avenue, BSRL 421

department: Cellular and Molecular Medicine

institute: University of Arizona

summary: Technical advances have enabled the collection of genome and transcriptome data sets with single-cell resolution. However, single-cell characterization of the epigenome has remained challenging. Furthermore, because cells must be physically separated prior to biochemical processing, conventional single-cell preparatory methods scale linearly. We applied combinatorial cellular indexing to measure chromatin accessibility in thousands of single cells per assay, circumventing the need for compartmentalization of individual cells. We report chromatin accessibility profiles from over 15,000 single cells and use these data to cluster cells on the basis of chromatin accessibility landscapes. We identify modules of coordinately regulated chromatin accessibility at the level of single cells both between and within cell types, with a scalable method that may accelerate progress toward a human cell atlas.

overall_design: A series of experiments involving mixtures of cells

first: Riza

last: Daza

first: Andrew

last: Adey

first: Hannah

last: Pliner

first: Lena

last: Christiansen

first: Kevin

last: Gunderson

first: Frank

middle: J

last: Steemers

first: Cole

last: Trapnell

first: Jay

last: Shendure


title: Tex10 Coordinates Epigenetic Control of Super-Enhancer Activity for Pluripotency and Reprogramming [ChIP-Seq]

first: Jianlong

last: Wang

state: New York

department: Department of Medicine

institute: Columbia University Irving Medical Center

summary: Super-enhancers (SEs) are large clusters of transcriptional enhancers that are co-occupied by multiple lineage specific transcription factors driving expression of genes that define cell identity. In embryonic stem cells (ESCs), SEs are highly enriched for Oct4, Sox2, and Nanog in the enhanceosome assembly and express enhancer RNAs (eRNAs). We sought to dissect the molecular control mechanism of SE activity and eRNA transcription for pluripotency and reprogramming. Starting from a protein interaction network surrounding Sox2, a key pluripotency and reprogramming factor that guides the ESC-specific enhanceosome assembly and orchestrates the hierarchical transcriptional activation during the final stage of reprogramming, we discovered Tex10 as a novel pluripotency factor that is evolutionally conserved and functionally significant in ESC self-renewal, early embryo development, and reprogramming. Tex10 is enriched at SEs in a Sox2-dependent manner and coordinates histone acetylation and DNA demethylation of SEs. Our study sheds new light on epigenetic control of SE activity for cell fate determination.

overall_design: Genome binding/occupancy profiling of Tex10 was performed in mouse embryonic stem cells by ChIP sequencing.

first: Junjun

last: Ding

first: Xin

last: Huang


title: Ion Transporter NKCC1 - Modulator of Neurogenesis in Murine Olfactory Neurons

city: Bochum

first: Claudia

last: Haering

address: Universitätsstr. 150

department: Biology

institute: Ruhr University Bochum

summary: Olfaction is one of the most crucial senses for vertebrates regarding foraging and social behavior. Therefore, it is of particular interest to investigate the sense of smell, its function on a molecular level, the signaling proteins involved in the process and the mechanism of required ion transport. In recent years, the precise role of the ion transporter NKCC1 in olfactory sensory neuron (OSN) chloride accumulation has been a controversial subject. NKCC1 is expressed in OSNs and is involved in chloride accumulation of dissociated neurons, but it had not been shown to play a role in mouse odorant sensation.  To characterize transporter gene expression in NKCC1-/- mice, we examined the OE gene profile (Supplementary Table 1) using Illumina RNA-Seq to generate OE transcriptomes from NKCC1-/- and wild type mice. We analyzed RNA from OEs of male and female NKCC1+/+ (12 ± 1 weeks) and NKCC1-/- mice (16.5 ± 3.5 weeks, NMRI background); each RNA sample was prepared from an OE pool of 4 (mixed-gender pool RNA isolation) or 2 (gender RNA pool) different mice for each condition. Our data demonstrated the absence of a highly expressed ion transporter that could compensate for NKCC1.

overall_design: The Illumina RNA-Seq protocol was utilized. In total, we amplified and sequenced up to 38 million 101 nt-long fragments from murine NKCC1+/+ and NKCC1-/- adult OEs.

first: Ninthujah

last: Kanageswaran

first: Pascal

last: Bouvain

first: Paul

last: Scholz

first: Janine

last: Altmüller

first: Christian

last: Becker

first: Günter

last: Gisselmann

last: Wäring-Bischof

first: Hanns

last: Hatt


title: Prospective derivation of a 'Living Organoid Biobank' of colorectal cancer patients

city: Utrecht

first: Lennart

last: Kester

address: Uppsalalaan 8

institute: Hubrecht Institute

summary: In Rspondin-based three-dimensional cultures, Lgr5 stem cells from multiple organs form ever-expanding epithelial organoids that retain their tissue identity. Here we report the establishment of tumor organoid cultures from 20 consecutive colorectal carcinoma (CRC) patients. For most, organoids were also generated from adjacent normal tissue. Organoids closely resemble the original tumor. The spectrum of genetic changes within the 'living biobank' agrees well with previous large-scale mutational analyses of CRC. Gene expression analysis indicates that the major CRC molecular subtypes are represented. Tumor organoids are amenable to high-throughput drug screens allowing detection of gene-drug associations. As an example, a single organoid culture was exquisitely sensitive to Wnt secretion (porcupine) inhibitors and carried a mutation in the negative Wnt feedback regulator RNF43, rather than in APC. Organoid technology may fill the gap between cancer genetics and patient trials, complement cell line- and xenograft-based drug studies and allow personalized therapy design.

overall_design: Self-renewal of the intestinal epithelium is driven by Lgr5 stem cells located in crypts. We have recently developed a long-term culture system that maintains basic crypt physiology. Wnt signals are required for the maintenance of active crypt stem cells. Indeed, the Wnt agonist R-spondin1 induces dramatic crypt hyperplasia in vivo. R-spondin-1 is the ligand for Lgr5. Epidermal growth factor (EGF) signaling is associated with intestinal proliferation, while transgenic expression of Noggin induces a dramatic increase in crypt numbers. The combination of R-spondin-1, EGF, and Noggin in Matrigel sustains ever-expanding small intestinal organoids, which display all hallmarks of the original tissue in terms of architecture, cell type composition, and self-renewal dynamics. We adapted the culture condition for long-term expansion of human colonic epithelium and primary colonic adenocarcinoma, by adding nicotinamide, A83-01 (Alk inhibitor), Prostaglandin E2 and the p38 inhibitor SB202190. Of note, a two-dimensional culture method for cells from normal and malignant primary tissue has been described by Schlegel and colleagues. Here, we explore organoid technology to routinely establish and phenotypically annotate ‘paired organoids’ derived from adjacent tumor and healthy epithelium from CRC patients.

first: Marc

last: van de Wetering

first: Alexander

last: van Oudenaarden

first: Hans

last: Clevers


title: Tex10 Coordinates Epigenetic Control of Super-Enhancer Activity for Pluripotency and Reprogramming [RNA-Seq]

overall_design: RNA sequencing analysis was performed in mouse embryonic stem cells with Luciferase and Tex10 knockdown. RNA-seq Experiments were carry out in two biological replicates.


title: Role of miRNAs in acute kidney injury recovery induced by mesenchymal stromal cells and derived extracellular vesicles.

city: Torino

first: Francesco

last: Neri

state: Italy

address: Via Nizza 52

country: Italy

institute: University of Torino

summary: Phenotypic changes induced by extracellular vesicles (EVs) have been implicated in the recovery of acute kidney injury (AKI) induced by mesenchymal stromal cells (MSCs). miRNAs are potential candidates for cell reprogramming towards a pro-regenerative phenotype. The aim of the present study was to evaluate whether miRNA de-regulation inhibits the regenerative potential of MSCs and derived-EVs in a model of glycerol-induced AKI in SCID mice. For this purpose, we generated MSCs depleted of Drosha, a critical enzyme of miRNA maturation, to alter miRNA expression within MSCs and EVs. Drosha knock-down MSCs (MSC-Dsh) maintained the phenotype and differentiation capacity. They produced EVs that did not differ from those of wild type cells in quantity, surface molecule expression and internalization within renal tubular epithelial cells. However, EVs derived from MSC-Dsh (EV-Dsh) showed global down-regulation of miRNAs. Whereas, wild type MSCs and derived EVs were able to induce morphological and functional recovery in AKI, MSC-Dsh and EV-Dsh were ineffective. RNA sequencing analysis showed that genes deregulated in the kidney of AKI mice were restored by treatment with MSCs and EVs but not by MSC-Dsh and EV-Dsh. Gene Ontology analysis showed that down-regulated genes in AKI were associated with fatty acid metabolism. The up-regulated genes in AKI were involved in inflammation, ECM-receptor interaction and cell adhesion molecules. These alterations were reverted by treatment with wild type MSCs and EVs, but not by the Drosha counterparts. In conclusion, miRNA depletion in MSCs and EVs significantly reduced their intrinsic regenerative potential in AKI, suggesting a critical role of miRNAs.

overall_design: RNA-seq

first: Federica

last: Collino

first: Stefania

last: Bruno

first: Danny

last: Incarnato

first: Daniela

last: Dettori

first: Salvatore

last: Oliviero

first: Ciro

last: Tetta

first: Peter

last: Quesenberry

first: Giovanni

last: Camussi


title: Gadd45a promotes DNA demethylation through TDG

city: Beijing

first: xiaomeng

last: liu

state: Beijing

address: Yiheyuan Road No.5

institute: Biodynamics Optical Imaging Center (BIOPIC), Peking University

summary: Growth arrest and DNA-damage-inducible protein 45 (Gadd45) family members have been implicated in DNA demethylation in vertebrates. However, it remained unclear how they contribute to the demethylation process. Here, we demonstrate that Gadd45a promotes active DNA demethylation through thymine DNA glycosylase (TDG) which has recently been shown to excise 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC) generated in Ten-eleven-translocation (Tet) -initiated oxidative demethylation. The connection of Gadd45a with oxidative demethylation is evidenced by the enhanced activation of a methylated reporter gene in HEK293T cells expressing Gadd45a in combination with catalytically active TDG and Tet. Gadd45a interacts with TDG physically and increases the removal of 5fC and 5caC from genomic and transfected plasmid DNA by TDG. Knockout of both Gadd45a and Gadd45b from mouse ES cells leads to hypermethylation of specific genomic loci most of which are also targets of TDG and show 5fC enrichment in TDG-deficient cells. These observations indicate that the demethylation effect of Gadd45a is mediated by TDG activity. This finding thus unites Gadd45a with the recently defined Tet-initiated demethylation pathway.

overall_design: The dataset includes RRBS anlysis of 2 WT ES cell samples and 2 Gadd45a/b DKO ES cell samples.

first: Zheng

last: Li

first: Tian

middle: P

last: Gu

first: Alain

middle: R

last: Weber

first: Jia

middle: Z

last: Shen

first: Bin

first: Zhi

middle: G

last: Xie

first: Ruichuan

last: Yin

first: Fan

last: Guo

first: Xiaomeng

last: Liu

first: Fuchou

last: Tang

first: Hailin

last: Wang

first: Primo

last: Schär

first: Guo

last: Xu


title: Transcriptome analysis of the effects of GSK-J4 and L-ascorbic acid in female embryonic stem cells

city: White Plains

first: Mary

last: Donohoe

state: NY

address: 785 Mamaroneck

department: Burme Medical Research Institute

institute: Weill Cornell Medical College

summary: The effects of a histone demethylase inhibitor, GSK-J4, and L-ascorbic acid for the transcriptome in female ES cells were analyzed by RNA-sequence. Total RNA was used for high-throughput sequence with Illumina HiSeq 2500 and mapped to mm10.

overall_design: Total RNA profile

first: M

first: Y

middle: F

last: Kamikawa


title: Genome wide map of H3K4me3 histone modification in C6 rat glioma cells [SOLiD]

city: Uppsala

first: Marcin

last: Kruczyk

address: Husargatan 3

country: Sweden

institute: Uppsala University

summary: We report a map of H3K4me3 - an activiting expression histone modification in C6 rat glioma cells. The data was obtained using whole genome high throughput technology. The sequencing was performed on Solid 5500xl platform.

overall_design: Examination of  H3K4me3 histone modification in C6 rat glioma cell line

first: Bozena

last: Kaminska

first: Jan

last: Komorowski


title: Digital Gene Expression profiling of Schmidtea mediterranea stem cells.

city: Barcelona

first: Gustavo

last: Rodriguez-Esteban

state: Catalonia

address: C. Baldiri Reixac, 4; Barcelona Science Park

department: Development

institute: Centro Nacional de Análisis Genómico

summary: The freshwater planarian Schmidtea mediterranea is well known by its amazing regeneration capabilities thanks to the presence of adult stem cells, the neoblasts, the only proliferative cells and responsible for the differentiation in all the cell types of the organism. This study involves the creation of separated transcript libraries from both, isolated neoblasts and differentiated cells, as well as their posterior sequencing and quantification through Digital Gene Expression (DGE) for the characterization of the neoblast transcriptome.

overall_design: Three DGE libraries were produced from FACS isolated cell populations X1 (proliferating stem cells, S/G2/M), X2 (a mix of proliferating stem cells and stem cell progeny, G0/G1) and Xin (differentiated cells, G0/G1). Cells were isolated from a pool of 32 regenerating animals two days after being cut off pre- and post-pharingeally to trigger regeneration and neoblast proliferation.

first: Emili

last: Saló

first: Josep

last: Abril


title: Integration of genome-wide Stat3 binding and epigenetic modifications with transcriptome allowed identification of novel Stat3 target genes in glioma cells

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

overall_design: Refer to individual Series


title: RNA-Seq gene expression profiling in the plant growth promoting Bacillus atrophaeus UCMB-5137 revealed an alternative strategy of plant colonization compared to Bacillus amyloliquefaciens endophytes

status: Public on May 06 2015

city: Pretoria

first: Oleg

last: Reva

address: Lynnwood Rd., Hillcrest

department: Biochemistry

country: South Africa

institute: University of Pretoria

summary: Plant growth promoting rhizobacteria (PGPR) of the genus Bacillus are successfully used as biofertilizers and biopesticides. They potentially can reduce the use of chemicals in agriculture as an ecologically safe alternative, but to optimize the application of PGPR, more profound knowledge on specific gene regulation and molecular mechanisms of interaction with plants is needed. Advance in sequencing technologies made it affordable to compare transcriptom profiles of relative organisms to check to which extend PGPR strains or closely related species differ in their strategies of plant colonization. This work aimed at analysis of gene regulation in a biotechnological strain Bacillus atrophaeus UCMB-5137 to compare it with the gene expression profile of a generally recognized PGPR strain B. amyloliquefaciens FZB42. It was found out that despite the close taxonomic relatedness, these two organisms developed ability to colonize plants independently and use different strategies of plant colonization. Root exudate has triggered in UCMB-5137 alteration in expression in many genes controlled by stress response transcription factors (TF) SigB and SigD, while SigF, SigH, SigW, CcpA and several other TFs regulated genes associated with quorum sensing and biofilm formation, and adjusted the carbohydrate metabolism. Counting to peculiarities of gene regulation in different PGPR strains will allow optimization of their practical application.

overall_design: Examination of 2 experimental samples grown on liquid medium with maize root exudate and 3 control samples

first: Liberata

last: Mwita


title: Multiplex Single Cell Profiling of Chromatin Accessibility by Combinatorial Cellular Indexing

first: Darren

last: Cusanovich

summary:
Technical advances have enabled the collection of genome and transcriptome data sets with single-cell resolution. However, single-cell characterization of the epigenome has remained challenging. Furthermore, because cells must be physically separated prior to biochemical processing, conventional single-cell preparatory methods scale linearly. We applied combinatorial cellular indexing to measure chromatin accessibility in thousands of single cells per assay, circumventing the need for compartmentalization of individual cells. We report chromatin accessibility profiles from over 15,000 single cells and use these data to cluster cells on the basis of chromatin accessibility landscapes. We identify modules of coordinately regulated chromatin accessibility at the level of single cells both between and within cell types, with a scalable method that may accelerate progress toward a human cell atlas.

This SuperSeries is composed of the SubSeries listed below.