title: Development of a microarray for Enchytraeus albidus (Oligochaeta): Preliminary tool with diverse applications

status: Public on Mar 30 2011

city: Aveiro

first: Mónica

last: Amorim

address: Campus Universitário de Santiago, Department of Biology, University of Aveiro

department: Biology

country: Portugal

institute: CESAM & University of Aveiro

summary:
Standard bioassays allow hazard assessment at the population level, but much remains to be learned about the molecular level
response of organisms to stressors. The main aim of this study was the development of a DNA microarray for Enchytraeus albidus, a
common soil worm species. Further, this microarray was tested using worms exposed to Cu, phenmedipham, and different soil types.
Hybridization onto the developed microarray revealed several genes with homology to known sequences. Genes of interest were
confirmed through real-time polymerase chain reaction. It was possible to discriminate between natural and chemical stressors
and chemical concentrations. Gene responses were detected under conditions known to have effects in the reproduction of individuals.
It was confirmed that the integration of different endpoints improves the assessment process and enhances the understanding of the
modes of action of stressors. The chemical stress–induced genes were related to factors such as immune response, stress response,
metabolic processes, and/or signal transduction. The present study represents the first step of a gene-level study in the ecologically
relevant and standard test species E. albidus. It demonstrates the usefulness of cDNA normalization in the production of cDNA
libraries of ecotoxicological standard organisms that are not genome models like E. albidus.

overall_design: Fluorescently labelled cDNA, from enchytraeids exposed during 2 days to control LUFA 2.2 (Cy3) soil and to the different exposure conditions (Cy5), was synthesized for microarray analysis and hybridizations were performed. After scanning,  spots were identified and ratios quantified using the Genepix software 5.0 (Axon Instruments). Statistical analysis of the microarrays was performed using limmaGUI package (1.18.0) (Smyth, 2005) in the R (2.8.0) software environment (http://www.R-project.org/). After being submitted to local background subtraction, microarrays were normalized using global loess method. To statistically evaluate the differential gene expression between the different conditions, a gene-per-gene linear model (limma – linear model for microarray analysis) and empirical Bayes methods were applied. The results were then corrected for multiple testing using the Benjamini-Hochberg’s method (adjusted p<0.05 was considered significant) (Benjamini and Hochberg, 1995).

last: Novais

first: Karlij

last: Van Der Ven

first: Tine

last: Vandenbrouck

first: Amadeu

last: Soares

first: Wim

last: De Coen


title: Genetic deletion of P2X7 receptors causes genome-wide alteration of gene expression and results in antidepressant phenotype in mice amygdale

status: Public on Mar 31 2011

city: Budapest

first: Cecilia

last: Katalin

address: Szigony 43

department: Pharmacology

country: Hungary

institute: IEM HAS

summary: The purpose of the study was to explore the role of P2X7 receptors (P2X7R) in mood related behavior we have employed whole genome microarray expression profiling as a discovery platform.To identify genes affected by the genetic deletion of P2X7 receptors in the mouse amygdala we performed gene expression microarray analysis. For validation of differentially expressed genes, we performed TaqMan real-time RT-PCR.Our microarray studies have identified 7217 transcripts that were significantly affected by the deficiency of P2X7 receptor. Validation experiments revealed that among these genes at least 30 genes could be associated to synaptic signaling, neuroplasticity and thereby to the pathogenesis of depression (e.g. beta and gamma subunits of GABAA receptor, ionotropic glutamate receptors, α2 adrenergic receptor, which were down regulated in KO mice). 287 transcripts were found be more than two-fold over-expressed in the LPS treated groups which can be classified according to their inflammatory biological function: Il4ra, Saa1, chemokine ligands.

overall_design: Intraperitoneal injection of LPS induced alteration of gene expression in mouse amygdala was measured at 6 hours after exposure to doses of 250µg/kg.  Four independent experiments were performed for each of the four experimental groups (wild type mice and P2X7R -/- KO and saline treatment and LPS treatment).

first: Cecília

last: Csölle

first: Beáta

last: Sperlágh


title: 3CB-induced gene expression of Pseudomonas putida KT2440/pSL1 in soil (with RNA amplification)

status: Public on Mar 29 2011

city: Tsukuba

first: yong

last: wang

state: Ibaraki

address: 3-1-3 Kannondai

department: Environmental Biofunction Division

country: Japan

institute: National Institute for Agro-Environmental Sciences

summary: Genome-wide scanning of gene expression by microarray techniques was successfully performed on RNA extracted from a sterilized soil inoculated with Pseudomonas putida KT2440/pSL1, which contains a chloroaromatic degrading plasmid, in the presence or absence of 3-chlorobenzoic acid (3CB). The genes showing significant changes in their expression in both triplicate microarray analyses using amplified RNA and single microarray analysis using unamplified RNA were investigated. Pathway analysis revealed that the benzoate degradation pathway underwent the most significant changes following treatment with 3CB. Analysis based on categorization of differentially expressed genes against 3CB revealed new findings about the cellular responses of the bacteria to 3CB, including upregulation of the genes specifically involved in transport of 3CB, and induction of a K+/H+ antiporter complex, an universal stress protein, two cytochrome P450 proteins and an efflux transporter. Downregulated expression of some genes involved in carbon metabolism and the genes belong to a prophage in the presence of 3CB was observed. This study demonstrated the applicability of the method of soil RNA extraction for microarray analysis through a proof-of-concept experiment using a sterilized soil inoculated with Pseudomonas putida KT2440/pSL1.

overall_design: A study using total RNA extracted from soil cultures of Pseudomonas putida KT2440/pSL1.  Each chip measures the expression level of 5,341 genes from the Pseudomonas putida KT2440 genome with two sets of six 60-mer probes per gene.

first: Yong

last: Wang

first: Sho

last: Morimoto

first: Naoto

last: Ogawa

first: Takeshi

last: Fujii


title: Chromosome wide analysis of parental allele specific chromatin and DNA methylation along mouse distal chr15

city: Duarte

first: Xiwei

last: Wu

state: CA

address: 1500 E. Duarte Rd.

department: Computational and Quantitative Medicine

country: USA

institute: City of Hope National Medical Center

type: Genome binding/occupancy profiling by genome tiling array

type: Methylation profiling by genome tiling array

summary:
Imprinted genes are monoallelically expressed according to parental inheritance. The maternally and paternally inherited alleles are distinguished epigenetically by DNA methylation and histone modifications.
Chromosome-wide Chromatin immunoprecipitation (ChIP) and MIRA analysis of  MatDup.dist7 and PatDup.dist7 MEFs provided a panoramic map of reciprocal allele-specific histone modifications and DNA methylation at imprinted genes along distal chromosome 7 and 15.

overall_design: ChIP-chip and MIRA-chip was done to map histone modifications and DNA methylation along distal chr15 in the maternal allele and paternal allele in Patdup.dist7=MatDup.dist15 and Patdup.dist7=MatDupdis15 MEFs, respectively, using Nimblegen tiling arrays for chr15.

first: Purnima

last: Singh

first: Piroska

middle: E

last: Szabo


title: Expression of the genetically engineered E. coli in response to histidine starvation

first: Bei-Wen

last: Ying

address: 1-1-1 Tennoudai

institute: University of Tsukuba

type: Expression profiling by genome tiling array

summary:
Stochastic switching of a monostable circuit mediated the adaptation of the engineered OSU12-hisC Escherichia coli strain to histidine starvation. In this strain, the hisC gene was deleted from the His operon and placed under the control of a monostable foreign promoter. In response to histidine depletion, the OSU12-hisC population shifted to a higher HisC expression level, which are beneficial under starving conditions but are not favoured by the monostable circuit.
Microarray analysis indicated that OSU12-hisC reorganised its transcriptome to reach the appropriate physiological state upon starvation. The study suggests that bacteria do not necessarily need to evolve signalling mechanisms to control gene expression appropriately, even for essential genes.

overall_design: The hisC rewired strain and the control strain exponetially grown in the presence and absence of histidine were collected for the transcriptional analysis. Every 3 biological replications for each condition were performed.

first: Nao

last: Yasuda

first: Shingo

last: Suzuki

first: Tetsuya

last: Yomo


title: Massive decrease of maternal polyA mRNA upon post-ovulatory aging in Xenopus tropicalis

city: Essen

first: Ludger

last: Klein-Hitpass

address: Virchowstr. 173

department: Universitaetsklinikum

country: Germany

institute: Institut fuer Zellbiologie

summary: We used microarrays to detail the global gene expression changes following aging of oocytes

overall_design: We compared fresh oocytes with post-ovulatory aged oocytes  by microarray analysis

first: L

first: A

last: Kosubek


title: 5' Cap-Isolated RNA Pol II Transcripts and ENCODE Tiling Arrays to Identify Gene Expression Changes in Hepatitis C Liver

city: SaltLake City

first: Christopher

middle: Ian

last: Maxwell

state: UT

department: Internal Medicine / Gastroenterology

institute: University of Utah

summary: Recent studies of the human transcriptome suggest that many transcripts have short or lack 3' poly(A) ends. In addition, the length of 3' poly(A) ends may vary for some transcript to regulate mRNA translation. Heterogeneous lengths of poly(A) ends may introduce variations in measuring RNA transcripts using current approaches. We hypothesized that affinity purifying Pol II RNAs by their unique 5' m7GpppN cap regardless of their polyadenylation status would add additional information to genomic expression analyses and possibly overcome the variation in signals sometimes observed with analysis of poly(A) selected mRNA. 5' capped and 3' poly(A) RNA were isolated from normal and hepatitis C cirrhotic (HCV) human liver using a high-affinity variant of eIF4E and standard oligo(dT) methods, respectively. Both populations of RNA were analyzed using ENCODE tiling arrays representing 1% of the genome. A total of 64 annotated genes were significantly increased in HCV as compared to normal liver; 27 of these genes were identified by analyzing 5' capped RNA. A total of 31 annotated genes were significantly decreased; 16 of these were identified by analyzing 5' capped RNA. Bioinformatic analysis showed that 5' capped RNA provided a more extensive expression profile of intronic regions and identified Pol II transcriptionally active regions in unannotated regions of the genome that were differentially expressed in normal and disease states. The analysis of 5' capped RNA isolated from biospecimens provides additional gene expression information and identifies novel Pol II transcripts that are differentially expressed. This approach may also be useful in studying of RNAs regulated by 3' polyadenylation or RNA degradation.

overall_design: Four-condition experiment: 5'Cap and 3'Poly(A) isolation in both Normal and HCV Cirrhotic liver. Biological samples: 1 experimental, 1 control. Experimental replicates: 4 of each sample, for a total of 16 samples.

first: Milan

last: Folkers

first: Don

middle: A

last: Delker

middle: I

first: Cassie

last: Nelson

first: David

last: Nix

first: Curt

middle: H

last: Hagedorn


title: Expression analysis of Shigella sonnei MS390 delta-rodZ mutant

city: Shinjuku

first: Jiro

last: Mitobe

state: Tokyo

address: 1-23-1 Toyama

department: Bacteriology

institute: National Institute of Infectious Diseases

summary:
Expression of type III secretion system (TTSS), a major determinant of virulence in Shigella, is markedly inhibited in a deletion mutant (cpxA) of the CpxAR two-component sensor at the level of post-transcriptional processing of the second TTSS regulator, InvE. A novel mutant of rodZ, which encodes a cytoskeletal protein involved in maintaining the rod-shaped morphology of bacilli, was isolated in a screen for mutations that restored TTSS expression in a cpxA mutant. The rodZ mutants of Shigella sonnei efficiently expressed InvE at 30°C, a temperature at which expression is normally repressed through temperature-dependent post-transcriptional regulation. Consistent with a marked increase in invE mRNA stability in the rodZ mutant, purified RodZ strongly bound to an invE RNA probe. Functional domain mapping indicated that the RNA binding activity of RodZ was dependent on a short basic region (KRRKKR) and multimer formation. Electron microscopy revealed that RodZ multimers formed filamentous superstructures, similar to another bacterial cytoskeletal protein, MreB. Our results indicate that RodZ functions as a novel membrane-bound RNA binding protein that provides a scaffold for post-transcriptional regulation.
Phenotypic analysis of deletion mutant for RNA binding protein RodZ. The mutants analyzed in this study are further described in Jiro Mitobe, Itaru Yanagihara, Kiyouhisa Ohnishi, Akira Ishihama and Haruo Watanabe 2010. Bacterial cytoskeleton RodZ regulates post-transcriptional processing of Shigella type III secretion system. EMBO-J ( in submission).

overall_design: Four samples. Wild-type Shigella sonnei strain and the deletion mutant of rodZ gene, with or without rifampicin treatment for kinetics of mRNA stability.

first: Itaru

last: Yanagihara

first: Akira

last: Ishihama

first: Kiyouhisa

last: Ohnishi

first: Haruo

last: Watanabe


title: (1) Murine CD4 T cells: naïve vs peptide treated and naïve vs peptide+LPS treated. (2) Murine CD8 T cells naïve vs HY-specific cells from tolerant mice and naïve vs cells from rejecting mice

status: Public on Mar 28 2011

city: London

first: Diane

middle: Marylin

last: Scott

address: Du Cane Road

department: Immunobiology

country: United Kingdom

institute: Imperial College London

summary:
(1) Transcriptional profiling of mouse CD4 T cells comparing naïve HY-specific TCR transgenic T cells with the same cells recovered after adoptive transfer into WT female C57BL/6 mice on day 5 following treatment with HY peptide (100microgrammes intranasally days 1,2 and 3) and comparing naïve HY-specific TCR transgenic T cells with the same cells recovered after adoptive transfer into WT female C57BL/6 mice on day 5 following treatment with HY peptide plus LPS (100microgrammes peptide and 20microgrammes LPS intranasally day 1)
(2)Transcriptional profiling of mouse CD8 T cells comparing naïve CD8 T cells with HY-tetramer positive cells recovered from tolerant peptide treated (see above) WT female C57BL/6 mice on day 22 after male skin graft or 10-15 days after treatment with male spleen cells and comparing naïve CD8 T cells with HY-tetramer positive cells recovered from untreated (rejecting) WT female C57BL/6 mice on day 22 after male skin graft or 10-15 days after treatment with male spleen cells.

overall_design: Two condition experiment comapring naïve cells with cells from tolerant or immunised mice - both CD4 and CD8 T cells. Biological replicates: CD4s: Naïve, 4 replicates from pools of at least 3 mice; peptide-treated 3 replicates from pools of cells from 18 mice; peptide+PLS-treated 4 replicated from pools of cells from 15 mice. CD8s: Naïve, 3 replicates from pools of at least 3 mice; peptide-treated tolerant, 4 replicates from pools of 14 mice; untreated rejecting, 3 replicates from pools of 10 mice.

first: Katy

last: Derbyshire

first: Caroline

last: Addey

first: Jason

last: Bubier

first: Daniel

middle: J

last: Shaffer

first: Derry

middle: C

last: Roopenian


title: PADI4 ChIP-chip from MCF-7 cells

city: Bronx

first: Matthew

middle: Jon

last: Gamble

state: NY

address: 1300 Morris Park Ave / Golding 203

department: Molecular Pharmacology

institute: Albert Einstein College of Medicine

summary: Peptidylarginine deiminase IV (PADI4) catalyzes the conversion of positively charged arginine and methylarginine residues to neutrally charged citrulline residues on histone tails. This activity has been linked to the repression of gene transcription on a limited number of genes. To broaden our knowledge of the regulatory potential of PADI4, we utilized chromatin immunoprecipitation coupled with promoter tiling array (ChIP-chip) to more comprehensively investigate the range of PADI4 target genes across the genome in MCF-7 cells. Results showed that PADI4 is enriched in the gene promoter regions near the transcription start sites (TSSs) and, surprisingly, this pattern of binding is primarily associated with actively transcribed genes. Computational analysis found Elk-1, a member of the ETS oncogene family, to be highly enriched around PADI4 binding sites and coimmunoprecipitation analysis then confirmed that Elk-1 physically associates with PADI4. The expression of two well characterized Elk-1 target genes, c-fos and egr-1, was then found to be inhibited following treatment of MCF-7 cells with a PADI4 specific inhibitor. The inhibitor also significantly reduced levels of acetylation at H4 lysine 5 at these promoters suggesting that the activating function of PADI4 at these target genes is mediated, in part, by interplay between histone citrullination and HAT-mediated acetylation. These findings greatly expand our knowledge of the role of PADI4 in gene regulation by defining a new role for PADI4 catalyzed histone citrullination in mediating gene transactivation.

overall_design: Two PADI4 ChIP-chip biological replicates from MCF-7 human breast cancer cells are included.

first: Xuesen

last: Zhang

first: Lee

last: Kraus

first: Scott

last: Coonrod


title: Transcriptomic analysis of C. jejuni grown in anaerobic jar with air mix and in Modular Atmospheric Controlled System

status: Public on Apr 01 2011

city: Leicestershire

first: Amy

last: John

address: Loughborough

institute: University of Nottingham

summary:
Transcriptomic analysis of C. jejuni HPC5 grown in anaerobic jar supplemented with air mix and in Modular Atmospheric Controlled System (MAC).
C. jejuni HPC5 is a Campylobacter strain isolated from broiler chickens. Majority of the Campylobacter strains are cultured in an atmosphere containing 5-10% (v/v) oxygen or 5% hydrogen and 5-10% (v/v) carbon dioxide and the rest with nitrogen. To set up a jar (3.5L, Oxoid) to the required microaerophilic conditions, a vacuum is created to -22 psi and then air mixture of 85% v/v nitrogen, 10% v/v carbon dioxide and 5% v/v hydrogen (Air Products, Crewe, UK) is introduced into the jar. This resulted in a microaerobic atmosphere containing approximately 5.6% oxygen, 3.6% hydrogen, 7.3% carbon dioxide and 83% nitrogen. The Modular Atmospheric Controlled System (MAC; Don Whitely Scientific) is an anaerobic chamber where air mixture of 85% nitrogen, 10% carbon dioxide and 5% oxygen is used for growing Campylobacter. Differences in growth pattern were observed when Campylobacters were grown in it and transcriptomic analysis was done to prove the difference in the regulation of genes in the two conditions.

platform_organism: Campylobacter jejuni subsp. jejuni NCTC 11168 = ATCC 700819

overall_design:
There are three biological replicates for each experiment. Seven independent cDNA preps of C. jejuni HPC5 were prepared and they were labelled independently using AF555 (Fisher, UK). The labelled cDNAs were mixed together and this acted as the control. C. jejuni HPC5 was grown both in the Jar and in the MAC and their cDNAs were labelled independently with AF645 (Fisher, UK). Hybridisations were done between C. jejuni HPC5 grown in jar and in MAC in triplicates.
The supplementary file (linked at the foot of this record) represents the averaged normalised values for each experimental condition (3replicates/experimental condition).

first: Andrew

last: Timms

first: Philippa

last: Connerton

first: Ian


title: Genome-wide analysis of H3K27me3 and H3K4me3 deposition in the fie mutant (h3k4me3_wt_col_seedlings_db)

city: Lyon

first: François

last: ROUDIER

address: 46 Allée d'Italie

department: Biologie

country: France

institute: Ecole Normale Supérieure de Lyon

summary:
a2e_fie - h3k4me3_wt_col_seedlings_db - Epigenomic mapping in fie - Genome-wide epigenomic mapping

Keywords: chip-chip

overall_design: 4 dye-swap - CATMA arrays

first: Vincent

last: Colot

first: Francois

last: Roudier

first: Marie-Laure

last: Martin-Magniette


title: Genome-wide analysis of H3K27me3 and H3K4me3 deposition in the fie mutant (h3k4me3_fie_col_seedlings_db)

summary:
a2e_fie - h3k4me3_fie_col_seedlings_db - Epigenomic mapping in fie - Genome-wide epigenomic mapping

Keywords: chip-chip


title: MicroRNA array analysis during the regenerative peak following rat liver resection

city: Berlin

first: Nathanael

last: Raschzok

address: Augustenburger Platz 1

department: Department of Surgery; Campus Charite Mitte / Campus Virchow Klinikum

institute: Charité - Universitätsmedizin Berlin

summary: The liver has the unique capacity to regenerate after surgical resection. However, the regulation of liver regeneration is not completely understood. We performed miRNA microarray analyses of liver tissue from Wistar rats at different time points after 70% partial hepatectomy (0, 2, 6, 12, 24, 48 hours, and 5 days) and after sham laparotomy (12, 24, and 48 hours). We demonstrate that miRNA expression patterns changed during liver regeneration and that these changes were most evident during the peak of DNA replication at 24 hours after resection. Expression of 13 miRNAs, including five members of the let-7 family, was significantly reduced 12-48 hours after resection, whereas 3 miRNAs were significantly upregulated (> 25% change). We provide a temporal miRNA expression dataset of the regenerating rat liver, which indicates a primary function for miRNA during the peak of DNA replication. These data will assist further functional studies on the role of miRNAs during liver regeneration.

overall_design: This project analyzed the miRNA expression in 30 different sample types of the organism rattus norvegicus. The miRNA expression was compared at 7 different growth time points after liver resection (0, 2, 6, 12, 24, 48 hours, 5 days) and at 3 different time points after sham laparotomy (12, 24, and 48 hours). Three biological replicates were used per time point.

first: Wiebke

last: Werner

first: Hannes

last: Sallmon

first: Nils

last: Billecke

first: Christof

last: Dame

first: Peter

last: Neuhaus

first: Igor

middle: M

last: Sauer


title: Expression data of Medicago truncatula Jemalong A17 roots treated with auxin transport inhibitors

status: Public on Mar 26 2011

city: Stanford

first: Adriana

middle: Parra

last: Rightmyer

address: 371 Serra Mall

institute: Stanford University

summary: Rhizobium and allied bacteria form symbiotic nitrogen-fixing nodules on legume roots. Plant hormones appear to play a role in nodule formation. We treated Medicago truncatula roots with auxin transport inhibitors (ATIs) N-(1-naphthyl)phthalamic acid (NPA) and 2,3,5-triiodobenzoic acid (TIBA) to induce the formation of pseudonodules. We compared the transcriptional responses of M. truncatula roots treated with ATIs to roots inoculated with Sinorhizobium meliloti. The transcriptional response of M. truncatula roots 1 and 7 days after ATI treatment were opposite to roots treated with S. meliloti.

platform_organism:
Sinorhizobium meliloti
Medicago sativa
Medicago truncatula

overall_design: Three independent biological replicates were performed at each time point (1 and 7 days after treatment) for each treatment (buffer and ATI).

middle: P

first: Sharon

last: Long


title: Effect of clp1 induction on b-dependent gene regulation

city: Karlsruhe

first: Jörg

middle: Thomas

last: Kämper

address: Hertzstrasse 16, Geb. 06.40

department: Department for Applied Biosciences

institute: Karlsruhe Institute of Technology

summary: The clp1 gene is a b-regulated gene that is required for clamp formation during the biotrophic growth phase of U. maydis. Clp1 expression blocks b-dependent filament formation and cell cycle arrest if co-induced with the b-heterodimer.

overall_design: Strains AB33 (control) and UMS84 were grown for the times indicated in array medium containing 3 g/l nitrate and 1% arabinose for the induction of nar1P- (bE1/bW2) and crg1P- (clp1) controlled gene expression. Samples were taken from two biological replicates each.

first: Mario

last: Scherer

first: Kai

last: Heimel

first: Joerg

last: Kaemper


title: Expression data from Pseudomonas putida KT2440 (Progenika oligo array data)

status: Public on Mar 25 2011

city: Hannover

first: Sarah

last: Frank

address: Carl-Neuberg-Str. 1

department: Clinic for Paediatric Pneumology and Neonatology

institute: Medical School Hannover

summary: We used Progenika oligonucleotide arrays to monitore the gene expression of P. putida after a cold shock from 30°C to 10°C

overall_design:
Culture was grown in minimal medium supplemented with succinate at 30°C. In mid-exp. phase cells were harvested for subsequent RNA extraction (standard condition) and medium was cooled down to 10°C. Two hrs after reaching the temperature, cells were again harvested for RNA extraction.
Data from four replicate hybridizations (two dye-swaps) were combined into a single ratio value. Individual normalized signal intensity VALUES for each channel of each array are provided in the FullNormalizedMatrix.txt file available on the Series record.


title: Genome-wide analysis of H3K27me3 and H3K4me3 deposition in the fie mutant (h3k27me3_wt_col_seedlings_db)

first: François

last: ROUDIER

summary:
a2e_fie - h3k27me3_wt_col_seedlings_db - Epigenomic mapping in fie - Genome-wide epigenomic mapping

Keywords: chip-chip

first: Vincent

last: Colot


title: Expression data from Pseudomonas putida KT2440 (Affymetrix data)

summary: We used Affymetrix microarrays to monitore the gene expression of P. putida after a cold shock from 30°C to 10°C

overall_design: Culture was grown in minimal medium supplemented with succinate at 30°C. In mid-exp. phase cells were harvested for subsequent RNA extraction (standard condition) and medium was cooled down to 10°C. Two hrs after reaching the temperature, cells were again harvested for RNA extraction.


title: Chromosome wide analysis of parental allele specific chromatin and DNA methylation along mouse distal chr7

first: Xiwei

last: Wu

overall_design: ChIP-chip and MIRA-chip was done to map histone modifications and DNA methylation along distal chr7 in the maternal allele and paternal allele in Matdup.dist7 and Patdup.dist7 MEFs, respectively, using Nimblegen tiling arrays for distal chr7.

first: Purnima

last: Singh

first: Piroska

last: Szabo


title: Incomplete X-inactivation initiated by a hypomorphic Xist allele in the mouse

first: Rieko

last: Ikeda

address: 3-1-1 Koyadai

department: BioResource Center

institute: RIKEN

summary: Mammals have evolved a mechanism to compensate for the difference in the number of X chromosome genes between XX females and XY males by inactivating one of the two X chromosomes in females. This X chromosome inactivation phenomenon is triggered by Xist gene, which is essential for X-inactivation to occur in cis. We demonstrate that an Xist mutant, XistIVS, is unique in that it retains a partial function to silence the X chromosome, which would provide an opportunity to dissect the molecular mechanism of Xist RNA-mediated chromosome silencing.

overall_design: To study the effect of XistIVS mutation on silencing of X-linked genes, global expression levels of X-linked genes were measured in mutant and control cells. We used female trophoblast stem (TS) cells carrying XistIVS mutation and gene expression was assayed using microarray containing 572 probes corresponding most of the X-linked genes. Then the result was compared with that obtained from wild-type control cells.

first: Yuko

last: Hoki

first: Nathan

last: Mise

first: Yuka

last: Sakata

first: Tatsuya

last: Ohhata

first: Hiroyuki

last: Sasaki

first: Kuniya

last: Abe

first: Takashi

last: Sado


title: High resolution temporal profiling of gene expression during Arabidopsis leaf development (senescence process)

city: Norwich

first: Jonathan

middle: David

last: Moore

address: Norwich Research Park

department: Platforms and Pipelines

institute: The Genome Analysis Centre

summary: Leaf senescence is an essential developmental process that involves altered regulation of thousands of genes and changes in many metabolic and signaling pathways resulting in massive physiological and structural changes in the leaf. The regulation of senescence is complex and although several senescence regulatory genes have been identified and characterized there is little information on how these individual regulators function globally in the control of the process. In this paper we use microarray analysis to obtain a high-resolution time course profile of gene expression during development of a single leaf over a three week period from just before full expansion to senescence. The multiple time points enable the use of highly informative clustering tools to reveal distinct time points at which signaling and metabolic pathways change during senescence. Analysis of motif enrichment in co-regulated gene clusters identifies clear groups of transcription factors active at different stages of leaf development and senescence.

overall_design: A novel experimental design strategy (A Mead et al, in preparation), based on the principle of the “loop design”, was developed to enable efficient extraction of information about key sample comparisons using a two-colour hybridisation experimental system. With 88 distinct samples (four biological replicates at each of 22 time points) to be compared, the experimental design included 176 two-colour microarray slides, allowing four technical replicates of each sample to be observed. Half of the slides were devoted to assessment of changes in gene expression between time points, using a simple loop design to link 11 samples from either the 7h time points or the 14h time points across the 11 sampling days, directly comparing samples collected on adjacent sampling days (i.e. 19 DAS with 21 DAS, 27 DAS with 29 DAS, etc.), and directly comparing the samples collected at 39 DAS with those collected at 19 DAS. Four separate loops were constructed for the 7h time points and for the 14h time points, using the arbitrary biological replicate labelling to identify the samples to be included in each loop. The remaining slides provided assessment of differences between the 7h and 14h samples and between the arbitrarily labelled biological replicates, with some further assessment of changes between sampling days. All direct comparisons (pairs of samples hybridised together on a slide) were between 7h and 14h samples collected on adjacent sampling days (i.e. 19 DAS with 21 DAS, etc.), including comparisons between samples collected at 39 DAS and at 19 DAS, and different arbitrarily labelled biological replicates.  These 88 comparisons formed a single loop connecting all 88 treatments, therefore ensuring that the design was fully connected (allowing each sample to be compared with every other sample).

first: Emily

last: Breeze

first: Ellizabeth

last: Harrison

first: Stuart

last: McHattie

first: Linda

last: Hughes

first: Richard

last: Hickman

first: Claire

last: Hill

first: Steven

last: Kiddle

first: Youn-sung

last: Kim

first: Christopher

last: Penfold

first: Dafyd

last: Jenkins

first: Cunjin

last: Zhang

first: Karl

last: Morris

first: Carol

last: Jenner

first: Stephen

last: Jackson

first: Brian

last: Thomas

first: Alex

last: Tabrett

first: Roxane

last: Legaie

middle: D

first: David

middle: L

last: Wild

first: Sascha

last: Ott

last: Rand

first: Jim

last: Beynon

first: Katherine

last: Denby

first: Andrew

last: Mead

first: Vicky

last: Buchanan-Wollaston


title: Chromosome wide analysis of parental allele specific chromatin and DNA methylation along the central part of mouse chr7

first: Xiwei

last: Wu

summary:
Imprinted genes are monoallelically expressed according to parental inheritance. The maternally and paternally inherited alleles are distinguished epigenetically by DNA methylation and histone modifications.
Chromosome-wide Chromatin immunoprecipitation (ChIP) and MIRA analysis of  MatDup.dist7 and PatDup.dist7 MEFs provided a panoramic map of reciprocal allele-specific histone modifications and DNA methylation at imprinted genes along distal chromosome 7 and 15.

overall_design: ChIP-chip and MIRA-chip was done to map histone modifications and DNA methylation along central chr7 in the maternal allele and paternal allele in Matdup.dist7 and Patdup.dist7 MEFs, respectively, using Nimblegen tiling arrays for central chr7.

first: Purnima

last: Singh

first: Piroska

last: Szabo


title: MicroRNAs are Transported in Plasma and Delivered to Recipient Cells by High-Density Lipoproteins (Mouse HDL signatures from WT and LDLR-/- high fat diet)

status: Public on Mar 24 2011

city: Bethesda

first: Kasey

last: Vickers

state: MD

address: 10 Center Dr. Bldg. 10 8N222

department: National Heart, Lung and Blood Institute

institute: National Institutes of Health

summary:
Circulating microRNAs (miRNA) are relatively stable in plasma and are a new class of disease biomarkers. Here we present evidence that human high-density lipoprotein (HDL) transports endogenous miRNAs and delivers them to recipient cells with functional targeting capabilities. Highly-purified fractions of human HDL contain small RNAs, and the HDL-miRNA profile from normal subjects is significantly different than familial hypercholesterolemia subjects. miRNAs were demonstrated to associate with both native and reconstituted HDL particles, and reconstituted HDL injected into mice retrieved distinct miRNA profiles from normal and atherogenic models. Cellular export of miRNAs to HDL was demonstrated to be regulated by neutral sphingomyelinase. HDL-mediated delivery of miRNAs to recipient cells was demonstrated to be scavenger receptor BI-dependent. Furthermore, HDL delivery of both exogenous and endogenous miRNAs resulted in the direct targeting of mRNA reporters. Notably, HDL-miRNA from atherosclerotic subjects induced differential gene expression, with significant loss of conserved mRNA targets in cultured hepatocytes. Collectively, these observations suggest that HDL participates in a novel mechanism of intercellular communication involving the transport and delivery of miRNAs.
Mouse HDL signatures from WT and LDLR-/- high fat diet

overall_design: Profiled HDL miRNA Signatures from N=4 WT; n=5 LDLR-/- HF mice

middle: T

last: Palmisano

first: Bassem

last: Shoucri

first: Robert

last: Shamburek

first: Alan

last: Remaley


title: MicroRNAs are Transported in Plasma and Delivered to Recipient Cells by High-Density Lipoproteins (Human exosome, LDL, and HDL miRNA Signatures)

summary:
Circulating microRNAs (miRNA) are relatively stable in plasma and are a new class of disease biomarkers. Here we present evidence that human high-density lipoprotein (HDL) transports endogenous miRNAs and delivers them to recipient cells with functional targeting capabilities. Highly-purified fractions of human HDL contain small RNAs, and the HDL-miRNA profile from normal subjects is significantly different than familial hypercholesterolemia subjects. miRNAs were demonstrated to associate with both native and reconstituted HDL particles, and reconstituted HDL injected into mice retrieved distinct miRNA profiles from normal and atherogenic models. Cellular export of miRNAs to HDL was demonstrated to be regulated by neutral sphingomyelinase. HDL-mediated delivery of miRNAs to recipient cells was demonstrated to be scavenger receptor BI-dependent. Furthermore, HDL delivery of both exogenous and endogenous miRNAs resulted in the direct targeting of mRNA reporters. Notably, HDL-miRNA from atherosclerotic subjects induced differential gene expression, with significant loss of conserved mRNA targets in cultured hepatocytes. Collectively, these observations suggest that HDL participates in a novel mechanism of intercellular communication involving the transport and delivery of miRNAs.
Human exosome, LDL, and HDL miRNA Signatures

overall_design: Profiled miRNA signatures from exosome, LDL, HDL in same subject


title: MicroRNAs are Transported in Plasma and Delivered to Recipient Cells by High-Density Lipoproteins (microRNA signatures retrieved from rHDL injected into WT, ApoE-/- chow, ApoE-/-high fat diet)

summary:
Circulating microRNAs (miRNA) are relatively stable in plasma and are a new class of disease biomarkers. Here we present evidence that human high-density lipoprotein (HDL) transports endogenous miRNAs and delivers them to recipient cells with functional targeting capabilities. Highly-purified fractions of human HDL contain small RNAs, and the HDL-miRNA profile from normal subjects is significantly different than familial hypercholesterolemia subjects. miRNAs were demonstrated to associate with both native and reconstituted HDL particles, and reconstituted HDL injected into mice retrieved distinct miRNA profiles from normal and atherogenic models. Cellular export of miRNAs to HDL was demonstrated to be regulated by neutral sphingomyelinase. HDL-mediated delivery of miRNAs to recipient cells was demonstrated to be scavenger receptor BI-dependent. Furthermore, HDL delivery of both exogenous and endogenous miRNAs resulted in the direct targeting of mRNA reporters. Notably, HDL-miRNA from atherosclerotic subjects induced differential gene expression, with significant loss of conserved mRNA targets in cultured hepatocytes. Collectively, these observations suggest that HDL participates in a novel mechanism of intercellular communication involving the transport and delivery of miRNAs.
microRNA signatures retrieved from rHDL injected into WT, ApoE-/- chow, ApoE-/-high fat diet

overall_design: Profiled rHDL miRNA signatures from n=3 WT; n=3 ApoE-/- chow, n=4 ApoE-/- high fat mice


title: Gene expression in Delta Sub1 with and without 4NQ0

city: Gif-sur-Yvette cedex

first: Christine

last: Conesa

address: CEA Saclay

department: iBiTeC-S

institute: CEA

summary:
Microarrays analysis was undertaken to compare mRNA levels in exponentially growing wild-type and sub1Δ cells, or after 1 h treatment with 4NQO. After spot quantification and normalization, only few genes were found to be differentially expressed when the two strains were exponentially grown in rich medium. In contrast, the distribution of gene expression ratios was clearly different upon 4NQO treatment showing that the transcriptional response to DNA damages was affected in the absence of Sub1. Most of the differentially expressed genes were up-regulated in sub1Δ cells, suggesting a global repression effect of Sub1. About half of them were bound by Sub1 demonstrating that Sub1 occupancy correlates with gene regulation. A GoTermFinder analysis revealed an enrichment in genes implicated in cell cycle process for the down-regulated genes. The up-regulated genes were enriched in ribosome biogenesis and cell wall structure GO categories, confirming that most of the genes bound by Sub1 were up-regulated in sub1Δ cells upon 4NQO treatment. A third GO category corresponded to genes encoding proteins involved in amino acid metabolism whose expression is under the control of the Gcn4 transcriptional activator. This GO category could reflect an indirect effect since the GCN4 gene is bound by Sub1, up-regulated in sub1Δ cells and induced at the translational level in response to DNA-damaging agents.
Keywords: Transcriptomic Genomic Comparative

overall_design:
Cell culture and RNA extraction.

Wild-type (yPH500) or sub1Δ cells were grown to exponential phase (OD600 = 1) in YPD medium at 30°C and an aliquot of cells was collected to serve as the time-zero reference. After the addition of 4NQO (1 µg/µl), the cells were further incubated for 1h. The cells were harvested by centrifugation and resuspended in 50 mM Na acetate, pH 5.3, 10 mM EDTA. Total RNA was isolated by heating and freezing the cells in the presence of SDS and phenol as described (Schmitt et al., 1990).

Preparation of labeled cDNAs

Labeled cDNAs were synthesized from 20 to 30μg total RNA using an indirect labeling protocol adapted from P.Brown
(http://cmgm.stanford.edu/pbrown/protocols/aadUTPCouplingProcedure.htm) with the Superscript II reverse transcription kit (Gibco-BRL) and amino-allyl dUTP. lAmino-allyl modified DNA was purified on Microcon filters (Amicon) recovered with 20 µL of H2O and the DNA was lyophilized. The DNA pellet was resuspended in 9 µL of 100 mM sodium bicarbonate, pH 9.0. This sample DNA was used to dissolve a dry pellet of monofunctional NHS-ester Cy3 or Cy5 (1/16e of the quantity delivered from the mono-reactive dye pack PA23001 or PA25001 respectively from Amersham) and the mixture was incubated for 1 hour at room temperature in the dark. The coupling reaction of the Cy dyes to the amino-allyl dUTP was stopped by quenching with the addition of 4.5 µL of 4M hydroxylamine (Sigma) and the solution was further incubated for 15 min in the dark. Cy3-immunoprecipitated DNA and Cy5-WCE control DNA were mixed and unincorporated/quenched Cy dyes were removed using a Microcon YM-30 filter (Amicon/Millipore). DNA was recovered with 60 µL of TE and ethanol precipitated.

Construction of ORF-microarrays

S. cerevisiae open reading frames (ORF), reamplified using a pair of universal primers from 6144 full-length ORFs (1 ng) amplified from genomic DNA by Research Genetics. The PCR products were purified by ethanol precipitation, and their size and concentration were measured by agarose gel electrophoresis. 92.4% of the ORFs were correctly amplified and purified. The purified DNAs were spotted onto amino-silane coated glass slides (GAPS II, Corning) using an automated arrayer (MicroGrid II, BioRobotics) with a spotting success percentage higher than 96 %..

Prehybridization and hybridization.

Prehybridization and hybridization conditions were those described on P. Brown's web site (http://brownlab.stanford.edu/protocols.html) with few minor modifications. The labeled precipitated DNA samples were recovered in hybridization buffer (50% formamide, 5X Denhardt’s, 0.5% SDS, 7X SSPE, 10 µg herring sperm DNA) and hybridized to the DNA microarray in a Corning slide chamber at 42°C overnight. Arrays were washed at room temperature once with 0.1X SSC + 0.1% SDS, twice with 0.1X SSC, then dried by centrifugation.

Data analysis

Hybridized arrays were scanned using a GenePix 4000A scanner (Axon Instruments, Inc.) and fluorescence ratio measurements were determined with the GenePix Pro 6.0 software (Axon Instruments, Inc.). Array analyses were undertaken using the Limma package (Smyth, 2005) from the R/Bioconductor software (R-Development-Core-Team, 2007). Microarray spot intensities have been normalized by subtracting the background and using the LOWESS method (Cleveland, 1979) with the smooth parameter set to 0.33 as recommended (Quackenbush, 2002). Data from four technical replicates corresponding to two independent experiments were compiled. Normalized measures served to compute the ratios of Cy3/Cy5 intensity and the associated log2-transform (denoted log2-ratios) for each gene. To identify genes with a log2-ratio significantly different between the mutant and wild-type strains, p-values were calculated for each gene using a moderated t-test. The moderated t test applied here was based on an empirical Bayes analysis and was equivalent to shrinkage (or expansion) of the estimated sample variances towards a pooled estimate, resulting in a more stable inference.

first: A

last: Suleau

first: M

last: Michaut

first: JC

last: Aude

first: C


title: A direct role for cohesin in genome-wide gene regulation and Ecdysone-response in Drosophila salivary gland

status: Public on Mar 23 2011

city: Amsterdam

first: Bas

last: van Steensel

address: Plesmanlaan 121

department: division of Molecular Biology

country: Netherlands

institute: Netherlands Cancer Institute

summary: Developmental abnormalities observed in Cornelia de Lange Syndrome (CdLS) have been genetically linked to mutations in the cohesin machinery. These findings raise the possibility that cohesin, in addition to its canonical function of mediating sister chromatid cohesion, might also be involved in regulating gene expression. We report that cleavage of cohesin’s kleisin subunit in post-mitotic Drosophila salivary glands induces major changes (both up and down) in the transcript levels of many genes. Kinetic analyses of changes in transcript levels upon cohesin cleavage reveal that a subset of genes responds to cohesin cleavage within a few hours. In addition, cohesin binds to most of these loci, suggesting that cohesin is directly regulating their expression. Amongst these genes are several that are regulated by the steroid hormone Ecdysone. Transcripts at EcR and Eip74EF, which encode an Ecdysone Receptor and an Ecdysone-regulated transcription factor, respectively, decline ten-fold within four hours of cohesin cleavage. Cytological visualization of transcription at selected Ecdysone-responsive genes reveals that puffing at Eip74EF ceases within an hour or two of cohesin cleavage, long before any decline in EcR associated with this locus. We conclude that cohesin regulates expression of a distinct set of genes, including those mediating the Ecdysone response

overall_design: DamID experiments for Rad21 were performed on gDNA from drosophila salivary glands. Samples were hybridized to 380k NimbleGen arrays with 300 bp probe spacing. The experiment was done in duplicate in the reverse dye orientation.

first: Andrea

last: Pauli

first: Joke

middle: G

last: van Bemmel

first: Raquel

last: Oliveira

first: Takehiko

last: Itoh

first: Katsuhiko

last: Shirahige

first: Kim

last: Nasmyth


title: Role of TGF-β in the Differential Effect of Neisseria gonorrhoeae on the Generation of Th1, Th2, and Th17 Immune Responses

status: Public on Mar 22 2011

city: Buffalo

first: Yingru

last: Liu

address: 3435 Main Street

institute: University at Buffalo

summary: The immune response to Neisseria gonorrhoeae is poorly understood, but its extensive antigenic variability and resistance to complement are thought to allow it to evade destruction by the host’s immune defenses.  We propose that N. gonorrhoeae also avoids inducing protective immune responses in the first place.  We previously found that N. gonorrhoeae induces IL-17-dependent innate responses in mice and suppresses Th1/Th2-dependent adaptive responses in murine cells in vitro through the induction of TGF-β.  In this study using a murine model of vaginal gonococcal infection, mice treated with anti-TGF-β antibody during primary infection showed accelerated clearance of N. gonorrhoeae with incipient development of Th1 and Th2 responses and diminished Th17 responses in genital tract tissue.  Upon secondary reinfection, mice that had been treated with anti-TGF-β during primary infection showed anamnestic recall of both Th1 and Th2 responses, with the development of anti-gonococcal antibodies in serum and secretions, and enhanced resistance to reinfection.  In knockout mouse strains defective in Th1 or Th2 responses, accelerated clearance of primary infection due to anti-TGF-β treatment was dependent on Th1 but not Th2 activity, whereas resistance to secondary infection resulting from anti-TGF-β treatment during primary infection was due to both Th1- and Th2-dependent memory responses.  We propose that N. gonorrhoeae proactively elicits Th-17-driven innate responses that it can resist, and suppresses Th1/Th2-driven specific adaptive immunity that would protect the host.  Blockade of TGF-β reverses this pattern of host immune responsiveness and facilitates the emergence of protective anti-gonococcal immunity.

overall_design:
We only did microarray assay for wild-type mice with or without anti-TGF-b treatment.
Experiment A: Totally there are three groups: Sham-infected mice without treatment; N.gonorrhoeae-infected with control IgG treatment; N.gonorrhoeae-infected with anti-TGF-β treatment. For each group, two mice were studied. Total RNA from mouse vagina were analysed.
Experiment B: Totally there are three groups: Sham-reinfected mice without treatment; N.gonorrhoeae-reinfected with control IgG treatment; N.gonorrhoeae-reinfected with anti-TGF-β treatment. For each group, two mice were studied. Total RNA from mouse vagina were analysed.

first: Michael

middle: W

last: Russell


title: Microarray studies of darkness stress and bleaching in the Caribbean coral Montastraea faveolata

city: San Francisco

last: DeSalvo

department: Department of Anesthesia

institute: UC San Francisco

summary: Coral bleaching occurs in response to numerous abiotic stressors, the ecologically most relevant of which is hyperthermic stress due to increasing seawater temperatures. Bleaching events can span large geographic areas and are currently a potent threat to coral reefs worldwide. Much effort has been focused on understanding the molecular and cellular events underlying bleaching, and these studies have mainly utilized heat and light stress regimes. In an effort to determine whether different stressors share common bleaching mechanisms, we used cDNA microarrays for the corals Acropora palmata and Montastraea faveolata (containing > 10,000 features) to measure differential gene expression during darkness stress. This is the first coral microarray experiment aimed at darkness stress, and the first for these species to interrogate gene expression at such a large scale. Our results reveal a striking transcriptomic response to darkness in A. palmata involving chaperone and antioxidant up-regulation, growth arrest, and metabolic modifications. As these responses were also measured during thermal stress, our results suggest that different stressors may share common bleaching mechanisms. Furthermore, our results point to ER stress as a critical cellular event involved in darkness-specific (and possibly more general) molecular bleaching mechanisms. On the other hand, we identified a meager transcriptomic response to darkness in M. faveolata where gene expression differences between host colonies and/or sampling locations were greater than differences between control and stressed fragments. To this end, we discuss the importance of factors related to host genotype, Symbiodinium genotype, and the abiotic environment that influence host gene expression and thereby can hinder an investigator’s ability to measure gene expression during a condition of interest.

overall_design: We employed a reference design where all control and dark-stressed samples were compared to a pooled reference aRNA sample composed of aRNA from all fragments. Since all RNA samples were compared to the reference sample, direct comparisons of gene expression across all time points and conditions can be performed.

middle: K

first: Andres

last: Estrada

first: Shinichi

last: Sunagawa

first: Monica

last: Medina


title: Murine: papillary thyroid cancer vs. poorly differentiated thyroid cancer

city: New York

first: Yupu

last: Liang

address: 1230 York Avenue

institute: The Rockefeller University

summary: Mice with thyroid-specific expression of oncogenic BRAF (Tg-Braf) develop papillary thyroid cancers (PTC) that are locally invasive and have well-defined foci of poorly differentiated thyroid carcinoma (PDTC).  To investigate the PTC-PDTC progression, we performed a microarray analysis using RNA from paired samples of PDTC and PTC collected from the same animals by laser capture microdissection. Analysis of 8 paired samples revealed a profound deregulation of genes involved in cell adhesion and intracellular junctions, with changes consistent with an epithelial-mesenchymal transition (EMT). This was confirmed by IHC, as vimentin expression was increased and E-cadherin lost in PDTC compared to adjacent PTC. Moreover, PDTC stained positively for phospho-Smad2, suggesting a role for transforming growth factor-beta (TGFbeta) in mediating this process. Accordingly, TGFbeta induced EMT in primary cultures of thyroid cells from Tg-Braf mice, whereas wild-type thyroid cells retained their epithelial features. TGFbeta-induced Smad2 phosphorylation, transcriptional activity and induction of EMT required MAPK pathway activation in Tg-Braf thyrocytes. Hence, tumor initiation by oncogenic BRAF renders thyroid cells susceptible to TGFbeta-induced EMT, through a MAPK-dependent process.

overall_design: Comparing the transcription profiles of 8 pairs of murine poorly differentiated thyroid cancer and papillary thyroid cancer collected from the same animals by laser capture microdissection. Co-hybridizations were done in triplicate with a single dye flip.

first: Jeffrey

middle: A

last: Knauf

first: James

last: Fagin

first: Maureen

last: Sartor

first: Mario

last: Medvedovic


title: Genome-wide analysis of H3K27me3 and H3K4me3 deposition in the fie mutant (h3k27me3_fie_col_seedlings_db)

first: François

last: ROUDIER

summary:
a2e_fie - h3k27me3_fie_col_seedlings_db - Epigenomic mapping in fie - Genome-wide epigenomic mapping

Keywords: chip-chip

overall_design: 4 dye-swap - CATMA arrays

first: Vincent

last: Colot

first: Francois

last: Roudier

first: Marie-Laure

last: Martin-Magniette


title: Transcriptional response of Saccharomyces cerevisiae to high glucose (20%) concentration

city: Valencia

first: Marcel·lí

last: del Olmo

address: Dr. Moliner, 50

country: Spain

institute: Universitat de València

summary: Yeast cells can be affected during their growth to several stress conditions. One of the most known and characterised is the osmotic stress and most of the studies about osmotic sterss response in yeast have been focused on salt or sorbitol stress. However, during yeast growth in industrially relevant processes (for instance throughout alcoholic fermentation on the must to produce alcoholic beverages) the osmotic stress is mainly due to the high sugar(in particular glucose) concentration (200-250 g/L).

overall_design:
In this study we want to know the transcriptional response of the Saccharomyces cerevisiae when it was grown in a medium with high glucose concentration. For this aim we have grown yeast in YP medium containing 2% of glucose in cultures overnight and after that we diluted this cultures to an OD600 of 0.1 in two differents mediums: YP containing 2% or 20% of glucose.One hour later of inoculation we collect the cells and quikly frozen in liquid nitrogen.
We extracted the total mRNA of the cells and after that we did the microarrays, comparing cells were grown in YP2 media against the cells were grown in YP20 media.

first: Elena

last: Jiménez-Martí


title: MicroRNA programs in normal and aberrant stem and progenitor cells

status: Public on Mar 21 2011

first: Christopher

middle: Paul

last: Arnold

address: 269 Campus Drive, CCSR building, Room 3205B

department: Microbiology and Immunology

summary: Emerging evidence suggests that microRNAs (miRNAs), an abundant class of ~22-nt small regulatory RNAs, play key roles in controlling the post-transcriptional genetic programs in stem and progenitor cells. Here we systematically examined miRNA expression profiles in various adult tissue-specific stem cells and their differentiated counterparts.  These analyses revealed miRNA programs that are common or unique to blood, muscle, and neural stem cell populations and miRNA signatures that mark the transitions from self-renewing and quiescent stem cells to proliferative and differentiating progenitor cells. Moreover, we found a stem/progenitor transition miRNA (SPT-miRNA) signature that predicts the effects of genetic perturbations, such as loss of PTEN and the Rb family, AML-ETO expression, and MLL-AF10 transformation, on self-renewal, proliferation, and/or differentiation potentials of mutant stem/progenitor cells. More importantly, some SPT-miRNAs can control rate of self-renewal in embryonic stem cells and the reconstitution potential of hematopoietic stem cells (HSCs). Finally, we found that some SPT-miRNAs may coordinately regulate genes that are known to play roles in controlling HSC self-renewal, such as Hoxb6 and Hoxa4.  Together, these analyses revealed the miRNA programs that control key processes in normal and aberrant stem and progenitor cells, setting the foundations for dissecting post-transcriptional regulatory networks in stem cells.

platform_organism:
Homo sapiens
Mus musculus

overall_design: We used a multiplex protocol to amplify miRNAs from 20-1000 sorted stem and/or progenitor cells and then analyzed the expression of 425 mature miRNAs using TaqMan miRNA quantitative PCR analyses

first: Chris

first: Chang-Zheng

last: Chen


title: Comparative profiling of primary colorectal carcinomas and liver metastases

first: Jonathan

last: Pollack

address: 269 Campus Drive, CCSR3245A

department: Pathology

summary:
PURPOSE: We sought to identify genes of clinical significance to predict survival and the risk for colorectal liver metastasis (CLM), the most common site of metastasis from colorectal cancer (CRC).
PATIENTS AND METHODS: We profiled gene expression in 31 specimens from primary CRC and 32 unmatched specimens of CLM, and performed Significance Analysis of Microarrays (SAM) of the expressed genes between these two groups.  To characterize the clinical relevance of two highly-ranked differentially expressed signature genes, we analyzed the expression of secreted phosphoprotein 1 (SPP1 or osteopontin) and lymphoid enhancer factor-1 (LEF1) by immunohistochemistry using a tissue microarray (TMA) representing an independent set of 154 patients with primary CRC.
Supervised analysis using SAM identified 963 genes with significantly higher expression in CLM compared to primary CRC, with a false discovery rate of <0.5%.  TMA analysis showed SPP1 and LEF1 protein overexpression in 60% and 44% of CRC cases, respectively. Subsequent occurrence of CLM was significantly correlated with the overexpression of LEF1 (chi-square p = 0.035), but not SPP1 (p = 0.14).  Kaplan Meier analysis revealed significantly worse survival in patients with overexpression of LEF1 (p < 0.01), but not SPP1 (p = 0.11).  Both univariate and multivariate analyses identified stage (p < 0.0001) and LEF1 overexpression (p < 0.05) as important prognostic markers, but not tumor grade or SPP1.
CONCLUSION: Among signature genes differentially expressed between CLM and primary CRC, we demonstrate overexpression of LEF1 in primary CRC to be a prognostic factor for poor survival and increased risk for liver metastasis.

overall_design: cDNA microarrays from the Stanford Functional Genomics Facility were used to perform mRNA transcript profiling of 31 freshly-frozen primary colorectal cancer specimens, and compared to published profiles of 32 unmatched colorectal liver metastases and 12 normal liver specimens.

first: Mei-Sze

last: Chua

first: William

last: Yeh

first: Xin


title: MicroRNAs are Transported in Plasma and Delivered to Recipient Cells by High-Density Lipoproteins (Human HDL miRNA Signatures)

first: Kasey

last: Vickers

summary:
Circulating microRNAs (miRNA) are relatively stable in plasma and are a new class of disease biomarkers. Here we present evidence that human high-density lipoprotein (HDL) transports endogenous miRNAs and delivers them to recipient cells with functional targeting capabilities. Highly-purified fractions of human HDL contain small RNAs, and the HDL-miRNA profile from normal subjects is significantly different than familial hypercholesterolemia subjects. miRNAs were demonstrated to associate with both native and reconstituted HDL particles, and reconstituted HDL injected into mice retrieved distinct miRNA profiles from normal and atherogenic models. Cellular export of miRNAs to HDL was demonstrated to be regulated by neutral sphingomyelinase. HDL-mediated delivery of miRNAs to recipient cells was demonstrated to be scavenger receptor BI-dependent. Furthermore, HDL delivery of both exogenous and endogenous miRNAs resulted in the direct targeting of mRNA reporters. Notably, HDL-miRNA from atherosclerotic subjects induced differential gene expression, with significant loss of conserved mRNA targets in cultured hepatocytes. Collectively, these observations suggest that HDL participates in a novel mechanism of intercellular communication involving the transport and delivery of miRNAs.
Human HDL miRNA Signatures

overall_design: Profiled miRNA Signatures from N=6 Normal Human HDL; n=5 FH Human HDL

first: Brian

last: Palmisano

first: Bassem

last: Shoucri

first: Robert

last: Shamburek

first: Alan

last: Remaley


title: Microarray studies of darkness stress and bleaching in the Caribbean coral Acropora palmata

first: Michael

last: DeSalvo

summary: Coral bleaching occurs in response to numerous abiotic stressors, the ecologically most relevant of which is hyperthermic stress due to increasing seawater temperatures. Bleaching events can span large geographic areas and are currently a potent threat to coral reefs worldwide. Much effort has been focused on understanding the molecular and cellular events underlying bleaching, and these studies have mainly utilized heat and light stress regimes. In an effort to determine whether different stressors share common bleaching mechanisms, we used cDNA microarrays for the corals Acropora palmata and Montastraea faveolata (containing > 10,000 features) to measure differential gene expression during darkness stress. This is the first coral microarray experiment aimed at darkness stress, and the first for these species to interrogate gene expression at such a large scale. Our results reveal a striking transcriptomic response to darkness in A. palmata involving chaperone and antioxidant up-regulation, growth arrest, and metabolic modifications. As these responses were also measured during thermal stress, our results suggest that different stressors may share common bleaching mechanisms. Furthermore, our results point to ER stress as a critical cellular event involved in darkness-specific (and possibly more general) molecular bleaching mechanisms. On the other hand, we identified a meager transcriptomic response to darkness in M. faveolata, where gene expression differences between host colonies and/or sampling locations were greater than differences between control and stressed fragments. To this end, we discuss the importance of factors related to host genotype, Symbiodinium genotype, and the abiotic environment that influence host gene expression and thereby can hinder an investigator’s ability to measure gene expression during a condition of interest.

first: Andres

last: Estrada

first: Shinichi

last: Sunagawa

first: Monica

last: Medina


title: MicroRNAs are Transported in Plasma and Delivered to Recipient Cells by High-Density Lipoproteins (Human HDL and Exosome miRNA Signatures)

summary:
Circulating microRNAs (miRNA) are relatively stable in plasma and are a new class of disease biomarkers. Here we present evidence that human high-density lipoprotein (HDL) transports endogenous miRNAs and delivers them to recipient cells with functional targeting capabilities. Highly-purified fractions of human HDL contain small RNAs, and the HDL-miRNA profile from normal subjects is significantly different than familial hypercholesterolemia subjects. miRNAs were demonstrated to associate with both native and reconstituted HDL particles, and reconstituted HDL injected into mice retrieved distinct miRNA profiles from normal and atherogenic models. Cellular export of miRNAs to HDL was demonstrated to be regulated by neutral sphingomyelinase. HDL-mediated delivery of miRNAs to recipient cells was demonstrated to be scavenger receptor BI-dependent. Furthermore, HDL delivery of both exogenous and endogenous miRNAs resulted in the direct targeting of mRNA reporters. Notably, HDL-miRNA from atherosclerotic subjects induced differential gene expression, with significant loss of conserved mRNA targets in cultured hepatocytes. Collectively, these observations suggest that HDL participates in a novel mechanism of intercellular communication involving the transport and delivery of miRNAs.
Human HDL and Exosome miRNA Signatures

overall_design: Profiled miRNA signatures from HDL and exosome in matched subjects


title: ChIP-chip of siGFP- or siHOTTIP-treated foreskin fibroblasts with anti-H3K4me3, anti-H3K27me3, anti-H3K4me2, anti-histone, anti-MLL1, and anti-WDR5 antibodies on HOX tiling array

first: Kevin

last: Wang

address: 269 Campus Drive CCSR 2142

department: Dermatology

institute: Stanford University School of Medicine

summary:
Coordinate loss of MLL1 and WDR5 occupancy caused by HOTTIP knockdown were observed in distal HOXA genes. These regions correspondingly lost H3K4me3 and H3K4me2, without changes in pan-histone or H3K27me3.
human foreskin fibroblasts (CRL2091), anti-H3K4me3 (Abcam ab8580, lot#1016899), anti-H3K4me2 (Abcam ab32356, lot#947550), anti-H3K27me3 (Abcam ab4729, lot#1021724), anti-histone H3 (Abcam ab1791, lot#1025144), anti-MLL1 (gift of R. Roeder, Rockefeller University), and anti-WDR5 (gift of W. Herr, UNIL)

overall_design: Comparison of occupancy of MLL1 and WDR5 of siGFP and siHOTTIP foreskin fibroblasts on HOX tiling array. Human foreskin fibroblasts are transfected with siGFP or siHOTTIP for 72 hrs. The cells are harvesed and ChIP performed with H3K4me3, H4K27me3, H3K4me2, MLL1, WDR5, and histone antibodies.


title: Nuclear Scaffold Attachment Sites in HeLa Cells Across 1% of the Human Genome

city: Charlottesville

first: Mignon

last: Keaton

state: VA

address: 1340 Jefferson Park Ave, Rm 1232

department: Biochemistry and Molecular Genetics

institute: University of Virginia

summary: The nuclear scaffold, consisting of structural and functional nuclear proteins, remains after extraction of nuclei and anchors loops of DNA.  In the search for cis-elements functioning as chromatin domain boundaries, we mapped 453 nuclear scaffold attachment sites purified by lithium-3,5-iodosalicylate from HeLa cells across 30 Mb of the human genome studied by the ENCODE pilot project.  The scaffold attachment sites recovered mapped predominately near expressed genes and localized near transcription start sites and the ends of genes.

overall_design: Nuclear scaffold associated DNA was purfied from lithium-diiodosalicylate extracted HeLa nuclei by digestion with EcoRI/HindIII/HaeIII restriction enzymes.  Nuclear scaffold DNA from three independent biological replicates and total genomic DNA digested with EcoRI and HindIII were biotin labeled and hybridized to ENCODE01-F (P/N 900543; Affymetrix, Santa Clara, CA) tiling microarrays.  Hybridization data were analyzed using Model-based analysis tool (MAT) for tiling arrays (Johnson et al., 2006) and genomic positions (using the hg17 build (May 2004) of the Human genome assembly) with a statistically significant enrichment (P <10-3 within a 1-kb window) of scaffold signal as compared to the genomic control were reported as scaffold attachment regions.  These sites were then remapped to hg18 build of the genome using the UCSC Genome Browser liftover tool (http://genome.ucsc.edu) .  To account for potential gaps in the data due to repetitive sequences that were not spotted on the microarray, intervals from the MAT analysis that were less than 2501 bp apart were subsequently joined yielding 453 SARs.

first: Anindya

last: Dutta


title: Chip-chip from HeLa S3 expressing epitope-tagged H2Abbd, macroH2A and H2A histones: H2Abbd versus H2A, and macroH2A versus H2A, distributions on 13 regions of the HG18 assembly

status: Public on Mar 19 2011

city: Dublin

first: Ana

last: Barat

state: Dublin

address: Collins Avenue

department: School of Computing

country: Ireland

institute: Dublin City University

summary: The distribution of histone variants H2Abbd and macroH2A in 13 regions of the HG18 assembly have been studied using a variant of the ChIP-on-Chip technique. HeLa S3  cell lines  expressing tagged histones H2A, H2Abbd or macroHA were obtained using retroviral transfer. DNA fractions associated with tagged histones were isolated using a two-step purification procedure that involved affinity chromatography on a column with anti-FLAG antibodies, followed by affinity chromatography on a Ni-agarose column. The obtained genomic DNA samples were analyzed by hybridization with custom NimbleGene genomic microarrays.

overall_design: Two samples. Test sample 1 is HeLa S3 cells expressing epitope-tagged histone H2Abbd and test sample 2 is HeLa S3 cells expressing epitope-tagged histone macroH2A . The control for both test sample 1 and test sample 2 is HeLa S3 expressing epitope-tagged histone H2A. Two copies of each probe per array were made.

first: Elena

middle: S

last: Ioudinkova

first: Vasily

last: Ogrizko

first: Yegor

last: Vassetzky

first: Serghey

middle: V

last: Razin

first: Andrey

last: Pichugin

last: Markova

first: Mark

last: Lipinsky


title: Analysis of the human endothelial cell response to Borrelia burgdorferi wild-type and deleted in the gene encoding the integrin ligand P66 [experiment B]

status: Public on Mar 18 2011

city: Milwaukee

first: Jenifer

last: Coburn

state: WI

address: 8701 Watertown Plank Rd.

department: ID/CBID

institute: Medical College of Wisconsin

summary: The overall goal of these experiments was to determine how human cells respond to ontegrin binding by Borrelia burgdorferi. Lyme disease is the most common vector-borne infection in the Northern hemisphere, yet little is understood about the pathogenic mechanisms of the causative agent, Borrelia burgdorferi. Cells in culture were infected with the bacteria that do and do not express the beta3 integrin ligand P66, which are otherwise isogenic. As additional controls, uninfected cells were also included in the analyses.

overall_design:
The cell line used for primary data analyses provided here was Ea.hy926, a macrovascular line (Edgell, C. J.,et al. 1990. Endothelium specific Weibel-Palade bodies in a continuous human cell line, EA.hy926. In vitro Cell. & Dev. Biol. 26:1167-1172, and Edgell, C. J., et al. 1983. Permanent cell line expressing human factor VIII-related antigen established by hybridization. Proc. Natl. Acad. Sci. 80:3734-3737). Epithelial cells HEK-293 (which does not express b3 integrins) and HEK-293 transfected to express integrin αvβ3 were also analyzed. Infection times were 1 hour or 3 hours for cells lifted with EDTA and washed prior to bacterial infection. RNA was harvested and reverse transcribed. Labeled cDNAs were used to probe 2 color arrays.
In each of at least three biological replicate experiments, for each time point, three comparisons were made. First, cells infected with B. burgdorferi with a marker inserted adjacent to the p66 gene (p66+) were compared to the cells infected with B. burgdorferi with the p66 gene interrupted by the marker (Coburn, J. and Cugini, C. (2003) Targeted mutation of P66 of Borrelia burgdorferi disrupts attachment to integrin αvβ3. Proc. Natl. Acad. Sci. 100:7301-7306). Second, the p66+-infected cells were compared to the uninfected cells. Third, the p66- mutant-infected cells were compared to the uninfected cells. Arrays used were generated at the Tufts University core faculity using Operon's human v2.1.2 plus v2.1.4 cDNA library (Operon Technologies, Inc., Alameda, CA). Only data generated from spots that in all three experiments were of sufficient quality to allow interpretation are included.
IEa.hy926 endothelial cell lines, or HEK-293 epithelial cells, or transfected HEK-293 derivative expressing integrin avb3


title: Systematic identification of genes involved in embyonic chicken muscle development

city: Beijing

first: Tingting

last: Li

address: 38 Xueyuan Rd

department: Medical informatics

country: China

institute: Peking University

summary: The genetic closeness and divergent muscle growth rates of broilers and layers make them great models for myogenesis study. In order to discover the molecular mechanisms determining the divergent muscle growth rates and muscle fiber sizes in different chicken lines, we systematically identified differentially expressed genes between broilers and layers during muscle development (embyonic day 10, 12, 14 and 18) by microarray hybridization experiment.

overall_design: Time-course studies of two different intra-species breeds

first: Yong

last: Zhang

first: Dahai

last: Zhu


title: Long non-coding RNAs detected in Plasmodium falciparum malaria using high-resolution DNA tiling microarray technology.

status: Public on Mar 16 2011

city: Cambridge

first: Kate

middle: Mariel

last: Broadbent

state: MA

address: FAS Center for Systems Biology Northwest Building, room 469 52 Oxford Street

department: Systems Biology

institute: Harvard University

type: Non-coding RNA profiling by genome tiling array

summary:
Survey of transcriptional activity across 22.6% of the P. falciparum strain 3D7 genome.
872 protein coding genes and 60 putative P. falciparum lncRNAs identified under developmental regulation during the parasite's pathogenic human blood stage.

overall_design:
Total RNA was isolated from synchronous in vitro parasite cultures at 18, 24, 30, and 36 hours post invasion (hpi) of red blood cells.
1 μg of total RNA was then subjected to poly-A selective amplification using Message Amp II (Ambion), substituting biased dNTP/NTP mixes (2A/T/U:1C/G).
The resulting aRNA was labeled with Superscript II reverse transcriptase (Invitrogen), using random hexamers, and either Cy3- or Cy5-dUTPs (Amersham).

first: Daniel

last: Park

first: Ashley

last: Wolf

first: Pardis

last: Sabeti

first: John

last: Rinn


title: Effect of HIV infection on co-stimulatory capacity of dendritic cells

status: Public on Mar 15 2011

city: Pune

first: Meera

middle: Vir

last: Singh

state: Maharashtra

address: 73-G, MIDC, Bhosari

department: Immunology

country: India

institute: National AIDS Research Institute

summary: HIV uses dendritic cells as a carrier to infect its target CD4+ T cells. At the same time, it also hampers DCs function in terms of their T cell stimulatory capacity and cytokine secretion. We have shown that HIV causes reduction in the expression of co-stimulatory molecules, CD80 and CD86 by DCs at the mRNA level irrespective of the viral subtype or the mode in DC-HIV interaction. The microarray experiments were performed to understand the changes in the transcription factor profile of HIV infected DCs which may in turn lead to reduced CD80/CD86 expression as well as the effect of HIV infection on other co-stimulatory molecules on DCs. The data suggests that TFs NFKB1, NFKB2, REL and MYB may be responsible for the observed decrease in CD80 and CD86 expression and that HIV infection also hampers the expression of other co-stimulatory molecules like CD70, CD30, 4-1BB, OX40L etc.

overall_design: The total RNA was isolated from LPS matured DCs (positive control) and HIV infected DCs using Qiagen RNeasy Minikit. Agilent Quick-Amp Labelling kit (p/n 5190-0444) was used to synthesize the labeled cDNA from 3 ug of total RNA and further process the hybridized cDNA on the array. All the steps were carried out according to manufacturer’s instructions (www.agilent.com/chem/dnamauals-protocol). The array slides were scanned immediately using high throughput Agilent scanner with SureScan technology. Agilent feature extraction software was used for normalization and statistical analysis,. Pathway and gene ontology analysis was done using Genespring GX v 10.0 and biointerpreter software.  The experiments were performed on DCs cultured from two independent donors. The differential expression was considered if the Log 2 mean of at least -1 for the down regulated genes and +1 for the upregulated genes.  We considered only the genes that were differentially regulated genes in both the donors.

first: Madhuri

last: Thakar

first: Ramesh

last: Paranjape


title: Induction of a peptide with activity against a broad spectrum of pathogens in the Aedes aegypti salivary gland, following infection with dengue virus

city: Montpellier

first: David

last: Piquemal

address: 1682, rue de la Valsière

institute: ACOBIOM

summary: Dengue viruses (DENV) are generally maintained in a cycle which requires horizontal transmission via their arthropod vector, Ae. Aegypti, to the vertebrate host. One important consequence of this process is the interference of these arboviruses with both invertebrate and vertebrate immune systems. While infection of vertebrates causes disease, the presence of DENV gives rise to life-long, persistent infection in mosquitoes. The results of a comparative transcriptome analysis between DENV-infected and uninfected salivary glands revealed activation of both the immune deficiency (IMD) and the Toll pathways, as well as involvement of the putative antibacterial cecropin-like peptide (AAEL000598), in controlling DENV infection in Ae. aegypti. The mature form of this peptide was found to be active against DENV and Chikungunya viruses, whereas its precursor also had a strong anti-Leishmania effect. This study is the first to establish a comparative transcriptome analysis of DENV-infected and uninfected salivary glands and demonstrates that certain DENV-induced peptides, that are part of the IMD pathway, possess broad-spectrum anti-pathogenic activity and may have therapeutic potential in human.

overall_design: Infectious blood meals were offered to 3-day-old, adult, female Ae. aegypti Liverpool mosquitoes using a silicone membrane feeder system (Alto et al., 2005). Human blood was combined with DENV-2 16681 to provide a blood meal titer of 5.106 plaque forming units (PFU)/ml. At different time-points after the blood meal, salivary glands were dissected in acid guanidium thiocyanate-phenol-chloroform (RNAble; Eurobio, France) or phosphate-buffered saline (PBS), and the samples were frozen at -70°C until use.

first: Dorothée

last: Misseé


title: Analysis of the human endothelial cell response to Borrelia burgdorferi wild-type and deleted in the gene encoding the integrin ligand P66 [experiment A]

overall_design:
The cell line used for primary data analyses provided here was Ea.hy926, a macrovascular line (Edgell, C. J.,et al. 1990. Endothelium specific Weibel-Palade bodies in a continuous human cell line, EA.hy926. In vitro Cell. & Dev. Biol. 26:1167-1172, and Edgell, C. J., et al. 1983. Permanent cell line expressing human factor VIII-related antigen established by hybridization. Proc. Natl. Acad. Sci. 80:3734-3737). In addition, the human microvascular endothelial cell line HMEC (Ades et al, 1992. HMEC-1: establishment of an immortalized human microvascular endothelial cell line. J Invest Dermatol. 99:683-690) was used. Epithelial cells HEK-293 (which does not express b3 integrins) and HEK-293 transfected to express integrin αvβ3 were also analyzed. Infection times were 1 hour or 3 hours for cells lifted with EDTA and washed prior to bacterial infection, or 3, 6, and 24 hours for samples in which the bacteria were added to adherent cells. RNA was harvested and reverse transcribed. Labeled cDNAs were used to probe 2 color arrays.
In each of at least three biological replicate experiments, for each time point, the following comparisons were made. First, cells infected with B. burgdorferi with a marker inserted adjacent to the p66 gene (p66+)  were compared to  uninfected cells. Second, cells infected with the p66 gene interrupted by the marker (Coburn, J. and Cugini, C. (2003) Targeted mutation of P66 of Borrelia burgdorferi disrupts attachment to integrin αvβ3. Proc. Natl. Acad. Sci. 100:7301-7306) the mutant-infected cells were compared to the uninfected cells. In some experiments, cells infected with p66+ were compared to cells infected with the p66- mutant. Arrays used were HEEBO arrays purchased from Microarrays Inc. (Nashville, TN).
Ea.hy926 or HMEC endothelial cell lines, or HEK-293 epithelial cells, or transfected HEK-293 derivative expressing integrin avb3


title: Identification of miRNAs involved in embyonic chicken muscle development

summary: In the current study, we expanded our previous work to identify miRNAs implicated in the myogenesis regulation through the comparison of miRNAs transcriptome in skeletal muscle tissues between broilers and layers.

overall_design: miRNA expression studies of two different intra-species breeds


title: High recombination rates and hotspots in a Plasmodium falciparum genetic cross.

city: Rockville

first: Hongying

last: Jiang

address: 12735 Twinbrook Parkway

institute: NIAID/NIH

type: Genome variation profiling by genome tiling array

summary: We used a high-density tiling array to estimate genetic recombination rate among 32 independent recombinant progeny of a P. falciparum genetic cross (7G8 × GB4). We detected 3184 segregating multi-probe single-feature polymorphisms (mSFPs) and 638 recombination events (496 excluding those from subtelomeric regions). These data, in combination with results from 254 previously reported microsatellites, enabled us to construct a high-resolution genetic map. Comparing genetic and physical maps, we obtained an overall recombination rate of 9.6 kb/cM (12.8 kb/cM excluding subtelomeric regions) and identified 54 hotspots, some of which occurred in genes encoding surface antigens or proteins with repetitive motifs that might play a role in genetic recombination in the parasite. Motifs enriched in hotspots were also identified. In agreement with results from a previous cross (HB3 ´ Dd2), there was positive correlation between sizes of individual chromosomes and their recombination events. These results show that the P. falciparum genome is highly recombinogenic, providing an important genetic basis for parasite survival under various selection pressures. GC-rich repetitive motifs identified in the hotspot sequences may play a role in the high recombination frequency observed.

overall_design:
Ten microgram of genomic DNA, extracted and purified from 3D7 (reference), thirty-two P. falciparum independent recombinant progeny of the 7G8 x GB4 cross, and the two parental lines (Hayton, 2008), were hybridized to the PFSANGER Genechip® (Affymetrix, Inc., Santa Clara, CA, USA). The scanned image CEL files were first processed using the RMA method, then averaged and compared with reference genome 3D7, and lastly assigned either 7G8 or GB4 alleles based on similarities to the two parental lines. Total of 35 genomic DNA samples (biological replicates: 6 for 3D7, 4 for 7G8, 4 for GB4, and 2 for Pf_WE2).

The supplementary file 'GSE25656_QuantNormData_Log2_AllSamples.txt' contains the RMA-normalized data for all of the samples. The supplementary files 'GSE25656_chr*' contain the parental allele assignment of each chromosome and include probe-level annotation.

first: Vivek

last: Gopalan

first: Martine

middle: M

last: Zilversmit

first: Sudhir

last: Varma

first: Vijayaraj

last: Nagarajan

first: Jian

first: Jianbing

last: Mu

first: Karen

last: Hayton

first: Bruce

last: Henschen

first: Ming

last: Yi

first: Robert

last: Stephens

first: Gilean

last: McVean

first: Philip

last: Awadalla

first: Thomas

middle: E

last: Wellems

first: Xin-zhuan

last: Su


title: Comprehensive across production systems assessment of the impact of in vitro micro-environment on the expression of messengers and long non-coding RNAs in the bovine blastocyst

status: Public on Mar 11 2011

city: Quebec

first: Claude

last: Robert

state: Quebec

address: Pavillon Comtois

department: Animal Sciences

country: Canada

institute: Laval University

summary: In vitro production of cattle embryos over the past two decades has revealed several negative impacts, which have been attributed to the artificial microenvironment. Studies of embryos produced in vitro clearly point to aberrant gene expression levels. So far, the causal association between phenotype and measured gene expression has not led to substantial improvement of in vitro production systems. The aim of the present study was to generate a unique dataset composed of microarray-derived relative transcript abundance values for blastocysts produced in ten in vitro systems differing primarily in culture medium formulation. Between-group comparisons allowed us to determine the level of overall similarity between systems relative to in vivo reference embryos. The use of the dataset to contrast all in vitro treatments with the in vivo blastocysts pointed to a single common gene network. The “boutique” array contained a panel of novel uncharacterized transcripts that were expressed variably depending on the medium in which the blastocysts were produced. These novel transcripts were expressed differentially in blastocysts even as carry-over from conditions encountered seven days earlier during oocyte maturation. All of the selected novel candidates that were thus expressed were from intergenic regions. The function of this long non-coding RNA remains unknown but points clearly to an additional level of complexity in early embryo development.

overall_design: For all ten in vitro systems and the in vivo counterparts, three biological replicates were done. Technical duplicates for each of these 33 samples were analyzed to generate a dataset composed of 65 microarray hybridizations (one of the replicated array failed). During analysis, the in vivo derived blastocysts were used as reference. Following data pre-processing (that includes, background subtraction, intra-array normalization (Loess) and inter-array normalization (Quantile), all technical replication (replicated spots on the array and technically replicated arrays) were combined to generate a single value (duplicated spots were averaged and the median was drawn from duplicated arrays) that was used for dowstream significance analysis.

first: Isabelle

last: Côté

first: Christian

last: Vigneault

last: Laflamme

first: Joanie

last: Laquerre

first: Eric

last: Fournier

last: Gilbert

first: Sara

last: Scantland

first: Dominic

last: Gagne

first: Patrick

last: Blondin


title: H3K27 tri-methylation analysis of meristematic tissue (Me) and young leaves (Le) isolated by manual dissection of clv3-9 plants

city: Duesseldorf

first: Daniel

last: Schubert

address: Universitaetsstr. 1

department: Institute for Genetics

institute: Heinrich-Heine-University Duesseldorf

summary: For the histone methylation analysis of undifferentiated and differentiated tissues a meristem enriched fraction and young leaves (up to 3 mm leaf blade length) were isolated from clavata3-9 plants. Clavata3-9 mutants harbour larger shoot apical meristems than wild type plants enabling the preparation of meristematic tissue by manual dissection. Samples from both tissues were used for expression and histone H3 lysine 27 trimethylation (H3K27me3) analysis. Tissues for expression and H3K27me3 ChIP-chip analyses were simultaneously harvested.

overall_design: Meristem enriched samples (Me1 and Me2) and young leaf samples (Le1 and Le2) of clavata3-9 plants were isolated and hybridised to whole genome tiling arrays [platform GPL3371]. Two biological replicates were performed.

first: Marcel

last: Lafos

first: Phillip

last: Kroll


title: Reference Design time-course

city: Ikoma

first: Naotake

last: Ogasawara

state: Nara

institute: Nara Institute of Science and Technology

summary:
E.coli K-12 W3110 was grown in LB medium and harvested at each time point. And time series microarray experiments were performed based on reference desgin. In reference design, the control sample is collected at one representative time point. Combining with data from sequential design, more acculate and reliable expression series could be collected.
Keywords: Reference design

overall_design: The control sample is collected at one representative time point.

first: Ryosuke

last: Ito

first: Hiroki

last: Takahashi

first: Taku

last: Oshima

first: Shigehiko

last: Kanaya


title: Reduction of Salmonella Typhimurium following exposure to multiple hurdle treatments of heated, acidified organic acid salt solutions occurs via membrane destabilization

status: Public on Mar 20 2011

city: Fayetteville

first: Arunachalam

last: Muthaiyan

state: AR

address: 2435 N Hatch Ave

department: Food Science

institute: University of Arkansas

summary: The antimicrobial activity of organic acids in combination with non-chemical treatments was evaluated for inactivation of Salmonella Typhimurium. It was observed that the effectiveness of the multiple hurdle treatments were temperature and pH dependent and corresponded to the degree of organic acid lipophilicity. This led to the hypothesis that the loss in viability was due to cell membrane disruption. Evaluation of osmotic response, potassium ion leakage, and transmission electron micrographs confirmed effects on the cell membrane following treatment. Interestingly, all treatments, even those with no affect on viability, such as with sodium acetate, resulted in measurable cellular stress. Microarray experiments explored the specific response of S. Typhimurium to sodium acetate and sodium propionate, the most similar treatments in terms of pKa and ionic strength, and found little difference in the changes in gene expression following either treatment, despite their very different effects on viability. Taken together, the results reported support our hypothesis that following treatment with heated, acidified, organic acid salts, the loss of S. Typhimurium viability is at least in part due to membrane damage and the more lipophilic the organic acid the more effective the treatment. Overall, the data presented here indicate that a combined thermal, acidified sodium propionate treatment can provide a simple, yet effective antimicrobial treatment to combat Salmonella.

sample_organism: Salmonella enterica subsp. enterica serovar Typhimurium

overall_design:
We have strategically designed a multiple hurdle intervention employing various organic acid salts [sodium acetate (SA), sodium lactate, and sodium propionate (SP)], pH, and temperature hurdles into one treatment against S. Typhimurium. Previous research in our lab has shown similar treatments to be highly effective against S. Typhimurium. Here, we further investigate the range of effectiveness as well as the mechanism of action behind this multiple hurdle treatment using viability, osmotic response, potassium leakage, and transmission electron microscopy studies.
Total bacterial RNA was isolated as previously described and the RNA samples were then converted to fluorescently-labeled cDNA and hybridized to S. Typhimurium microarrays version 8 (NIAID's Pathogen Functional Genomics Resource Center).

first: S

middle: R

last: Milillo

first: E

last: Martin

first: A

middle: C

last: Ricke


title: Peripheral profiles from patients with cancerous and non cancerous lung diseases

city: Homburg

first: Andreas

last: Keller

address: Building 60

institute: Saarland University

summary: This project analyzes peripheral miRNA blood profiles of patients with lung diseases. Since miRNAs are known to be valuable diagnostic markers we asked whether respective patterns of lung cancer patients and COPD patients can be detected in peripheral blood samples rather than in biopsies. The project aimed at an impoved understanding of complex profiles rather than single markers. Thus, a high-throughput technique was necessary, profiling all known miRNAs integratively.

overall_design: n = 19 normal controls, n = 28 lung cancer patients and n = 24 COPD samples have been screened for the complete miRNA repertoire.  Please note that each miRNA has been measured in seven replicates and the median of the replica has been computed.

first: Petra

last: Leidinger


title: PU.1 and C/EBPalpha synergistically program distinct response to NF-kappaB activation through establishing monocyte-specific enhancers (ChIP-chip data)

city: Cleveland

first: Fulai

last: Jin

state: OH - OHIO

address: 10900 Euclid Avenue

institute: Case Western Reserve University

summary: Unraveling the complexity of transcriptional programs coded by different cell types has been one of the central goals of cell biology. Using genome-wide location analysis, we examined how two different cell types generate different responses to the NF-kappaB signaling pathway. We showed that, after tumor necrosis factor-alpha (TNF-alpha) treatment, NF-kappaB p65 subunit binds to distinct genome locations and subsequently induces different subsets of genes in human monocytic THP-1 cells versus HeLa cells . Interestingly, the differential p65 binding in two cell types correlates with pre-existing cell-type specific enhancers prior to TNF-alpha stimulation, marked by histone modifications. We also found that two transcription factors, PU.1 and C/EBPalpha, appear to synergistically mediate enhancer creation and affect NF-kappaB target selection in THP-1 cells. In HeLa cells, co-expression of PU.1 and C/EBPalpha conferred TNF-alpha responsiveness to a subset of THP-1 specific NF-kappaB target genes. These results suggest that the diversity of transcriptional programs in mammalian cells arises, at least in part, from pre-existing enhancers that are established by cell-specific transcription factors.

overall_design: We performed ChIP-chip for p65, p300, H3K4me1 and H3K4me3 using NimbleGen human whole genome tiling arrays in these cells with or without TNF-alpha treatment. In addition, we also mapped PU.1 and C/EBPalpha in untreated THP-1 cells with specific antibodies against these transcription factors.

first: Yan

last: Li

first: Rama

last: Natarajan

first: Bing

last: Ren


title: Gene expression during seed coat development in Arabidopsis

status: Public on Mar 10 2011

city: Saskatoon

first: Raju

last: Datla

state: SK

address: 110 Gymnasium Place

institute: NRC-PBI

summary: Transcriptional profiling during Arabidopsis seed coat development at 3 key developmental timepoints by using 2 mutant lines and their wild types. The data provides a globe view of seed coat development in arabidopsis can be used for identification of new gene candidates for seed coat development.

overall_design: 3 seed coat development stages, 4 lines (2 wild type + 2 mutants) of arabidopsis were sampled. 4 biological replicates.

first: Gillian

last: Dean

first: Yongguo

last: Cao

first: Daoquan

last: Xiang

first: Nicholas

middle: J

last: Provart

first: Larissa

last: Ramsay

first: Abdul

last: Ahad

first: Rick

last: White

first: Gopalan

last: Selvaraj

first: George

last: Haughn


title: Broad epigenetic signature of maternal care in the brain of adult rats (DNA methylation and H3K9 acetylation profiles)

status: Public on Mar 09 2011

city: Montreal

first: Moshe

last: Szyf

address: McIntyre Medical Building, 3655 Promenade Sir William Osler, Room 1309

department: Pharmacology

institute: McGill University

summary: Maternal care is associated with long-term effects on behavior and epigenetic programming of the NR3C1 (GLUCOCORTICOID RECEPTOR) gene in the hippocampus of both rats and humans. However, epigenetic changes at a single gene promoter are unlikely to account for the range of responses to the early life environment and the persistent change in expression of hundreds of additional genes in adult rats in response to differences in maternal care. Here we show that natural variations in maternal care are associated with coordinate changes in DNA methylation, chromatin, and gene expression spanning over a hundred kilobase pairs in the hippocampus of adult rats. The offspring of high compared to low maternal care mothers show epigenetic changes in promoters, exons, and gene ends and higher transcriptional activity across many genes, suggesting that a broad epigenetic regulation of gene expression may form part of a coordinated response to early maternal care.

overall_design: We obtained hippocampal samples from adult offspring of rat mothers that differed in the frequency of pup licking/grooming in the first week of life.  Using custom-designed microarrays with probes tiling the 7 million base pair region of rat chromosome 18 centered at the NR3C1 gene at 100 bp spacing, we obtained DNA methylation and H3K9 acetylation profiles by ChIP-on-chip.  Each profile was generated in triplicate, each type of profile from 3 high frequency and 3 low frequency mothers.

first: Patrick

last: McGowan

first: Matthew

last: Suderman

first: Michael

last: Meaney