title: Blood mRNA profile during EPO treatment

status: Public on Jan 01 2010

city: Montpellier

last: Piquemal

address: 1682, rue de la Valsière

institute: ACOBIOM

summary: In the present study, we obtained a global view from the transcriptome of total blood cells with or without darbepoetin alpha -an Erythropoiesis-Stimulating Agent (ESAs)- treatment using the SAGE method, which allows large-scale quantitative gene expression measurements.

overall_design: We sequenced 60,000 tags from 3 SAGE libraries.


title: C. elegans microarrays: daf-2(-) vs. daf-2(-) xbp-1(-)

status: Public on Feb 03 2010

city: Ramat Gan

first: sivan

last: korenblit

address: Ramat Gan

department: life sciences

country: Israel

institute: Bar-Ilan University

summary: Transcriptional profiling of day 1 animals comparing untreated daf-2(-) animals and daf-2(-) xbp-1(-) animals. Goal was to identify genes whose expression in daf-2 mutants is dependent (directly or indirectly) on xbp-1.

overall_design: Two-condition experiment, daf-2(-) animals and daf-2(-) xbp-1(-) animals. 4 Biological replicates in daf-2(e1370) background: 4 sets of daf-2(e1370) animals and their matching daf-2(e1370) xbp-1(zc12) animals. 4 Biological replicates in daf-2(e1368) background: 4 sets of daf-2(e1368) animals and their matching daf-2(e1368) xbp-1(zc12) animals.

first: Sivan

last: Henis-Korenblit

first: Cynthia

last: Kenyon


title: Paramecium tetraurelia autogamy series 1

status: Public on Jul 07 2010

city: Villerubanne

first: Jean-François

last: Gout

address: 43 Bld du 11 Novembre 1918

country: France

institute: Laboratoire Biométrie et Biologie Evolutive

type: Expression profiling by genome tiling array

summary: In the ciliate Paramecium tetraurelia, autogamy is a self-fertilization process, during which the zygotic nucleus results from the fusion of two identical gametic nuclei. This phenomenon occurs in response to starvation. It starts with meiosis of the germline nuclei (micronuclei or MIC) and fragmentation of the parental somatic nucleus (macronucleus or MAC). This is followed by mitotic division of one haploid nucleus issued from meiosis to yield two identical gametic nuclei, then karyogamy takes place, followed by mitosis of zygotic nucleus and differentiation of new MICs and MACs from the resulting copies of the zygotic nucleus. Within the developing new MACs, developmentally programmed DNA amplification and extensive genome rearrangements (precise excision of short non coding Internal Eliminated Sequences and chromosome fragmentation associated with the imprecise elimination of repetitive DNA) give rise to the highly polyploid somatic genome. To gain further insight into the complex regulation of these successive steps, we used whole genome microarrays to study the different gene networks that become activated throughout autogamy.

overall_design: Total RNA was extracted before the begining of autogamy (reference sample) and at 5 different time points during the process of autogamy. There are two biological replicates for each point.

first: Mireille

last: Betermier


title: LongSAGE transcriptome comparison on Botrytis cinerea conidia germination with and without resveratrol treatment

status: Public on Apr 06 2010

city: Beijing

first: Shangwu

last: Chen

address: Qing Hua Dong Lu No. 17

country: China

institute: China Agricultural University

summary:
Two long serial analysis of gene expression (LongSAGE) libraries of germinating conidia of Botrytis cinerea were generated with and without resveratrol treatments for unraveling the antifungal mechanisms of this phytoalexin by global transcriptome analysis – 82,786 total tags and a minimum of 15,665 unitags were obtained respectively. By tag-to-gene mapping, about 55% of the total unitags were matched to known genes. There were 109 unitags that showed significant differences in the two LongSAGE libraries, corresponding to 43 up- and 44 down-regulated genes, and 22 unmatched unitags as new gene candidates. These results showed that resveratrol inhibited expression of genes related to the glycolysis-Krebs cycle and mitochondrial electron transport chains at multiple sites in conidia germination, with induction of modified Entner–Doudoroff pathway as metabolic compensation. The chaperone-mediated protein folding and ubiquitin proteolysis systems were inhibited, while detoxification of resveratrol via ABC transporters was activated in this fungal pathogen.
Keywords: Botrytis cinerea; LongSAGE; metabolic pathway; resveratrol; transcriptome

overall_design:
Botrytis cinerea Pers. Ex was identified and isolated from stage III fruit of grapevines (Vitis vinifera cv. Merlot) in Huailai, Beijing. The most highly pathogenic strain M4 was chosen for these experiments. PSA (300g potato, 20g sugar, 20g agar per liter) was used for Botrytis culture at 20-23°C (Adrian et al. 1997). Conidia were aseptically harvested from 14 d cultures by washing with PSM (37.5g potato, 20g sugar per liter).

Conidia treatments and germination: 5 x 105 cfu/ml conidia in PSM was added with resveratrol (for BCB library treatment) to a final concentration of 140 µg/ml tran-resveratrol (Sigma, >99% purity), with no addition of resveratrol as a control (for BCA library treatment) (Adrian et al. 1997). After 12-14 h incubation in the dark at 21-22°C with shaking in shallow Petri dishes, 5 x 106 conidia from each treatment were collected and washed with H2O by centrifugation at 1,600g for 20 min at 21°C. Conidia were then frozen and stored at -80°C for RNA isolation.

first: Chuanlin

last: Zheng

first: Bing

last: Zhang

first: Huiqin

last: Ma

first: Songnian

last: Hu


title: Identification of Prognostic Genes for Recurrent Risk Prediction in Triple Negative Breast Cancer Patients in Taiwan

status: Public on Feb 02 2012

city: Taipei

first: Liang-Chuan

last: Lai

state: Taiwan

address: 1 Jen-Ai Road, Sec 1

department: Physiology

country: Taiwan

institute: National Taiwan University

type:
Third-party reanalysis
Expression profiling by array

summary: Discrepancies in the prognosis of triple negative breast cancer exist between Caucasian and Asian populations.  Yet, the gene signature of triple negative breast cancer specifically for Asians has not become available.  Therefore, the purpose of this study is to construct a prediction model for recurrence of triple negative breast cancer in Taiwanese patients.

overall_design: Whole genome expression profiling of breast cancers from 185 patients in Taiwan from 1995 to 2008 was performed, and the results were compared to the previously published literature to detect differences between Asian and Western patients.  Pathway analysis and Cox proportional hazard models were applied to construct a prediction model for the recurrence of triple negative breast cancer. Most expression data of samples (181/185) were reanalyzed from previous studies already uploaded to GEO (see "reanalysis of" links below). Four additional gene expression profiling data of triple negative breast cancer sample were added to this study.

first: Lee

middle: H

first: Wen-Hung

last: Kuo

first: Mong-Hsun

last: Tsai

first: Pei-Chun

first: Chuhsing

middle: K

last: Hsiao

first: Eric

middle: Y

last: Chuang

first: Li-Yun

last: Chang

first: Fon-Jou

last: Hsieh

first: King-Jen


title: Expression data from postharvest Ponkan (Citrus reticulata Blanco) fruit under cold storage

status: Public on Dec 18 2009

city: Wuhan

first: An

middle: dan

last: Zhu

state: Hubei

address: Shizishan street NO.1

department: College of Horticulture and Forestry Science

institute: Huazhong agricultural university

summary: Low temperature storage of citrus fruits are susceptible to chilling disorder symptoms that impact fruit quality. Understanding the molecular frame underlying the cold storage process will provide a basic guidance for practical control. We used Affymetrix Citrus GeneChip to examine the transcriptional changes in cold-stored Citrus reticulata Blanco cv. ‘Ponkan’ pulp tissue for three successive months.

overall_design: Ponkan postharvest fruits were under cold storage for three successive months. The pulp tissue was used for RNA extraction and hybridization on Affymetrix Citrus Genome microarrays.

first: Yunjiang

last: Cheng

first: Andan

first: Wenyun

last: Li

first: Junli

last: Ye

first: Xiaohua

last: Sun

first: Yuduan

last: Ding

first: Xiuxin

last: Deng


title: INTACT method for gene expression and chromatin profiling of individual cell types within a tissue

status: Public on Jun 14 2010

city: Seattle

first: Jorja

last: Henikoff

state: WA

address: 1100 Fairview AV N, A1-162

department: Basic Sciences

country: USA

institute: Fred Hutchinson Cancer Research Center

type: Genome binding/occupancy profiling by genome tiling array

summary:
We used in vivo biotin labeling of a nuclear envelope protein in individual cell types followed by affinity isolation of labeled nuclei to measure gene expression and chromatin features of the hair and non-hair cell types of the Arabidopsis root epidermis.

Keywords: Chromatin affinity-purification on microarray

overall_design: All experiments were done using two channels per chip. Two types of experiments were performed. Expression profiling was performed by comparing cDNAs synthesized from nuclear RNA isolated from INTACT-prepared Arabidopsis nuclei to total sonicated genomic DNA isolated from Arabidopsis plants. Chromatin profiling was performed using INTACT-prepared Arabidopsis nuclei. DNA isolated by Chromatin immunoprecipitation (ChIP) using antibodies to modified histones was compared to histone H3 ChIP DNA from the same sample.

first: Steven

first: Roger

middle: B

last: Deal


title: miRNA Profiling from total blood of Multiple Sclerosis and control samples

status: Public on Sep 24 2009

city: Homburg

first: Andreas

last: Keller

address: Building 60

country: Germany

institute: Saarland University

summary: This project aims at the detection of specific patterns of miRNAs in peripheral blood samples of MS patients. As controls, blood of donors without known affection have been tested. Using the miRNA patterns we hope to detect a diagnostic pattern for the non-invasive diagnosis of MS.

overall_design: In this study, we compared 20 Multiple Sclerosis samples to 21 control samples using the sanger miRBAse 12.0 miRNA biochip manufactured by febit. Samples were analyzed with the Geniom Realtime Analyzer (GRTA, febit gmbh, Heidelberg, Germany) using the Geniom Biochip miRNA homo sapiens. Each array contains 7 replicates of 866 miRNAs and miRNA star sequences as annotated in the Sanger miRBase 12.0. To benchmark the platform, we tested technical replicates using bought total RNA of brain and liver samples (ambion), achieving a correlation of 0.97.

first: Petra

last: Leidinger

first: Anke

last: Wendschlag

first: Matthias

last: Scheffler

first: Eckart

last: Meese

first: Frank

last: Wucherpfennig

first: Klemens

last: Ruprecht

first: Hannah

last: Schroers

first: Anne

last: Borries


title: Peripheral blood transcriptomics_Human_Single supratherapeutic dose of acetaminophen

status: Public on Oct 06 2009

city: RTP

first: NIEHS

last: Microarray Core

state: NC

address: 111 T.W. Alexander Drive

department: DIR

institute: NIEHS

summary: This was a controlled pilot study of 9 healthy adults admitted to the General Clinical Research Center at the University of North Carolina Hospital. After 3 days of clinical acclimatization, 6 subjects received a single 4g bolus dose or a placebo. Peripheral blood was collected on each day preceeding dosage, and then 6, 18, 24, 48, 72, and 96 hours post dosing. Total RNA was extracted and analyzed for differential gene expression on Agilent Human 1vA2 microarray chips.

overall_design: 7.5 mls (2.5 ml in 3 PAXgene tubes) of blood were collected at each time point. Total RNA was amplified and labeled according to Agilent protocols and a single chip was run using a referenced design with Stratagenes Universal Human reference RNA. Each patient was normalized to there own 0 hr data. Significant differential expression was determined by comparing  APAP treated subjects to placebos.

first: Rick

middle: D

last: Fannin

first: Mark

last: Russo

first: Thomas

middle: M

last: O'Connell

first: Kevin

last: Gerrish

first: Jeffrey

last: MacDonald

first: Jack

last: Newton

first: Shahid

last: Malik

first: Stella

middle: O

last: Sieber

first: Joel

last: Parker

first: Ruchir

last: Shah

first: Tong

last: Zhou

first: Paul

last: Watkins

first: Richard

middle: S

last: Paules


title: Effects of exercise training and antioxidant supplementation on endothelial cell gene expression

city: Brisbane

first: Aya

last: Matsumoto

state: Qld

address: Blair Drive

department: Hunan Movement Studies

country: Australia

institute: University of Queensland

summary: The molecular mechanisms of exercise-induced cardiovascular protection are poorly understood. There is growing evidence that reactive oxygen species (ROS) are necessary for some of these adaptations and antioxidants may be used to investigate this effect. This study aimed to determine the effects of exercise and/or antioxidant supplementation on myocardial and vascular endothelium gene expression. Male Wistar rats were divided into four groups: i) endurance exercise (90min of treadmill running 4d/week, 14 weeks); ii) antioxidant-treated; iii) antioxidant and endurance exercise and iv) control. The supplemented animals received Vitamin E (1000 IU/kg diet) and alpha-lipoic acid (1.6 g/kg diet) mixed with rat chow. cDNA microarray analysis was performed using purified endothelial RNA from myocardial and coronary artery endothelial cells and showed that the expression levels of 35, 40 and 40 genes were altered for groups i, ii, and iii respectively compared to control. Differentially expressed genes were analysed using the KEGG pathway database, hierarchical cluster and DAVID analysis. These analyses revealed that a gene involved in cardiovascular disease progression, Ras homolog gene family member A (RhoA) was down-regulated by exercise, upregulated by antioxidant supplementation and the combination of exercise and antioxidant blunted both effects. These findings were confirmed by real-time PCR. In summary, exercise and antioxidant supplementation affect endothelial cell gene expression and ROS appear necessary for some of these adaptations.

overall_design: 48 Male Wistar rats were divided into four groups: i) endurance exercise (90min of treadmill running 4d/week, 14 weeks); ii) antioxidant-treated; iii) antioxidant and endurance exercise and iv) control. The supplemented animals received Vitamin E (1000 IU/kg diet) and alpha-lipoic acid (1.6 g/kg diet) mixed with rat chow. cDNA microarray analysis was performed using purified endothelial RNA from myocardial and coronary artery endothelial cells. Based on the limited material available, it was necessary to combine the RNA into three pools per treatment group for CAEC (approx 400 ng of total RNA), and four pools for LVEC (approx 1 μg of total RNA) with an additional reference pool from each control group.

first: Steven

middle: R

last: Mason

first: Trudi

last: Flatscher-Bader

first: Leigh

middle: C

last: Ward

first: Susan

middle: A

last: Marsh

first: Peter

last: Wilce

first: Robert

middle: G

last: Fassett

first: Jeff

last: Coombes


title: Global transcriptome response of recombinant E. coli to heat-shock and dual heat-shock recombinant protein induction

status: Public on Aug 07 2009

city: Clemson

first: Sarah

middle: W

last: Harcum

state: SC

address: 105 Collings Street

department: Bioengineering

institute: Clemson University

summary:
Recombinant Escherichia coli cultures are used to manufacture numerous therapeutic proteins and industrial enzymes, where many of these processes use elevated temperatures to induce recombinant protein production.  The heat-shock response in wild-type E. coli has been well studied.  In this study, the transcriptome profiles of recombinant E. coli subjected to a heat-shock and to a dual heat-shock recombinant protein induction were examined.  Most classical heat-shock protein genes were identified as regulated in both conditions.  The major transcriptome differences between the recombinant and reported wild-type cultures were heavily populated by hypothetical and putative genes, which indicates recombinant cultures utilize many unique genes to respond to a heat-shock.  Comparison of the dual stressed culture data with literature recombinant protein induced culture data revealed numerous differences.  The dual stressed response encompassed three major response patterns: induced-like, in-between, and greater than either individual stress response.  Also, there were no genes that only responded to the dual stress.  The most interesting difference between the dual stressed and induced cultures was the amino acid-tRNA gene levels.  The amino acid-tRNA genes were elevated for the dual cultures compared to the induced cultures.  Since tRNAs facilitate protein synthesis via translation, this observed increase in amino acid-tRNA transcriptome levels, in concert with elevated heat-shock chaperones, might account for improved productivities often observed for thermo-inducible systems.  Most importantly, the response of the recombinant cultures to a heat-shock was more profound than wild-type cultures, and further, the response to recombinant protein induction was not a simple additive response of the individual stresses.
The objective of the present work is to gain a better understanding of the heat-shock response in recombinant cultures and how this response might impact recombinant protein production.  To accomplish this objective, the transcriptome response of recombinant cultures subjected to a heat-shock and a dual heat-shock recombinant protein induction were analyzed.  The transcriptome levels were determined using Affymetrix E. coli Antisense DNA microarrays, such that the entire genome was evaluated.  These two transcriptome responses were also compared to recombinant cultures at normal growth temperature that were not over-expressing the recombinant protein and a set of literature recombinant culture data that were chemically induced to over-express the recombinant protein.  Additionally, the heat-shock response of the recombinant cultures was compared to the literature report of the heat-shock response in wild-type cultures.  The results of the global transcriptome analysis demonstrated that recombinant cultures respond differently to a heat-shock stress than wild-type cultures, where the transcriptome response of the recombinant cultures is further modified by production of a recombinant protein.

overall_design: The heat-shock and recombinant protein production phases were synchronized to the cell density of 11.5 OD, which is referred to as Sample Time 0.  For the heat-shocked cultures, the temperature was increased from 37°C to 50°C over 8 minutes beginning at Sample Time 0.  The temperature and duration used in this study are the same conditions used to evaluate the heat-shock in wild-type cultures.  The temperature was then decreased from 50°C to 37°C over 4 minutes. For the dual heat-shocked recombinant protein production cultures, 5 mM IPTG was added 8 minutes after Sample Time 0. The unstressed recombinant cultures were conducted similarly, except without the heat-shock or IPTG-addition.  Each sample condition was obtained from at least two separate fermentations (two biological replicates).  RNA from each biological replicate was purified and processed independently.  Prior to hybridization, where only two biological replicates existed, one of the processed samples was divided (two technical replicates), resulting in three separate hybridized chips.  The heat-shocked and dual stressed culture samples all consisted of three technical replicates from two biological duplicates.  For the unstressed culture samples, triplicate samples were obtained for the 11.5 OD and duplicates for the 14 OD conditions.  There were no statistical differences between the 11.5 and 14 OD unstressed samples (p ≤ 0.001).  Thus, the unstressed culture transcriptome profile consisted of six technical replicates from five biological replicates and four independent fermentations.

first: Fu'ad

middle: T

last: Haddadin


title: Absolute expression level of Saccharomyces cerevisiae genes

status: Public on Jul 06 2010

city: Boston

first: Alex

last: Tsankov

state: MA

address: 155 Bay State

institute: MIT

summary: Total RNA versus genomic DNA hybridization on custom arrays designed for all Saccharomyces cerevisiae genes

overall_design: Total RNA was collected in mid-log phase from Saccharomyces cerevisiae cells grown in rich medium (abbreviated CM, in house recipe).   RNA was then converted to cDNA, Cy3-labeled and hybridized competitively against Cy5 labeled genomic DNA from Saccharomyces cerevisiae.

first: Alexander

first: Oliver

last: Rando

first: Aviv

last: Regev


title: Primary human umbilical vein and arterial endothelial cells

status: Public on Dec 31 2009

city: Hsinchu

first: Yung-Jen

last: Chuang

address: No. 101, Sec. 2, Kuang Fu Road

department: Department of Medical Science

institute: National Tsing Hua University

summary:
Human umbilical cords were obtained from the Lucille Packard Children Hospital. In this study, we devised a rapid isolation scheme to preserve the in vivo phenotype of each endothelial subtype. ECs were first isolated from umbilical cords by collagenase perfusion through the vein or artery as described. Cells were further purified using a Percoll density gradient (Amersham Biosciences, Piscataway, NJ) to remove residual erythrocytes and platelets. 1-5 x 106 ECs were then cultured overnight on gelatin-coated T12.5 flasks in Clonetics EGM-2 media (Cambrex Bio Science, Walkersville, MD) in which the ECs generally reached confluence the next morning. The confluent EC were trypsinized and subjected to two rounds of immuno-magnetic beads purification, adapted from a published purification protocol. There was a negative selection step using CD14, CD45 and CD64 to remove residual contaminating leukocytes, followed by positive selection using a mouse anti-endothelial cell monoclonal antibody (anti-CD146/clone P1H12 purchased from Chemicon, Temecula, CA). Total processing time was limited to 20 to 24 hours. The homogeneous, viable, primary ECs were used immediately to construct the library. The construction of SAGE libraries was performed with the I-SAGE kit (Invitrogen, Carlsbad, CA) per manufacturer’s instructions
Keywords: other

first: Kyunga

last: Seo

first: John

last: Gray

first: Oana

last: Lonel

first: Maurice

last: Druzin

first: Calvin

last: Kuo

first: Lawrence

last: Leung


status: Public on Jul 01 2009

city: Denver

first: Ivana

middle: V

last: Yang

state: CO

address: 1400 Jackson Street

department: Medicine

institute: National Jewish Health

summary:
Molecular Pathways and Transcriptional Networks Involved in the Macrophage Response to LPS, poly(I:C) and CpG DNA Stimulation

Background:  Toll-like family of receptors recognizes pathogen-associated molecular patterns (PAMPs) from different organisms.  TLR4 is the receptor for bacterial lipopolysaccharide (LPS), dsRNA viral mimic poly(I:C) binds to TLR3, and bacterial CpG DNA signals through TLR9.  TLR4 signaling is mediated by adaptor molecules Myd88 and TRIF while TLR3 pathway involves only the TRIF adaptor and TLR9 signals solely through Myd88.      Methods:  To identify genes other than those in TLR pathways that mediate macrophage response to different PAMPs, RAW264.7 cells were stimulated with LPS, poly(I:C), or CpG DNA, and RNA was profiled on microarrays 6 hrs and 24 hrs post-treatment.  Gene expression data were analyzed to determine genes, pathways and transcriptional networks that are in common and unique to each of the three TLR stimuli.  Potentially novel candidates revealed by this analysis were tested for their role in innate immunity using RNA interference.    Results:  Many genes are differentially regulated by LPS and poly(I:C) at both 6 hrs and 24 hrs following treatment, while CpG DNA elicits a much less pronounced transcriptional response.  By analyzing gene expression data for networks and pathways, we prioritized differentially expressed genes that are in common to all three PAMPs as well as those shared by LPS and poly(I:C).  Knockdown by RNA interference of two genes, Plec1 and TPST1, inhibited production of IL-6 in response to LPS in cultured macrophages.      Conclusions:  We have identified novel innate immunity genes that may be important in macrophage response to LPS, poly(I:C), and CpG DNA stimuli.  Our results provide potential biomarkers and therapeutic targets that should be further investigated in mice and human populations.

overall_design: For each treatment (LPS, polyIC, CpG DNA, media only), three biological replicates (separate macrophage cultures and RNA isolations) were profiled.  Each sample was labeled with Cy3 and Cy5 and co-hybridized with Stratagene Universal Mouse Reference (dye flip techical replicates).  Expression at 2 timepoints (6 and 24 hours post-treatment) was assessed.

first: Wayne

last: Jiang

first: David

last: Pisetsky

last: Schwartz


title: Cadmium probe selection experiment for custom array production

city: Antwerpen

first: Dries

last: Knapen

address: Groenenborgerlaan 171

department: Biology

country: Belgium

institute: University of Antwerp

summary: Microarrays have evolved from low-density cDNA or oligonucleotide arrays to high-density platforms, for several study species even covering the complete transcriptome. At the same time, transcriptomics experiments have become more complex and multifactorial in nature, requiring many microarrays to assess multiple biologically relevant hypotheses. Scientists using this technology are therefore painfully aware of the high financial cost of a typical microarray experiment. Unfortunately, this often leads to either a suboptimal experimental design in an effort to reduce the cost by using fewer microarrays, or to abandoning microarray technology altogether. In this study, we argue that for many studies high-density full genome microarrays are in fact technical overkill. By selectively reducing full genome probe sets to a lower number of probes, it is possible to significantly reduce the total cost of a microarray experiment. The study consists of four microarray analyses: a cadmium probe selection experiment, a temperature probe selection experiment, a cadmium validation experiment and a cadmium validation experiment.

overall_design: Samples were hybridized in a simple loop design, in which every sample was labelled once with the red dye and once with the green dye. The hybridization design for the cadmium probe selection experiment was (C0D28R1 > C5D28R1 > C1D28R1 > C0D4R1 > C5D28R2 > C0D28R2 > C1D28R2 > C0D28R1 + C0D28R2 > C5D28R1). Arrows (>) indicate individual microarrays; heads represent the green dye and tails represent the red dye. Sample names are of the form CxDyRz (cadmium concentration x in µM, day y, biological replicate z).

first: Lucia

last: Vergauwen

first: Ronny

last: Blust


title: Effect of aging on the meiotic cell cycle

status: Public on Jun 16 2009

city: Cambridge

first: Monica

last: Boselli

address: 40 Ames street

department: Center for cancer research

institute: Koch Inst. (KI) MIT

summary:
Identify which  stages of meiosis are impaired in replicative aging cells
Transcriptional profile of aging yeast cells (young and old) incubated in sporulation-inducing conditions compared to new-born cells grown in vegetative conditions.

overall_design: Two conditions experiments. Young and Old cells incubated in sporulation-inducing conditions. Control strain was same strain used for the experimental samples but grown in vegetative conditions.

first: Angelika

last: Amon

first: Stuart

last: Levine


title: Identification of Hedgehog Signaling and Novel Transcription Factors Involved in Regulation of Systemic Response to LPS

summary:
Background:
Our understanding of the role host genetic factors play in the initiation and severity of infections caused by Gram negative bacteria is incomplete.
Methods:
To identify novel regulators of the host response to lipopolysaccharide (LPS), 11 inbred murine strains were challenged with LPS systemically.  RNA from lung, liver, and spleen tissue was profiled on oligonucleotide microarrays to determine if unique transcripts differentiate susceptible and resistant strains of mice.  Gene expression data were analyzed to identify over-represented pathways and transcription factors (TFs).  The role of several TFs in innate immune response to LPS was examined by RNA interference in a mouse macrophage cell line.  Mouse lines with targeted mutations were utilized to further confirm involvement of two novel genes in innate immunity.
Results:
In addition to two strains lacking functional TLR4 (C3H/HeJ and C57BL/6JTLR4-/-), three murine strains with functional TLR4 (C57BL/6, 129/SvlmJ, and NZW/LacJ) were found to be resistant to systemic LPS challenge; the other six strains were classified as sensitive.  Gene expression analysis supports the involvement of a number of previously identified genes, molecular pathways, and TFs in the host response to LPS but also identifies Hedgehog signaling as a novel pathway activated by LPS.  B6;129Ptch1+/+ wild-type mice were shown to be more sensitive to systemic LPS than B6;129Ptch1+/- heterozygote littermates further supporting the role of Hedgehog signaling in systemic LPS response.  RNA interference-mediated inhibition of 6 TFs (C/EBP, Cdx-2, E2F1, Hoxa4, Nhlh1, and Tead2), out of 15 tested, was found to diminish production of IL-6 and TNFalpha protein in murine macrophages.  The role of E2F1 was confirmed by showing that B6;129E2F1-/- knockout mice are more sensitive to systemic LPS than wild-type controls.
Conclusions:
Our analysis of gene expression data identified novel pathways and transcription factors that regulate the host response to systemic LPS.  Our results provide potential sepsis biomarkers and therapeutic targets that should be further investigated in human populations.

Keywords: disease state analysis

overall_design: For each strain of mice, 6 mice were treated with LPS and 6 with saline.  RNA of 3 animals was pooled to make two pools per condition (two biological replicates).  Each pool was labeled with Cy3 and Cy5 and co-hybridized with Stratagene Univeral Mouse Reference (dye flip techical replicates). RNA from lung, liver, and spleen tissue was profiled on oligonucleotide microarrays


title: Low temperatures impact dormancy status, flowering competence, and transcript profiles in crown buds of leafy spurge.

status: Public on Dec 01 2009

city: Fargo

first: Munevver

last: Dogramaci

state: ND

address: 1605 Albrecht Blvd N

institute: USDA-ARS

summary: Leafy spurge (Euphorbia esula) is an herbaceous perennial weed that produces vegetatively from an abundance of underground adventitious buds. The objectives of this study were to determine how mimicking natural seasonal conditions (photoperiod and temperature) under controlled environmental conditions affect dormancy and flowering competence; to determine molecular mechanisms associated with well-defined phases of seasonal dormancy transitions based on transcript profiles obtained by microarray analysis; and to link mechanisms regulating induction and release of endodormancy and flowering competence. Reduction in temperature (27 to 10°C) and photoperiod (16 to 8 h) over a three-month period induced a para- to endo-dormant transition in crown buds. An additional eleven weeks of prolonged cold (5-7°C) and short-photoperiod treatment resulted in accelerated shoot growth from crown buds, and 99% floral competence when plants were returned to growth promoting conditions. Exposure of paradormant plants to short-photoperiod and prolonged cold treatment alone had minimal affect growth potential or on flowering (~1%); whereas endodormant crown buds without prolonged cold treatment, had delayed shoot growth and approximately 2% flowering when returned to growth promoting conditions. Transcriptome analyses revealed that 373 and 260 genes were differentially expressed (p<0.005) during para- to endo-dormant and endo- to eco-dormant transitions, respectively. Transcripts from flower competent vs. non-flower competent crown buds identified 607 differentially expressed genes, and genes involved in cell cycle and DNA processing, oxidative stress, flower regulation, and proteolysis were over-represented. Further, sub-network analysis identified expression targets and binding partners associated with circadian clock, dehydration/cold signaling, phosphorylation cascades, and response to abscisic acid, ethylene, gibberellic acid, and jasmonic acid, suggesting these central regulators affect well-defined phases of dormancy. Potential genetic pathways associated with these dormancy transitions and flowering were used to develop a proposed conceptual model.

platform_organism:
Manihot esculenta
Euphorbia esula

overall_design: Leafy spurge is an herbaceous perennial weed that undergoes vegetative reproduction from an abundance of underground adventitious buds which exhibit phases of para-, endo-, and eco-dormancy (defined by Lang et al. 1987) during summer, fall, and winter, respectively (Anderson et al. 2005). In this study a population of leafy spurge plants were propagated through random cuttings from the genetically uniform biotype 1984-ND001. Paradormant crown buds from three-month old greenhouse plants were collected after one week of acclimation under growth chamber conditions. Induction of endodormancy in crown buds was accomplished by subjecting greenhouse grown and growth chamber acclimated plants to a ramp down (RD) treatment consisting of a reduction in temperature of 1.42°C week-1 (27°C → 10°C) and a decreasing photoperiod of 40 min week-1 (16h → 8h light) for 12 weeks as previously established by Foley et al. (2009). These conditions mimic the average seasonal environmental conditions experienced in Fargo, ND (46°54′ N, 96°48′ W) during the transition from para- to endo-dormancy (Anderson et al. 2005). To induce a transition of crown buds from endo- to eco-dormancy, plants subjected to the RD treatment were given prolonged cold treatment for 11 weeks at 5-7°C, under constant 8 h:16 h day:night cycle; light fluencies were approximately 250 μmol m-2 s-1. To compare the effects of the prolonged cold with or without RD treatment on dormancy status and flowering competence in crown buds, greenhouse-grown and growth chamber acclimated plants were subjected directly to an extended cold treatment for 11 weeks at 5-7°C, as previously described, without a RD treatment. For microarray hybridizations, labeled cDNAs were prepared from 30 µg of total RNA using the Alexa Fluor cDNA labeling kit (Invitrogen, Carlsbad, CA, USA) according to manufacturer's protocols. Labeled cDNAs were hybridized to a custom made 23K element microarray that contained 19,808 unigenes from a leafy spurge EST database (Anderson et al. 2007) and an additional 4,129 unigenes from a cassava EST database (Lokko et al. 2007). Comparison of gene expression between samples was accomplished using a rolling circle dye swap hybridization scheme (Churchill 2002) to provide every biological replicate with technical replicates. In our microarray analyses experiment, each of the four biological replicates included four technical replicates using two different dyes, resulting in a total of 16 technical replicates for each treatment. Microarray hybridization was visualized using a GenePix 4000B scanner and probe intensities and background were quantified using GenePix 6.0 software (Molecular Devices, Sunnyvale, California, USA). A quality control value of “1” was assigned to all probes that had intensity values greater than 2 times the standard deviation over average of the negative control and empty probe intensities (after deletion of 1% of the most intense negative/empty probe values). Hybridization intensities were log2 transformed, and arrays were centered and normalized against each other.

middle: P

last: Horvath

first: Wun

last: Chao

first: Michael

last: Foley

middle: J

last: Christoffers

first: James

last: Anderson


title: Csac growth on monosaccharides found in lignocellulose

status: Public on Dec 29 2009

city: Raleigh

first: Amy

last: VanFossen

address: 840 Main Campus Dr.  Rm3305

department: Chemical and Biomolecular Engineering

institute: NC State Unversity

summary: The co-utilization of hexoses and pentoses derived from lignocellulose is an attractive trait for in microorganisms considered for consolidated biomass processing to biofuels. This issue was examined for the H2-producing, extremely thermophilic bacterium Caldicellulosiruptor saccharolyticus (Csac) growing on individual monosaccharides (arabinose, fructose, galactose, glucose, mannose and xylose), mixtures of these sugars. Based on the whole-genome transcriptional response analysis and comparative genomics, carbohydrate specificities for transport systems could be proposed for most of the 24 putative carbohydrate ATP-binding cassette (ABC) transporters in the C. saccharolyticus genome.  Transcriptome contrasts for monosacchride growth showed that minimal changes were observed in some cases (e.g., 31 ORFs changed >/=2-fold for glucose/galactose) while substantial changes occurred for cases involving mannose (e.g., 363 ORFs >/=2-fold for glucose/mannose). No evidence for catabolite repression was noted for either growth on multi-sugar mixtures or in the corresponding transcriptomes.

overall_design: C. saccharolyticus was subcultured (overnight) seven times on the substrate of interest in modified DSMZ 640 medium before inoculating a 1-liter batch containing 0.5 gram substrate per liter. Cells were grown at 70 °C until mid-logarithmic phase (3-5*107) and harvested by rapid cooling to 4 °C and centrifugation and then stored at -80 °C. To elucidate the transporters plus the central carbon metabolic pathways and their regulation utilized on the different sugars, transcriptome analysis was performed after growth on arabinose, fructose, galactose, glucose, mannose, xylose and a mixture of all six substrates.

middle: L


title: Paramecium tetraurelia reciliation (stationary phase)

status: Public on Nov 01 2009

first: Jean-François

last: Gout

summary:
Paramecium cells in stationary phase were treated for deciliation and total mRNA extracted at two time points (45 and 130 minutes) after deciliation.

Keywords: Time course analysis of expression during reciliation

overall_design: Experimental design has two biological replications for the 45 and 130 minutes time points and only one sample for the reference (T0) time point.

first: Cohen

last: Jean

first: Bouhouch

last: Khaled


title: C. elegans embryos: control vs. mes-2 mutant

status: Public on May 26 2009

city: Salt Lake City

first: Susan

last: Mango

state: UT

address: 2000 Circle of Hope

department: Oncological Sciences

institute: University of Utah

summary: Transcriptional profiling of early C. elegans embryos comparing control (N2) embryos with mes-2 mutant embryos at different developmental stages: 2E (24-40 cells), 4E (50-90 cells) and 8E stage (100-200 cells). Goal was to determine the effects of mes-2 loss on global gene expression as embryos transit from a developmentally plastic state (2E stage) to the onset of differentiation (8E stage). Our microarray data showed that early-expressed genes remain active, differentiation genes fail to reach wild-type levels in mes-2 mutant embryos at the 8E stage.

overall_design: Two-condition experiment, wild type vs. mes-2 embryos. Biological replicates: 3 control replicates, 3 mes-2 replicates for each stage.

first: Tatiana

middle: N

last: Yuzyuk


title: INO80-regulated genes in S-phase after MMS treatment

status: Public on Oct 25 2009

city: Smithville

first: Xuetong

last: Shen

state: TX

address: 1808 Park Road 1-C

institute: MD Anderson Cancer Center

summary: This is transcription analysis of genes regulated by the INO80 chromatin remodeling complex in S-phase of the cell cycle after MMS treatment.

overall_design: Wildtype and ino80 mutant cells were first synchronized at G1 using alpha-factor, the cells were released in to S-phase in the presence of 0.02% MMS. After 45 minutes, total RNA was isolated and analyzed to identify genes that are regulated by INO80 under such conditions.

first: K

last: Falbo

first: R

last: Basom

first: J

last: Delrow

first: X


title: Protein Kinase C Alpha Promotes Tumor Growth by Initiating a Feed-forward Network Involving miR-15a and Cyclin E

status: Public on Mar 22 2009

city: Nashville

first: Christine

middle: H.

last: Chung

state: TN

address: 777 PRB

department: Internal Medicine

institute: Vanderbilt University

summary:
Protein Kinase C alpha (PKC) is associated with progression and poor prognosis in head and neck cancer.  Previous studies have demonstrated that PKC sustains the proliferative signal by increasing cyclin E expression, leading to enhanced E2F target gene transcription and DNA synthesis. Here we show that PKC increases DNA synthesis through inhibition of the microRNA, miR-15a, upregulating translation of its target cyclin E. Importantly, gene expression and qRT-PCR analysis of primary squamous cell carcinoma tumors of the head and neck (SCCHN) reveals a significant negative correlation between PKC alpha and miR-15a levels. In contrast to normal cell cycle initiation, PKC decreases microRNA expression, leading first to increased cyclin E protein followed by enhanced transcription of cyclin E and other DNA synthesis mediators.  These results identify a signaling network regulated by PKC whereby constitutive kinase activation switches the system to feed forward, overriding normal regulation of cell cycle progression through a post-transcriptional mechanism involving microRNAs.  This reprogramming of the network is likely a more general phenomenon that can account for the oncogenic potency of established signaling pathways.
Keywords: miRNA; dose response; cancer

platform_organism:
Homo sapiens
Mus musculus
Rattus norvegicus

overall_design: 6 treated samples across two time points hybridized to paired time 0, untreated controls

middle: H

first: Joel

last: Parker


title: Serial Analysis of Gene Expression molecular signature for disease progression in Idiopathic Pulmonary Fibrosis

status: Public on Apr 07 2009

city: Research Triangle Park

first: Kathy

last: Boon

address: 111 TW Alexander Dr

department: Division of Intramural Research

institute: NHLBI/NIEHS

summary: Idiopathic pulmonary fibrosis (IPF) is a chronic and progressive fibrosing interstitial lung disease that is unresponsive to current therapy. While it carries a median survival of less than 3 years its rate of progression varies widely between patients. We hypothesized that studying the gene expression profiles of physiologically stable patients and those in which the disease progressed rapidly after the initial diagnosis would aid in the search for biomarkers and contribute to the understanding of disease pathogenesis.

overall_design: We generated 12 Idiopathic Pulmonary Fibrosis (IPF) lung parenchyma SAGE profiles. Initial cluster analysis including 8 other public available lung SAGE libraries verified that the IPF transcriptome is distinct from normal lung tissue and other lung diseases like COPD. In order to identify candidate markers of disease progression we segregated the IPF SAGE profiles in two groups based on clinical parameters regarding lung volume and lung function.

first: Nathaniel

last: Bailey

first: Jun

last: Yang

first: Kervitsky

last: Dolly

first: Kevin

middle: K

last: Brown

first: Schwartz

last: David


title: Cancer Genome Anatomy Project (CGAP) Human SAGE

status: Public on Mar 20 2009

city: Bethesda

first: Cancer Genome Anatomy Project

last: CGAP

state: MD

department: Cancer Genome Anatomy Project

institute: National Cancer Institute

summary:
This series represents the Cancer Genome Anatomy Project SAGE library collection.  Libraries contained herein were either produced through CGAP funding, or donated to CGAP.
The Cancer Genome Anatomy Project (CGAP: http://cgap.nci.nih.gov) is an interdisciplinary program established and administered by the National Cancer Institute (NCI: http://www.nci.nih.gov) to generate the information and technological tools needed to decipher the molecular anatomy of the cancer cell.

SAGE libraries are named according to the following convention:
* SAGE_Organ_histology_code_unique identifier, e.g., SAGE_Colon_adenocarcinoma_CL_Caco2
* Codes: B = bulk; CL = cell line; CS = short-term cell culture; MD = micro-dissected; AP = antibody purified.

overall_design: 324 libraries

last: Strausberg

first: D

last: Gerhard

first: Gregory

last: Riggins


title: Liver gene expression in a panel of laboratory inbred mouse strains

status: Public on Feb 28 2009

city: Chapel Hill

first: Daniel

middle: M

last: Gatti

address: 135 Dauer Dr.

department: Environmental Sciences & Engineering

institute: University of Noth Carolina

summary:
This study aimed to perform gene expression quantitative trait locus mapping (eQTL) in the livers of a panel of 36 laboratory inbred strains.  Mice were dosed with a saline solution by oral gavage and sacrificed at 24 hours. Livers were removed at sacrifice, RNA was extracted and gene expression was assayed using the Agilent G4121A array.

Keywords: eQTL, mouse, liver

overall_design: 73 arrays were run on 36 laboratory inbred strains, with one to three arrays per strain.

first: Alison

last: Harrill

first: David

last: Threadgill

first: Ivan

last: Rusyn


title: Paramecium tetraurelia reciliation (log phase)

city: Villerubanne

first: Jean-François

last: Gout

address: 43 Bld du 11 Novembre 1918

institute: Laboratoire Biométrie et Biologie Evolutive

summary:
Paramecium cells in log-phase growth were treated for deciliation and total mRNA extracted at two time points after deciliation. This is usefull to study genes involved in cilia biogenesis.
Time points are :
T0 : mRNA extracted before deciliation.
T30 : mRNA extracted 30 minutes after deciliation.
T120 : mRNA extracted 120 minutes after deciliation.

overall_design: Experimental design has two biological replications of each time point (T0, T30 and T120) plus two supplementary replications of control time point (T0).

first: Jean

last: Cohen

first: Khaled

last: Bouhouch


title: Differential microRNA Expression during Myogenic Progenitor Cell Proliferation and Differentiation

status: Public on Jun 13 2009

city: San Antonio

first: Yongxin

last: Chen

address: 7703 Floyd Curl Drive

department: Surgery

institute: University of Texas Health Science Center at San Antonio

summary: MicroRNAs (miRNAs) are important in the regulation of many biological processes such as growth and development. To evaluate the role of miRNAs in skeletal muscle regeneration, global miRNA expression was measured during muscle cell growth and differentiation. Primary cultures of murine myogenic progenitor cells (MPC) were studied for miRNA expression using quantitative PCR-array. During MPC differentiation or proliferation, 139 or 16 miRNAs, respectively, exhibited significant >2-fold changes. Cluster analysis revealed 5 distinct miRNA expression patterns at different stages of differentiation. Fourteen miRNAs exhibiting >10-fold change during differentiation included miR-1, 10b, 96, 98, 133a, 139-5p, 330, 335-3p, 339-5p, 344, 486, 499, 504, and 598. Ten of these miRNAs were located in introns of protein coding genes, such as miR-499 located in the myosin heavy chain isoform Myh7b. In silico analysis of possible miRNA-mRNA interactions indicated that many of these miRNAs targeted mRNA critically involved in muscle differentiation. Interestingly, several miRNAs targeted different sites in a given mRNA, suggesting coordinated expression of multiple miRNAs to ensure the regulation of essential genes. These results identify differentially expressed miRNAs that could represent new regulatory elements in MPC proliferation and differentiation.

overall_design: Myogenic progenitor cell (MPC) growth and differentiation are key elements duing muscle regeneration. Using defined culture conditions to promote proliferation or differentiation, we profiled miRNA expression in primary cultures of murine MPC.


title: Expression data from Neisseria gonorrhoeae (FA19)

status: Public on Dec 26 2008

city: Oklahoma City

first: Lydgia

last: Jackson

state: OK

address: 975 NE 10th Street

department: Microbiology and Immunology

institute: University of Oklahoma Health Sciences Center

summary:
Naturally occurring mtrR mutants of gonococci displaying clinically relevant levels of antibiotic resistance are often isolated from patients and mtrR mutants have been reported to be more fit than the wild type parent strain in a murine vaginal infection model. DNA-binding proteins, such as MtrR, that negatively regulate bacterial efflux pump genes have been considered to be “local” gene regulators, although there is increasing evidence that they can directly or indirectly influence expression of other genes. To define the regulatory properties of MtrR we employed microarray analysis of isogenic MtrR-positive and MtrR-negative gonococci.

Keywords: single time point

overall_design: RNA was isolated at mid-log growth for expression analysis

first: Dave

last: Dyer

first: Jason

last: Folster

first: Paul

last: Johnson

first: Vijaya

last: Dhulipali

first: William

last: Shafer


title: Comparison of TCF hyper-activated and control Huh7 cells - SAGE

status: Public on Sep 30 2009

city: Istanbul

first: Ersen

last: Kavak

address: Bogazici Uni. North Campus, North otopark building, 212

department: Molecular Biology and Genetics

country: Turkey

institute: Bogazici University

summary: The Wnt signaling pathway is involved in many differentiation events during embryonic development and can lead to tumor formation after aberrant activation of its components. Β-catenin, a cytoplasmic component, plays a major role in the transduction of the canonical wnt/ β-catenin signaling. The aim of this study was to identify novel genes that are regulated by active β-catenin/TCF signaling in hepatocellular carcinoma. We selected and expanded isogenic clones from hepatocellular carcinoma-derived Huh7 cells with high and low β-catenin/TCF activities. We showed that, high TCF activity Huh7 cells lead to bigger and more aggressive tumors when xenografted into nude mice. We used SAGE (Serial Analysis of Gene Expression), genome-wide microarray and in silico promoter analysis in parallel, to compare gene expression between low (basal) and high (transfected) β-catenin/TCF activity clones, those had been xenografted into nude mice. We compared and contrasted SAGE and genome-wide microarray data, in parallel. Finally; after combined analysis, we identified BRI3 and HSF2 as novel targets of Wnt/β-catenin signaling in hepatocellular carcinoma.

overall_design: High TCF activity Huh7 cell line (Huh7-S33Y) was compared to control Huh7 cell line (Huh7-Vec) by using 25 ug of total RNA isolated from each sample

first: Kader

last: Cavusoglu

first: Tahsin

last: Saygili

first: Adil

middle: D

last: Duru

first: Mahmut

last: Hiz

first: Ahmet

last: Koman


title: Transcriptome of ang3 mutant shoot apices

status: Public on Oct 14 2009

city: Leuven

first: Rekin's

last: Janky

address: Herestraat 49 Box 816

department: Nucleomics Core

institute: VIB

summary: The role of translation in the regulation of higher plant growth and development is not well understood. Mutational analysis is a powerful tool to identify and study the function of genes related to a biological process, such as growth. Here we analyzed functionally the angusta3 (ang3) narrow leaf mutant. The ANG3 gene was cloned by fine-mapping combined with candidate gene sequencing and it corresponded to the ribosomal protein gene RPL5B. Based on amino acid sequence homology, promoter DNA sequence homology and in silico gene expression analysis, RPL5B was found to be putatively functionally redundant with RPL5A. The morphological analysis of ang3 mutants showed that the leaf lamina area was significantly reduced from the third rosette leaf on, mainly because of decreased width. Cellular analysis of the abaxial epidermal cell layer of the third leaf indicated that the cell number in the mutant was similar to that of the wild type, but the cell size was significantly reduced. We postulate that the reduced cell expansion in the epidermis contributes to the narrow shape of ang3 leaves. Growth was also significantly impaired in hypocotyls and primary roots, hinting at a general role for RPL5B in organ growth, unrelated to dorsi-ventral axis formation. Comparison of the transcriptome of the shoot apices of the mutant and the wild type revealed a limited number of differentially expressed genes, such as MYB23 and MYB5, of which the lower expression in the ang3 mutant correlated with reduced trichome density. Our data suggest that translation is an important level of control of growth and development in plants.

overall_design:
Four hybridizations that each compare ang3 versus the control Ler-genotypes.  Hybridizations were performed as 2 dye swaps with biological replicates.

ang3 and Ler seedlings were sown in vitro on half-strength Murashige and Skoog medium, vernalized for 3 days and germinated at 21C and under a 16-h light regime. The green parts of stage 1.03 plants were harvested (Boyes et al. 2001), consisting of the SAM, the first and second young leaves, the third leaf larger than 1 mm, the fourth leaf smaller than 1 mm and the cotyledons. Per genotype, four replicates were harvested and per replica three repetitions. Total RNA was extracted with the RNeasy Plant Mini Kit (Qiagen, Hilden, Germany) and the samples analyzed In the VIB MicroArrays Facility with the Agilent microarray platform. The processed Cy3 and Cy5 intensities were normalized and statistically analyzed with the Feature Extraction Software (version 10.1.1.1; Agilent Technologies, Santa Clara, CA, USA). P values had to be lower than 0.05 and the absolute value of the log2 ratio had to be higher than one. To visualize overrepresented Gene Ontology (GO) categories, upregulated and downregulated data sets were analyzed with BiNGO (Maere et al. 2005; Fleury et al. 2007).

first: Annemie

last: Van Minnebruggen

first: Pia

last: Neyt

first: Steven

last: De Groeve

first: Griet

last: Coussens

first: Maria

last: Ponce

first: Jose

last: Micol

first: Mieke

last: Van Lijsebettens


title: Identification of Differential Gene Expression During Transition of Bovine Corpus Luteum from Early To Mid–Phase.

status: Public on Feb 23 2009

city: MORGANTOWN

first: MADHUSUDHAN

last: GORAVANAHALLY

state: WV

address: LIFE SCIENCES BUILDING

department: DEPARTMENT OF BIOLOGY

institute: WEST VIRGINIA UNIVERSITY

summary:
Prostaglandin F2 alpha (PGF2alpha) brings about regression of the bovine corpus luteum (CL). This luteolytic property of PGF2alpha is used in the beef and dairy cattle to synchronize estrus. A limitation of this protocol is an insensitivity of the early CL to luteolytic actions of PGF2alpha. The mechanisms underlying luteal sensitivity (LS) are poorly understood. The early CL has maximum number of PGF2alpha receptors; therefore differences in signaling events might be responsible for LS. Hence differential gene expression at two developmental stages, days 4 (D-4) and 10 (D-10) post estrus, might account for differences in signal transduction pathways associated with LS. This possibility was examined in these studies. Microarray analysis (n=3 per stage) identified 167 genes that were differentially expressed (p<0.05). These were categorized into genes involved in cell signaling, steroidogenesis and metabolism, protein degradation, transcription regulation and DNA biosynthesis, protein biosynthesis and modification, extracellular matrix and cytoskeletal proteins, antioxidant property, miscellaneous, and unknown functions. Real-time PCR confirmed the differential expression of 9 randomly selected genes, including protein kinase C inhibitor protein-1 (KCIP-1) and regulator of G-protein signaling 2 (RGS2), observed in microarray. Further, the in vivo effect of exogenous PGF2alpha (n=3 per stage) on selected genes that were found differentially expressed during this developmental transition was examined. PGF2alpha increased the expression of a guanine nucleotide binding protein beta polypeptide 1 (GNB1) in D-4 CL and Ca2+/calmodulin dependent kinase kinase 2 beta (Camkk2) in D-10 CL. Therefore, GNB1, Camkk2, KCIP-1, and RGS2 are candidate genes that might play significant role in acquisition of luteal sensitivity to PGF2alpha.
Keywords: Direct comparison between Day-4 and Day-10 CL, Developmental type comparision

overall_design:
The present experiment made a direct comparision of gene expression in early (Day-4) vs mid-phase CL (Day-10). Each experiment contained two technical relicates per slide. Total of three biological replicates were used ( 3 animals for Day-4 and 3 animals for Day-10). Two Day-4 samples were labelled Cy5 and one Day-10 with Cy5.
Two Day-10 samples were labelled with Cy3 and one Day-4 with Cy3.
The Day-4 sample was control for our experiments.

first: MADHUSUDAN

first: MOHAMED

last: SALEM

first: JIANBO

last: YAO

first: KEITH

last: INSKEEP

first: JORGE

last: FLORES


title: Bioligical pathways and in vitro anti-proliferative activity of Hsp90 inhibition in adult T cell leukemia cells

status: Public on May 27 2009

city: Tokyo

first: Junko

last: Ohyashiki

address: 6-7-1, Nishishinjuku, Shinjuku-ku

department: Intractable Immune System Diseases Research Center

country: Japan

institute: Tokyo Medical University

summary:
Background.  Heat shock protein 90 (Hsp90) is essential for the stability and the function of many client proteins, such as ERB2, C-RAF, CDK4, HIF-1 aplha and AKT. Recent reports demonstrated that inhibition of Hsp90 modulates multiple functions required for survival of human cancer, such as myeloma (Mitsiades et al, Blood:107, 1092, 2006), however, the precise mechanism of anti-cancer effect of Hsp90 inhibition is still uncertain.
Aim. The aim of this study is evaluate the effect of Hsp90 inhibition, and to identify molecular pathways responsible for anti-proliferative effect on ATL cells.
Method. For Hsp90 inhibition, Geldanamycin derivates, 17AAG (17-allylamino -17-demethoxygeldanamycin) and 17DMAG (17-(dimethylaminoethylamino) 17-demethoxygeldanamycin) were used in this study. Interleukin 2-independent ATL cell lines (MT-2 and MT-4) and an interleukin 2-dependent ATL cell line (TaY) were incubated, with or without Hsp90 inhibitors. Fresh ATL cells obtained from patients were also used after obtaining informed consent. Cell numbers at 48 h after incubation with or without Hsp90 inhibitors were assessed with the Cell Counting Kit-8 assay (Dojindo Molecular Technologies, Gaithersburg, MD, USA). For detection of apoptosis, we used Annexin V-biotin apoptosis detection kit (Calbiochem, La Jolla, CA, USA). Gene expression analysis was done using a DNA microarray (NCBI Gene expression omnibus; GPL 2531) and statistical analysis was done by a GeneSifter (VizXlabs, Seattle, WA, USA).
Results. We found cell death induced by Hsp90 inhibitors in all the 3 ATL cell lines as well as patient specimens. Inhibitory concentration (IC50) of 17AAG in 3 ATL cell lines was 300 to 700 nM, and that of 17DMAG was 150 to 200 nM. Fresh ATL cells obtained from patients were more sensitive for either 17AAG or 17DMAG. Gene expression analysis of ATL cells revealed that up-regulation of HSPA1A encoding Hsp70, and genes related to cell cycle arrest (i.e. CDKN1A). Genes regulating cell proliferation or anti-apoptosis (i.e. MYC, BCL2 and Cyclin C), genes related to cytokine or chemokine (i.e. IL9, CCL 17, and CCL27), and notably, genes involved in Wnt/beta-catenin signaling pathway (i.e.TCF7L2 and TCF4), were remarkably repressed.  Inhibition of AKT at the protein level was also evident, suggesting the possibility that AKT may down-regulates beta-catenin/ TCF7L2 pathways in response to Hsp90 inhibitors in ATL cells.
Conclusion. Our results have provided new insights into the complex molecular pharmacology of Hsp90 inhibitors, and suggest that Hsp90 inhibitors might be beneficial as anti-proliferative agent in treating ATL patients.

overall_design: Six samples treated with Hsp90 inhibitors (17-AAG or 17-DMAG) were analyzed in biological duplicate.

first: Ryosuke

last: Kurashina

first: Toshihiko

last: Hirano

first: Kazuma


status: Public on Oct 03 2008

city: Evanston

first: San Ming

last: Wang

state: IL

address: 1001 University Place

department: Center for Functional Genomics

institute: ENH Research Institute, Northwestern University

summary: We performed a genome-wide analysis of gene expression in primary human CD151 myeloid progenitor cells. By using the serial analysis of gene expression (SAGE) technique, we obtained quantitative information for the expression of 37,519 unique SAGE-tag sequences. Of these unique tags, (i) 25% were detected at high and intermediate levels, whereas 75% were present as single copies, (ii) 53% of the tags matched known expressed sequences, 34% of which were matched to more than one known expressed sequence, and (iii) 47% of the tags had no matches and represent potentially novel genes. The correct genes were confirmed by application of the generation of longer cDNA fragments from SAGE tags for gene identification (GLGI) technique for high-copy tags with multiple matches. A set of genes known to be important in myeloid differentiation were expressed at various levels and used different spliced forms. This study provides a normal baseline for comparison of gene expression in myeloid diseases. The strategy of using SAGE and GLGI techniques in this study has broad applications to the genome-wide identification of expressed genes. (Lee S et al. The pattern of gene expression in human CD15+ myeloid progenitor cells. Proc Natl Acad Sci U S A. 98(6):3340-5, 2001;  Lee S et al. Gene expression profiles in acute myeloid leukemia with common translocations using SAGE. Proc Natl Acad Sci U S A. 103(4):1030-5, 2006)

overall_design: Human bone-marrow mononuclear cells were isolated from bone marrow with FicollyPaque solution and stored in liquid nitrogen. Cells from three donors were thawed at 37°C, pooled, and used immediately for the isolation of myeloid progenitor cells with CD15 magnetic beads (Dynal, Oslo, Norway). The cells isolated by CD15 magnetic beads were lysed directly with TRIzol reagent for isolation of total RNA. mRNA was purified from 5 mg of total RNA with oligo(dT)25 beads. cDNA was synthesized with a cDNA synthesis kit, and SAGE was performed according to the SAGE protocol with the exceptions described in the original paper. SAGE-tag sequences were collected with the Big-Dye sequencing kit and ABI377 sequencer, and tag sequences were extracted with SAGE 300 software.

first: Sanggyu

last: Lee

first: Guolin

last: Zhou

first: Terry

last: Clark

first: Jianjun

last: Chen

first: Janet

last: Rowley

first: San


title: The Pseudomonas fluorescens Crc affects the length of the transition phase during diauxic growth

status: Public on Sep 11 2008

city: San Diego

first: Hongfan

last: Jin

state: CA

address: 5501 Oberlin Drive

department: Biotechnology

institute: The Dow Chemical Company

summary:
Pseudomonads, unlike enteric bacteria, prefer to utilize tricarboxylic acid cycle intermediates over glucose. The crc gene has been implicated to impart repression on glucose and other metabolic enzymes in various Pseudomonas species when grown in a mixture of glucose and various organic acids.  The crc gene of P. fluorescens strain MB101 has been identified and a crc deletion derivative was constructed. Unlike a crc mutant of P. aeruginosa described earlier, the P. fluorescens deletion mutant crc did not begin to catabolize glucose until most of the succinate has been utilized in a medium containing both carbon sources.  The transition phase during diauxic growth in succinate/glucose media was found to be shorter in a strain lacking crc and considerably longer in a strain overproducing Crc as compared to the wild type. DNA microarray analyses were performed with wild-type and crc mutant strains during the transition phase from succinate to glucose.  In the wild-type strain, high expression signals were obtained for genes normally induced in cells that approach or are in stationary phase.  By contrast, deletion mutant crc cells showed higher expression signals for genes associated with rapid growth than the wild-type strain.  Taken together, this data indicates that Crc influences the length of the lag during the transition phase from succinate to glucose by directly or indirectly affecting the mRNA levels of target genes.
Keywords: time course strain comparison

overall_design: These are time course experiments to compare crc mutant strain to the wildtype MB101 strain at 5, 6, 7, 8, and 9 ht time points. Dye swaps were included in each time point comparison.  There are 10 pairs in total.

first: Laura

last: Block-Alper

first: Charles

last: Squires

first: Tom

last: Ramseier


title: Expression Profiling of Dll1 mutant mouse lines on different genetic background

status: Public on Jan 01 2009

city: Neuherberg

first: Martin

last: Irmler

state: Bayern

address: Ingolstaedter Landstrasse 1

department: Institute of Experimental Genetics

institute: Helmholtz Zentrum München GmbH

summary:
The evolutionarily conserved Delta-Notch cell-cell signal-transduction pathway regulates the determination of various cell fates, which are important for the morphogenesis and development of numerous organs and tissues in many vertebrates and invertebrates.

The Dll1 gene was mutated by homologous recombination in embryonic stem cells such that amino acids 2-116 were replaced with an in-frame fusion of the lacZ gene of E.coli.
Mouse Dll1 genomic clones were isolated from a strain 129/SvPas library. The replacement vector was constructed by inserting a 4 kb ClaI/XhoI fragment, which encodes a part of the beta-galactosidase gene followed by the PGK-neo cassette, and a 2.6 kb XhoI/EcoRI fragment as the 3’ homologous region of the replacement vector into pKS+. This vector was linearized with SalI and ClaI, and a 4.1 kb SalI/NcoI as the 5’ homologous region plus an NcoI/ClaI fragment (800 bp) completing the b-galactosidase gene, were inserted, resulting in the complete replacement vector. This vector was linearized with SalI and electroporated in R1 embryonic stem cells. Correctly targeted clones were identified and verified by Southern blot analysis using external probes from the 3’ and 5’ region of the targeted area, and were injected into C57BL/6J embryo to obtain germline transmission.

The two analysed Dll1 mutant mouse lines carry the same mutation but are on different genetic background: 1. Heterozygous F1-animals were backcrossed several times to the 129SV/J wild type; 2. Heterozygous F1-animals were outcrossed 11 generation to C3HeB/FeJ wild type

overall_design: Four organs (liver, spleen, thymus, brain) of two Dll1 mutant mouse lines on different genetic background carrying the same mutation were analysed by cDNA microarray technology. Experiment include 4-5 biological replicates for reference (wildtype) and mutant animals. Up to 4 technical replicates for each mutant mouse were performed. As reference pooled RNA was used. 50% of the chip hybridisations are dye sway experiments.

first: Johannes

last: Beckers

first: Marion

last: Horsch

first: Gerhard

last: Przemeck

first: Isabel

last: Rubia-Aliaga


title: Correlation of Survival with Gene Expression in Oesophageal Cancer

status: Public on Aug 15 2008

city: Sutton

first: Ian

last: Giddings

state: Surrey

address: 15 Cotswold Road

department: Section of Molecular Carcinogenesis

country: United Kingdom

institute: Institute of Cancer Research

summary:
We evaluated and correlated gene expression profiles of pre-treatment endoscopic biopsies in a pre-defined patient population of resectable oesophageal adenocarcinoma who had received neoadjuvant platinum and fluoropyrimidine based chemotherapy with clinical outcome, the primary end-point being overall survival.  At the time of pre-treatment staging EUS, tumour biopsies were collected (approximately 1.5 mm3).  and immediately snap frozen and stored in liquid nitrogen. Total RNA was extracted from the biopsies and amplified using a T7 bacteriophage RNA polymerase based linear amplification protocol, then expression profiling was carried out. The expression levels of 47 genes selected by biological function, fold change and degree of significance were further validated using quantitative reverse transcription-PCR

Keywords: Tumour vs normal comparison

overall_design: 35 samples  - All samples compared to Universal Human Reference


title: Effects of hypothermia on gene expression in zebrafish gills

status: Public on Sep 01 2008

city: Taipei

first: Ming Yi

last: Chou

state: Taiwan

address: 128 Sec. 2 Acdemia Rd.

department: Cellular and Organismic Biology

institute: Acdemia Sinica

summary: Ectothermic vertebrates are different from mammals that are sensitive to hypothermia and they have to maintain core temperature for survival. Why and how ectothermic animals can survive, grow and reproduce in low temperature have been for a long time a scientifically challenging and important inquiry to biologists. We used a microarray to profile the gill transcriptome in zebrafish (Danio rerio) after exposure to low temperature. Adult zebrafish were acclimated to a low temperature of 12 °C for 1 (1-d) and 30 d (30-d), and the gill transcriptome was compared to wild types by oligonucleotide microarray hybridization. Results showed 11 and 22 transcripts were found to be upregulated by low-temperature treatment for 1-d and 30-d respectively, while 56 and 70 transcrips were downregulated. The gill transcriptome profiles revealed that ionoregulation-related gene was highly upregulated in cold-acclimated zebrafish. This observation encouraged us to investigate the role of ionoregulatory genes in zebrafish gills during cold acclimation. Cold acclimation caused upregulation of genes that are essential for ionocyte specification, differentiation, ionoregulation, and acid/base balance, and also increased the numbers of cells expressing these genes. mRNA expression of epithelial Ca2+ channel (ECaC), one of these genes, was increased in parallel with the level of Ca2+ influx, revealing a functional compensation after long-term acclimation to cold. Phospho-histone H3 and TUNEL staining showed that the cell turnover rate was retarded in cold-acclimated gills. These results suggest that gills may sustain their functions by yielding mature ionocytes from preexisting undifferentiated progenitors in low-temperature environments.

overall_design:
The AB strain of zebrafish (D. rerio) was originally obtained from the University of Oregon, and were kept in the zebrafish stock center at Academia Sinica, Taipei, Taiwan. Fish were reared in local tap water at 28 °C and a photoperiod regime of 14-h L/10-h D. Adult zebrafish were acclimated to 12 °C with a gradually reduced temperature at a gradient of 4 °C/h in order to prevent temperature shock and reduce mortality. After 30 d of acclimation, surviving (over an 80% survival rate) fish appeared to be feeding and behaving normally compared with control fish. The experimental protocols were approved by the Academia Sinica Institutional Animal Care and Utilization Committee (approval no. RFiZOOHP2006083). After the low-temperature treatment, gill tissues dissected from 6 individuals were pooled as a sample and then homogenized in 5 ml Trizol reagent (Invitrogen, Carlsbad, CA). Thirty six individuals (18 for low-temperature treatment and 18 for control) were sacrificed for each microarray hybridization experiment, and another 60 individuals were used for quantitative reverse-transcription polymerase chain reaction. After chloroform extraction, RNA precipitation and ethanol washing, the RNA samples were purified and treated with DNase1 to remove the genomic DNA by using RNeasy Mini Kit (Qiagen, Huntsville, Alabama). The quantity and quality of total RNA were assessed by spectrophotometry and agarose gel electrophoresis, respectively.

The commercial zebrafish 14K oligonucleotides set (MWG Biotech AG, Ebersbach, Germany) were obtained and were printed on UltraGAPS Coated slide (Corning, New York, NY ) with use of the OmniGrid 100 microarrayer (Genomic Solutions, Ann Arbor, MI) according to the manufacture's instructions. The 14,067 oligonucleotides represent 9666 genes (7009 singlet genes and 2657 redundant genes), and the redundancy of this chip is 31 %. The detailed description of the oligonucleotides information can be obtained on the Ocimun Biosolution website (http://www.ocimumbio.com/web/default.asp). cDNA probes were synthesized by reverse transcription of 20 ug total RNA using a SuperScript indirect cDNA labeling system (Invitrogen) and were labeled with Cy5 (cold treatment groups) and Cy3 (control groups)(Amersham Bioscience, Buckinghamshire, UK) , respectively. The zebrafish 14K OciChip array chip was pretreated with 1% bovine serum albumin (BSA) (fraction V), 4x saline-sodium citrate (SSC), and 1% sodium dodecylsulfate (SDS) at 42 °C for 45 min, and then hybridized overnight in a cocktail containing 5x Denhardt's solution, 6x SSC, 0.5% SDS, 50% formamide, 50 mM sodium phosphate, and 2 µg/µl yeast tRNA. Slides were washed with 2x SSC and 0.1% SDS (5 min), 1x SSC and 0.1% SDS (5 min), 0.5x SSC (5 min), and twice with 0.1x SSC (2 min each). Scanning was performed with a Genepix scanner (Molecular Devices, Sunnyvale, CA). The acquired images were analyzed using ScanArray Express 3.0 (PerkinElmer, Waltham, MA) and Genespring (Aglient Technologies, Foster City, CA) software. The measurements of spots were filtered by flags, and the lowest normalization was performed after subtraction of the median background. To assess the differential expressions of genes, we identified the ratio of Cy5/Cy3 intensity > 2 or < 0.5. Each experiment contained 3 biological replicates (including 1 dye swap) with different samples, and the differentially expressed genes were selected from those with at least 2 of 3 significant signals (ratio > 2 or < 0.5).

first: MY

first: CD

last: Hsiao

first: SC

first: IW

first: ST

last: Liu

first: PP

last: Hwang


title: Partial de-repression of the hyphal program does not restore hyphae formation in absence of a functional Arp2/3 complex

status: Public on Jan 23 2010

city: Montréal

first: Elias

last: Epp

address: 1205 Docteur Penfield

department: Biology

country: Canada

institute: McGill University

summary: Deleting components of the Arp2/3 complex in Candida albicans resulted in a global lack of hyphal specific gene induction. This observation suggests that the failure in hyphal growth of Arp2/3 complex mutants could be a result of failure to activate hyphal specific genes. If the hyphal defect was primarily due to failure to activate gene expression, de-repressing hyphal-specific gene expression by deleting the NRG1 repressor could potentially suppress the defect, as deletion of NRG1 leads to constitutive filamentous growth even in the absence of any hyphal induction signals (Garcia-Sanchez et al., 2005, Kadosh & Johnson, 2005). We therefore created an nrg1Δ/Δarp2Δ/Δ mutant. When grown under non-inducing conditions, nrg1Δ/Δarp2Δ/Δ cells showed the arp2Δ/Δ mutant morphology of round and swollen cells. When induced for hyphal growth, nrg1Δ/Δarp2Δ/Δ cells also exhibited the arp2Δ/Δ cell morphology and did not form hyphae even after extended overnight incubation times.

overall_design: To determine if the hyphal-specific genes are de-repressed in the nrg1Δ/Δarp2Δ/Δ mutant, we performed transcript profiling. We compared the nrg1Δ/Δarp2Δ/Δ mutant grown under hyphal conditions to the arp2Δ/Δ mutant grown under the same conditions (10% serum, 37C, three hours), and found that a significant number of hyphal-specific genes that are normally induced when WT cells are undergoing the yeast to hyphae switch (WT-HY) showed greater expression in the nrg1Δ/Δarp2Δ/Δ mutant compared to arp2Δ/Δ cells (p-value 4.9x10-9). When we examined the set of NRG1-dependent hyphal-specific genes previously identified (Kadosh & Johnson, 2005), we found that seven of 28 genes (HYR1, SAP5, SAP4, KIP4, ORF19.6079, ALS3 and UME6) showed significantly increased expression (≥ 2 fold) in nrg1Δ/Δarp2Δ/Δ cells compared to arp2Δ/Δ cells, while a further four genes (IHD1, CBP1, ORF19.6705 and ALS10) showed moderately increased expression between 1.5 and 2-fold. Thus, while deleting a transcriptional repressor of the filamentation program leads to de-repression of many hyphal genes, the entire regulated gene set is not de-repressed; this presumably reflects the complex interplay that different transcriptional (co-) repressors exert on the yeast-to-hyphae transition (Garcia-Sanchez et al., 2005, Kadosh & Johnson, 2005). We further found that despite the increased induction of some hyphal genes in the nrg1Δ/Δarp2Δ/Δ mutant, a few of those genes are not as highly induced as in WT cells. One gene that was induced in both the ‘nrg1Δ/Δarp2Δ/Δ vs arp2Δ/Δ’ and the ‘nrg1Δ/Δarp2Δ/Δ vs WT’-comparisons is UME6, a recently identified key regulator of the hyphal program (Banerjee et al., 2008, Zeidler et al., 2009). Interestingly, although constitutive over-expression of UME6 in WT cells resulted in constitutive filamentous growth even in the absence of hyphae signals (Carlisle et al., 2009), the increased expression level of UME6 in the nrg1Δ/Δarp2Δ/Δ mutant is not sufficient to restore filamentation in the absence of a functional Arp2/3 complex. Thus despite partial de-repression of the hyphal program, hyphae do not form, making it likely other roles of the Arp2/3 complex, such as its function in actin patch formation and actin branching, are required for hyphal development.

first: Andrea

last: Walther

first: Guylaine

last: Lépine

first: Zully

last: Leon

first: Alaka

last: Mullick

first: Martine

last: Raymond

first: Jürgen

last: Wendland

first: Malcolm

last: Whiteway


title: 7 Brain Regions in 20 Inbred Strains of Laboratory Mice

status: Public on Feb 05 2009

first: Hanwen

last: Huang

state: NC

address: University of North Carolina

institute: UNC

summary:
Survey of gene expression in 7 brain regions at 6-8wks of age in 20 inbred strains of laboratory mice.
Autism is a highly heritable neurodevelopmental disorder with no consistent neuropathological hallmarks, diagnosed through behavioral inventory rather than a biomarker.  The genetic etiology of autism is complex and likely has a significant interaction with the environment.  To investigate the molecular and genetic nature of autism, we modeled the core symptom of social deficits in inbred strains of mice.  Using gene expression profiles and behavioral phenotyping relevant to social deficits, we have identified basic cellular functions that may be dysregulated in the brain in autism.  Correlations between these two complex datasets indicate cell morphology, proliferation, and migration may be perturbed in disorders of altered social behavior.  Furthermore, these data in the mouse show significant overlap with published expression profiling in lymphoblastoid lines from autistic individuals.
Keywords: Comparison between inbred strains and brain regions.

overall_design: Each sample consists of tissue pooled from three individual animals.  Each pool was hybridized against the Stratagene RNA standard and is represented by a single array in the dataset.  There are three biological replicates of each strain by region condition.

first: Jessica

last: Nadler

first: Terry

last: Magnuson

first: Fei

last: Zou

first: William

last: Barry

first: Fred

last: Wright


title: Normal Oral Transcriptome

status: Public on Nov 22 2008

city: Vancouver

first: Greg

last: Vatcher

address: 675 West 10th Ave.

institute: BC Cancer Research Centre

summary:
Three SAGE libraries brushed from the surface of the tongue and three SAGE libraries extracted from oral biopsies define the normal oral transcriptome.
Keywords: SAGE, normal oral

overall_design: Six SAGE libraries isolated from the oral cavity were used to define the oral transcriptome.

first: Wan

last: Lam

first: Miriam

last: Rosin


title: Fibrotic Myofibroblasts Manifest Genome-Wide Derangements of Translational Control

status: Public on Nov 21 2008

city: Stockholm

first: Ola

last: Larsson

department: Department of oncology-pathology

country: Sweden

institute: Karolinska Institutet

summary:
As a group, fibroproliferative disorders of the lung, liver, kidney, heart, vasculature and integument are common, progressive and refractory to therapy. They can emerge following toxic insults, but are frequently idiopathic. Their enigmatic propensity to resist therapy and progress to organ failure has focused attention on the myofibroblast – the primary effector of the fibroproliferative response. A central unanswered question is whether these myofibroblasts have acquired a distinct pathological phenotype - or whether they are normal myofibroblasts with a pathological phenotype that depends upon residing in a sea of pro-fibrotic cytokines and an abnormal extracellular matrix.
Using a systems approach to study this question in a prototype fibrotic disease, Idiopathic Pulmonary Fibrosis (IPF); here we show organized changes in the gene expression pathway of primary lung myofibroblasts that persist for up to 9 sub-cultivations in vitro.  When comparing IPF and control myofibroblasts in a 3-dimensional type I collagen matrix, more genes differed at the level of ribosome recruitment than at the level of transcript abundance, indicating pathological translational control as a major characteristic of IPF myofibroblasts. To determine the effect of matrix state on translational control, myofibroblasts were permitted to contract the matrix. Ribosome recruitment in control myofibroblasts was relatively stable. In contrast, IPF cells manifested large alterations in the ribosome recruitment pattern. Pathological studies suggest an epithelial origin for IPF myofibroblasts through the epithelial to mesenchymal transition (EMT). In accord with this, we found systems-level indications for TGF-beta -driven EMT as one source of IPF myofibroblasts.
Keywords: cell type comparison

overall_design: Comparison of lung myofibroblasts from patients with Idiopatic Pulmonary Fibrosis (IPF, 6 donors) to control (6 donors) in contractile and non-contractile matrices using both total and polyribosomal RNA

first: Peter

last: Bitterman


title: N-Tera2 differentiated human neuronal model: Mock infected vs HCoV-OC43 infected

status: Public on Dec 11 2008

city: Laval

first: Dominique

middle: Jean

last: Favreau

state: Quebec

address: 531 boul. des prairies, Bl.18

department: Neuroimmunovirology

institute: INRS-IAF

summary:
Transcriptional profiling of N-Tera2 differentiated human neuronal cells, comparing control uninfected cells to HCoV-OC43 infected cells at 24, 48 and 72 hour post-infection

Keywords: Cell response to viral infection

overall_design: Two-condition experiment, N-Tera2 differentiated human neuronal cell mock infected vs. N-Tera2 differentiated human neuronal cell HCoV-OC43 infected at 24, 48 and 72 hours. Biological replicates: 2 at each time-course point. Technical replicate: 2 dye-swap at each time-point. 2 arrays hybridized with mock(cy3) vs infected(cy5) and 2 array with infected(cy3) vs mock(cy5).

first: Pierre

last: Talbot

first: Marc

last: Desforges


title: Induction of 1C Aldoketoreductases and Other Drug Dose-dependent Genes Upon Acquisition of Anthracycline Resistance

status: Public on Jul 16 2008

city: Sudbury

first: Baoqing

last: Guo

state: ON

address: 41 Ramsey Lake Road

department: Tumor Biology

institute: Advanced Medical Research Institute of Canada

summary:
Recent studies have suggested that elevated expression of aldoketoreductase (AKR) 1C1 or 1C2 in tumour cells is associated with increased resistance to DNA damaging agents such as cisplatin and doxorubicin.  However, it has not been shown whether selection of tumour cells for resistance to DNA-damaging anthracyclines actually results in increased expression of AKRs and increased conversion of anthracyclines to 10-fold less toxic 13-hydroxy metabolites.  It is also unclear whether the induction of aldokeoreductases is temporally correlated with the onset of anthracycline resistance and whether there is a direct relationship between the level of AKR expression or activity and the magnitude of drug resistance.  Through microarray profiling of MCF-7 breast cancer cells selected for progressive resistance to doxorubicin or epirubicin, we have identified several genes whose expression has been correlated with both the onset and magnitude of drug resistance, including a “1C” AKR.  AKR 1C overexpression was verified by quantitative PCR.  Also associated with the onset of anthracycline resistance were genes involved in drug transport (ABCB1), cell signaling and transcription (RDC1, CXCR4), cell proliferation or apoptosis (BMP7, CAV1), ROS protection (TXNRD1, MT2A), and structural or immune system proteins (IFI30, STMN1).  Consistent with the role of AKRs in anthracycline resistance, doxorubicin- and epirubicin-resistant breast tumour cells exhibited 2.2-fold and 6.1-fold higher levels of the 13-hydroxy metabolite of doxorubicin (doxorubicinol) than wildtype MCF-7 cells.  In addition, an inhibitor of AKR 1C2 (5beta-cholanic acid) almost completely restored sensitivity to doxorubicin in Abcb1-deficient doxorubicin-resistant cells, while having no effect on Abcb1-expressing epirubicin-resistant cells.  Taken together, our findings strongly suggest the involvement of multiple genes in the acquisition of anthracycline resistance in breast tumor cells---in particular redox genes such as the 1C AKRs.

Keywords: Drug resistance of breast cancer cells

overall_design: In order to identify genes whose expression strongly correlates with the acquisition or magnitude of drug resistance or are temporally related with the acquisition of resistance, we selected MCF-7 breast tumor cells for survival in increasing concentrations (doses) of doxorubicin or  epirubicin.   Panels  of  cells   exhibiting  progressive   resistance  to  either  doxorubicin (MCF-7DOX-2) or epirubicin (MCF-7EPI) were obtained.  Cells were also “selected” in the absence of drug at each step during selection to serve as co-cultured control (MCF-7CC) cells.  In this study, we have used cDNA microarray analysis of these cell lines to identify a variety of “redox” genes whose expression can be correlated with the acquisition or magnitude of drug resistance in MCF-7DOX-2 and MCF-7EPI cells, including a “1C” aldoketoreductase (AKR).

first: Zachary

last: Veitch

first: Stacey

last: Hembruff

first: David

last: Villeneuve

first: Jamei

last: Eng

first: Monique

last: Laberge

first: Stephanie

last: Cull

last: Maclean

first: Amadeo

last: Parissenti


title: Gene expression profiling of 35S:AtMYB102-SRDXplant under salt stress treatment

status: Public on Jan 21 2010

city: Tsukuba

first: Tomomi

last: Mito

state: Ibaraki

address: Central 4 Higashi 1-1-1

institute: Greensogna,Inc.

summary:
Transcriptional profiling of Arabidopsis transgenic 35S:AtMYB102-SRDX plant under MS medium containing 200mM NaCl for 24h.
Transgenic Arabidopsis (background : Col-0) overexpressing AtMYB102 fused to an active transcriptional repression domain (SRDX) showed strong salt tolerance.

overall_design: Two-condition experiment, Control condition (3-week old plant was transfered to MS medium for 24h) and Salt stress condition (plant was transferred to MS containg 200mM NaCl for 24h), Biological replicates: 3 control replicates and 3 salt stress condition replicates.

first: Seki

last: Motoaki

first: Kazuo

last: Shinozaki

first: Masaru

last: Ohme-Takagi

first: Kyoko

last: Matsui


title: Simultaneous gene expression profiling in human macrophages infected with Leishmania major parasites using SAGE

status: Public on May 22 2008

city: Tunis-Belvedere

first: Dhafer

last: Laouini

address: 13, place Pasteur. BP74

country: Tunisia

institute: Institut Pasteur de Tunis

summary:
Leishmania (L) are intracellular protozoan parasites which are able to survive and replicate within the harsh and potentially hostile phago-lysosomal environment of mammalian mononuclear phagocytes. A complex interplay then takes place between the macrophage (MΦ) striving to eliminate the pathogen and the parasite struggling for its own survival.
To investigate, at the transcriptional level, this host-parasite conflict in the context of monocyte-derived human MΦs (MDM) infection by L.  major metacyclic promastigotes, the quantitative technique of serial analysis of gene expression (SAGE) was used.
After extracting mRNA from resting human MΦs, Leishmania-infected human MΦs and L. major parasites, three SAGE libraries were constructed and sequenced generating up to 28,173; 57,514 and 33,906 tags respectively (corresponding to 12,946; 23,442 and 9,530 unique tags). Using computational data analysis and direct comparison to 394,059 publicly available experimental human tags, the parasite and the host cell transcriptomes were then simultaneously characterized from the mixed cellular extract, allowing to confidently discriminate host from parasite transcripts. This procedure led us to reliably assign 3,814 tags to MΦs’ and 3,666 tags to L. major parasites’ transcripts. We focused on those, showing significant changes in their expression that are likely to be relevant to the pathogenesis of parasite infection: (i) human MΦs genes, belonging to key immune response proteins (i.e. IFNγ pathway, S100 and chemokine families) and (ii) a group of Leishmania genes showing a preferential expression at the intra-cellular developing stage of the parasite.
Dual SAGE transcriptome analysis provided a useful, powerful and accurate approach to discriminate between genes of human or parasitic origin in Leishmania-infected human MΦs. The findings presented in this work suggest that the Leishmania parasite is modulating key transcripts in the human MΦs that may be beneficial for its establishment and survival and provided an overview of gene expression at two developmental stages of the parasite, namely metacyclic promastigotes and intracellular amastigotes, indicating a broad difference between their transcriptomic profiles. Finally, our reported set of expressed genes could deserve future rounds of data mining and gene annotation.
Keywords: Leishmania major, Human macrophages, in vitro, infection, transcriptome, SAGE

sample_organism:
Leishmania major
Homo sapiens

platform_organism:
Leishmania major
Homo sapiens

overall_design: Human monocyte derived macrophages (MDM) from four healthy donors were infected in vitro for 24 hours with metacyclic Leishmania major parasites (ratio 1:5) and the pool was used to construct SAGE library. Non infected MDM from the same donors and from metacyclic Leishmania major parasites were used to construct the two controls' SAGE libraries.

first: Fatma

middle: Z

last: Guerfali

first: Lamia

last: Guizani-Tabbane

first: Florence

last: Ottones

first: Khadija

last: Ben-Aissa

first: Ons

last: Mghirbi

first: Laurent

last: Manchon

first: Sondos

last: Smandi

first: Alia

last: Benkahla

first: Therese

last: Commes

last: Piquemal

first: Jacques

last: Marti

first: Koussay

last: Dellagi


title: Female adult hornfly gene expression vs male adult hornfly gene expression

status: Public on Jun 19 2008

city: Lubbock

first: Scot

last: Dowd

address: 4321 Marsha Sharp Fwy

department: MBRI

institute: MBRI

summary:
Background: The horn fly, Haematobia irritans (L.), is an obligate blood-feeding parasite of cattle and control of this pest is a continuing problem in the United States and other parts of the world. Worldwide annual economic losses attributable to this pest surpass $1 billion. The fly is becoming resistant to pesticides and new control technologies are needed by cattle producers.
Results: Dominant conditional lethal gene systems are being investigated as population control technologies against agricultural insect pests. One of the critical components of these systems is a highly expressed female-specific gene promoter which can be used to drive expression of a lethality-inducing gene. To identify candidate genes to supply this gene promoter, microarrays were designed from a recently developed horn fly EST database and probed to identify female-specific and larval-specific differential gene expression. Analysis of dyeswap experiments found 432 and 417 transcripts which were over- and under-expressed in adult female flies, respectively, compared to adult male flies. Additionally, 419 and 871 transcripts were over- and under-expressed in first instar larvae compared to adult flies. Three transcripts were identified which were over-expressed in adult females flies compared to adult males and which also were over-expressed in the first instar larval lifestage compared to adult flies.
Conclusion: We have identified 3 strong candidates for further evaluation as a gene promoter source for the development of a female-specific conditional lethality system in the horn fly. One of these candidates, the putative nanos orthologue, has a high female-to-male expression ratio, has a moderate expression level in first instar larvae, and has been well characterized in D. melanogaster. Further investigations leading from this microarray analysis will include transformation of the horn fly and evaluation of the female conditional lethal system components for applicability to the specific biological parameters of natural populations of the horn fly.
Keywords: adult male vs female hornfly gene expression

overall_design: In short, RNA from female horn flies was labeled and hybridized against labeled RNA from male horn flies in a dyeswap design. Similarly, RNA from first instar larvae was labeled and hybridized against labeled a 50:50 mix of RNA from adult male and female flies. All labeling, hybridization and washing procedures were performed according to respective manufacturer protocols. For each sample, 10 ug of total RNA and 50-fold dilution of Agilent two color spike-in control RNA kit (Agilent Technologies Inc.) were labeled with either CyDye3-dCTP or CyDye5-dCTP (Amersham Biosciences, Piscataway, NJ) using the LabelStar kit (Qiagen Inc., Valencia, CA) and oligo-dT and Random nonamers (Sigma-Aldrich Inc., St. Louis, MO). Labeled cDNA was hybridized to the Agilent microarrays using the Gene Expression Hybridization kit (Agilent Technologies Inc.) following manufacturer’s protocols. Arrays were washed with Gene Expression Wash Buffer kit (Agilent Technologies Inc.) A total of 8 arrays, including a dye swap for each RNA replicate (4 biological replicates), were analyzed to obtain genes that were consistently regulated while limiting false discover rate (FDR) below 2.5% [26].

middle: E

first: Felix

last: Guerrero


title: In vivo gene expression in granulosa cells during pig terminal follicular development- oligo

status: Public on Apr 30 2008

city: Castanet-Tolosan Cedex

first: Agnes

last: Bonnet

address: Chemin de Borde-Rouge  BP 52627

institute: INRA

summary:
These vector oligonucleotide hybridizations were performed  to determine the quality of the spotting process. EST with low signal values of the vector array were excluded from the analysis, since they probably were mis-amplified or mis-spotted
Keywords: spotting control

overall_design:
cDNA arrays (GPL3978) were first hybridized with a vector oligonucleotide labeled (T7 probe) with {alpha}33P-ATP. After washing, the arrays were exposed 6 hours to radioisotopic-sensitive imaging plates (BAS-2025, Fujifilm, Raytest France S.A.R.L., Courbevoie, France). The imaging plates were scanned thereafter with a phosphor imaging system at a 25 µm resolution (BAS-5000, Fujifilm, Raytest France S.A.R.L., Courbevoie, France). The images were quantified using semi-automated software, BZScan (Lopez F, Rougemont J, Loriod B, Bourgeois A, Loi L, Bertucci F, Hingamp P, Houlgatte R, and Granjeaud S. Feature extraction and signal processing for nylon DNA microarrays. BMC Genomics 5: 38, 2004). Fixed circle segmentation, i.e. a grid process with a fixed spot diameter was applied. The hybridization images were quantified by the extraction of the intensity for each spot. A new version of BZsan (AGscan solfware) is available at :
http://mulcyber.toulouse.inra.fr/projects/agscan/
The experimental data were managed using the free BASE software (Saal LH, Troein C, Vallon-Christersson J, Gruvberger S, Borg A, and Peterson C. BioArray Software Environment (BASE): a platform for comprehensive management and analysis of microarray data. Genome Biol 3: SOFTWARE0003, 2002.) modified by Sigenae team to accept radioactive experiments.
After stripping, the arrays were hybridized with a complex target.
The data correspond to the hybridization of the first membrane DEV-1 and the last one DEV-20.
The arrays contained PCR products from 2848 pig cDNA clones (GPL3978), spotted in duplicate on two separate fields of the same membrane (last number 1 of the local ID corresponds to the left field and 2 corresponds to right field).
Finally, data consisted in four data files and were used to delete clones with a bad quality of spotting.


title: Drosophila 16-20 hrs embryos: DHR3G60S/Df(2R)12 vs. w1118

status: Public on Dec 01 2009

city: Salt Lake City

first: Carl

last: Thummel

department: Human Genetics

institute: University of Utah

summary: Transcriptional profiling of 16-20 hrs Drosophila embryos comparing control (w1118) and DHR3 mutants (DHR3G60S/Df(2R)12)

overall_design: Two-condition experiment, control vs. DHR3 mutant embryos. Biological replicates: 4 replicates.

first: Anne-Françoise

last: Ruaud

middle: S


title: Gene expression profiles of head and neck squamous cell carcinoma

status: Public on Sep 30 2008

city: Ribeirão Preto

first: Daniel

middle: Guariz

last: Pinheiro

state: São Paulo

address: Rua Tenente Catão Roxo, 2501

department: Departamento de Genética

country: Brazil

institute: Faculdade de Medicina de Ribeirão Preto (Universidade de São Paulo)

summary: Sage performed on microdissection of Head and Neck tumor, and Head and Neck normal tissue

overall_design: comparative analysis of gene expression profiles of head and neck squamous cell carcinoma and Head and Neck normal tissue