title: Histone H2B R102A

status: Public on Feb 28 2010

city: Pullman

first: John

middle: J.

last: Wyrick

state: WA

address: Biotechnology Life Sciences 241

department: School of Molecular Biosciences

institute: Washington State University

summary: Total RNA from three replicate cultures of wild-type and mutant strains was isolated and the expression profiles were determined using Affymetrix arrays. Comparisons between the sample groups allow the identification of genes regulated by histone H2B R102A mutant.

overall_design: Comparison of genes whose mRNA levels are affected in a histone H2B R102A mutant yeast strain compared to wild-type yeast strain.

first: McKenna

last: Kyriss


title: Histone H2B K111A

country: USA

summary: Total RNA from three replicate cultures of wild-type and mutant strains was isolated and the expression profiles were determined using Affymetrix arrays. Comparisons between the sample groups allow the identification of genes regulated by histone H2B K111A mutant.

platform_organism:
Schizosaccharomyces pombe
Saccharomyces cerevisiae

overall_design: Comparison of genes whose mRNA levels are affected in a histone H2B K111A mutant yeast strain compared to wild-type yeast strain.

middle: J


title: Expression data from rat cardiac fibroblasts

status: Public on Mar 23 2010

city: Kawasaki

first: Shihori

last: Tanabe

state: Kanagawa

address: 3-25-26, Tonomachi, Kawasaki-ku

department: Division of Risk Assessment

country: Japan

institute: National Institute of Health Sciences

summary:
The overall data contains expression profile of rat genes of cardiac fibroblasts. G-protein signaling is regulated in cardiac fibroblasts. The expresssion of angiotensin II receptor or cytokine such as interleukin is regulated by PTX (which is suggested to suppress Gi function) treatment.
Keywords: control or PTX treatment

overall_design: Rat cardiac fibroblasts were primarily isolated and plated onto 6 well plate. Cells were treated with water (Ctl 1-3) or PTX for 24 hr (PTX 1-3) and lysed by Buffer RLT. Total RNA was isolated using QIAGEN kit and gene expression was analysed using Affymetrix Gene Chip system and GCOS analysis.

first: Motohiro

last: Nishida

first: Yoji

last: Sato

first: Hitoshi

last: Kurose

first: Taku

last: Nagao


status: Public on Mar 01 2010

city: Zurich

first: Sarah-Maria

last: Fendt

country: Switzerland

institute: ETH Zurich

summary: Expression data from wild-type FY4 and GCR2 deletion strain. Impact of the transcription factor Gcr2p on mRNA expression was investigated in the corresponding deletion strain in exponentially growing glucose minimal medium batch cultures.

overall_design: Both streins were grown on glucose minimal medium and harvested during exponential growth for transcript analysis

first: Ana

middle: P

last: Oliveira

first: Uwe

last: Sauer


title: Glucose uptake is required for glucose-stimulated transcriptional response in Saccharomyces cerevisiae

status: Public on Mar 02 2010

city: Göteborg

first: Goutham

middle: N

last: Vemuri

address: Kemivagen 10

department: Chemical and Biological Engineering

country: Sweden

institute: Chalmers University of Technology

summary:
Saccharomyces cerevisiae developed elegant mechanisms to monitor nutrient availability and trigger adaptative responses to nutrient deficiency. Nutrient sensing requires close coordination of cell surface sensors with intracellular mechanisms. This yeast senses the presence of glucose by two modified hexose transporters, Rgt2 and Snf3 (regulating expression of genes encoding hexose transporters) and the G-protein coupled receptor Gpr1 (modulating Protein Kinase A (PKA) activity).. It has been difficult to differentiate between cellular responses mediated by cell surface and intracellular sensors, respectively. Using a strain that is devoid of glucose uptake, we show that the mere presence of glucose does not elicit any glucose-dependent transcriptional responses. This indicates that signals generated by surface sensors are not sufficient to mediate glucose-dependent transcriptional responses. Instead, intracellular glucose or metabolites derived from it are required for transcriptional changes associated with glucose exposure.
We used microarrays from biological triplicate samples to measure the global transcriptional response to sudden addition of glucose to yeast cells growing at steady state on ethanol. The experiment was conducted using a strain that is devoid of glucose uptake and compared with an isogenic strain.

overall_design: The CEN.PK strain was used in this research. A strain with all known hexose transporters deleted (Null strain) was compared with an isogenic reference. The two strains were grown in a chemostat (D = 0.1 h-1) using ethanol as the carbon source. Transcriptional responses between the strains were measured in biological triplicates at steady state and then pulsed with glucose at time t = 0. Transcriptional response was measured again after t = 20 min to determine changes in gene expression chanes only in response to the presence of glucose.

first: G


title: Responses to ethanol in haploid and diploid strain

status: Public on Jan 30 2010

city: Tianjin

first: Bing-Zhi

last: Li

address: Weijin Road 92#

country: China

institute: Tianjin University

summary:
Diploid and haploid strains often exhibit different tolerance to variety of stresses. Transcriptome  of acclimation to ethanol stress in diploid and haploid strain of Saccharomyces cerevisiae was analyzed.
We analyzed transcriptome profiles of diploid and haploid strains in the presence of ethanol.

overall_design: Haploid and diploid strains were cultured in YEPD media with 0%, 3% and 7% ethanol(v/v) in fermentors. The samples were collected at the growth stage for each strain under different conditions.

first: Ying-Jin

last: Yuan


title: Eos1 involvement in zinc homeostasis

status: Public on Mar 31 2010

city: Tsukuba

first: Jun

last: Shima

state: Ibaraki

address: 2-1-12 Kannondai

institute: National Food Research Institute

summary: EOS1 is required for tolerance to oxidative stress in Saccharomyces cerevisiae; mutants are defective in the gene sensitive to hydrogen peroxide and tolerant to tunicamycin. To clarify the function of Eos1, we screened yeast genomic DNA libraries for heterologous genes that, when overexpressed from a plasmid, can suppress the hydrogen peroxide-sensitive eos1 mutation. We identified such a gene as IZH2, which has previously been reported to be a Zap1-regulated gene. However, the EOS1 and IZH2 genes do not themselves appear to be functionally interchangeable. Double disruption of the EOS1 and IZH2 genes yielded a slow-growth phenotype, suggesting that the two proteins are involved in related cellular processes. DNA microarray analysis revealed decreased expression of Zap1-regulated genes in the eos1-deletion mutant (delta eos1). Thus, it is likely that Eos1 is involved in zinc homeostasis.

overall_design: Yeast cells were cultivated in YPD at 30 oC with shaking at 150 rpm, and then the cells were harvested at the log phase (OD600=1.0). Affymetrix Yeast Genome 2.0 arrays (Affymetrix, Santa Clara, CA, USA) were used as the DNA microarrays. Statistical analysis of the expression data were performed using GeneSpring ver. 7.3.1 (Agilent Technologies, Palo Alto, CA, USA) based on the gene expression data from two independent experiments.

first: Toshihide

last: Nakamura

first: Shunsuke

last: Takahashi

first: Hiroshi

last: Takagi


title: S. pombe genome-wide nucleosome mapping reveals positioning mechanisms distinct from S. cerevisiae

status: Public on Feb 01 2010

city: Martinsried

first: Tobias

last: Straub

address: Großhadener Str. 9

department: Biomedical Center, Bioinformatics

country: Germany

institute: LMU Munich

type: Genome binding/occupancy profiling by genome tiling array

summary: Positioned nucleosomes limit the access of proteins to DNA and implement regulatory features encoded in eukaryotic genomes. Here we generated the first genome-wide nucleosome positioning map for Schizosaccharomyces pombe and annotated transcription start and termination sites genome-wide. Using this resource we found surprising differences compared to the nucleosome organization in the distantly related yeast Saccharomyces cerevisiae [the cerevisiae data has been published by others (PMID: 17873876) and the raw data is deposited at ArrayExpress(E-MEXP-1172)]. DNA sequence guides nucleosome positioning differently, e.g., poly(dA:dT) elements are not enriched in S. pombe nucleosome-depleted regions (NDRs). Regular nucleosomal arrays emanate more asymmetrically, i.e., mainly co-directionally with transcription, from promoter NDRs, but promoters harbouring the histone variant H2A.Z show regular arrays also upstream. Regular nucleosome phasing in S. pombe has a very short repeat length of 154 base pairs, and requires a remodeler, Mit1, conserved in humans but not found in S. cerevisiae. Nucleosome positioning mechanisms are evidently not universal but evolutionarily plastic.

overall_design:
For GSM401392-400: Nucleosomal DNA vs. Genomic Input DNA in wildtype, Mit1 mutant and Fft3 mutant S. pombe.

For GSM468642-6: Wildtype and Mit1 mutant S. pombe total RNA. 3 biological replicates for wildtype and 2 for mutant have been collected.

For GSM468647-8: Nucleosomal DNA vs. Genomic Input DNA in wildtype, Mit1 mutant, Fft3 and Tup11/12 mutant S. pombe.

first: A

last: Lantermann

first: T

last: Strålfors

middle: C

first: K

last: Ekwall

first: P

last: Korber


status: Public on Jan 01 2010

city: Potsdam-Golm

first: Dirk

last: Walther

address: Am Mühlenberg 1

department: Bioinformatics

institute: Max Planck Institute for Molecular Plant Physiology

summary:
Understanding the response processes in cellular systems to external perturbations is a central goal of large-scale molecular profiling experiments. We investigated the molecular response of yeast to increased and lowered temperatures relative to optimal reference conditions across two levels of molecular organization: the transcriptome using a whole yeast genome microarray and the metabolome applying the GC/MS technology with in-vivo stable-isotope labeling for accurate relative quantification of a total of 50 different metabolites. The molecular adaptation of yeast to increased or lowered temperatures relative control conditions at both the metabolic and transcriptional level is dominated by temperature-inverted differential regulation patterns of transcriptional and metabolite responses and the temporal response observed to be biphasic. The set of previously described general environmental stress response (ESR) genes showed particularly strong temperature-inverted response patterns. Among the metabolites measured, trehalose was detected to respond strongest to the temperature stress and with temperature-inverted up and downregulation relative to control, mid-temperature conditions. Although associated with the same principal environmental parameter, the two different temperature regimes caused very distinct molecular response patterns at both the metabolite and the transcript level. While pairwise correlations between different transcripts and between different metabolites were found generally preserved under the various conditions, substantial differences were also observed indicative of changed underlying network architectures or modified regulatory relationships. Gene and associated gene functions were identified that are differentially regulated specifically under the gradual stress induction applied here compared to abrupt stress exposure investigated in previous studies, including genes of as of yet unidentified function and genes involved in protein synthesis and energy metabolism.

Reference: Strassburg K, Walther D, Takahashi H, Kanaya S, and Kopka J. Dynamic transcriptional and metabolic responses in yeast adapting to temperature stress. OMICS JIB, 2010, 14(3), in press.

overall_design: Time-series with 8 time points for three different ambient temperatures. Single replicate per time point and temperature series.

first: Katrin

last: Strassburg

first: Joachim

last: Kopka


title: Expression data from kidney transplantions in a fisher-lewis rat model

status: Public on Oct 01 2009

city: Neuherberg

first: Christine

last: von Toerne

address: Ingoldstädter Landstraße 1

department: Proteinanalytik

institute: HelmholtzZentrum

summary:
Most kidney allograft losses are caused by chronic allograft dysfunction (CAD) that is characterized by interstitial fibrosis, tubular atrophy and a smoldering inflammatory process. The aim of the study was to correlate changes in gene expression over time, as evidenced by effects on regulatory pathways linked to the development of fibrosis and inflammation during the development of chronic damage. Renal allografts were harvested for time points 0d, 7d, 14d and 56d (n=3-5) and examined for steady state mRNA expression using Affymetrix microarray RG-U34A. A select group of genes previously associated with chronic fibrosis was then examined in the context of progressive dysfunction. In order to verify the microarray analysis, qPCR has been performed.
Microarray data was used to obtain transcriptomic changes reflecting signaltransduction pathway dysregulation

Timecourse results (log2 placebo/control) linked below as supplementary files.
Keywords: timecourse analysis

overall_design: Progression of rejection of transplanted kidneys was observed in untreated animals at timepoints 7d,14d and 56d after transplantation

first: Herrmann

last: Gröne

first: Peter

last: Nelson


title: The metabolic responce to iron deficiency in Saccharomyces cerevisiae

status: Public on Nov 13 2009

city: Bethesda

first: Caroline

last: Philpott

state: MD

address: 10 Center Dr, Bldg.10-Rm.9B-05

department: NIDDK

institute: National Institutes of Health

summary: Iron is an essential cofactor for enzymes involved in numerous cellular processes. We analyzed the metabolomes and transcriptomes of yeast grown in iron-rich and iron-poor media to determine which biosynthetic processes are altered when iron availability falls.

overall_design: Saccharomyces cerevisiae DBY7286 strain was grown from very low density to mid-log phase (A600 = 0.5, approximately 18 hrs.) in defined-iron SD minimal medium containing only the supplements necessary to meet auxotrophic requirements. Defined-iron SD minimal media were prepared with yeast nitrogen base lacking iron and copper, supplemented with 1 µM copper sulfate, 25 mM MES pH 6.1, 1 mM Ferrozine (Fluka), and the indicated concentrations of ferrous ammonium sulfate 10 µM (low iron) or 300 µM (high iron). All cells were grown at 30°C with shaking and four independent cultures were prepared for each growth condition

first: Minoo

last: Shakoury-Elizeh

first: Olga

last: Protchenko


title: Time-course gene expression profiles of a Saccharomyces cerevisiae wild type and long-lived sch9-delta mutant

status: Public on Dec 25 2009

city: Los Angeles

first: Huanying

last: Ge

state: CA

address: 1050 Childs Way, RRI building 408D

department: Biology

institute: Computational biology and bioinformatics

summary:
The SCH9 null strain has smaller cell size, grows at a slower rate and survives three times longer than wide-type yeast.
This study aims to dissect the mechanisms that lead to the yeast life span extension of  sch9-delta.

overall_design: We measure gene expression profiles of the S. cerevisiae wild type and the long-lived sch9∆ strain every 12 hours from 12 to 120 hours. At each time point, one sample is hybridized to one Affymetrix GeneChip Yeast 2.0 array. In total, ten time points are measured for each strain. When we perform a check of the data quality, all arrays look normal except the 120-hour sch9∆ array. Thus we abandon this array in the further data analysis.

first: Min

last: Wei

first: Paola

last: Fabrizio

first: Chao

last: Cheng

first: Valter

middle: D

last: Longo

first: Lei

middle: M

last: Li


title: High-level furfural resistance in S.  cerevisiae is based on NADPH-dependent reduction by at least two oxireductases

status: Public on Sep 29 2009

first: Dominik

last: Heer

address: Wolfgang-Pauli-Str. 16

department: IMSB

summary:
Resistance of Saccharomyces cerevisiae to high furfural concentration is based on NADPH-dependent reduction by at least two oxireductases.
Biofuels derived from lignocellulosic biomass hold promises for a sustainable fuel economy, but several problems hamper their economical feasibility. One important problem is the presence of toxic compounds in processed lignocellulosic hydrolysates with furfural as a key toxin. While Saccharomyces cerevisiae has some intrinsic ability to reduce furfural to the less toxic furfuryl alcohol, higher resistance is necessary for process conditions. By comparing an evolved, furfural resistant strain and its parent in micro-aerobic, glucose-limited chemostats at increasing furfural challenge, we elucidate key mechanism and the molecular basis of both natural and high-level furfural resistance. At lower furfural concentrations, NADH-dependent oxireductases are the main defence mechanism. At concentrations above 15 mM, however, [1-13C]-flux and global array-based transcript analysis demonstrated that the NADPH-generating flux through pentose-phosphate pathway increases and that NADPH-dependent oxireductases became the major resistance mechanism. The transcript analysis further revealed that iron transmembrane transport is up-regulated in response to furfural. While these responses occur in both strains, high-level resistance in the evolved strain was based on strong induction of ADH7, the uncharacterised ORF YKL071W and 4 further, likely NADPH-dependent oxireductases. By overexpressing the ADH7 gene and the ORF YKL071W, we inverse engineered significantly increased furfural resistance in the parent strain, thereby demonstrating these two enzymes to be key elements of the resistance phenotype.

overall_design: RNA levels were measured in glucose limited, micro-aerobic chemostat cultures with different concentrations of the growth inhibitor furfural. Two strains were compared: TMB3400-FT30-3 is a strain that has been evolutionary adapted to withstand high furfural concentrations. TMB3400 is its less resistant parent. Number of biological replicates: 2-3.

first: Daniel

last: Heine

first: Uwe

last: Sauer


title: Global analysis of gene expression in REF52 cells expressing SV40 T antigens

status: Public on Sep 01 2009

city: Pittsburgh

first: Ulka

last: Sachdev-Ost

address: 300 Halket Street Suite 5414

institute: University of Pittsburgh School of Medicine

summary: SV40 transforms cells through the action of two oncoproteins, large T antigen and small t antigen. Small t antigen targets phosphatase PP2A, while large T antigen stimulates cell proliferation and survival by action on multiple proteins, including the tumor suppressors Rb and p53. Large T antigen also binds components of the transcription initiation complex and several transcription factors. We examine global gene expression in SV40-transformed REF52 cells. SV40 transformation alters the expression of approximately 480 cellular genes. Much of this regulation is consistent with T antigen action on the Rb-E2F pathway.

overall_design: We used the Affymetrix RGU34A microarrays to probe the global mRNA expression from two wild-type REF52neo and three REF52neoT stable cell lines. Cells were grown to two days post-confluence and harvested.

first: Paul

middle: G

last: Cantalupo


title: A Core MYC Gene Expression Signature is prominent in basal-like cancer

status: Public on Sep 22 2009

city: Chapel Hill

first: Charles

middle: M.

last: Perou

state: NC

address: 12-044 Lineberger Comprehensive Cancer Center CB# 7295

department: Professor of Genetics, and Pathology & Laboratory Medicine; Lineberger Comprehensive Cancer Center

institute: University of North Carolina at Chapel Hill

summary:
BACKGROUND: The MYC oncogene contributes to induction and growth of many cancers but the full spectrum of the MYC transcriptional response remains unclear.

METHODOLOGY/PRINCIPAL FINDINGS: Using microarrays, we conducted a detailed kinetic study of genes that respond to MYCN or MYCNDeltaMBII induction in primary human fibroblasts. In parallel, we determined the response to steady state overexpression of MYCN and MYCNDeltaMBII in the same cell type. An overlapping set of 398 genes from the two protocols was designated a 'Core MYC Signature' and used for further analysis. Comparison of the Core MYC Signature to a published study of the genes induced by serum stimulation revealed that only 7.4% of the Core MYC Signature genes are in the Core Serum Response and display similar expression changes to both MYC and serum. Furthermore, more than 50% of the Core MYC Signature genes were not influenced by serum stimulation. In contrast, comparison to a panel of breast cancers revealed a strong concordance in gene expression between the Core MYC Signature and the basal-like breast tumor subtype, which is a subtype with poor prognosis. This concordance was supported by the higher average level of MYC expression in the same tumor samples.

CONCLUSIONS/SIGNIFICANCE: The Core MYC Signature has clinical relevance as this profile can be used to deduce an underlying genetic program that is likely to contribute to a clinical phenotype. Therefore, the presence of the Core MYC Signature may predict clinical responsiveness to therapeutics that are designed to disrupt MYC-mediated phenotypes.

overall_design: reference X sample

first: Sanjay

last: Chandrian


title: S. pombe acetyltransferase mutants identifies redundant pathways of gene regulation, Affymetrix dataset

status: Public on Sep 20 2009

first: Susan

last: Forsburg

address: 1050 Childs Way, RRI 201B

department: Molecular and Computational Biology

institute: University of Southern California

summary: Analysis of histone acetyl transferases (HATs) from the MYST and GNAT families in S. pombe to identify functional differences or overlap with regard to gene expression. Mutations were made to Elp3 and Gcn5 (GNAT family), and to Mst2 (MYST family). Mutants showed distinct phenotypes which were repressed or enhanced by mutant combinations.

overall_design: Mutations were made to Elp3 and Gcn5 (GNAT family), and to Mst2 (MYST family) in S. pombe. RNA was extracted from the various mutants and hybridized on Affymetrix arrays.

first: Rebecca

last: Nugent

first: Anna

last: Johnsson

first: Brian

last: Fleharty

first: Madelaine

last: Gogol

first: Yongtao

last: Xue-Franzén

first: Chris

last: Seidel

first: Anthony

last: Wright

middle: L


title: Expression profiling of S. pombe acetyltransferase mutants identifies redundant pathways of gene regulation

summary:
Analysis of histone acetyl transferases (HATs) from the MYST and GNAT families in S. pombe to identify functional differences or overlap with regard to gene expression. Mutations were made to Elp3 and Gcn5 (GNAT family), and to Mst2 (MYST family). Mutants showed distinct phenotypes which were repressed or enhanced by mutant combinations.

This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Insufficiency of Copper Ion Homeostasis Causes Freeze-Thaw Injury of Yeast Cells

status: Public on Aug 31 2009

first: Jun

last: Shima

summary: Saccharomyces cerevisiae is exposed to freeze-thaw stress in commercial processes including frozen dough baking.  The cell viability and fermentation activity after freeze-thaw were dramatically decreased due to freeze-thaw injury.  Because freeze-thaw injury involves complex phenomena, the mechanisms of it are not fully understood.  We attempted to analyze the mechanisms of freeze-thaw injury by indirect gene expression analysis during post-thaw incubation after freeze-thaw treatment using DNA microarray profiling.  The results showed that a high frequency of the genes involved in the homeostasis of metal ions were up-regulated depending on the freezing period.  The phenotype of the deletion mutants of the up-regulated genes extracted by indirect gene expression analysis was assessed.  The deletion strains of the MAC1 and CTR1 genes involved in copper ion homeostasis exhibited freeze-thaw sensitivity, suggesting that copper ion homeostasis is required for freeze-thaw tolerance.  Supplementation with copper ions during post-thaw incubation increased intracellular superoxide dismutase activity.  Inverse correlated with intracellular superoxide dismutase activity, intracellular levels of reactive oxygen species were decreased.  Moreover, cell viability increased by supplementation with copper ions under specific assessment conditions.  This study suggested that insufficiency of copper ion homeostasis may be one of the causes of freeze-thaw injury.

overall_design: Total RNA was extracted from the stress-treated yeast cells by using a hot phenol method. Poly(A)+ RNA was enriched from total RNA by using an Oligotex dT30 (Super) mRNA purification kit (Takara Bio, Ohtsu, Japan). cDNA synthesis, cRNA synthesis, and labeling were performed according to the Affymetrix user’s manual (Affymetrix, Santa Clara, USA). Biotinyated cRNA was fragmented and then used as a probe.Affimetrix Yeast Genome 2.0 arrays (Affymetrix) were used as DNA microarrays. All experiments were done in triplicate independently.

first: Shunsuke

last: Takahashi

first: Hiroshi

last: Takagi


title: Gene expression analysis of rat livers treated with pharmaceutical development compounds

status: Public on Jul 29 2009

city: Basel

first: Adrian

middle: B

last: Roth

address: Grenzacherstr 124

department: NCS

institute: Roche

summary:
We used microarrays to analyze gene expression changes in liver after treatment of rats with two compounds from drug development (R1, R2) to identify potential effects related to hepatotoxicity.
The development cpds have been described in:
Pierson et al., J Med Chem 52: 3855-62.
R1 corresponds to 36, R2 corresponds to 38

overall_design: Compound R1 or R2, or vehicle control, was given for either 24h or 2 weeks (336h) at different doses.

first: Guido

last: Steiner


title: Cell-type Specific Regulation of Gene Expression by Simian Virus 40 T antigens

first: Ulka

last: Sachdev-Ost

summary: SV40 transforms cells through the action of two oncoproteins, large T antigen and small t antigen. Small t antigen targets phosphatase PP2A, while large T antigen stimulates cell proliferation and survival by action on multiple proteins, including the tumor suppressors Rb and p53. Large T antigen also binds components of the transcription initiation complex and several transcription factors. We examined global gene expression in SV40-transformed mouse embryo fibroblasts, and in enterocytes obtained from transgenic mice. SV40 transformation alters the expression of approximately 800 cellular genes in both systems. Much of this regulation is observed in both MEFs and enterocytes and is consistent with T antigen action on the Rb-E2F pathway. However, the regulation of many genes is cell-type specific, suggesting that unique signaling pathways are activated in different cell types upon transformation, and that the consequences of SV40 transformation depends on the type of cell targeted.

sample_organism:
Mus musculus
Rattus norvegicus

platform_organism:
Mus musculus
Rattus norvegicus

overall_design: We used the Affymetrix RGU34A microarrays to probe the global mRNA expression from two wild-type REF52neo and three REF52neoT stable cell lines. For the MEF microarrays, we used a dye-flip reference design (Churchill GA, 2002, Nature Genetics, Vol. 32, p.490-495) to compare a pool of MEF total RNA to three independent T antigen MEF cell lines. All REF52 and MEF cells were grown to two days post-confluence and harvested. For the mouse small intestine microarrays, we used three different intestinal preparations: Whole, Villi and Vlcm in a replicated dye-flip design (Churchill GA) using three (Vlcm) or four (Whole and Villi) T+ mice and the same number of their wild-type littermates for each intestinal preparation. We used the Agilent Mouse cDNA (G4104A) array for the MEF and small intestine experiments.


title: Toxicogenomic Comparison of TCDD and PCB 126 Responsiveness in Primary Rat Hepatocytes

status: Public on Aug 19 2009

city: Niskayuna

first: Erik

middle: A

last: Carlson

state: NY

address: 1 Research Circle

institute: General Electric

summary: (Abstract) Toxicogenomics has great potential for enhancing our understanding of environmental chemical toxicity, hopefully leading to better-informed human health risk assessments. This study employed toxicogenomic technology to reveal species differences in response to two prototypical aryl hydrocarbon receptor (AHR) agonists, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and the polychlorinated biphenyl (PCB) congener PCB 126. Dose responses of primary cultures of rat and human hepatocytes were determined using species-specific microarrays sharing over 4,000 gene orthologs. Forty-seven human and 79 rat genes satisfied dose response criteria for both chemicals and were subjected to further analysis including the calculation of EC50 and the relative potency (REP) of PCB 126 for each gene. Only 5 responsive orthologous genes were shared between the two species, yet the geometric mean of the REPs for all rat and human modeled responsive genes were 0.06 (95% Confidence Interval (CI); 0.03-0.1) and 0.002 (95% CI; 0.001-0.005), respectively, suggesting broad species differences in the initial events that follow AHR activation but precede toxicity. This indicates that there are species differences in both the specific genes that responded and the agonist potency and relative potency for those genes. This observed insensitivity of human cells to PCB 126 is consistent with more traditional measurements of AHR activation (i.e., CYP1A1 enzyme activity) and suggests that the species difference in PCB 126 sensitivity is likely due to certain aspects of AHR function. That a species divergence also exists in this expanded AHR-regulated gene repertoire is a novel finding and should help when extrapolating animal data to humans.

overall_design: Primary hepatocyte cultures, separated into 2 pools of 3 female Sprague Dawley rats each, were treated with vehicle (0.5% DMSO), TCDD (-14 to -6.5 log10 M) , or PCB 126 (-12 to -5 log10 M) for 48h. Total RNA was extracted and screened with RG-U34A microarrays for dose response.

first: Colin

last: McCulloch

first: Aruna

last: Koganti

first: Shirlean

last: Goodwin

first: Thomas

middle: R

last: Sutter

first: Jay

last: Silkworth


title: Transcriptional Profiling of Saccharomyces cerevisiae Upon Exposure to Saxitoxin

status: Public on Jun 18 2009

city: Knoxville

first: Kathleen

last: Cusick

state: TN

address: 676 Dabney Hall

department: Ctr for Environmental Biotechnology

institute: University of Tennessee

summary: Saxitoxin is a potent neurotoxin produced by several species of dinoflagellates and cyanobacteria. The molecular target of saxitoxin in higher eukaryotes is the voltage-gated sodium channel; however, its target in lower eukaryotic organisms remains unknown. The goal of this study was to obtain the transcriptional fingerprint of the model lower eukaryote Saccharomyces cerevisiae upon exposure to saxitoxin to identify potential genes suitable for biomarker development. Microarray analyses identified multiple genes associated with copper and iron homeostasis and sulfur metabolism as significantly differentially expressed upon exposure to saxitoxin; these results were verified with quantitative reverse-transcriptase PCR (qRT-PCR). Additionally, the qRT-PCR assays were used to generate expression profiles in a subset of the differentially regulated genes across multiple exposure times and concentrations, the results of which demonstrated that overall, genes tended to respond in a consistent manner to the toxin. In general, the genes encoding the metallothioneins CUP1 and CRS5 were induced following exposure to saxitoxin, while those encoding the ferric/cupric reductase FRE1 and the copper uptake transporter CTR1 were repressed. The gene encoding the multicopper ferroxidase FET3, part of the high-affinity iron uptake system, was also induced in all treatments, along with the STR3 gene, which codes for the cystathionine beta-lyase found in the methionine biosynthetic pathway.

overall_design: Cultures of S. cerevisiae S288C were grown to OD600=1.0 in YPD (1% yeast extract, 2% peptone, 2% dextrose). Four hundred microliters from individual 10 mL overnight cultures were transferred to sterile tubes and saxitoxin added to a final concentration of 16 µM. Sample were mixed gently and incubated at 30° C for 45 min. Controls consisted of deionized water. Three biological replicates were performed under these conditions for microarray analysis. Cells were harvested via sequential centrifugation and placed immediately in liquid nitrogen.Total RNA was extracted from frozen cell pellets using a protocol that combined the hot acid phenol method with the RNEasy kit. Total RNA was processed by the Affymetrix Core Facility at the University of Tennessee (Knoxville, TN) following the protocols of the Affymetrix GeneChip Expression Analysis Technical Manual. MAS5.0 was used to provide present/absent calls; genes absent in all six samples were then removed from the data set. Intensity values for all transcripts were obtained from the scanned image files of the chips. The intensity values of the six arrays were background-corrected and normalized with the GC-RMA algorithm.

first: Gregory

last: Boyer

first: Steven

middle: W

last: Wilhelm

first: Gary

middle: S

last: Sayler


title: Expression data from S. cerevisiae under serial stresses

status: Public on Aug 07 2009

city: Rehovot

first: Amir

last: Mitchell

address: Hertzel

country: Israel

institute: Weizmann Inst.

summary:
S. cerevisae cells were exposed to different series of mild stresses. Stress type include heat shock, oxidative and osmotic stresses.
Microarrays were used to follow the genome-wide transcriptional response to the stresses and to identify genes that can underlie the cross protection phenotype between heat shock and oxidative stress.

Keywords: time course

overall_design: Cell sample at different time points after stress application were used for RNA extraction and hybridization on Affymetrix microarrays.

first: Yitzhak

last: Pilpel

first: Orna

last: Dahan


title: Responses to inhibitors in an industrial strain of yeast and the adapted strains

first: Bing-Zhi

last: Li

summary: Furfural, phenol and acetic acid, generated during cellulosic material pretreatment, are the representative inhibitors to yeast used for ethanol production. The responses to these inhibitors in industrial yeast and the corresponding adapted strains were analyzed.

overall_design: We analyzed the transient response to inhibitors and the different transcriptions in industrial yeast and furfural-, phenol-, and acetic acid-adapted strains. Industrial yeast and the adapted strains were collected at 20 minutes after inhibitor addition. The reference samples for industrial yeast and adapted strains were collected at the same time without inhibitor addition. 2 replicates for each strain/treatment were analyzed.

first: Ying-Jin

last: Yuan


title: Involvement of ergosterol contents in the tolerance to vanillin in Saccharomyces cerevisiae

status: Public on Jun 29 2009

first: Jun

last: Shima

summary: The vanillin tolerance Saccharomyces cerevisiae was screened and compared intracellular ergosterol levels with several laboratory yeast strains, to study potential relationship between ergosterol contents and vanillin tolerance.  S. cerevisiae NBRC1950 was selected as a vanillin tolerant strain.  Its ergosterol contents were higher than those of laboratory strains.  The results of DNA microarray and quantitative RT-PCR analysis showed that 5 genes involved in ergosterol biosynthesis (ERG28, HMG1, MCR1, ERG5 and ERG7) were up-regulated in NBRC 1950 compared with strain X2180, suggested that high expressions of genes involved in ergosterol biosynthesis may cause for the high ergosterol content in strain NBRC 1950.  S. cerevisiae HX strain, which was a high ergosterol content strain derived from X2180, became more tolerant to vanillin compared with the parental strain.  It is suggested that high ergosterol contents may be in part responsible for vanillin tolerance.  These findings provide a biotechnological basis for the molecular engineering of S. cerevisiae with increased tolerance to vanillin.

overall_design: Total RNA was extracted from cells in YPD media with shakin by using a hot phenol method. Poly(A)+ RNA was enriched from total RNA by using an Oligotex dT30 (Super) mRNA purification kit (Takara Bio, Ohtsu, Japan). cDNA synthesis, cRNA synthesis, and labeling were performed according to the Affymetrix user’s manual (Affymetrix, Santa Clara, USA). Biotinyated cRNA was fragmented and then used as a probe.Affimetrix Yeast Genome 2.0 arrays (Affymetrix) were used as DNA microarrays. All experiments were done in duplicate independently. Statistical analysis after data acquisition and normalization of expression data were performed using GeneSpring ver.7.3.1 (Agilent Technologies, Palo Alto, CA, USA) based on the gene expression data from two independent experiments. After data transformation to GeneSpring, per-chip normalization to the 50th percentile was performed, and per-gene normalization to the specific samples (X2180 samples) was applied to the per-chip normalized data. Quality control was performed based on experimental confidence levels (each condition in which all samples were present or marginal) and on statistical confidence levels (condition in which P-value of T-test comparisons between X2180 and NBRC 1950 was less than 0.05).

first: Ayako

last: Endo

first: Toshihide

last: Nakamura


title: Mat-Lylu cell line compared to G cell line

status: Public on May 12 2009

city: Valhalla

first: Ismael

last: Reyes

address: Microbiology department BSB room 311

institute: New York Medical College

summary:
we analyzed the gene expression profiles of Mat-Lylu cell lines (in duplicate) compared to G cell lines (in duplicate) using Affymetrix tools and dChip software. The objective was to find metastasis-associated genes in prostate cancer, using this in vitro model.
Keywords: cell line comparison

overall_design: we used Mat_lylu cell line samples (duplicates) as the Experiment group and G cell line samples (duplicates) as control group.


title: Analysis of the effect of L-carnitine supplementation on global expression of Sachharomyces cerevisiae cultures

city: Stellenbosch

first: Jaco

last: Franken

state: Western Cape

address: Victoria Street

department: Institute for Winebiotechnology

country: South Africa

institute: University of Stellenbosch

summary:
Microarray analysis of the effect of L-carnitine supplementation on global expression of Sachharomyces cerevisiae cultures in logarithmic growing conditions and after exposure to H2O2 induced oxidative stress

L-Carnitine plays a well documented role in eukaryotic energy homeostasis by acting as a shuttling molecule for activated acyl residues across intracellular membranes. This activity is supported by carnitine acyl-transferases and transporters, and is referred to as the carnitine shuttle. However, several pleiotropic and often beneficial effects of carnitine in humans have been reported that appear to be unrelated to the shuttling activity, but little conclusive evidence regarding the molecular networks that would be affected by carnitine exist. We have recently demonstrated a protective role of carnitine in oxidative stress in yeast that is independent of the carnitine shuttle. A DNA microarray-based global gene expression analysis identified Cyc3p, a cytochrome c heme lyase, as being important for carnitine's protective impact in oxidative stress conditions. These findings establish a direct genetic link to a carnitine-related phenotype that is independent of the shuttle system. The data suggest that the yeast Saccharomyces cerevisiae should provide a useful model for further elucidation of carnitine's physiological roles.

overall_design: Yeast cultures was grown to mid-log phase with and without carnitine supplementation to a final concentration of 100 mg/L.  A second set was grown to mid log phase, with and without carnitine supplementation and exposed to 0.4 mM H2O2 for 30 min.  The experiments were performed using biological duplicate.

first: Florian

middle: F

last: Bauer


title: Cardiac 12/15-lipoxygenase-induced inflammation is involved in heart failure

status: Public on Jun 06 2009

city: Chiba

first: Yosuke

last: Kayama

state: Chiba

address: 1-8-1 Inohana, Chuo-ku

department: Cardiovascular Science and Medicine

institute: Chiba University Graduate School of Medicine

summary: To identify a novel target for the treatment of heart failure, we examined gene expression in the failing heart. Among the genes analyzed, 12/15 lipoxygenase (12/15-LOX) was markedly up-regulated in heart failure. To determine whether increased expression of 12/15-LOX causes heart failure, we established transgenic mice that overexpressed 12/15-LOX in cardiomyocytes. Echocardiography showed that 12/15-LOX transgenic mice developed systolic dysfunction. Cardiac fibrosis increased in 12/15-LOX transgenic mice with advancing age, and was associated with the infiltration of macrophages. Consistent with these observations, cardiac expression of monocyte chemoattractant protein-1 (Mcp-1) was up-regulated in 12/15-LOX transgenic mice compared with wild-type mice. Treatment with 12-hydroxy-eicosatetraenotic acid, a major metabolite of 12/15-LOX, increased MCP-1 expression in cardiac fibroblasts and endothelial cells, but not in cardiomyocytes. Inhibition of Mcp-1 reduced the infiltration of macrophages into the myocardium and prevented both systolic dysfunction and cardiac fibrosis in 12/15-LOX transgenic mice. Likewise, disruption of 12/15-LOX significantly reduced cardiac Mcp-1 expression and macrophage infiltration, thereby improving systolic dysfunction induced by chronic pressure overload. Our results suggest that cardiac 12/15-LOX is involved in the development of heart failure and that inhibition of 12/15-LOX could be a novel treatment for this condition.

overall_design: Heart failure is still one of the leading causes of death worldwide. Therefore, it is important to elucidate the underlying mechanisms of heart failure and develop more effective treatments for this condition. To clarify the molecular mechanisms of heart failure, we performed microarray analysis using cardiac tissue samples obtained from a hypertensive heart failure model (Dahl salt-sensitive rats). ~300 genes showed significant changes of expression in the failing hearts compared with control hearts. Among the genes analyzed, 12/15-lipoxygenase (12/15-LOX) was most markedly up-regulated in failing hearts compared with control hearts .


title: Profiling the transcriptional response upon deletion of PUN1

status: Public on Feb 17 2010

city: Ann Arbor

first: Tao

last: Xu

state: MI

address: 210 Washtenaw Ave

department: Life Science Institute

institute: University of Michigan

summary: Yeast filamentous growth is a stress response to conditions of nitrogen deprivation, wherein yeast colonies form pseudohyphal filaments of elongated and connected cells.  As proteins mediating adhesion and transport are required for this growth transition, the protein complement at the yeast cell periphery plays a critical and tightly regulated role in enabling pseudohyphal filamentation.  To identify proteins differentially abundant at the yeast cell periphery during pseudohyphal growth, we generated quantitative proteomic profiles of plasma membrane protein preparations under conditions of vegetative growth and filamentation.  By iTRAQ chemistry and two-dimensional LC-MS/MS, we profiled 2,463 peptides and 356 proteins, from which we identified eleven differentially abundant proteins that localize to the yeast cell periphery.  This protein set includes Ylr414cp, herein renamed Pun1p, a previously uncharacterized protein localized to the plasma membrane compartment of Can1 (MCC).  Pun1p abundance is increased two-fold under conditions of nitrogen stress, and deletion of PUN1 abolishes filamentous growth in haploids and diploids; pun1D mutants are non-invasive, lack surface-spread filamentation, grow slowly, and exhibit impaired cell adhesion.  Conversely, overexpression of PUN1 results in exaggerated cell elongation under conditions of nitrogen stress.  PUN1 contributes to yeast nitrogen signaling, as pun1D mutants misregulate amino acid biosynthetic genes during nitrogen deprivation.  By chromatin immunoprecipitation and RT-PCR, we find that the filamentous growth factor Mss11p directly binds to the PUN1 promoter and regulates its transcription.  In total, this study provides the first profile of protein abundance during pseudohyphal growth, identifying a previously uncharacterized MCC protein required for wild-type nitrogen signaling and filamentous growth.

overall_design: For this study, we constructed a homozygous diploid strain in the filamentous Σ1278b background deleted for PUN1; a wild-type diploid strain of the same background served as the control.  Both strains were grown under conditions of low nitrogen, and RNA was extracted from three biological replicates of each strain after identical culturing.  The RNA samples were analyzed using affymerix DNA microarrays.

first: Christian

last: Shively

first: Owen

last: Ryan

first: Rui

last: Jin

first: Matthew

last: Eckwahl

first: Craig

last: Dobry

first: Qingxuan

last: Song

first: Charles

last: Boone

first: Anuj

last: Kumar


title: A compact VEGF signature associated with distant metastases and poor outcomes

status: Public on Mar 31 2009

last: Perou

summary:
Results

When compared with each other, primary tumors and regional metastases showed statistically indistinguishable gene expression patterns. Supervised analyses comparing patients with distant metastases versus primary tumors or regional metastases showed
that the distant metastases were distinct and distinguished by the lack of expression of fibroblast/mesenchymal genes, and by the high expression of a 13-gene profile (that is, the ‘vascular endothelial growth factor (VEGF) profile’) that included VEGF, ANGPTL4, ADM and the monocarboxylic acid transporter SLC16A3. At least 8 out of 13 of these
genes contained HIF1α binding sites, many are known to be HIF1α-regulated, and expression of the VEGF profile correlated with HIF1α IHC positivity. The VEGF profile also showed prognostic significance on tests of sets of patients with breast and lung cancer and glioblastomas, and was an independent predictor of outcomes in primary breast cancers when tested in models that contained other prognostic gene expression
profiles and clinical variables.

Conclusions

These data identify a compact in vivo hypoxia signature that tends to be present in distant metastasis samples, and which portends a poor outcome in multiple tumor types.

overall_design: Microarrays and immunohistochemistry were used to analyze primary breast tumors, regional (lymph node) metastases, and distant metastases in order to identify biological features associated with distant metastases.

first: Zhiyuan

last: Hu


title: Expression data for UV time course in Saccharomyces cerevisiae wild-type and mutant strains

status: Public on Aug 18 2009

city: Charlottesville

first: Kunal

last: Poorey

state: VA

address: 1340 Jefferson Park Avenue

department: Biomedical Sciences: Biochemistry, Molecular Biology & Genetics

institute: University of Virginia Health System

summary: Treatment of cells with DNA damaging agents leads to large-scale gene expression changes. Proper transcriptional regulation is important for cells to arrest, repair damage and adjust cellular processes such as metabolism in order to survive the damaging assault. Damage-induced transcription is a highly regulated response. This study establishes a novel role for two factors, Snf1 and Rad23, in regulation of the UV-induced transcriptional response.

overall_design: Yeast strains were grown to log phase and irradiated with 100 J/m2 UV-C light. Unirradiated samples were also taken. Three different strains (WT, snf1Δ, mig3Δ) were analyzed in the absence of damage and at 15, 30 and 60 minutes following irradiation. Three other strains (rad23Δ, snf1Δ rad23Δ, rad4Δ) were analyzed in the absence of damage and at 30 and 60 minutes following irradiation for a total of 21 different strain/condition combinations. Experiments were performed in duplicate (two biological replicates for each condition). Samples were hybridized to Affymetrix Yeast 2.0 arrays. Undamaged WT samples were used as a control.

first: Staton

last: Wade

first: Stefan

last: Bekiranov

first: David

middle: T

last: Auble


title: Gene expression in neurons or astrocytes treated with grape seed extract

status: Public on Apr 09 2009

city: Kawasaki

first: Shihori

last: Tanabe

address: 3-25-26, Tonomachi, Kawasaki-ku

institute: National Institute of Health Sciences

summary:
We examined the effect of grape seed extract (GSE), which are known to protect neurons against oxidative stress, on primary cultures of hippocampal astrocytes. GSE increased interleukin-6 (IL-6) expression.
Microarrays are used to examine the effects of GSE on primary cultures of hippocampal neurons and astrocytes.

overall_design: Primary cultures of hippocampal neurons were treated with 0, 1, or 10 ug/ml of GSE for 24 hrs. Primary cultures of hippocampal astrocytes were treated with 0, 1, 10, or 100 ug/ml of GSE for 24 hrs.

first: Kayoko

last: Fujishita

first: Yoji

last: Sato

first: Schuichi

last: Koizumi

first: Tetsuro

last: Ozawa

first: Keisuke

last: Shibata

first: Toru

last: Okuda

first: M

last: Hisamoto


title: Differential yeast gene transcription during brewery propagation

status: Public on Jun 02 2009

city: Loughborough

last: Nottingham Arabidopsis Stock Centre (NASC)

address: Sutton Bonington Campus

department: School of Biosciences, University of Nottingham

country: United Kingdom

summary:
The aim of this study was to determine the transcriptomic response of Saccharomyces pastorianus to environmental change during brewery propagation. Utilization of amino acids and fermentable carbohydrates during full-scale brewery propagation was compared to the simultaneous changes occurring in the yeast transcriptome. Transcription profiles were observed to fall within one of four different groups and the greatest changes were observed to occur within the first eight hours following inoculation. Nutrient uptake was at its greatest during this period and many genes involved in the utilization of amino acids and carbohydrates were activated. There was also a significant derepression response following monosaccharide exhaustion. A number of stress response genes were activated following inoculation, indicating a possible osmotic stress response. Nutrient limitation did not appear to initiate a significant stress response, but did activate genes involved in pseudohyphal growth and meiosis, despite the fact that neither biological process occurs in the strain utilized. The first hours after inoculation into propagation wort are particularly active with respect to nutrient utilization and transcriptional change. It is likely that this period is particularly stressful for the yeast cells though there is no evidence of viability loss and it may be concluded that the yeast cell can cope with this level of stress, possibly aided by the dynamic response of the transcriptome.
Keywords: amino acid, brewing, carbohydrate, microarray, propagation, transcription, yeast

overall_design: GeneChip analyses were performed to determine the transcriptomic response of Saccharomyces pastorianus to environmental change during brewery propagation. Triplicate samples were taken immediately after inoculation and at intervals 4h, 8h, and 30h.

first: Katherine

last: Smart

first: Brian

last: Gibson

first: Neil

last: Graham

first: Chris

last: Boulton

first: Wendy

last: Box

first: Stephen

last: Lawrence

first: Robert

last: Linforth

first: Sean

last: May


title: Effect of copper ion on recovery process from freeze-thaw injury in Saccharomyces cerevisiae

status: Public on Feb 28 2009

city: Tsukuba

first: Jun

last: Shima

address: 2-1-12 Kannondai

institute: National Food Research Institute

summary: In frozen dough baking technology, baker’s yeast Saccharomyces cerevisiae encounter freeze-thaw injury. After thawing, dramatically decrease in cell viability and fermentation activity is caused by freeze-thaw injury. The freezing period is critical factor in freeze-thaw injury, thus we focused and investigated time-dependent gene expression profiles in recovery process from freeze injury. First, changes in gene expression profiles in S. cerevisiae in recovery process from freeze-thaw injury were analyzed using a DNA microarray. The results showed the genes which were involved in homeostasis of metal ions were time-dependent up-regulated 2-fold or more in a series. Then we examined whether these genes were related to tolerance in freeze-thaw injury by using deletion strain. The results showed that deletion of MAC1, CTR1, and PCA1 genes which involved in copper ion transport exhibited freeze-thaw sensitivity in compared with wild type. These genes are involved in copper ion uptake to a cell under a copper deficiency condition or in copper ion homeostasis, suggesting that it may be related between freeze-thaw injury and copper ion transport. To determine the effect of supplementation of copper ion on cells after freeze-thaw treatment, cell viability, intracellular superoxide dismutase (SOD) activity, and intracellular levels of reactive oxygen species (ROS) were examined by various copper ion condition medium. The results showed that intracellular SOD activity was increased and intracellular levels of ROS were decreased by supplementation of copper ion, but there was no significant difference in cell viability. These results of the present study may suggest that copper ion concentration in yeast cell after freeze-thaw treatment is important to recovery from freeze-thaw injury due to redox control of intracellular levels of ROS, but copper ion did not directly affect cell viability.

overall_design: Total RNA was extracted from the stress-treated yeast cells by using a hot phenol method. Poly(A)+ RNA was enriched from total RNA by using an Oligotex dT30 (Super) mRNA purification kit (Takara Bio, Ohtsu, Japan). cDNA synthesis, cRNA synthesis, and labeling were performed according to the Affymetrix user’s manual (Affymetrix, Santa Clara, USA). Biotinyated cRNA was fragmented and then used as a probe.Affimetrix Yeast Genome 2.0 arrays (Affymetrix) were used as DNA microarrays. All experiments were done in duplicate independently

first: Shunsuke

last: Takahashi


title: Temporal Profiling of Rat Transcriptomes in Retinol-Replenished Vitamin A-Deficient Testis

status: Public on Feb 27 2009

city: Pullman

first: Timothy

middle: J.

last: Doyle

address: 100 Dairy Road

department: School of Molecular Biosciences

institute: Washington State University

summary: We used microarrays to examine the temporal transcriptome changes of vitamin a deficient rats testes replenished with retinol

overall_design: 20-day old rats were put on a vitamin A deficient diet, after nine weeks the animals were treated with retinol for 0-(VAD), 4-, 8-, 24-hr to determine the global transcriptome changes induced by retinol replenishment of the vitmain A deficient testis

first: Asa

last: Oudes

first: Kwanhee

last: Kim


title: Genetic Control of Cellular Quiescence in S. pombe

status: Public on Apr 30 2009

city: Uruma

first: Kenichi

last: Sajiki

address: 12-22 Suzaki

institute: OIST-IRP

summary:
Transition from proliferation to quiescence brings about extensive changes in cellular behavior and structure. However, genes critical for establishing and/or for maintaining quiescence are largely unknown. The fission yeast S. pombe is found as an excellent model for studying this problem, because it becomes quiescent under nitrogen starvation. Here we characterize 610 temperature-sensitive (ts) mutants, and identify 33 genes required for entry into and the maintenance of quiescence. These genes cover a broad range of cellular functions in the cytoplasm, membrane and the nucleus, encoding proteins for stress-responsive and cell cycle kinase signaling pathway, actin-bound and osmo-controlling endosome formation, RNA transcription, splicing and ribosome biogenesis, chromatin silencing, biosynthesis of lipid and ATP, cell wall and membrane morphogenesis, protein trafficking and vesicle fusion. We specifically highlight Fcp1, CTD phosphatase of RNA polymerase II, which differentially affects transcription of genes involved in quiescence and proliferation. We propose that the transcriptional role of Fcp1 is central to differentiate quiescence from proliferation.

Keywords: cell cycle control, protein trafficking, cortex actin, RNA polymerase II, protein phosphatase

overall_design: Gene expression profile under nitrogen rich or starved condition at 26ºC or 37ºC in WT (L972) or fcp1-452 ts mutant strain of fission yeast.

first: Mitsuko

last: Hatanaka

first: Takahiro

last: Nakamura

first: Kojiro

last: Takeda

first: Mizuki

last: Shimanuki

first: Tomoko

last: Yoshida

first: Yuichiro

last: Hanyu

first: Takeshi

last: Hayashi

first: Yukinobu

last: Nakaseko

first: Mitsuhiro

last: Yanagida


title: Expression data from livers of rats treated with control, D3T, OLT, or TBD

status: Public on Dec 30 2008

city: Memphis

first: Thomas

last: Sutter

address: 3774 Walker Ave.

institute: University of Memphis

summary:
The activities of the dithiolethione analogs, D3T, OLT, and TBD are pharmacologically well-understood.  These compounds act as chemopreventive agents whose ability is to block or diminish early stages of carcinogenesis.  In addition, the three compounds are classified as monofunctional Phase II enzyme inducers and activate the same pathway, namely the Keap1-Nrf2 signal pathway. The three dithiolethiones were showed to ameliorate the AFB1-induced toxicity through increasing phase II enzymes including glutathione S-transferase (GST).  The parent D3T was observed to be the most potent chemoprotective agent.  Oltipraz, a clinically approved drug, has been shown to exhibit less efficacy than its analogs for inhibition of aflatoxin-induced hepatic foci.TBD was suggested to be better than OLT as a chemopreventive agent because of its reduced toxicity profile.
Thus based on gene expressions, our goal is to study the closely related structure activities that lie within the three compounds and the activity of OLT versus TBD at the 5 position of their dithiole rings and to determine whether the activity of TBD is closely related to OLT or its parent D3T.
Keywords: treatment effects

overall_design: Male Fischer F344 rats (80-100g) were obtained from Harlan at 6 wk of age. General procedures for animal care and housing were in accordance with the Guide for the Care and use of Laboratory Animals, National Research Council, 1996.  The animals were singly housed in polycarbonate cages in a temperature (22 ± 1°C)- and humidity (30-70%)-controlled room with a 14:10-h light-dark cycle, respectively.  Rat AIN 76A chow (Harlan Teklad) and tap water were provided at libitum. Sixteen animals were randomly assigned into four groups (n=16) and treated by gavage with 100?l of either vehicle (saturated sucrose), 3H-1,2-dithiole-3-thione (D3T), (0.3 mmol/kg body wt), 4-methyl-5-pyrazinyl-3H-1,2-dithiole-3-thione (oltipraz-OLT), (0.3 mmol/kg), and 5-tert-butyl-3H-1,2-dithiole-3-thione (TBD) , (0.3mmol/kg body wt),  once every other day for 5 days.  Animals were killed 24h after the third dose.  Liver tissues were harvested and snap frozen.  The protocol for this study was approved by the University of Memphis Animal Care Committee and Johns Hopkins University.


title: Urinary bladder post-spinal cord injury: time-points

status: Public on Dec 25 2008

city: Pittsburgh

first: Silvia

last: Wognum

state: PA

address: 100 Technology Drive

department: Bioengineering

institute: University of Pittsburgh

summary: Previously, we demonstrated using a rat model of spinal cord injury (SCI) that bladder wall tissue compliance significantly increased within the first 2 weeks following injury. In order to explore the potential molecular-level mechanisms of this event, the present study quantified molecules pertinent to bladder tissue remodeling and changes in mechanical properties. An initial gene array analysis followed by real-time qPCR revealed that the message levels for tropoelastin and lysyl oxidase were as high as 8-fold in SCI rats compared to normal. Furthermore, both the message and protein levels of TGF-beta1 and IGF-1, known stimulators of elastin synthesis, in SCI rat bladders were significantly higher compared to those of normal rats. Taken together, it can be speculated that functional changes of the bladder associated with SCI induce release of select growth factors, which, in turn, stimulate elastogenesis that lead to alteration of biomechanical properties of the wall tissue.

overall_design:
One bladder in each of the four groups. One normal control, one 3 days post-SCI, one 7 days, and one 25 days. Only one sample per group for initial screening analysis.
The microarray data was used in an additional study using Ingenuity Pathways Analysis to elucidate remodeling, inflammation and infection events.

first: Jiro

last: Nagatomi

first: Fernando

last: DeMiguel

first: Kazumasa

last: Torimoto

first: Michael

last: Chancellor

first: Robert

middle: H

last: Getzenberg

last: Sacks

first: Claudio

middle: E

last: Lagoa

first: Yoram

last: Vodovotz


title: A Supervised Risk Predictor of Breast Cancer Based on Biological Subtypes

status: Public on Feb 11 2009

city: Chapel Hill

first: Charles

last: Perou

address: 12-044 Lineberger Comprehensive Cancer Center CB# 7295

institute: University of North Carolina at Chapel Hill

summary:
Purpose: The biological subtypes of breast cancer designated as Luminal A, Luminal B, HER2+/ER-, and Basal-like are clinically important for prognosis and planning treatment strategies. Recognizing that there is a continuum in both the spectrum of breast cancer disease and the risk of survival, we sought to develop a clinical test for the biological subtypes using a supervised risk classier.Methods: Microarray and real-time quantitative RT-PCR (qRT-PCR) data from 189 samples, procured as fresh-frozen and formalin-fixed, paraffin-embedded tissues, were used to statistically select prototypical samples and genes for the biological subtypes of breast cancer. Predictions for biological subtype and risk of recurrence were determined for different stages of disease, treatments, and across analytical platforms. Results: The biological subtype predictions on a large combined microarray test set showed prognostic significance across all patients (1244 subjects; p<0.0001), on node negative patients with no adjuvant systemic therapy (738 subjects; p<0.0001), and on patients treated with endocrine therapy (404 subjects; p=0.001). Analysis of a neoadjuvant chemotherapy study revealed a high pathologic complete response (pCR) rate in HER2+/ER- and Basal-like patients. The subtype and risk predications were also highly significant when using the qRT-PCR assay from archived FFPE breast cancers. Conclusion: Our risk predictor based on distance to biological subtype centroids provides a continuous risk score that applies to all stages of breast cancer given current therapies. The assay can be performed using archived breast tissues and a real-time qRT-PCR assay, thus facilitating application to retrospective cohorts and clinical samples.
Keywords: reference x sample

overall_design: Comparison of reference samples against treatment

first: Joel

last: Parker

first: Philip

last: Bernard


title: Transcriptomic profiling of wine yeast strains during colombar fermentation

status: Public on May 01 2009

first: Debra

last: Rossouw

address: Cnr Neethling and Victoria rd

department: Institute for Wine Biotechnology

summary:
Comparative gene expression analysis of two wine yeast strains at three time points (days 2, 5 and 14) during fermentation of colombar must.

In our study we conducted parallel fermentations with the VIN13 and BM45 wine yeast strains in two different media, namely MS300 (syntheticmust) and Colombar must. The intersection of transcriptome datasets from both MS300 (simulated wine must;GSE11651) and Colombar fermentations should help to delineate relevant and ‘noisy’ changes in gene expression in response to experimental factors such as fermentation stage and strain identity.

overall_design: Two industrial wine yeast strains (BM45 and VIN13) grown micro-aerobically in Colombar must. Microarray analysis was performed at three time points during fermentation (days 2, 5 and 14), representing the exponential, early and late stationary growth phases respectively.

first: Florian

last: Bauer


title: Gcn4p-mediated transcriptional stress response

status: Public on Feb 01 2009

city: Cambridge

last: Moxley

state: MA

address: 77 Massachusettes Ave

institute: MIT

summary:
The transcriptional data from an integrative analysis of transcriptional and metabolic stress responses that provides a more complete understanding of the mechanisms by which genetic regulatory circuits mediate metabolic phenotype.
Keywords: stress response, genetic modification

overall_design: Our investigation disrupts native transcriptional control of amino acid biosynthesis via the Gcn4p- mediated transcriptional stress response.  Specifically, genetic perturbation of transcriptional regulator Gcn4p in a stress condition is achieved via wild-type S288C and gcn4-delta strains in a pH-controlled chemostat environment in which a titrated inhibitor creates near histidine starvation and glucose supply limits growth to a pre-defined rate (0.10 hr^-1).  The gene expression chip titles indicate the condition.

last: Jewett

first: Silas

last: Villas-Boas

first: Maciek

last: Antoniewicz

first: Hal

last: Alper

last: Wheeler

first: George

last: Stephanopoulos

first: Alan

last: Hinnebusch

first: Trey

last: Ideker

first: Jens

last: Nielsen

first: Gregory


title: Alpha-factor treatment of Saccharomyces cerevisiae

status: Public on Dec 01 2008

city: Aurora

first: Jay

middle: R.

last: Hesselberth

state: CO

department: Biochemistry and Molecular Gentetics

institute: University of Colorado School of Medicine

summary:
We measured the response of S. cerevisiae to arrest in the presence of alpha factor.  These were collected in support of a related DNaseI-sequencing study.
Keywords: Alpha-factor arrest

overall_design: S.cerevisiae R276 (MATa ura3Δ0 leu2Δ0 his3Δ1 met15Δ0 bar1Δ::KanMX) (C. Boone, University of Toronto; S288c background derived from BY4741), was cultured overnight in 50 ml rich medium (YPD) at 30°C, diluted into 500 ml fresh YPD to an OD660 of ~0.8, and treated with yeast α-factor (Sigma-Aldrich) at a final concentration of 50 ng / ml. This culture was incubated at 30°C with shaking for 3 hours (final OD660 ~1).  After this treatment, approximately 90% of the cells had formed mating projections when checked by light microscopy.  Total RNA from these cells was isolated using hot acidic phenol.   50 μg of total RNA was treated with Turbo Dnase (Ambion), and checked for integrity using a Bioanalyzer 2100 (Agilent).  Total RNA was labeled according to the manufacturer’s protocol and applied to Affymetrix Yeast 2.0 arrays.  Data were analyzed using the “affy” package from Bioconductor.


title: Expression data for rat CNS mixed glial cultures treated with cytokines

status: Public on Nov 07 2008

city: Detroit

first: wenbo

last: xu

address: 3127 Scott Hall ,540 East Canfield

institute: wayne state university

summary:
To examine changes in gene expression that might occur in CNS glial cells in response to the secreted products of immune cells, we used gene array analysis to assess the early effects of different cytokine mixtures on rat mixed CNS glia in culture. We compared effects at 6 hours of cytokines typical of Th1 and Th2 lymphocytes, and monocyte marophages (M/M)..  We found unique patterns of changes in gene expression for each of the three cytokine mixtures, including changes in immune-related molecules, neurotrophins, growth factors, proteins involved in axon/glial interactions, ion channels, neurotransmitters, mitochondrial function and apoptosis.  These changes may have relevance in neuroprotective or damaging mechanisms in neurodegenerative diseases such as multiple sclerosis, specifically with regard to formation, repair or inhibition of lesion formation.
Keywords: Treatment with different cytokine mixtures

overall_design: Mixed CNS glia cultures from newborn rat brain were treated for 6 hours with cytokine mixtures representative of Th1, Th2 or monocyte/macrophage cytokines

first: Robert

middle: P

last: Lisak

first: Joyce

last: Benjamins


title: Changes in Saccharomyces cerevisiae mRNA abundance following oxidative stress and DNA damage stress

status: Public on Oct 01 2008

city: Rehovot

first: Ophir

last: Shalem

address: Hertzel

summary:
We subjected yeast to two stresses, oxidative stress, which under current settings induces a fast and transient response in mRNA abundance, and DNA damage, which  triggers a slow enduring response. Using microarrays we performed a conventional quantification of change in mRNA abundance.

Keywords: Two separate time courses

overall_design: We used Affymetrix microarrays to quantify changes in mRNA abundance following oxidative stress (using hydrogen peroxide) and DNA damage stress (using methyl methanesulfonate) during a three-hour time course (0, 30 min, 60 min, 100 min, 140 min, 180 min).

first: Orna

last: Dahan

first: Michal

last: Levo

first: Maria

last: Martinez

first: Itay

last: Furman

first: Eran

last: Segal

first: Yitzhak

last: Pilpel


title: Transient transcriptional responses to stress are generated by opposing effects of mRNA production and degradation

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

overall_design: Refer to individual Series


title: Decay profiles of Saccharomyces cerevisiae mRNAs following oxidative stress and DNA damage

summary:
We subjected yeast to two stresses, oxidative stress, which under current settings induces a fast and transient response in mRNA abundance, and DNA damage, which triggers a slow enduring response. Using microarrays, we performed a transcriptional arrest experiment to measure genome-wide mRNA decay profiles under each condition. Genome-wide decay kinetics in each condition were compared to decay experiments that were performed in a reference condition (only transcription inhibition without an additional stress) to quantify changes in mRNA stability in each condition. We found condition-specific changes in mRNA decay rates and coordination between mRNA production and degradation. In the transient response, most induced genes were surprisingly destabilized, while repressed genes were somewhat stabilized, exhibiting counteraction between production and degradation. This strategy can reconcile high steady-state level with short response time among induced genes. In contrast, the stress that induces the slow response displays the more expected behavior, whereby most induced genes are stabilized, and repressed genes destabilized. Our results show genome-wide interplay between mRNA production and degradation, and that alternative modes of such interplay determine the kinetics of the transcriptome in response to stress.

Keywords: Four separate time courses

overall_design: We used Affymetrix microarrays to measure the decay profiles of all genes following transcription inhibition in four separate experiments. In two reference experiments, only transcription inhibtion was applied. In two other experiments, an additional stress was applied prior to transcription inhibition: oxidative stress (0.3mM hydrogen peroxide) or DNA damage (0.1% methyl methanesulfonate).


title: Significant and Systematic Expression Differentiation in Long-Lived Yeast Strains

status: Public on Nov 01 2008

city: Los Angeles

first: Huanying

last: Ge

address: 1050 Childs Way, RRI building 408D

institute: Computational biology and bioinformatics

summary:
The three yeast mutants sch9Δ, ras2Δ, tor1Δ show extended chronological life span up to three folds.
This study aims to dissect the mechanisms that lead to the yeast life span extension.

Keywords: Expression comparison between wild type and long-lived yeast mutant strain

overall_design:
We obtain gene expression profiles of sch9Mutant, ras2Mutant, tor1Mutant as well as that for a wild type at day 2.5 in SDC medium using Affymetrix Yeast2.0 arrays. Each sample has three replicates.

We use a complicated pre-processing method and get the expression differntiation directly. We consider the issue of processing robustness and expression large differentiation. The normalization is carried out in a pairwise fashion. Namely, for each wild type sample (W1, W2, W3), three replicates of a mutant are normalized with respect to each reference. Take sch9Mutant (triplicates: S1, S2 and S3) for example, the normalized arrays with respect to W1, W2, W3 are respectively denoted by S1\W1, S2\W1, S3\W1, S1\W2, S2\W2, S3\W2, S1\W3, S2\W3, S3\W3. We group the wild type and normalized mutant arrays by the reference, and then summarize each group. We summarize the four arrays (the reference plus three normalized) W1, S1\W1, S2\W1, S3\W1 together. This leads to three estimates of expression fold changes of the mutant versus the wild type. In total, we have nine estimates from three wild type references, and we take their median difference as the final estimate.

Expression fold change estimates (generated by the analysis) are given in the Series table.

first: Chao

last: Cheng

first: Paola

last: Fabrizio

first: Min

last: Wei

first: Valter

middle: D

last: Longo

first: Lei

last: Li


title: Transcriptomes for different level of glucose

status: Public on Oct 02 2008

city: Seoul

first: Cheol-Koo

last: Lee

address: Korea University, Biotechnology #318 West building, Anam-dong, Seongbuk-gu

department: Biotechnology

country: South Korea

institute: Korea University

summary: To characterize gene expression that is dependent on the strength of calorie restriction (CR), we obtained transcriptome at different levels of glucose, which is a major energy and carbon source for budding yeast. To faithfully mimic mammalian CR in yeast culture, we reconstituted and grew seeding yeast cells in fresh 2% YPD media before inoculating into 2%, 1%, 0.5% and 0.25% YPD media to reflect different CR strengths. We collected and characterized 160 genes that responded to CR strength based on the rigorous statistical analyses of multiple test corrected ANOVA (adjusted é value < 0.1 or raw é value < 0.0031) and Pearson correlation (|r| > 0.7). Based on the individual gene studies and the GO Term Finder analysis of 160 genes, we found that CR dosedependently and gradually increased mitochondrial function at the transcriptional level. Therefore, we suggest these 160 genes are markers that respond to CR strength and that might be useful in elucidating CR mechanisms, especially how stronger CR extends life span more.

overall_design:
Different glucose levels: 2%, 1%, 0.5%, and 0.25%
Three biological replicates in each level

first: Yae-Lim


title: Loss of the retinoblastoma tumor suppressor is a common event in Basal-like and Luminal B breast carcinomas

status: Public on Sep 18 2008

first: Charles

last: Perou

summary:
Breast cancers can be classified using whole genome expression into distinct subtypes that show differences in patient prognosis.  One of these groups, the basal-like carcinomas, are poorly differentiated, highly metastatic, and genomically unstable.  These tumors also contain specific genetic alterations with one example being frequent p53 mutations. The loss of the tumor suppressor gene encoded by the retinoblastoma (RB1) locus is a well-characterized occurrence in many tumor types, however, its role in breast cancer is less clear with many reports demonstrating a Loss of Heterozygosity (LOH) that does not correlate with loss of RB1 protein expression.  Here we report that LOH of the RB1 locus was observed at a high frequency in basal-like and luminal B tumors.  These tumors also concurrently have low expression of RB1 mRNA as assessed by DNA microarray.  As in previous reports, we also did not see a significant correlation between RB1 LOH and protein expression as measured by immunohistochemistry (IHC).  p16INK4a, however, was highly expressed both by microarray and IHC, in basal-like tumors only presumably due to a previously reported feedback loop caused by RB1 loss.  These results suggest that the functional loss of RB1 is a common event in the progression of basal-like and luminal B breast tumors, which may play a key role in dictating therapeutic responses
Keywords: reference x sample

first: Jason

middle: I

last: Herschkowitz

middle: M


title: Response of Yeast (Saccharomyces cerevisiae) to Oxygen, Heme and Cobalt

status: Public on Sep 10 2008

city: NEW YORK CITY

first: ANSHUL

middle: BHARAT

last: KUNDAJE

address: 110 MORNINGSIDE DRIVE, APT. 31A

department: COMPUTER SCIENCE

institute: COLUMBIA UNIVERSITY

summary: Deciphering gene regulatory mechanisms through the analysis of high-throughput expression data is a challenging computational problem. Previous computational studies have used large expression datasets in order to resolve fine patterns of coexpression, producing clusters or modules of potentially coregulated genes. These methods typically examine promoter sequence information, such as DNA motifs or transcription factor occupancy data, in a separate step after clustering. We needed an alternative and more integrative approach to study the oxygen regulatory network in Saccharomyces cerevisiae using a small dataset of perturbation experiments. Mechanisms of oxygen sensing and regulation underlie many physiological and pathological processes, and only a handful of oxygen regulators have been identified in previous studies. We used a new machine learning algorithm called MEDUSA to uncover detailed information about the oxygen regulatory network using genome-wide expression changes in response to perturbations in the levels of oxygen, heme, Hap1, and Co2+. MEDUSA integrates mRNA expression, promoter sequence, and ChIP-chip occupancy data to learn a model that accurately predicts the differential expression of target genes in held-out data. We used a novel margin-based score to extract significant condition-specific regulators and assemble a global map of the oxygen sensing and regulatory network. This network includes both known oxygen and heme regulators, such as Hap1, Mga2, Hap4, and Upc2, as well as many new candidate regulators. MEDUSA also identified many DNA motifs that are consistent with previous experimentally identified transcription factor binding sites. Because MEDUSA's regulatory program associates regulators to target genes through their promoter sequences, we directly tested the predicted regulators for OLE1, a gene specifically induced under hypoxia, by experimental analysis of the activity of its promoter. In each case, deletion of the candidate regulator resulted in the predicted effect on promoter activity, confirming that several novel regulators identified by MEDUSA are indeed involved in oxygen regulation. MEDUSA can reveal important information from a small dataset and generate testable hypotheses for further experimental analysis.

overall_design:
Yeast cell growth and treatment:
Yeast strains used were L51 (MATa, ura3-52, leu2-3, 112, his4-519, ade1-100, trp1::HisG, hap1::LEU2) and MHY100 (MATa, ura3-52, leu2-3, 112, his4-519, ade1-100, hem1-delta100).  L51 was used for studies of oxygen regulation, and MHY100 was used for studies of heme regulation.  To avoid variations from the differences accumulated after many generations of growth of strains, we transformed the L51 strain with the HAP1 gene deleted for studies of Hap1 function.  Hap1 protein was expressed in L51 cells by transforming an ARS-CEN plasmid bearing the complete HAP1 genomic sequence.  For comparison with cells without Hap1 expressed, an empty vector was transformed into L51 cells.  The use of Hap1 expression plasmid generated much more reproducible results than the use of different strains.  Yeast cells with or without Hap1 expressed grew at similar rates under both anaerobic and aerobic conditions.

We chose to use a low oxygen level (~10 ppb) in this study, in order to identify all oxygen-regulated genes.  Previous studies have shown that most, if not all, oxygen-regulated genes are affected a low concentrations, but some genes are not affected at higher oxygen levels (for example, > 1 ppm).  Anaerobic (~10 ppb O2, measured by using an oxygen monitor and confirmed by CHEMetrics oxygen kits) growth condition was created by using an anaerobic chamber (Coy Laboratory, Inc.) and by filling the chamber with a mixture of 5% H2 and 95% N2 in the presence of palladium catalyst [61].  L51 cells bearing the Hap1 expression or empty vector were grown under normoxic or anaerobic conditions for 1.5 or 6 hours.  The UAS1/CYC1-lacZ reporter plasmid was transformed into yeast cells to confirm the expression of Hap1 and the oxygen levels.  Cells were grown in yeast synthetic complete media.  Co2+-induced cells were grown in the presence of 400 microM cobalt chloride for 6 hours, as described previously.  MHY100 cells were grown in medium containing 2.5 microg/ml (heme-deficient) or 250 microg/ml (heme-sufficient) 5-aminolevulinic acid.  For RNA preparations, yeast cells were inoculated so that the optical density of yeast cells was in the range of 0.8-1.0 immediately before the collection of cells.

RNA preparation and microarray gene expression profiling:
RNA was extracted from yeast cells exactly as previously described.  RNA samples were prepared from 8 different experimental conditions:  (1) L51 yeast cells bearing the Hap1 expression plasmid maintained under aerobic conditions, (2) L51 yeast cells bearing the empty expression plasmid maintained under aerobic conditions, (3) L51 yeast cells bearing the Hap1 expression plasmid maintained under anaerobic conditions for 1.5 hours, (4) L51 yeast cells bearing the Hap1 expression plasmid maintained under anaerobic conditions for 6 hours, (5) L51 yeast cells bearing the empty expression plasmid maintained under anaerobic conditions for 6 hours, (6) L51 yeast cells bearing the Hap1 expression plasmid in the presence of 400 microM cobalt chloride for 6 hours, (7) MHY100 cells grown in medium containing 250 microg/ml (heme-sufficient) 5-aminolevulinic acid, and (8) MHY100 cells grown in medium containing 2.5 microg/ml (heme-deficient) 5-aminolevulinic acid.  For each condition, three replicates were generated by preparing RNA samples from three batches of independently grown cells.  Microarray expression analyses were performed by using three batches of replicate RNA samples.  The quality of RNA was high as assessed by measuring absorbance at 260 and 280 nm, by gel electrophoresis, and by the quality of microarray data (see below).

The synthesis of cDNA and biotin-labeled cRNA were carried out exactly as described in the Affymetrix GeneChip Expression Analysis Technical Manual (2000).  The yeast Saccharomyces cerevisiae genome 2.0 arrays were purchased from Affymetrix, Inc.  Probe hybridization and data collection were carried out by the Columbia University Affymetrix GeneChip processing center.  Specifically, the Affymetrix GeneChip Hybridization Oven 640 and the next generation GeneChip Fluidics Station 450 were used for hybridization and chip processing.  Chip scanning was performed by using the GeneChip scanner 3000.  Initial data acquisition, analysis was performed by using the Affymetrix Microarray suite.  By using GCOS1.2 with the advanced PLIER (probe logarithmic intensity error) algorithm, we calculated and examined the parameters reflecting the image quality of the arrays.  Arrays with a high background level in any region were discarded and replaced.  The average noise or background level was limited to less than 5%.  The average intensity for those genes judged to be present was at least 10-fold higher than those judged to be absent.  Also, arrays that deviated considerably in the percentage of present and absent genes from the majority of the arrays were replaced.  Arrays with a beta-actin 3’/5’ ratio greater than 2 were replaced.

Normalization of microarray data:
For each microarray, we converted the  .DAT image files into  .CEL files using the Affymetrix GCOS software.  These raw  .CEL files were further processed into expression values using the RMA express software by Bolstad.  This software uses the robust multiarray average method by Irrizary et al., which involves a background correction and a quantile-based normalization scheme.

first: Anshul

last: Kundaje

first: Xiantong

last: Xin

first: Changgui

last: Lan

first: Steve

last: Lianoglou

first: Mei

last: Zhou

first: Christina

last: Leslie

first: Li

last: Zhang


title: Alterations in Neuronal Gene Expression Profiles in Response to Experimental Demyelination and Axonal Transection

status: Public on Aug 05 2008

city: Bethesda

first: Joe

last: Nielsen

address: 49 Convent Drive BD49,5A75

institute: NIH/NINDS

summary: The pathological basis of multiple sclerosis involves damage to both myelin sheaths and axons. Demyelination and axonal transection are considered to cause reversible and irreversible neurological deficits respectively, gradually destroying the neuronal circuitry of the CNS. In order to analyse the individual effects of the pathological hallmarks of multiple sclerosis on neurons, the pontocerebellar pathway was targeted with either lysolecithin-induced chemical demyelination or complete pathway transection. Transcriptional changes in the pontocerebellar neuronal nuclei were investigated with microarrays at days 4, 10 and 37 post-intervention to identify underlying molecular responses.  A common as well as unique set of injury response genes was identified in the transection and the demyelination conditions. The increased expression of activating transcription factor 3 (Atf3) and thyrotropin-releasing hormone (Trh) in both injury paradigms was validated by immunohistochemistry. Expression of Atf3 in a patient with Marburg’s variant of multiple sclerosis was also detected in the pons with large cerebellar demyelination, confirming the activation of the Atf3 pathway in a human disease sample as well.   This experimental approach may be useful for the identification of pathways that could be targeted for remyelinative and neuroprotective drug development.

overall_design:
The pontine nuclei containing the neuronal cell bodies of the projecting lesioned axons (saline, lysolecithin or transection) were collected at different time points post-intervention (day 4, day 10, and day 37), and assayed for changes in gene expression caused by the distal injury site. The contralateral pontine nuclei were used as controls.

CEL files lost

first: Gabor

last: Lovas

first: Joseph

first: Kory

last: Johnson

first: lynn

last: Hudson


title: Expression data from yeast strain containing CDC34tm allele compared to WT

status: Public on Jun 21 2008

city: Indianapolis

first: Mark

last: Goebl

state: IN

address: 1345 West 16th St.

department: Biochemistry and Molecular Biology

institute: Indiana University School of Medicine

summary:
Cdc34 is an essential E2 ubiquitin conjugating enzyme found in nearly all eukaryotes.  It contains a highly conserved motif composed of S73/S97/12 amino acid insert near the active site cysteine.  This motif is unique to Cdc34/Ubc7 type E2s while other E2s contain K/D/no insert at these positions.  To better understand the function of this motif we mutated Cdc34 S73/S97/insert to be K/D/no insert and observed changes in transcript levels in mid-log phase yeast cells.  ABSTRACT [Cdc34 is a ubiquitin conjugating enzyme necessary for the ubiquitylation of substrates by the SCF family of ubiquitin ligases.  Previous work has shown that the Cdc34 protein is phosphorylated in vivo on serine residues.  Cdc34 contains two serines within its catalytic domain, S73 and S97, that together with a 12 amino acid acidic loop, constitute a highly conserved motif (serine, serine, insert) among all members of the Cdc34 family of E2 enzymes. Using phosphospecific antibodies, we show that the essential serine S97 is indeed phosphorylated in vivo.  Furthermore, this phosphorylation event is regulated by treatment with pheromone in yeast.  Consistently, expression of a Cdc34 mutant lacking this motif (serine, serine, insert) leads to misregulation of the SCF substrates, Sic1, Far1, Cln1 and Cln2 and suppresses the cell cycle arrest brought about by an activated mating pathway.  We further explored the function of this motif by microarray analysis and show that the transcripts of nearly the entire Sic1 cluster of co-transcribed genes is altered in a strain the expresses Cdc34 lacking this motif.  Our data reveals that this highly conserved motif in Cdc34 and its phosphorylation are important for modulating SCF substrate abundance both transcriptionally and post-transcriptionally.]
Keywords: genetic modification

overall_design: The Saccharomyces cerevisiae CDC34tm strain (RRC85, MATα, CDC34tm(NatR)) which is marked by a nourseothricin resistance gene (NatR) and an isogenic wild type strain (DBY2059, MATα, leu2-3, 112) were inoculated from stationary phase cultures into a synthetic defined medium containing 2% dextrose, 0.17% yeast nitrogen base minus amino acids and ammonium sulfate, 0.25% L-glutamine, 0.025% magnesium sulfate and 25.2 mg/L L-Leucine. Four separate cultures of each strain were grown at 30°C, allowing between three and four doublings and cells were collected at approximately 8 x 10e6cells/ml.  Cells were centrifuged and immediately frozen in liquid nitrogen.  Total RNA was extracted using a hot acid phenol-chloroform protocol as previously described (Schmitt et al. 1990).  RNA quality was verified with OD260/280 readings and a 1.5% agarose gel.  An Affymetrix Genechip Scanner 3000 and Affymetrix Yeast 2.0 arrays were used for the microarray.

middle: G

first: Ross

last: Cocklin


title: Transcriptomic profiling of five industrial wine yeast strains at three time points during allcoholic fermentation

status: Public on Jul 01 2008

city: Stellenbosch

first: Debra

last: Rossouw

institute: University of Stellenbosch

summary:
Our study involves a transcriptomic approach to the analysis of industrial yeast metabolism. Historically, among the hundreds of yeast species, Saccharomyces cerevisiae has played an important role in scientific investigations and industrial applications, and it is universally acknowledged as one of the model systems for eukaryotic organisms. Yeast is also an important component of the wine fermentation process and determines various attributes of the final product.

Our research takes a holistic approach to the improvement of industrial yeast strains by integrating large data sets from various yeast strains during fermentation. This means that analysis can be done in such a way as to co-evaluate several parameters simultaneously to identify points of interest and target genes for metabolic engineering.

Eventually we hope to construct an accurate information matrix and a more complete cellular map for the fermenting yeast. This will enable accurate model-building for industrial yeast and facilitated the design of intelligent yeast improvement strategies which can be applied via traditional avenues of molecular biology.

overall_design: Five different Saccharomyces cerevisiae strains used in industrial winemaking processes were used in synthetic must (MS300) fermentations. All fermentations were carried out in triplicate, so each sample is represented by three completely independent biological repeats. Samples for microarray analysis were taken at three different time points during fermentation, representative of the exponential (day2), early stationary (day5) and late stationary (day14) growth stages.

first: Florian

last: Bauer


title: Changes in Global Gene Expression in Rat Myometrium in the Transition from Late Pregnancy to Parturition

status: Public on Sep 23 2008

first: Enrico

last: Stefani

address: 650 Charles Young Dr

department: Anesthesiology

institute: David Geffen School of Medicine at UCLA

summary: The process of parturition involves the complex interplay of factors that changes the excitability and contractile activity of the uterus. We have compared the relative gene expression profile of myometrium from rats before parturition (21 days pregnant) and during delivery using high-density DNA microarray. Of 8740 sequences available in the array a total of 3782 were detected as present. From the sequences that were significantly altered, 59 genes were upregulated and 82 genes were downregulated. We were able to detect changes in genes described to have altered expression level at term including connexin 43 and 26, cyclo-oxygenase 2 and oxytocin receptor as well as novel genes that have been not previously associated with parturition.  Quantitative real time PCR on selected genes further confirmed the microarray data. Here we report for the first time that aquaporin5 (AQP5), a member of aquaporin (AQP) water channel family, was dramatically downregulated during parturition (~100 fold by microarray and ~50 fold by Real Time PCR). The emerging profile highlights biochemical cascades occurring in a period of about 36 hours that triggers parturition and the initiation of myometrium reverse remodeling after partum. The microarray analysis uncovered genes that were previously suspected to play a role in parturition. This regulation involves genes from immune/inflammatory response, steroid/lipids metabolism, calcium homeostasis, cell volume regulation, cell signaling, cell division, and tissue remodeling, suggesting the presence of multiple and redundant mechanisms altered in the process of birth.

overall_design: Uteri were dissected from late pregnant rats (21 days, LP) (three individuals, two replicates of sample 1) and rats during labor (DL) after the expulsion of the second pup (three individuals, two replicates of sample 1).  After dissection, uteri were placed immediately in cold Ca2+-free PBS buffer and peripheral large blood vessels, external surrounding connective tissue and placenta were removed. The endometrium superficial layer was separated by gentle scraping.  The myometrium was weighted, finely minced and homogenized with a polytron (Polytron PT3000, Brinkmann, Switzerland).

first: Gustavo

last: Helguera

first: Mansoureh

last: Eghbali

first: Daniel

last: Sforza

first: Tamara

middle: Y

last: Minosyan

first: Ligia

last: Toro


title: sua5D expression profiles

city: shanghai

first: feilong

last: meng

address: 320 yuyang road

department: SIBCB

institute: Shanghai institutes for biology sciences

summary:
To decipher the function of SUA5, we compare the expression profiles of sua5D and wildtype BY4742.  We also compared the expression profile changed during the passage of sua5D strain.
Keywords: genetic modification

overall_design:
Sua5 disruptioin strains were obtained by tetrad dissection of a sua5D/SUA5 diploid strain.  The spores were restreaked on YPD plate for 12 times. Each restreaking is suggested to 25 generaions. The first restreaking, 12th restreaking and wildtype strains were compared.
Four indepedent clones (First restreaking), three independent clones (12th restreaking), and three independent clones (wildtype control) were used.

first: Feilong

last: Meng

first: Jinqiu

last: Zhou


title: Transcriptional response in laboratory and wine strains of S. cerevisiae to growth temperature

status: Public on Jul 25 2008

city: Santiago

first: Francisco

middle: J

last: Pizarro

address: Av. Vicuna Mackenna 4860

department: Chemical and Bioprocess Engineering

country: Chile

institute: Universidad Catolica de Chile

summary: Laboratory strains of Saccharmoyces cerevisiae have been widely used as a model for studying eukaryotic cells and mapping the molecular mechanisms of many different human diseases. Industrial wine yeasts, on the other hand, have been selected over hundreds of years on the basis of their adaptation to stringent environmental conditions and the organoleptic properties they confer to wine. Here, we applied a two-factor design to study the response of a standard laboratory strain, CEN.PK.113-7D, and an industrial wine yeast-strain, EC1118, to growth temperature at 15°C and 30°C under 12 nitrogen-limited, anaerobic steady-state chemostat cultures.  Physiological characterization revealed that growth temperature strongly impacted biomass yields in both strains.  Moreover, we observed that the wine yeast is better adapted to mobilizing resources for biomass and that the laboratory yeast exhibited higher fermentation rates.  To elucidate mechanistic differences controlling the growth temperature response and underlying adaptive mechanisms between strains, DNA microarrays and targeted metabolome analysis were used.  We identified 1007 temperature dependent genes and 473 strain dependent genes. The transcriptional response was used to identify highly correlated subnetworks of significantly changing genes in metabolism.  We show that temperature differences most strongly affect nitrogen metabolism and the heat shock response.  Lack of STRE mediated gene induction, coupled with reduced trehalose levels, indicates a decreased general stress response at 15°C relative to 30°C.  Between strains, differential responses are centred around sugar uptake, nitrogen metabolism and expression of genes related to organoleptic properties.  Our study provides global insight into how growth temperature exerts a differential physiological and transcriptional response in laboratory and wine strains of S. cerevisiae.

overall_design: We used a two-factor design with two S.cerevisiae strains, grown at two temperatures (ab strain: CEN.PK 113-7D and Industrial Wine strain EC1118, 15ºC and 30ºC, respectively) in anaerobic continuous cultures (D=0.05) in biological triplicates.

first: Michael

middle: C

last: Jewett

first: Jens

last: Nielsen

first: Eduardo

last: Agosin