title: A molecular function map of Ewing’s Sarcoma

city: Vienna

first: Maximilian

last: Kauer

address: Zimmermannplatz 10

institute: CCRI, St.Anna Children Cancer Research Institute

summary:
EWS-FLI1 is a chimeric ETS transcription factor that is, due to a chromosomal rearrangement, specifically expressed in Ewing’s sarcoma family tumors (ESFT) and is thought to be the initiating event in the development of the disease. Previous genomic profiling experiments have identified a number of EWS-FLI1 regulated genes and genes that discriminate ESFT from other sarcomas, but so far a comprehensive analysis of EWS-FLI1 dependent molecular functions characterizing this aggressive cancer is lacking. In this study a molecular function map of ESFT was constructed based on an integrative analysis of gene expression profiling experiments on a uniform microarray platform following EWS-FLI1 knockdown in a panel of five ESFT cell lines, and on gene expression data from the same platform of 59 primary ESFT tumors. Based on the assumption that EWS-FLI1 is the driving transcriptional force in ESFT pathogenesis, we predicted an inverse correlation of gene expression for EWS-FLI1 regulated genes between the putative tissue of origin and the cell lines under EWS-FLI1 knockdown conditions. Consistent with recent reports, mesenchymal progenitor cells (MPC) were found to fit this hypothesis best and were therefore used as the reference tissue for the construction of the molecular function map in ESFT.
The interrelations of molecular pathways were visualized by measuring the similarity among annotated gene functions by gene sharing. The molecular function map highlighted distinct clusters of activities for EWS-FLI1 regulated genes in ESFT and revealed a striking difference between EWS-FLI1 up- and down-regulated genes: EWS-FLI1 induced genes mainly belong to cell cycle regulation, proliferation and response to DNA damage, while repressed genes were associated with differentiation and cell communication.
This study revealed that EWS-FLI1 combines by distinct molecular mechanisms two important functions of cellular transformation in one protein, growth promotion and differentiation blockage. By taking MPC as a reference tissue a significant EWS-FLI1 signature was discovered in ESFT that only partially overlapped with previously published EWS-FLI1 dependent gene expression patterns, identifying a series of novel targets for the chimeric protein in ESFT. Our results may guide target selection for future ESFT specific therapies.

Keywords: Ewing's Sarcoma, EWS-FLI1, RNAi knock down and controls

overall_design: EWS-FLI1 knock down was performed by expressing specific siRNAs against EWS-FLI1 as small hairpin (sh) RNAs from pSUPER-based retroviral expression constructs in 5 different Ewing's sarcoma cell lines (WE68, SK-N-MC, TC252, STA-ET-1, STA-ET-7.2). As a  negative control in each cell line, pSTNeg (Ambion, Applied Biosystems, Brunn am Gebirge, Austria) encoding a scrambled shRNA with no significant similarity to human sequences was used.

first: Jozef

last: Ban

first: Heinrich

last: Kovar

first: Bob

last: Walker

first: Sean

last: Davis

first: Paul

last: Meltzer

first: Reinhard

last: Kofler


title: Integrated genomics approach to detect allelic imbalances in multiple myeloma

status: Public on Apr 14 2009

city: MILAN

first: Luca

last: Agnelli

address: Venezian 1

department: Department of Advanced Diagnostics

country: Italy

institute: IRCCS Istituto Nazionale dei Tumori

summary:
Multiple myeloma (MM) is characterized by marked genomic instability. Beyond structural rearrangements, a relevant role in its biology is represented by allelic imbalances leading to significant variations in ploidy status. To better elucidate the genomic complexity of MM, we analyzed a panel of 45 patients using combined FISH and microarray approaches. Using a self-developed procedure to infer exact local copy numbers for each sample, we identified a significant fraction of patients showing marked aneuploidy. A conventional clustering analysis showed that aneuploidy, chromosome 1 alterations, hyperdiploidy and recursive deletions at 1p and chromosomes 13, 14 and 22 were the main aberrations driving samples grouping. Then, we integrated mapping information with gene and microRNAs expression profiles: a multiclass analysis of the identified clusters showed a marked gene-dosage effect, particularly concerning 1q transcripts, also confirmed by correlating gene expression levels and local copy number alterations. A wide dosage effect affected also microRNAs, indicating that structural abnormalities in MM closely reflect in their expression imbalances. Finally, we identified several loci in which genes and microRNAs expression correlated with loss-of-heterozygosity occurrence. Our results provide insights into the composite network linking genome structure and gene/microRNA transcriptional features in MM.

Keywords: Integrated genomics approach based on SNP microarray and FISH procedures to detect allelic imbalances in multiple myeloma.

overall_design: This series of microarray experiments contains the gene expression profiles of purified plasma cells (PCs) obtained from 5 normal donors (N), 11 monoclonal gammopathy of undetermined significance (MGUS), 133 multiple myeloma (MM) and 9 plasma cell leukemia (PCL) at diagnosis. PCs were purified from bone marrow specimens, after red blood cell lysis with 0.86% ammonium chloride, using CD138 immunomagnetic microbeads. The purity of the positively selected PCs was assessed by morphology and flow cytometry and was > 90% in all cases. 5 micrograms of total RNA was processed and, in accordance with the manufacturer's protocols, 15 micrograms of fragmented biotin-labelled cRNA were hybridized on GeneChip Human Genome U133A Arrays (Affymetrix Inc.). The arrays were scanned using the Agilent GeneChip Scanner G2500A. The images were acquired using Affymetrix MicroArray Suite (MAS) 5.0 software and the probe level data converted to expression values using the Bioconductor function for the Robust Multi-Array average (RMA) procedure (Irizarry et al, 2003), in which perfect match intensities are background adjusted, quantile-quantile normalised and log2 transformed.

first: Laura

last: Mosca

first: Sonia

last: Fabris

first: Marta

last: Lionetti

first: Adrian

last: Andronache

first: Ivo

last: Kwee

first: Katia

last: Todoerti

first: Donata

last: Verdelli

first: Cristina

last: Battaglia

first: Francesco

last: Bertoni

first: Giorgio

middle: D

last: Lambertenghi

first: Antonino

last: Neri


title: ngatha mutant inflorescences

status: Public on Apr 11 2009

city: Rehovot

first: Idan

middle: D.

last: Efroni

address: Weizmann

country: Israel

institute: Weizmann Institute

summary: Floral organ identities are specified by few transcription factors which act as master regulators. Subsequently, specification of organ axes programs the distribution of distinct tissue types within the organs that themselves develop unique identities. The C-class, AGAMOUS-clade MADS box genes are primary promoters of the gynoecium which is divided into a distal style and a subtending ovary along the apical-basal axis. We show that members of a clade of B3-domain transcription factors, NGATHA1 to NGA4 (NGA1-4), are expressed distally in all lateral organs, and all four have a redundant and essential role in style development. Loss of all four genes results in gynoecia where style is replaced by valve-like projections and a reduction in style-specific SHATTERPROOF1 (SHP1) expression. In agreement, floral misexpression of NGA1 promotes ectopic style and SHP1 expression. STYLISH1, an auxin biosynthesis inducer, conditionally activated NGA genes, which in turn, promoted distal expression of other STY genes in a putative positive feed back loop. Inhibited auxin transport or lack of YABBY1 gene activities resulted in a basally expanded style domain and broader expression of NGA genes. We speculate that early gynoecium factors delimit NGA gene response to an auxin-based signal, elicited by STY gene activity, to restrict the activation of style program to a late and distal carpel domain.

overall_design: RNA was extracted from inflorescences of 30 day old plants using Qiagen RNAEasy kit. cRNA was synthesized and hybridized to Affymetrix ATH1 array according to manufacture recommendation. Seven repeats of wild type, two repeats of nga quadruple mutants and two repeats of 35S:amiR-NGA164a were collected. Signal values were obtained and normalized using the GC-RMA protocol, using R 2.7.2 (www.r-project.org) and Bioconductor 2.2 (www.bioconductor.org/). Average correlation values between repeats was 0.985, while the average correlation between wild type and mutant samples was only 0.972. As the average correlation between samples the nga quadruple mutants and 35S:amiR-NGA164a plants was 0.984, and given the identical phenotype, we grouped the two genotypes together.

first: John

middle: P

last: Alvarez

first: Alexander

last: Goldshmidt

last: Bowman

first: Yuval

last: Eshed


title: Expression data from Non small cell lung cancer cell lines

status: Public on Apr 15 2009

city: Cologne

first: Thomas

last: Zander

address: Kerpenerstr. 62

country: Germany

institute: University Hospital Cologne

summary:
Non small cell lung cancer is the leading cause of cancer related mortality in the western world. RNA expression profiles have been demonstrated to be associated with a specific clinical course of the disease.
We used microarray analysis to capture the whole transcriptome of a series of lung cancer cell lines to extract RNA profiles associated with specific genomic lesions

Keywords: steady state

overall_design: Lung cancer cell lines were exponentially grown and harvested during this phase of exponential growth

first: Roman

middle: K

last: Thomas


title: Proteomic and Genomic Profiling of Bronchial Epithelial Cells in Never and Current Smokers

status: Public on Apr 10 2009

city: Boston

first: Avrum

last: Spira

state: MA

address: 715 Albany Street, R304

department: Pulmonary and Critical Care Medicine

country: USA

institute: Boston University Medical Center

summary:
Comparison of gene and protein expression in the large airway epithelium of never and current smokers.
Keywords: gene expression array-based (RNA / in situ oligonucleotide)

overall_design: Large airway bronchial epithelial cells were obtained from never and current smokers by brushing the right mainstem bronchus during fiberoptic bronchoscopy.

first: Katrina

middle: A

last: Steiling

first: Agnes

last: Bergerat

first: Aran

middle: Y

last: Kadar

first: Jim

last: Flanigon

first: Jerome

middle: S

last: Brody

first: Martin

last: Steffen

first: Avi


title: Gene signatures of progression and metastasis in renal cell cancer

status: Public on Apr 28 2009

first: Hasan

middle: Huseyin

last: Otu

address: 4 Blackfan Circle Rm. 238

department: Medicine

institute: Harvard Medical School

summary: In order to address the progression, metastasis, and clinical heterogeneity of renal cell cancer (RCC), transcriptional profiling with oligonucleotide microarrays (22,283 genes) was done on 49 RCC tumors, 20 non-RCC renal tumors, and 23 normal kidney samples. Samples were clustered based on gene expression profiles and specific gene sets for each renal tumor type were identified. Gene expression was correlated to disease progression and a metastasis gene signature was derived. Gene signatures were identified for each tumor type with 100% accuracy. Differentially expressed genes during early tumor formation and tumor progression to metastatic RCC were found. Subsets of these genes code for secreted proteins and membrane receptors and are both potential therapeutic or diagnostic targets. A gene pattern ("metastatic signature") derived from primary tumors was very accurate in classifying tumors with and without metastases at the time of surgery. A previously described "global" metastatic signature derived by another group from various non-RCC tumors was validated in RCC. Unlike previous studies, we describe highly accurate and externally validated gene signatures for RCC subtypes and other renal tumors. Interestingly, the gene expression of primary tumors provides us information about the metastatic status in the respective patients and has the potential, if prospectively validated, to enrich the armamentarium of diagnostic tests in RCC. We validated in RCC, for the first time, a previously described metastatic signature and further showed the feasibility of applying a gene signature across different microarray platforms. Transcriptional profiling allows a better appreciation of the molecular and clinical heterogeneity in RCC.

overall_design:
We used the following tissue samples to obtain transcriptional profiling of kidney tumors using Affymetrix HGU-133A chips: 23 Normal, 32 clear cell RCC (cRCC), 11 papillary RCC (pRCC), 6 chromophobe RCC (chrRCC), 12 Oncocytoma (OC), and 8 transitional cell carcinoma (TCC).

The supplementary file 'GSE15641_mas5_data.txt' contains MAS5 signal values for the Samples included in Series GSE15641.

first: Jon

last: Jones

middle: H

first: Dimitrios

last: Spentzos

first: Mehmet

last: Inan

first: Wolf

last: Beecken

first: Christian

last: Fellbaum

first: Xuesong

last: Gu

first: Marie

last: Joseph

first: Allan

middle: J

last: Pantuck

first: Dietger

last: Jonas

first: Towia

last: Libermann


title: Expression data from 1h red light versus dark 7-day-old Arabidopsis whole seedlings

city: Gainesville

first: Melanie

last: Correll

state: FL

address: 142 Frazier Rogers Hall

department: Agricultural and Biological Engineering

institute: University of Florida

summary:
Red light can affect a variety of responses in Arabidopsis.  We characterize the early gene expression patterns of seedlings exposed to 1 hour of red light using a small sized sample of 5, 7-day-old seedlings and also performed dark controls.
Methods that were developed for tissue extraction and labeling for microarrays were tested using these samples

Keywords: 1 time point

overall_design: Only 5, 7 day-old dark-grown Arabidopsis seedlings were treated as described and harvested for RNA extraction and hybridization on Affymetrix microarrays. We pooled samples from one Petri plate with MS medium 2% sucrose.  Treatment was 1 hour of red light 630 nm (max) and dark controls

first: Alex

last: Stimpson


title: A complementary role for ELF3 and TFL1 in the regulation of flowering time by ambient temperature.

status: Public on Apr 18 2009

city: Ciudad de Buenos Aires

first: Pablo

middle: Diego

last: Cerdan

address: Av. Patricias Argentinas 435

country: Argentina

institute: Fundacion Instituto Leloir

summary: Plants regulate their time to flowering by gathering information from the environment. Photoperiod and temperature are among the most important environmental variables. Suboptimal, but not near-freezing, temperatures regulate flowering through the thermosensory pathway, which overlaps with the autonomous pathway. Here we show that ambient temperature regulates flowering by two genetically distinguishable pathways, one that requires TFL1 and another that requires ELF3. The delay in flowering time observed at lower temperatures was partially suppressed in single elf3 and tfl1 mutants, whereas double elf3 tfl1 mutants were insensitive to temperature. tfl1 mutations abolished the temperature response in cryptochrome mutants that are deficient in photoperiod perception, but not in phyB mutants that have a constitutive photoperiodic response. Contrary to tfl1, elf3 mutations were able to suppress the temperature response in phyB mutants, but not in cryptochrome mutants. The gene expression profile revealed that the tfl1 and elf3 effects are due to the activation of different sets of genes and identified CCA1 and SOC1/AGL20 as being important cross talk points. Finally, genome-wide gene expression analysis strongly suggests a general and complementary role for ELF3 and TFL1 in temperature signalling.

overall_design: Three genotypes, WT (Columbia), elf3-7 and tfl1-1 mutants. Three biological replicates for each condition (genotype X temperature combination). RNA prepared independently for each sample.

first: Bárbara

last: Strasser

first: Mariano

last: Alvarez

first: Andrea

last: Califano


title: Transcriptome Profiles of  Differentiated arabidopsis Leaf Cells and Dedifferentiating Protoplasts of Ler and kyp mutant

status: Public on Apr 02 2009

city: beer sheva

first: inbar

last: plaschkes

address: BGU univarsity

institute: NIBN-BGU

summary: Dedifferentiation signifies the capacity of somatic cells to acquire stem cell-likeproperties. This process characterizes the transition of differentiated plant cells toprotoplasts (plant cells devoid of cell walls), a transition accompanied by widespreadchromatin decondensation. Transcriptome profiling of dedifferentiating cells revealedstriking similarities with senescing cells; both display a large increase in the expressionof genes of specific transcription factor (TF) families including ANAC, WRKY, bZIPand C2H2. We further showed that leaves induced to senesce by exposure to dark displaycharacteristic features of dedifferentiating cells including widespread chromatindecondensation, disruption of the nucleolus and condensation of rRNA genes.Considering that premature senescence can be induced by various stress conditions, ourresults suggest that plant cells response to certain stresses converges on cellulardedifferentiation whereby cells first acquire stem cell-like state prior to acquisition of anew cell fate (e.g., reentry into the cell cycle, or death).

overall_design:
Labeled cRNA was prepared and hybridized to Affymetrix ATH1 GeneChips, 4 biological repeats (2 Ler and 2 kyp-2) for protoplasts and 3 biological repeats for leaves (2 Ler and 1 kyp-2).
Partek data processing results linked below as supplementary file.

first: Gideon

last: Grafi


title: Transcriptional response of Drosophila cells to FHV RNA1 replicon expression (replicon experiment)

status: Public on Apr 01 2009

city: Ann Arbor

first: David

last: Miller

state: MI

address: 3560B MSRB II, 1150 W. Medical Center Dr.

department: Internal Medicine

institute: University of Michigan

summary:
Virus infections induce cellular gene up and down regulation, and these changes often provide clues to cellular pathways utilized by viruses.
We used microarrays to examine the transcriptional responses of cultured Drosophila S2 cells to Flock House virus (FHV) replicon induction.

overall_design: Cultured S2 cells stably transfected with either a control replicon (pS2F1fs) or an FHV RNA1 replicon (pS2F1) were induced with 1 mM copper and we measured global transcript levels at 18 h after induction using Affymetrix Drosophila Genome 2.0 microarray chips.


title: Transcriptional response of Drosophila cells to FHV RNA replication

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

overall_design: Refer to individual Series


status: Public on Apr 06 2009

city: SAS Nagar

first: Ram

middle: Kishor

last: Yadav

state: Punjab

address: Sector 81

department: Biology

country: India

institute: Indian Institute of Science Education and Research Mohali

summary:
A gene expression map of Arabidopsis thaliana shoot apical meristem stem cell niche was generated by isolating the specific cell type using the cell sorting methods.  We used ap1-1;cal1-1 mutant background to enrich the sufficient number of cells for microarray analysis.

Spatial and temporal regulation of gene expression is critical for stem-cell homeostasis. The shoot apical meristems of Arabidopsis thaliana harbor a small set of stem-cells which are surrounded by several million differentiating cells, imposing a severe limitation on the genomic analyses of stem-cell homeostasis. We have employed cell type-specific gene expression profiling that allowed us to generate a high-resolution gene expression map and it has revealed gene expression networks specific to the cell types of the stem-cell niche. We demonstrate that the expression map can be used to predict in vivo gene expression domains to identify novel gene expression patterns. Furthermore, it has revealed molecular pathways that are conserved among plant and animal stem-cell populations. The expression map should guide future reverse genetics experiments, high-resolution analyses of cell-cell communication networks and epigenetic modifications.

Keywords: cell type comparison

overall_design: Three replicates were used CLV3p, CLV3n and FILp, while for WUSp only two replicates were used. CLV3n cells lacks CLV3p cell type.

first: Thomas

last: Girke

first: Sumana

last: Pasala

first: G.

middle: V

last: Reddy


title: Expression profiling of transgenic Drosophila: neurons

status: Public on Mar 31 2009

city: Duarte

first: Paul

middle: M.

last: Salvaterra

state: CA

address: 1500 E. Duarte Rd

department: Neuroscience

institute: Beckman Research Institute. City of Hope National Medical Center

summary:
Expression profiling of Drosophila neurons visualized with transgenic reporter genes, 7.4 kb Cha-Gal:UAS-GFP or 2.1 kb Gad1(s)-RFP.
Keywords: neuronal cell type comparison

overall_design: Transgenic Drosophila embryo cultures (5-7 day old) incubated at 25 C, dissociated and harvested for Fluorescent Active Cell Sorting (FACS) prior to microarray analysis.

middle: M


title: Buffering of segmental and chromosomal aneuploidies in Drosophila melanogaster

status: Public on Mar 30 2009

city: Umea

first: Jan

last: Larsson

address: Umea University

department: Molecular Biology

country: Sweden

institute: Umea University

summary:
Chromosomal instability which involves deletion and duplication of chromosomes or chromosome parts is a common feature of cancers, and deficiency screens are commonly used as a method to find genes involved in different biological pathways. Still, how gene expression from whole chromosomes or large chromosomal domains is affected by deficiencies, duplications or chromosome loss is largely unknown. Using expression microarrays of deficiency hemizygotes and a duplication hemizygote we show that expressed genes are significantly buffered when present in a deficiency hemizygote and that the buffering effect is general and not mainly caused by feedback regulation of individual genes. Differentially expressed genes are in general better buffered than ubiquitously expressed genes when present in one copy. When present in three copies, differentially expressed genes are in general less buffered than ubiquitously expressed genes. Furthermore, we show that the 4th chromosome is compensated in response to dose differences. Our results suggest that general mechanisms exist to stimulate and to repress gene expression of aneuploidy regions and on the 4th chromosome this compensation is mediated by POF (Painting of Fourth).

Keywords: dose response

overall_design: We prepared total RNA from flies heterozygous for three different deletions, Df(2L)J-H, Df (2L)ED4470, Df(2L)ED4651, flies heterozygous for the duplication Dp(2;2)Cam3 and flies with only one chromosome 4, or three copies of the 4th chromosome, as well as from wild type control flies. Three biological replicates of all genotypes were prepared.

first: Per

last: Stenberg


title: Pof D119 mutant expression analysis in Drosophila melanogaster testis

summary:
To study the influence of POF on transcription in testis we made expression arrays from dissected testes (Pof mutants and wild type control). It is clear that POF mainly alters the expression of genes on the fourth chromosome

Keywords: mutant analysis

overall_design: Three replicates, each of 60 testes from 0-24h old males (Pof mutants and wild type control), were dissected and total RNA prepared

first: Malin

last: Svensson


title: Baxter_etal_esb1_mapping

city: St. Louis

first: Ivan

last: Baxter

state: MO

address: 975 North Warson Rd

department: PLant Genetics Research Unit

institute: USDA-ARS

type: Genome variation profiling by genome tiling array

summary:
ABSTRACT: Background: Though central to our understanding of how roots perform their vital function of scavenging water and solutes from the soil, no direct genetic evidence currently exists to support the foundational model that suberin acts to form a chemical barrier limiting the extracellular, or apoplastic, transport of water and solutes in plant roots.
Methodologies/Principle Findings: Using the newly characterized enhanced suberin1 (esb1) mutant, we established a connection in Arabidopsis thaliana between suberin in the root, and both water movement through the plant, and solute accumulation in the shoot. Esb1 mutants, characterized by increased root suberin, were found to have reduced day time transpiration rates, and increased water use efficiency during their vegetative growth period. Furthermore, these changes in suberin and water transport were associated with decreases in the accumulation of Ca, Mn and Zn, and increases in the accumulation of Na, S, K, As, Se and Mo in the shoot.
Conclusions/Significance: Here we present direct genetic evidence establishing that suberin in the roots plays a critical role in controlling both water and mineral ion uptake and transport to the leaves. The changes observed in the elemental accumulation in leaves are also interpreted as evidence that a significant component of the radial root transport of Ca, Mn and Zn occurs in the apoplast.

Keywords: genomic hybridization bulked segregant analysis

overall_design: Hybridizations from a set of Bulk Segregant analysis. An F2 population from 14501 in the col-0 background crossed to Ler-0 was analyzed. This series contains the 3 hybs from Ler used to identify Single Feature Polymorphisms, the 3 hybs of Col-0 they were compared to, and 1 hyb for each pool from the BSA mapping (High Ca/B pool, low Ca/B Pool). It also included 2 tiling array hybs: a wt (Col-0) and a mutant (esb1-1).


title: ABA and ethylene signaling crosstalk

city: Taipei

first: Wan-Hsing

last: Cheng

address: 128, Section 2, Academia Road, Nankang

department: Institute of Plant and Microbial Biology

country: Taiwan

institute: Academia Sinica

summary: Our current study showed that the ABA and ethylene signal transduction pathways function in parallel and have antagonistic interaction during seed germination and early seedling growth. To further address the possible mechanism by which these two hormones crosstalk, microarray analysis was performed. By microarray analysis we found that an ACC oxidase (ACO) was significantly up-regulated in the aba2 mutant, whereas the 9-CIS-EPOXYCAROTENOID DIOXYGENASE (NCED3) gene in ein2, and both the ABSCISIC ACID INSENSITIVE1 (ABI1) and cytochrome P450, family 707, subfamily A, polypeptide 2 (CYP707A2) genes in etr1-1 were up- and down-regulated, respectively. These data further suggest that ABA and ethylene may control the hormonal biosynthesis, catabolism or signaling of each other to enhance their antagonistic effects upon seed germination and early seedling growth.

overall_design: Cold pre-treated seeds of wild type (Col), aba2 (or gin1-3), etr1-1, and ein2-1, were grown on 1% sucrose agar plates for 12 to 14 days. Total RNAs were extracted from which 10 μg aliquots were used for cDNA synthesis, labeling by in vitro transcription followed by fragmentation according to the GeneChip Expression Analysis Technical Manual rev5, Affymetrix. Eleven μg of each labeled samples was hybridized to an ATH1 GeneChip at 45oC for 16.5 hours. The washing and staining steps were performed using a Fluidic Station-450 and the ATH1 slides were scanned using the Affymetrix GeneChip Scanner 7G. Subsequent data processing and analysis was performed using the Affymetrix Microarray Suit Software version 5.0. Two independent sets of microarray analyses were performed in this study. The wild type in this study was used as a control.

first: Ming-Hau

last: Chiang

first: San-Gwang

last: Hwang

first: Pei-Chi

last: Lin


title: Early Iron Deficiency Induced Changes in Arabidopsis Roots

status: Public on Mar 13 2009

first: Wolfgang

last: Schmidt

address: No. 128, Sec 2, Academia Rd, Nankang

summary: Due to limited bio-availability of Fe, plants evolved adaptive alterations in development regulated at the transcriptional level. We investigated the early transcriptional response to Fe deficiency.

overall_design: A hydroponic system was used that allows bulk transferring of plantsand rapid Fe removal with little mechanical damage to the root systems.

last: Buckhout

last: Yang


title: Transcriptional profiling after inhibition of cellulose synthesis by TA and IXB in Arabidopsis thaliana suspension cells

status: Public on Aug 27 2009

city: Sherbrooke

first: Nathalie

last: Beaudoin

state: Quebec

address: 2500, Blvd Universite

department: Faculte des Sciences

country: Canada

institute: Universite de Sherbrooke

summary:
Transcriptional profiling after inhibition of cellulose synthesis by thaxtomin A and isoxaben in Arabidopsis thaliana suspension cells
Perturbations in the cellulose content of the plant cell wall lead to global modifications in cellular homeostasis, as seen in cellulose synthase mutants or after inhibiting cellulose synthesis. In particular, application of inhibitors of cellulose synthesis such as thaxtomin A (TA) and isoxaben (IXB) initiates a programmed cell death (PCD) in Arabidopsis thaliana suspension cells that is dependent on de novo gene transcription.
To further understand how TA and IXB activate PCD, a whole genome microarray analysis was performed on mRNA isolated from Arabidopsis suspension cells exposed to TA and IXB. More than 75% of the genes upregulated by TA were also upregulated by IXB, including genes encoding cell wall-related and calcium-binding proteins, defence/stress-related transcription factors, signalling components and cell death-related proteins. Comparisons with published transcriptional analyses revealed an important subset of genes generally induced in response to various biotic and abiotic stress.

overall_design: TA,  IXB and methanol (control) were added to Arabidopsis thaliana suspension cells three days after sub-culture. Cells were harvested for RNA isolation and frozen in liquid nitrogen after 6 hours of contact with the inhibitors. Samples consisted of four replicates for each condition.  A total of 12 Affymetrix GeneChips® were used in this study, which correspond to 12 RNA samples from the four biological replicates for each of the TA, IXB or methanol addition.

first: Isabelle

last: Duval


title: A genomic approach to bud mutation in sweet orange (Citrus sinensis L. Osbeck) fruit

status: Public on Mar 10 2009

city: Wuhan

first: Qing

last: Liu

address: Shizishan street NO.1

department: National Key Laboratory of Crop Genetic Improvement

country: China

institute: Huazhong Agricultural University

summary:
Bud mutations arise often in citrus. The selection of mutants is one of the most important breeding methods in citrus. However, the molecular bases of bud mutation have rarely been studied. To identify the potential important or novel genes involved in bud mutation, different transcriptomic techniques combing suppression subtractive hybridization (SSH) and microarray were performed between a lycopene accumulated mutant, ‘Hong Anliu’ sweet orange (Citrus sinensis L. Osbeck), and its wild-type during fruit maturation. Microarray analysis revealed that differentially expressed clones are extensively coordinated with the initiation of lycopene accumulation. After sequencing of the differentially expressed clones, a total of 267 non-redundant transcripts were obtained, 182 (68.2%) of which share homology (E-value ≤ 1×10-10) with known gene products or known protein domains. A list of candidate genes which involved in cellular metabolic process, primary metabolic process, localization, macromolecular metabolic process was obtained. Out of these genes, 12 share homology with previously described signal transduction or transcription factors, suggesting complex regulatory control. These results demonstrate profound effect on gene expression of bud mutation in citrus fruits and provide new insights into the molecular basis of bud mutation.
Keywords: bud mutation, candidate genes, Citrus, cNDA microarray, suppression subtractive hybridization (SSH)

overall_design: Fruits from the mutant and its wild type were collected at five time points from August to December. Total RNA extracted from the mutant was hybridized to the array together with RNA from the wild type. Each hybridization was performed in duplicate by dye swap.

first: Andan

last: Zhu

first: Lijun

last: Cai

first: Wenjing

last: Zhou

first: Keqin

last: Yu

first: Xiuxin

last: Deng


title: PAX3-FKHR regulated genes differentially expressed between alveolar and embryonal rhabdomyosarcoma

city: Philadelphia

first: Frederic

last: Barr

state: PA

address: 422 Curie Blvd

institute: University of Pennsylvania

summary: The overall goal and objective is to study the degree to which PAX3-FKHR accounts for differences between ARMS and ERMS by expressing a construct (termed P3FK/ER) consisting of PAX3-FKHR joined to the estrogen receptor ligand binding domain in an ERMS cell culture system.

overall_design: In the study design, three populations of RD ERMS cells expressing P3FK/ER were treated with or without 30 nM 4-hydroxytamoxifen and three control populations transduced with vector only (pK1) were similarly treated.

first: Gabriela

middle: E

last: Mercado

middle: G


title: Logjam p24 mutant microarray analysis

status: Public on Mar 16 2009

city: College Station

first: Ginger

middle: E.

last: Carney

state: TX

address: 3258 TAMU

institute: Texas A&M University

summary:
We performed genome-wide profiling experiments on tissues from Drosophila melanogaster adult females with a mutation in the p24 gene logjam (loj) to test the hypothesis that loss of loj function causes a transcriptional response characteristic of ER stress activation.
Keywords: logjam mutant compared to control

overall_design: We compared expression profiles from loj mutant females to those of controls. We compared mutant and control abdomen tissue as well as mutant and control tissue preparations from the remainder of the fly (the head/thorax). Three mutant and three control RNA samples for each set of tissue were used for array hybridization (12 total array samples).

first: Kara

last: Boltz


title: Expression data from Arabidopsis seed compartments at the heart stage.

status: Public on Mar 12 2009

city: Davis

first: John

last: Harada

address: One Shields Avenue

department: Plant Biology

institute: University of California, Davis

summary: We collected heart stage seed compartments from 7 micron paraffin sections using the Leica LMD6000 system in order to identify the mRNAs present in different compartments from seeds containing heart stage embryos. For the purposes of this study we captured 6 compartments: embryo proper, micropylar endosperm, peripheral endosperm, chalazal endosperm, chalazal seed coat and seed coat, as well sets of serial sections encompassing the entire heart stage seed.

overall_design: Heart stage seed compartments were isolated using the LMD6000 system. Total RNA was amplified and hybridized with Affymetrix ATH1 Arabidopsis array for 16 samples (embryo proper, micropylar endosperm, peripheral endosperm, seed coat and whole seeds, 2 biological replicates each; chalazal endosperm, chalazal seed coat, 3 biological replicates each).

first: Sandra

middle: L

last: Stone

first: Julie

last: Pelletier

first: Ryan

middle: C

last: Kirkbride

first: Mark

middle: F

last: Belmonte

first: Jiong

last: Fei

first: Meryl

last: Hashimoto

first: Chen

last: Cheng

first: Robert

middle: B

last: Golberg


title: Alterations of gene expression in human peripheral lymphocytes after exposure to cigarette smoke condensate in vitro

status: Public on Aug 28 2009

city: Washington

first: Jinguo

last: Chen

state: DC

address: 3800 Reservoir Road NW

department: Cancer Genetics and Epidemiology

institute: Georgetown University Lombardi Cancer Center

summary:
Little is known about alteration of the global gene expression by cigarette smoke (CS) and few biomarkers for smoking-related harm are available.
We used Affymetrix HG-U133A GeneChips to measure the transcriptomes in eight cultured lymphocyte samples exposed to cigarette smoke condensate (CSC) in vitro . The in vitro exposure of lymphocytes to CSC significantly changed expression levels of 2,266 genes many of which biologically interacted. They included genes encoding for xenobiotic metabolism and oxidative stress-response (e.g. Nrf2 and AhR signaling pathways), inflammation/immune response (e.g. cytokines), apoptosis, cell cycle and tumorigenesis. However, the magnitude of expression responses for some genes showed high inter-individual variability.

Keywords: toxic effects on gene expression

overall_design: The goals of this study were to evaluate novel gene expression profiles and pathways affected by cigarette smoke condensate (CSC), and to identify potential biomarkers for cigarette smoke exposure and harm. To this end,  we isolated the PBMC from eight light smokers, cultured the cells in vitro and exposed them to 2R4F CSC, then determined the gene expression profiles with Affymetrix microarray and analyzed alteration of global gene expression after exposure to CSC.

first: Peter

last: Shields


title: The effects of PTIP on Polycomb mediated repression in Drosophila Kc167 cells.

status: Public on Mar 04 2009

first: Greg

last: Dressler

address: 109 Zina Pitcher Drive

department: Pathology

summary: Drosophila Kc167 cells were used to test the function of PTIP in regulating gene expression before and after loss of polycomb function.  Four condtions were tested using dsRNA knockdowns (RNAi)

overall_design: Kc167 cells alone, Kc cells with loss of PTIP, cells with loss of polycomb and polyhomeotic, and cells with loss of PTIP, polycomb, and polyhomeotic. Performed in triplicate.

first: Ming

last: Fang

first: Gregory

middle: R


title: The differentially expressed genes identified in the microarray analysis using myb5 and wild-type (Col) seeds

status: Public on Mar 02 2009

city: Melbourne

first: Song

middle: Feng

last: Li

state: Victoria

address: Plenty Rd

department: Botany

country: Australia

institute: La Trobe University

summary: The MYB gene family encodes transcription factors with a diverse range of functions in Arabidopsis. This study demonstrated that MYB5, which is expressed in trichomes and seeds, plays a central role in trichome and seed development. A microarray analysis of myb5 seeds identified other members of the MYB5 regulatory network.

overall_design: Four wild-type biological replicates and three myb5 biological replicates were used for microarray analysis.


title: Positive fluorescent selection permits precise, rapid and in-depth over-expression analysis in plant protoplasts

status: Public on Mar 01 2009

city: New York

first: Kenneth

middle: David

last: Birnbaum

state: NY

address: 12 Waverly Place

institute: New York University

summary:
Transient genetic modification of plant protoplasts is a straightforward and rapid technique for the analysis of numerous aspects of plant biology. One drawback in the analysis of transformed protoplast suspensions is that they are a heterogeneous mix of cells that have and have not been successfully transfected. To overcome this problem, we have developed a system that employs a fluorescent positive selection marker in combination with flow cytometric analysis as well as fluorescence activated cell sorting (FACS) to isolate responses in the transfected protoplasts exclusively. This recombinase-compatible system enables high-throughput screening of genetic circuitry. Moreover, the use of FACS allows in depth downstream analysis. Lastly, over-expression is an effective means to dissect regulatory networks, especially where redundancy exists. Here, this system has been applied to the study of auxin signaling in order to investigate reporter gene activation and genome-wide transcriptional changes in response to manipulation of the auxin-response network. We have transiently over-expressed dominant negative mutant isoforms of Aux/IAA transcription factors (IAA7mII and IAA19mII; Tiwari et al., 2001) in Arabidopsis Pwer::GFP root protoplasts, making use of a RFP fluorescent positive selection marker and FACS to isolate the dually labeled (IAAnmII expressing and Pwer::GFP-positive) cells. We have compared the transcriptional differences between an empty vector control, IAA7mII and IAA19mII protoplasts that had either been treated with 5microM IAA or mock-treated for 3 hours.

Keywords: hormone treatment, genetic modification

overall_design: 18 samples with 3 replicates for each condition and transformation vector: 3x empty vector mock treated, 3x empty vector IAA treated, 3x IAA7mII over-expressor mock treated, 3x IAA7mII over-expressor IAA treated, 3x IAA19mII over-expressor mock treated and 3x IAA19mII over-expressor IAA treated.

middle: D

first: Bastiaan

middle: O

last: Bargmann


title: Expression data from Arabidopsis Seedlings treated with Salicylic Acid

status: Public on Feb 24 2009

city: Lucknow

first: Samir

last: Sawant

state: U.P, India

address: Rana Pratap Marg

department: Centre of Plant Molecular Biology

institute: National Botanical Research Institute

summary:
Disease resistance is associated with a plant defense response that involves an integrated set of signal transduction pathways. Gene Expression studies were performed using Arabidopsis wild type seedlings treated with and without salicylic acid (SA). In this study we examined pathogenesis related genes are upregulated.
We used Affymetrix GeneChip to study the genome wide gene expression with and without induction of defense by treatment with SA

overall_design: Arabidopsis seedlings Col-0 were grown for 9 days in controlled environment rooms at 22°C temperature and a photoperiod of 16 h light 100 µm m2 s_1 cool-white fluorescent illumination. After 9 days seedlings were transferred into water as a mock treatment and with the defense related signaling molecules salicylic acid (SA) for one day. The concentration of SA (Sigma, St. Louis) was 2mM and pH 6.5. RNA extracted from SA and mock treated seedling and hybridize with Affymetrix GeneChip to examine the change in plant defence gene expression due to induction of SA.

first: Rakesh

last: Tuli

last: Srivastava

first: Meenal

first: Deepti

last: Nigam


title: Transcriptome changes triggered by the synthetic defense elicitors DCA and INA in Arabidopsis thaliana

status: Public on Mar 20 2009

city: Riverside

first: Thomas

last: Eulgem

address: University of California, Department of Botany and Plant Sciences

department: Botany and Plant Sciences

institute: University of California

summary: DCA (3,5-Dichloroanthranilic acid) is a newly identified synthetic defense elicitor. To perform a comparative analysis of defense responses triggered by DCA and the structurally related defense inducer INA (2,6-Dichloroisonicotinic acid) Affymetrix chip experiments were performed with Arabidopsis thaliana seedlings treated with one of these two compounds.

overall_design: Arabidopsis thaliana plants (accession Col-0) were grown on soil in a semi-sterile growth chamber under fluorescent lights (14 hours light, 10 hours dark, 21 centi grades, 100 Einstein/m2/s). Wild type Col-0 plants and the npr1-3 mutant were used in this study. Aerial tissues of 2 week old soil grown Arabidopsis seedlings were sprayed with 100uM 3,5-Dichloroanthranilic acid (DCA), or 100uM 2,6-Dichloroisonicotinic acid (INA) or mock solution and harvested 48h or 6 days after the treatment. Final DMSO concentrations never exceeded 0.002%.  Mock treatments were application of 0.002% DMSO in water. For harvesting the aerial plant parts were shock-frozen in liquid nitrogen. Total RNA was isolated from seedlings using TRIZOL (Invitrogen) follwing the manufacturer’s instructions. RNA was processed and hybridized to Affymetrix Arabidopsis ATH1 genome array GeneChip following manufacturer’s instructions (Affymetrix) by the University of California, Riverside Core Instrument Facility.  Three independent biological replicates were performed for each treatment.

first: Colleen

last: Knoth

last: Girke


title: Immunogenic males: a genome-wide analysis of reproduction and the cost of mating in Drosophila melanogaster females

status: Public on Feb 19 2009

city: Uppsala

first: Paolo

last: Innocenti

address: Norbyvägen 18D

department: Dept. of Animal Ecology

institute: Uppsala university

summary:
In Drosophila melanogaster, mating radically transforms female physiology and behavior. Post-mating responses include an increase in the oviposition rate, a reduction in female receptivity, and an activation of the immune system . The fitness consequences of mating are similarly dramatic – females must mate once in order to produce fertile eggs, but additional matings have a clear negative effect. Previously, microarrays have been used to examine gene expression of females differing in their reproductive status with the aim of identifying genes influenced by mating. However, since only virgin and single mated females were compared, transcriptional changes associated with reproduction (under natural selection) and the effects of male-induced harm (under sexually antagonistic selection) cannot be disentangled. We partitioned these fundamentally different effects by instead examining the expression profiles of virgin, single mated and double mated females. We found substantial effects relating to reproduction and further effects that are only attributable to a second mating. Immune response genes dominate this male-induced harm effect indicating that the cost of mating may be due partly to this system's activation. We propose that both sexually antagonistic and natural selection have been important in the evolution of the innate immunity genes, thereby contributing to the sexual dimorphismand rapid evolution at these loci.

Keywords: Female response to mating

overall_design: Female flies were flash frozen in liquid nitrogen either as virgins or 6 hours after mating and stored at -80°C until RNA extraction was performed (not more than 2 days). 8 whole flies – randomly selected within each treatment – were pooled for each extraction. Total RNA was extracted using Trizol (Invitrogen) and purified with an RNeasy Mini Kit (Qiagen). RNA quantity and quality was checked with an Agilent Bioanalyzer. According to the manufacturer's instructions, samples were prepared and hybridized to Affymetrix GeneChip Drosophila Genome 2.0 (Affymetrix, Santa Clara, CA, USA) by the Uppsala Array Platform (Uppsala, Sweden). Each experimental treatment consisted of 4 independent RNA extractions and hybridizations, giving a total of 12 arrays.

first: Edward

middle: H

last: Morrow


title: Comparison of gene expression between wild type and gl3 egl3 trichomeless mutant green seedling tissues

city: Columbus

first: Kengo

last: Morohashi

state: OH

address: 1060 Carmack Rd

department: Molecular Genetics

institute: The Ohio State University

summary: The development of trichomes (leaf hairs) from pluripotent epidermal cells in Arabidopsis provides a powerful system to investigate the regulatory motifs involved in plant cell differentiation. Genetic studies have revealed that a bHLH transcription factor, GL3, activates downstream genes required for trichome initiation by interacting with a R2R3-MYB protein, GL1. We have taken advantage of several mutants in the trichome developmental pathway and gene expression analyses to identify a set of genes expressed predominantly in Arabidopsis trichomes.

overall_design: We compared the gene expression between the wild type and trichome mutant, gl3 egl3 using green tissue (not include root).

first: Manli

last: Davis

first: Rebecca

middle: S

last: Lamb

first: Erich

last: Grotewold


title: Inducible expression of trichome regulators, GL1 and GL3

summary: The development of trichomes (leaf hairs) from pluripotent epidermal cells in Arabidopsis provides a powerful system to investigate the regulatory motifs involved in plant cell differentiation. Genetic studies have revealed that a bHLH transcription factor, GL3, activates downstream genes required for trichome initiation by interacting with a R2R3-MYB protein, GL1. In order to investigate genome-wide regulatory functions of GL1 and GL3, we  performed genome-wide expression analyses using GR inducible systems of GL1 and GL3.

overall_design: Transgenic plants carrying pGL1::GL1-GR and pGL3::GL3-GR were treated with mock or Dex for 4 and 24 hours.


title: Expression data from Arabidopsis gapcp mutant

status: Public on Feb 10 2009

city: Burjassot

first: Roc

last: Ros

state: valencia

address: Vicent Andres Estelles

department: Biologia Vegetal

country: Spain

institute: Universitat de Valencia

summary: Glycolytic Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) catalyzes the conversion of glyceraldehyde 3-phospate to 1,3-bisphosphoglycerate by coupling with the reduction of NAD+ to NADH. Both cytosolic and plastidial isoforms of GAPDH has been described but the in vivo functions of the plastidial isoforms is unresolved. We generated mutants of the Arabidopsis plastidial GAPDH isoforms (At1g79530, At1g16300; GAPCp1, GAPCp2) and performed a microarray analysis comparing gapcp double (gapcp1 gapcp2) mutant and wild type seedlings

overall_design: 15-day old Arabidopsis seedlings (Col 0)  and gapcp1gapcp2 double mutants were used  for RNA extraction and hybridization on Affymetrix microarrays.


title: Gene expression analysis of spargel mutant versus genetically matched wildtype control

status: Public on Feb 13 2009

city: Zurich

first: Stefanie

last: Tiefenboeck

address: Schafmattstrasse 18

country: Switzerland

institute: ETH Zurich

summary: Mammalian PGC-1alpha, PGC-1beta and PRC are structurally related transcriptional coactivators, and are involved in multiple metabolic functions, including the regulation of mitochondrial biogenesis. However, due to redundancy, their in vivo roles are still poorly understood. By a genome-wide microarray study, we show that in the Drosophila larval fat body, Spargel (CG9809), the only fly PGC-1 family homologue, is required for proper expression of multiple genes encoding mitochondrial proteins.

overall_design: Six samples were analyzed in total: three biological replicates of the control (wt) and three biological replicates of the mutant (KG).

middle: K

last: Tiefenböck

first: Nicole

middle: A

last: Egli

first: Claudia

last: Baltzer

first: Christian

last: Frei


title: Global gene expression changes including drug metabolism and disposition induced by three-dimensional culture of HepG2

city: Setagaya

first: Seiichi

last: Ishida

state: Tokyo

address: 1-18-1, Kamiyoga

department: Pharmacology

country: Japan

institute: National Institute of Health Sciences

summary:
We found constitutive upregulation and higher degree induction of drug metabolism and disposition-related genes in a three-dimensional HepG2 culture. The upregulated genes are those believed to be regulated by different regulatory factors. The global gene expression analysis by Affymetrix GeneChip indicated that altered expressions of microtubule-related genes may change expressed levels of drug metabolism and disposition genes.  Stabilization of the microtubule molecules with docetaxel, a tubulin stabilizing agent, in the two-dimensional culture showed gene expression patterns similar to those in the three-dimensional culture, indicating that culture environment affects drug metabolism functions in HepG2 cells.

Keywords: radial flow bioreactor cell culture system, three dimensional culture, HepG2, GeneChip U133A,

overall_design: We compared the gene expression data from HepG2 cells cultured in the radial flow bioreactor (RFB) cell culture system for six days to those from HepG2 cells cultured in tissue culture plates for five days. Both cell cultures (RFB culture and plate culture) were performed twice independently. In each culture, three samples were collected from three different portions of the bioreactor or from three different tissue culture plates. Microarray analyses were performed in duplicate for each sample using the Affymetrix human genome U133A GeneChip.

first: Shin-ichiro

last: Horiuchi

first: Tomokatsu

last: Hongo

first: Youichi

last: Ishikawa

first: Atsuko

last: Miyajima

first: Jun-ichi

last: Sawada

first: Yasuo

last: Ohno

first: Ken

last: Nakazawa

first: Shogo

last: Ozawa


title: Effect of Spargel on transcriptional changes mediated by the insulin-signalling pathway

department: Biology

summary: Insulin signalling activity is well known to mediate transcriptional changes in response to nutrients and it is changed in the spargel mutant. We performed a  genome-wide microarray study to investigate the role of Spargel as a transcriptional coregulator downstream of insulin receptor signalling.

overall_design: To investigate the effect of Spargel on transcriptional changes mediated by the insulin-signalling pathway we analyzed 12 samples in total: three biological replicates of the control with or without UAS-Inr (Inr_wt and ctrl_wt) and three biological replicates of the spargel mutant with or without UAS-Inr (Inr_KGand ctrl_KG).


title: Expression data from Drosophila histone methyltransferase mutants

city: St. Louis

middle: Christina

last: Riddle

address: 1 Brookings Dr.; CB1137

institute: Washington University

summary:
H3K9 methylation is a mark of inactive chromatin.  In D.melanogaster, three enzymes (SU(VAR)3-9, EGG, and dG9A) are responsible for creating the H3K9 methyl mark.  We have investigated the effects of loss of SU(VAR)3-9 and EGG on gene expression in adult heads using microarray analysis.

Keywords: Expression profiling by microarray

overall_design: Heads from adults flies were used for RNA extraction and hybridization on Affymetrix microarrays.  Four genotypes were used: yw; +/CyO,  yw; egg235/CyO,   yw; +/CyO; Su(var)3-906;   yw; egg235/CyO; Su(var)3-906.  For each genotype, three biological replicates were analyzed.  For the yw; +/CyO control, an additional technical replicate (same RNA sample processed twice) is included.  While source files for both technical replicates are included, only one was used for the final analysis.


title: A Prognostic Gene Expression Index in Ovarian Cancer

status: Public on Feb 09 2009

city: Berlin

first: Jan

last: Budczies

address: Chariteplatz 1

department: Institute of Pathology

institute: Charite - Universitaetsmedizin Berlin

summary:
Ovarian carcinoma has the highest mortality rate among gynecological malignancies. In this project, we investigated the hypothesis that molecular markers are able to predict outcome of ovarian cancer independently of classical clinical predictors, and that these molecular markers can be validated using independent data sets.   We applied a semi-supervised method for prediction of patient survival. Microarrays from a cohort of 80 ovarian carcinomas (TOC cohort) were used for the development of a predictive model, which was then evaluated in an entirely independent cohort of 118 carcinomas (Duke cohort).  A 300 gene ovarian prognostic index (OPI) was generated and validated in a leave-one-out approach in the TOC cohort (Kaplan-Meier analysis, p=0.0087). In a second validation step the prognostic power of the OPI was confirmed in an independent data set (Duke cohort, p=0.0063). In multivariate analysis, the OPI was independent of the postoperative residual tumour, the main clinico-pathological prognostic parameter with an adjusted hazard ratio of 6.4 (TOC cohort, CI 1.8 – 23.5, p=0.0049) and 1.9 (Duke cohort, CI 1.2 – 3.0, p=0.0068). We constructed a combined score of molecular data (OPI) and clinical parameters (residual tumour), which was able to define patient groups with highly significant differences in survival.  The integrated analysis of gene expression data as well as residual tumour can be used for optimised assessment of prognosis. As traditional treatment options are limited, this analysis may be able to optimise clinical management and to identify those patients that would be candidates for new therapeutic strategies.

Keywords: disease state analysis

overall_design: RNA from 80 frozen ovarian cancer samples was analysed with oligonucleotide microarrays

first: Carsten

last: Denkert

first: Manfred

last: Dietel

first: Hermann

last: Lage


title: Severe Preeclampsia-Related Changes in Gene Expression at the Maternal-Fetal Interface Include Siglec-6 and Pappalysin-2

status: Public on Feb 06 2009

city: SAN FRANCISCO

first: Susan

last: Fisher

address: 513 PARNASSUS AVE

department: Obstretrics, Gynecology and Reproductive Sciences

institute: UCSF

summary:
Preeclampsia (PE), which affects 4-8% of human pregnancies, causes significant maternal and neonatal morbidity and mortality. Within the basal plate, placental cytotrophoblasts (CTBs) of fetal origin invade the uterus and extensively remodel the maternal vasculature. In PE, CTB invasion is often shallow, and vascular remodeling is rudimentary. To better understand possible causes, we conducted a global analysis of gene expression at the maternal-fetal interface in placental samples from women with PE (n = 12; 24-36 wk) vs. samples from women who delivered due to preterm labor with no evidence of infection (n = 11; 24-36 wk), a condition that our previous work showed is associated with normal CTB invasion. Using the HG-U133A&B Affymetrix GeneChip platform, and statistical significance set at log odds-ratio of B >0, 55 genes were differentially expressed in PE. They encoded proteins previously associated with PE [e.g. Flt-1 (vascular endothelial growth factor receptor-1), leptin, CRH, and inhibin] and novel molecules [e.g. sialic acid binding Ig-like lectin 6 (Siglec-6), a potential leptin receptor, and pappalysin-2 (PAPP-A2), a protease that cleaves IGF-binding proteins]. We used quantitative PCR to validate the expression patterns of a subset of the genes. At the protein level, we confirmed PE-related changes in the expression of Siglec-6 and PAPP-A2, which localized to invasive CTBs and syncytiotrophoblasts. Notably, Siglec-6 placental expression is uniquely human, as is spontaneous PE. The functional significance of these novel observations may provide new insights into the pathogenesis of PE, and assaying the circulating levels of these proteins could have clinical utility for predicting and/or diagnosing PE.

Keywords: disease state analysis

overall_design: Basal plate biopsies of preterm labor (24-36 weeks; n=11) and preterm severe preeclampsia (24-36 weeeks; n=12) were isolated and the global gene expression profiles determined using Affymetrix Human GeneChips.  Comparisons between the preeclampsia samples and the preterm labor controls revealed genes differentially expressed in preeclampsia.

first: Virginia

last: Winn

first: Matthew

last: Gormley

middle: J


title: Expression data from Arabidopsis seed compartments at the mature green stage.

first: John

last: Harada

summary: We collected mature green seed compartments from 7 micron paraffin sections using the Leica LMD6000 system in order to identify the mRNAs present in different compartments from seeds containing mature green-stage embryos. For the purposes of this study we captured 6 compartments: embryo proper, micropylar endosperm, cellularized peripherial endosperm, chalazal endosperm, chalazal seed coat and seed coat, as well sets of serial sections encompassing the entire mature green stage seed.

overall_design: Mature green stage seed compartments were isolated using the LMD6000 system. Total RNA was amplified and hybridized with Affymetrix ATH1 Arabidopsis array for 14 samples (embryo proper, micropylar endosperm, peripherial endosperm, chalazal endosperm, chalazal seed coat, seed coat and whole seeds, 2 biological replicates each).

first: Sandra

last: Stone

first: Julie

last: Pelletier

first: Ryan

last: Kirkbride

first: Mark

last: Belmonte

first: Jiong

last: Fei

first: Meryl

middle: M

last: Hashimoto

first: Chen

last: Cheng

first: Robert

last: Golberg


title: Gene expression analysis of delg_mut versus Df_plus control

first: Claudia

last: Baltzer

state: Zurich

department: Cell biology

institute: ETH Zürich

summary: Nuclear transcription factors drive mitochondrial mass by regulating the expression of genes encoding mitochondrial proteins. Among these factors, nuclear respiratory factor 2 (NRF-2/GABP) has been proposed to be critical for mitochondrial mass in mammalian cells, yet there is little genetic evidence to support this function in vivo. Here, we show that mutants of the Drosophila melanogaster NRF-2alpha/GABPalpha homologue Delg (CG6338) have reduced expression of multiple genes encoding mitochondrial proteins, leading to reduced mitochondrial mass.

overall_design: Six samples were analyzed in total: three biological replicates of the control (Df_plus) and three biological replicates of the mutant (Df_delg).

first: Stefanie

last: Tiefenboeck

last: Marti

first: Christian

last: Frei


title: Gene expression profiles of poorly differentiated, undifferentiated and metastatic cancers

status: Public on Mar 29 2009

city: Sunnyvale

first: Ljubomir

last: Buturovic

address: 1196 Borregas Avenue

department: Application Development

institute: Pathwork Diagnostics

summary:
The tissue of origin form metastatic tumors is sometimes difficult to identify from clinical and histologic information.  Gene expression signatures are one potential method for identifying the tissue of origin. In the development of algorithms to identify tissue of origin, a collection of human tumor metastatic specimens with known primary sites or primary tumors with poor differentiation are very useful in identifying gene expressions signatures that can classify unknown specimens as to the tissue of origin.  Here we describe a series of 276 such tumor specimens used for this purpose. The specimens are poorly differentiated, undifferentiated and metastatic specimens from tumors of the following types/tissues of origin: breast, liver, non-Hodgkin's lymphoma, non-small cell lung cancer, ovary, testicular germ cell, thyroid, kidney, pancreas, colorectal cancer, soft tissue sarcoma, bladder, gastric cancer, prostate and melanoma.  This data combined with other series (GSE2109) was used to validate a proprietary tumor classification algorithm of Pathwork Diagnostics. The results of this validation set (N = 545 CEL files) showed that the algorithm correctly identified the tissue of origin for 89.4% of the specimens.

Guidelines for commercial use:
http://pathworkdx.com/GSE12630_request.html

overall_design:
All tissue samples were fresh frozen in liquid nitrogen. The gene expression was profiled using Affymetrix GeneChip platform.

Instructions for opening the Pathwork Pathchip CEL files are appended below.


title: Golovinomyces orontii time course with Col-0 and eds16-1

status: Public on Feb 04 2009

city: Berkeley

first: Mary

last: Wildermuth

address: 221 Koshland Hall MC3102

department: Plant and Microbial Biology

institute: University of California Berkeley

summary:
Salicylic acid (SA) is a critical molecule mediating plant innate immunity with an important role limiting the growth and reproduction of the virulent powdery mildew (PM) Golovinomyces orontii on Arabidopsis thaliana. To investigate this later phase of the PM interaction, and the role played by SA, we performed replicated global expression profiling for wild type and SA biosynthetic mutant ics1 Arabidopsis from 0 to 7 days post infection. We found that ICS1-impacted genes comprise 3.8% of profiled genes with known molecular markers of Arabidopsis defense ranked very highly by the multivariate empirical Bayes statistic (T2 statistic ((Tai and Speed, 2006)). Functional analyses of T2-selected genes identified statistically significant PM-impacted processes including photosynthesis, cell wall modification, and alkaloid metabolism that are ICS1-independent. ICS1-impacted processes include redox, vacuolar transport/secretion, and signaling. Our data also supports a role for ICS1 (SA) in iron and calcium homeostasis and identifies components of SA crosstalk with other phytohormones. Through our analysis, 39 novel PM–impacted transcriptional regulators were identified. Insertion mutants in one of these regulators, PUX2, results in significantly reduced reproduction of the powdery mildew in a cell death independent manner. Though little is known about PUX2, PUX1 acts as a negative regulator of Arabidopsis CDC48 (Rancour et al., 2004; Park et al., 2007), an essential AAA-ATPase chaperone that mediates diverse cellular activities including homotypic fusion of ER and Golgi membranes, ER-associated protein degradation, cell cycle progression, and apoptosis. Future work will elucidate the functional role of the novel regulator PUX2 in PM  resistance.

Keywords: time course, pathogen response

overall_design: Arabidopsis whole leaves were harvested at 6h, 1 day, 3 days, 5 days and 7 days after Golovinomyces orontii infection for RNA extraction and hybridization to Affymetrix Arabidopsis ATH1 microarrays. Gene expression profiles were obtained for wild type Columbia-0 and enhanced disease susceptibility mutant eds16-1, a null isochorismate synthase 1 (At1g74710) mutant. In parallel, uninfected samples were collected at 0 hr and 7days from wild type and mutant plants.  The experiment includes 4 biological replicates.

first: Frederick

last: Ausubel


title: A Gene Signature for Chronic Fatigue Syndrome

status: Public on Jan 27 2009

city: Glasgow

first: Suzanne

last: Hagan

state: Lanarkshire

address: Cowcaddens Road

department: Biological Sciences

country: United Kingdom

institute: Glasgow Caledonian University

summary:
Human genome-wide Affymetrix GeneChip arrays were used to compare the levels of gene expression in the peripheral blood mononuclear cells (PMBCs) of male patients with post-viral chronic fatigue (n=8) and male healthy control subjects (n=7). Patients and healthy subjects differed significantly in the level of expression of 366 genes. Analysis of the differentially expressed genes indicated functional implications in immune modulation, oxidative stress and apoptosis. Prototype biomarkers were identified on the basis of differential levels of gene expression and possible biological significance. Differential expression of key genes identified in this study offer an insight into the possible mechanism of chronic fatigue following infection. The representative biomarkers identified in this research appear promising as potential biomarkers for diagnosis and treatment.

Keywords: comparison between two groups of samples

overall_design: Two groups of patients: Chronic Syndrom Fatigue (CFS) patients (n=8) and control patients (n=7).

first: J

middle: W

last: Gow

first: S

first: P

last: Herzyk

first: C

last: Cannon

last: Behan

first: A

last: Chaudhuri


title: Expression profiling of HNF4-dependent transcripts in Drosophila larvae

status: Public on Jan 24 2009

city: Salt Lake City

first: Carl

last: Thummel

state: UT

department: Human Genetics

institute: University of Utah

summary:
We show that dHNF4 null mutant larvae are sensitive to starvation. Starved mutant larvae consume glycogen normally, but retain lipids in their midgut and fat body, and have increased levels of long chain fatty acids, suggesting that they are unable to efficiently mobilize stored fat for energy. HNF4 dependent transcripts were identified under fed and starved conditions. Microarray studies indicate reduced expression of genes that control lipid catabolism and beta-oxidation. Late second instar control and HNF4 mutant larvae were collected and fed or starved for 24 hours, and then RNA was isolated for analysis. Taken together, our results support a feed-forward model for dHNF4, in which fatty acids released from triglycerides activate the receptor, inducing enzymes that drive fatty acid oxidation for energy production.

Keywords: expression profiling, HNF4, mutants, Affymetrix, Drosophila 2.0, starvation, TAG, fatty acid, beta-oxidation

overall_design: Staged late second instar control larvae that were transheterozygous for precise excisions of the EP2449 and KG08976 P-elements, and dHNF4D33/dHNF4D17 mutant larvae, were either fed or starved for 24 hrs in a 55mm petri dish containing Whatman paper soaked with PBS. RNA was extracted from the apparently healthy animals that molted to the third instar using Trizol (Gibco) and purified on RNAeasy columns (Qiagen). All samples were prepared in triplicate to facilitate subsequent statistical analysis. Probe labeling, hybridization to Affymetrix GeneChip® Drosophila Genome 2.0 Arrays.

first: Laura

last: Palanker

first: Geanette

last: Lam

first: Jason

last: Tennessen


title: Chromatin structure marks cell-type and gender specific replication of the Drosophila genome

city: Vienna

first: Michaela

last: Schwaiger

address: Althanstr. 14

country: Austria

institute: University of Vienna

type: Genome binding/occupancy profiling by genome tiling array

summary:
Duplication of eukaryotic genomes during S phase is coordinated in space and time. In order to identify zones of initiation and cell-type as well as gender-specific plasticity of DNA replication, we profiled replication timing, histone acetylation and transcription throughout the Drosophila genome. We observed two waves of replication initiation with many distinct zones firing in early and multiple, less defined peaks at the end of S phase, suggesting that initiation becomes more promiscuous at the end of S phase.   A comparison of different cell types revealed widespread plasticity of replication timing on autosomes. Most occur in large regions but only half coincide with local differences in transcription. In contrast to confined autosomal differences, a global shift in replication timing occurs throughout the single male X chromosome. Unlike in females, the dosage compensated X chromosome replicates almost exclusively early. This difference occurs at sites which are not transcriptionally hyperactivated, but show increased acetylation of lysine 16 of histone H4. This suggests a transcription-independent, yet chromosome-wide process related to chromatin. Importantly, H4K16ac is also enriched at initiation zones as well as early replicating regions on autosomes during S phase. Together, our data reveal novel organizational principles of DNA replication of the Drosophila genome and imply chromatin structure as a determinant of replication timing locally and chromosome-wide.

Keywords: cell type comparison, chip-chip, replication timing

overall_design: RNA was isolated from Kc and Cl8 cells (5 replicates each) and hybridized to gene expression arrays. RNA was isolated from kc ane Cl8 cells (2 replciates each) and hybridised to tiling arrays. Early and late replicating DNA was isolated from Kc (5 replicates), Cl8 (2 replicates) and S2 (2 replicates) cells and hybridized to tiling arrays. Replicating DNA from four S phase fractions ( 2 replicates each) was isolated from Kc cells and hybridized to tiling arrays, with genomic DNA (2 replicates) from Kc cells as a control. Nascent DNA was isolated from Kc cells and hybridized to tiling arrays, with genomic DNA as a control (3 replicates each). ChIP-chip was perfomred for RNA Polymerase II in Kc (2 replicates) and Cl8 (3 replicates) cells. ChIP-chip was performed for H4K16ac in Kc  (3 replicates) and Cl8 (4 replicates) cells. ChIP-chip for H4K16ac was performed in early and late S phase Kc cells (2 replicates each).

first: Michael

last: Stadler

first: Oliver

last: Bell

first: Hubertus

last: Kohler

first: Edward

last: Oakeley

first: Dirk

last: Schübeler


title: Predicting Relapse in Favorable Histology Wilms Tumor Using Gene Expression

status: Public on Jan 30 2009

city: Chicago

first: Elizabeth

last: Perlman

state: IL

address: 2373 North Lincoln Avenue Room A.204

department: Department of Pathology and Laboratory Medicine

institute: Children's Memorial Research Center

summary:
The gene expression patterns of favorable histology Wilms tumors (FHWT) that relapsed were compared with those that did not relapse using oligonucleotide arrays

Description:  250 FHWT of all stages enriched for relapses treated on National Wilms Tumor Study 5 passed quality parameters and were suitable for analysis using oligonucleotide arrays. Relapse risk stratification utilized Support Vector Machine; two and ten fold cross-validation was applied. The number of genes associated with relapse was less than that predicted by chance alone for 106 patients (32 relapses) with stages I and II FHWT and no further analyses were performed.  This number was greater than expected by chance for 76 local stage III patients.  Cross validation including an additional 68 local stage III patients (total 144 patients, 53 relapses) demonstrated that classifiers for relapse composed of 50 genes were associated with a median sensitivity of 47%, specificity 70%, and total error rate of 38%.  Analysis of genes differentially expressed in relapse patients revealed apoptosis, Wnt signaling, IGF pathway, and epigenetic modification to be mechanisms important in relapse. Potential therapeutic targets include FRAP/MTOR and CD40.
Keywords: Classification by microarray analysis

overall_design:
144 stage 3 FHWT, with fifty-three relapses (cases) and ninety-one non-relapses (controls) with a minimum of three years follow-up, included all relapses and a 30% random selection of non-relapses for which frozen tumor tissue was available who passed all quality control parameters.  The NWTS-5 protocol was approved by the review boards of institutions that registered patients. Histological diagnosis and local stage were confirmed by central review.

Quality control steps taken:
1. Samples were snap frozen immediately following surgery and were mailed on dry ice to the Tumor Bank and retained at -80C.
2. Frozen sections were evaluated histologically and tumors with less than 80% viable tumor cellularity were excluded.
3. Array images were assessed by eye to confirm scanner alignment and the absence of significant bubbles or scratches.
4. Samples for which the 3'/5' ratios for GAPDH were greater than 3.2 were excluded.
5. The BioB spike controls were confirmed as present; BioC, BioD and cre were confirmed as increasing intensity.
6. When scaled to a target intensity of 2500, scaling factors were between 12 and 53; background levels were 34 – 115: Q values were 1.3 – 3.7 and mean intensities were within acceptable limits.
7. The range of percent present calls was from 38% to 52%.
8. Verification of gene expression was performed utilizing quantitative RT-PCR for five genes.

Statistical Analysis:  Positional-dependent-nearest-neighbor model (PDNN) software was used to translate the scanned images into expression analysis files and to normalize the data across all arrays (http://odin.mdacc.tmc.edu/~zhangli/PerfectMatch/).  Genes with maximum expression less than a log scale of 6 across all tumors and Affymetrix control genes were excluded, resulting in 20,931 probe sets for analysis.  Support Vector Machine (SVM) as developed by Chang and Lin (http://www.csie.ntu.edu.tw/~cjlin/libsvm) and implemented in an R software package, e1071 was chosen for relapse risk stratification using the p-value of the t-test comparison between case and control to select the genes.  Using all 144 tumors, 109 genes were identified with p-value <0.001 and are provided in the data table labelled "t-test comparison between case and control" (ie, Supplemental table 2, in related publication)."    Two and ten fold cross validations were utilized to investigate the ability of classifiers established in randomly selected  training set to predict relapse in an independent test set comprised of the remaining tumors.  For two-fold cross validation, the dataset was randomly divided 500 times into training and corresponding test sets of equal size, each including half the patients who relapsed. A classifier for relapse was identified for each training set and used to assign tumors in the corresponding test set to low and high risk categories.  The training and test sets were then swapped. The number of top K genes in each classifier evaluated ranged from 1-150. Therefore, a total of 150,000 different classifiers were developed, one for each value of K from 1-150, for each of the 1,000 (500*2) training sets. For ten-fold cross validation the dataset was randomly divided 500 times into ten groups of approximately equal size.  Each group included approximately the same number of relapses. For each such group, a classifier was built with the remaining 9/10 of the samples and then used to categorize tumors in the group as low or high risk; the process was repeated until all tumor samples were categorized as low or high risk. For all the cross validation procedures, to avoid gene-selection bias, classifiers were completely rebuilt in each cross validation iteration.

Keywords: Tumor cell type comparison, oligonucleotide arrays, favorable histology Wilms tumor, pediatric renal tumors

first: Chiang

last: Huang


title: The Arabidopsis BRAHMA Chromatin Remodelling ATPase Is Involved in Direct Repression of Embryonic Traits in Leaves

status: Public on Jan 21 2009

city: London

first: Mohan

last: babu

state: Ontario

address: 1391 Sandford

department: Virology

institute: AAFC

summary:
Synthesis and accumulation of seed storage proteins (SSPs) is an important aspect of the seed maturation program. Genes encoding SSPs are specifically and highly expressed in the seed during maturation. However, the mechanisms that repress the expression of these genes in leaf tissue are not well understood. To gain insight into the repression mechanisms, we have performed a transgenic screening for mutants that express SSPs in leaves. Here we show that mutations of BRAHMA (BRM), a SNF2 chromatin remodelling ATPase, cause the ectopic expression of a subset of SSPs and other embryogenesis related genes in leaf tissue. Consistent with the notion that such SNF2-like ATPases form protein complexes in vivo, we observed similar phenotypes for mutations of AtSWI3C, a BRM interacting partner, and BSH, a SNF5 homolog and essential SWI/SNF subunit. Further, we present chromatin immunoprecipitation evidence that BRM is recruited to the promoters of a number of embryogenesis genes including the 2S genes, which are expressed/elevated in brm leaves. Consistent with its role in nucleosome remodelling, BRM appears to control the chromatin structure of the At2S2 promoter. These results show that a BRM-containing chromatin remodelling ATPase complex is involved in the direct repression of SSPs in leaf tissue.

A matrix comprising the signal intensity value of each gene per replicate hybridization and the averaged data of each gene generated from three replicate hybridizations of the wild type and mutant samples, respectively, is linked below as a supplementary file.

overall_design: Total RNA was isolated from three independent biological replicates of essp3 mutant and Wild type (ProβCG:GUS) respectively. Three ATH1 chips were used for the mutant and three for the wild type.

first: Xurong

last: Tang

first: Anfu

last: Hou

last: Babu

first: Vi

last: Nguyen

first: Lidia

last: Hurtado

first: Qing

last: Lu

first: Jose

last: Reyes

first: Aiming

last: Wang

first: Wilfred

last: Keller

last: Tsang

first: Yuhai

last: Cui


title: Gene expression profiling of trophoblast cells

status: Public on Jan 22 2009

first: Martin

last: Bilban

institute: Medical University of Vienna

summary: Invasion of cytotrophoblasts into uterine tissues is essential for placental development. To identify molecules regulating trophoblast invasion, mRNA signatures of purified villous (CTB, poor invasiveness) and extravillous (EVT, high invasiveness) trophoblasts isolated from first trimester human placentae and villous explant cultures, respectively, were compared using GeneChip analyses yielding 991 invasion/migration related transcripts. Several genes involved in physiological and pathologic cell invasion, including ADAM-12,-19,-28 as well as Spondin-2, were upregulated in EVT. Pathway prediction analyses identified several functional modules associated with either the invasive or the non-invasive trophoblast phenotype. One of the genes which were downregulated in the invasive mRNA pool, heme oxygenase-1 (HO-1), was selected for functional analyses. Real-time PCR analyses, Western blottting, and immunofluorescene of first trimester placentae and differentiating villous explant cultures demonstrated downregulation of HO-1 in invasive EVT as compared to CTB. Modulation of HO-1 expression in loss-of as well as gain-of function cell models (BeWo and HTR8/SVneo, respectively) demonstrated an inverse relationship of HO-1 expression with trophoblast migration in transwell and wound healing assays. Importantly, HO-1 expression led to an increase in protein levels and activity of the nuclear hormone receptor PPARgamma. Pharmacological inhibition of PPARgamma abrogated the inhibitory effects of HO-1 on trophoblast migration. Collectively, our results demonstrate that gene expression profiling of EVT and CTB can be used to unravel novel regulators of cell invasion. Accordingly, we identify heme oxygenase-1 as a negative regulator of trophoblast motility acting via upregulation of PPARgamma.

overall_design:
To identify genes potentially regulating cell invasion trophoblast cells of early human gestation with distinct invasive properties were profiled.
Distinct gene expression signatures of highly invasive EVT (n = 6) and poorly invasive CTB (n = 5) of different first trimester placentae using Affymetrix U133A GeneChips interrogating >20,000 genes were determined.

first: Gernot

last: Desoye

last: Knoefler


title: Gene expression by human splenic B-cell subsets

status: Public on Dec 31 2008

city: Darlinghurst

first: Stuart

last: Tangye

state: NSW

address: 384 Victoria St

department: Immunology

institute: Garvan Institute of Medical Research

summary:
Enhanced secondary Ab responses are a vital component of adaptive immunity, yet little is understood about the intrinsic and extrinsic regulators of naïve and memory B cells that results in differences in their responses to Ag. Microarray analysis, together with surface and intracellular phenotyping, revealed that memory B cells have increased expression of members of the TNF receptor, SLAM, B7 and Bcl2 families, as well as the TLR-related molecule CD180 (RP105). Accordingly, memory B cells exhibited enhanced survival, proliferation and Ig secretion, as well as entered division more rapidly than naïve B cells in response to both T-dependent and T-independent stimuli. Furthermore, both IgM and isotype switched memory B cells, but not naïve B cells, co-stimulated CD4+ T cells in vitro through a mechanism dependent on their constitutive expression of CD80 and CD86. This study demonstrates that upregulation of genes involved in activation, co-stimulation and survival provides memory B cells with a unique ability to produce enhanced immune responses and contributes to the maintenance of the memory B cell pool.

Keywords: cell type comparison

overall_design: Four subsets of human splenic B cells (naïve, IgM-memory, Ig isotype switched memory and plasma cells); sort-purified and analysed immediately ex vivo; performed in duplicate.

first: Kim

middle: L

last: Good

middle: G


title: Global gene expression of Arabidopsis lines with altered ANAC102 expression under normal and low oxygen conditions

status: Public on Jan 16 2009

city: Acton

first: Jed

last: Christianson

state: ACT

address: Clunies Ross Road

institute: CSIRO Plant Industry

summary:
Global gene expression was compared between Arabidopsis lines with altered expression of ANAC102 (over-expressed and knocked-out) and wild-type. ANAC102 is a putative NAC domain transcription factor. Gene expression was compared between an ANAC102 over-expressing line and parental ecotype C24 under ambient atmosphere to determine which genes ANAC102 is capable of regulating. Gene expression was also compared between three week old plants of an ANAC102 knock-out line and parental ecotype Col-0 under 0.1% Oxygen and ambient atmosphere conditions to determine which genes may require ANAC102 for appropriate expression under these conditions. Gene expression was also compared between imbibed seeds of an ANAC102 knock-out line and parental ecotype Col-0 following a 0.1% Oxygen treatment.

Keywords: Genetic modification

overall_design: Three related experiments were performed. In the first two, three week old Arabidopsis seedlings (4-6 leaves) were used. In the first experiment one line of ANAC102 over-expressing plants were compared to ecotype C24 (the parental line used in the creation of the ANAC102 over-expressing line). Two biological replicates were used for each line and for each biological replicate, five Arabidopsis seedlings grown on the same petri dish were bulked. RNA was extracted from whole plants. In the second experiment, one line of plants bearing a T-DNA insertion in the second exon of the ANAC102 gene was compared to ecotype Col-0 (the parental line used for the T-DNA mutagenesis) both without treatment and following treatment with 0.1% Oxygen for four hours. Two biological replicates were used for each line and for each biological replicate, five Arabidopsis seedlings grown on the same petri dish were bulked. RNA was extracted from root tissue only. In the third experiment imbibed seeds of one Arabiopsis line bearing a T-DNA insertion in the second exon of the ANAC102 gene was compared to seeds of ecotype Col-0 (the parental line used for the T-DNA mutagenesis) following treatment with 0.1% Oxygen for six days. Three biological replicates were used for each line and for each biological replicate, ~5mg (pre-imbibition) of seed were bulked. RNA was extracted from whole seed.

first: Iain

last: Wilson

first: Danny

last: Llewellyn

last: Dennis


title: Comparison between human normal and glaucomatous lamina cribrosa cells

status: Public on Jan 06 2009

city: Dublin

first: Ruaidhri

middle: Patrick

last: Kirwan

state: Leinster

address: Belfield

department: Conway Institute of Biomolecular and Biomedical Research & School of Medicine and Medical Science

country: Ireland

institute: University College Dublin

summary: Purpose: Marked extracellular matrix (ECM) remodeling occurs in the human optic nerve head in primary open angle glaucoma (POAG). The glial fibrillary acid protein (GFAP) negative lamina cribrosa cell may play an important role in this remodeling process. The authors report the first study of global and ECM-focused gene transcription differentials between GFAP-negative negative lamina cribrosa (LC) cells from normal and POAG human donors.  Methods: GFAP-negative LC cell lines were generated from the optic nerve tissue of three normal (n=3) and three POAG (n=3) human donors. Using Affymetrix U133A arrays the transcriptional profile between the normal and diseased groups were compared. Bioinformatic analysis was carried out using robust multichip average (RMA Express) and EASE/David.   Real time TaqMan PCR and immunohistochemistry analyses were performed to validate the microarray data. Results: 285 genes were up regulated by greater than 1.5 fold and 413 were down regulated by greater than 1.5 fold in the POAG LC cells versus normal controls. Upregulated genes in POAG LC cells included, SPARC, periostin, thrombospondin, CRTL-1, CTGF and collagen types I, III, V and VIII.   Downregulated ECM genes in POAG included MMP-1, fibulin, decorin and tenacsin XB.   All TaqMan PCR validation assays were significant (*p<0.05) and consistent with the array data. Immunohistochemistry of one target (periostin) confirmed its differential expression at the protein level in POAG optic nerve head tissue compared with non-glaucomatous controls. Functional annotation and over-representation analysis identified ECM genes as a statistically over-represented class of genes in POAG LC cells compared with normal LC cells. Conclusions: This study reports for the first time that POAG LC cells in-vitro demonstrate up regulated ECM and pro-fibrotic gene expression compared with normal LC cells. This may be a pathological characteristic of this cell type in POAG in-vivo. We believe that the LC cell may be a pivotal regulator of optic nerve head ECM remodeling and an attractive target for future therapeutic strategies in POAG.

overall_design: 8 human donors used (four normal and four glaucoma). Pooled RNA. N=2 biological replication of microarray analysis.

middle: P

first: Robert

last: Wordinger

first: Abbot

middle: F

last: Clark

first: Colm

last: O'Brien


title: Transcript profiling of Lymphangioleiomyomatosis (LAM) nodules in Human Patients

city: Bethesda

first: Eric

last: Billings

state: MD

address: 10 Center Drive

institute: NHLBI

summary:
Lymphangioleiomyomatosis (LAM) is characterized by cystic lung destruction caused by smooth, muscle-like LAM cells which have mutations in the tumor suppressor genes Tuberous Sclerosis Complex (TSC) 1 or 2, and the capacity to metastasize. Since chemokines and their receptors function in chemotaxis of metastatic cells, we hypothesized that LAM cells may be recruited by chemokine(s) in the lung.  Quantification of 25 chemokines in bronchoalveolar lavage fluid from LAM patients and healthy volunteers revealed that concentrations of MCP-1/CCL2, GROa/CXCL1 and ENA-78/CXCL5 were significantly higher in samples from LAM patients than healthy volunteers.  In this transcript analysis, expression of chemokine and chemokine receptor mRNA in LAM cells differed from those in melanoma and smooth muscle cells.  Subsequent immunohistochemistry of lung sections from 30 LAM patients confirmed protein expression of chemokines and these receptors varied among LAM patient and differed from that seen in breast cancer and melanoma cells. .   In vitro, MCP-1/CCL2 induced selective migration of cells showing loss of heterozygosity of TSC2 from a heterogeneous populations of cells grown from explanted LAM lungs. In addition, the frequencies of single-nucleotide polymorphisms in the MCP-1 gene promoter region differed significantly in LAM patients and healthy volunteers (p=0.018), and one polymorphism was associated significantly more frequently with the decline of lung function. These observations are consistent with the notion that chemokines such as MCP-1 may serve to specify site of LAM cell metastasis.

Keywords: Human patient sample comparison with cell lines

overall_design: The study is of case/control design with biological replication.  Biopsies of nodules from 14 LAM patients (cases) are compared with cultured cell lines (controls) with similar properties.

first: Yi

last: Zhang

first: Gustavo

last: Pacheco-Rodriguez


title: Expression of mRNAs Regulating Synaptic Function and Neuroplasticity in Incipient AD

status: Public on Jan 10 2009

city: Los Angeles

first: Celia

last: Williams

state: CA

address: 2011 Zonal Ave

department: Pathology

institute: Keck School of Medicine, USC

summary: In Alzheimer’s disease (AD), early deficits in learning and memory are a consequence of synaptic modification which are likely induced by toxic beta-amyloid oligomers (oAβ).  To identify molecular targets downstream of oAβ binding  we prepared synaptoneurosomes from frontal cortex of control and IAD patients, and isolated mRNAs for comparison of gene expression.  This approach elevated synaptic mRNAs above the threshold necessary for expression changes to be discriminated and also reduced other cellular mRNAs.  In patients with minimal cognitive impairment (MCI) termed incipient AD (IAD) global measures of cognition declined with increasing levels of dimeric Aβ (dAβ).  These patients also showed increased expression of neuroplasticity related genes, many encoding 3' UTR  consensus sequences that regulate local translation in the synapse.  One such gene, GluR2, displayed elevated mRNA and protein expression in IAD.  Other neurotransmitter-related genes were also upregulated.  Overexpressed genes may induce compensatory as well as negative effects on cognition and provide targets for intervention to moderate the response to dAβ.

overall_design:
Patient information
CNS tissues were obtained from the USC Alzheimer’s Disease Research Center (ADRC) Neuropathology Core, NIA AG05142. Clinical information, including neurological examination, neuropsychological testing such as cognitive assessment, family history, and medications were provided by the USC ADRC Clinical Core.  The groups did not differ significantly in age or years of education.  Medication histories indicated one IAD patient had used a selective serotonin uptake inhibitor.  Post-mortem interval (PMI) ranged from 2 to 9 hours (mean 5.1 hrs.). The study included 8 normal controls and 6 patients with Incipient Alzheimer's Disease.
To augment studies  comparing homogenates of  control to AD brains, we used synaptoneurosome preparations  to enrich for synaptic mRNAs.  Synaptoneurosomes were prepared from the prefrontal cortices of control and IAD patients by a simple, rapid, but gentle method using sequential mesh screens.

first: Ruty

last: Mehrian Shai

first: Yongchun

last: Wu

first: Ya-Hsuan

last: Hsu

first: Traci

last: Sitzer

first: Bryan

last: Spann

first: Carol

last: McCleary

last: Mo

last: Miller


title: Effects of Sxl on gene expression in the male head of Drosophila

status: Public on Dec 30 2008

first: Sha

last: Sun

address: 16 Barker Hall

department: Molecular and Cell Biology

institute: University of California, Berkeley

summary:
Sex-lethal (Sxl), a feminizing switch gene in Drosophila, also generates male-specific proteins and non-sex-specific mRNAs, despite the fact that deleting the gene from males causes no obvious phenotypic abnormalities.  We specifically investigate how Sxl may affect gene expression in the male head, where SXL isoforms have been detected.  The information may help us understand how Sxl acquired its functional roles in sex determination during evolution in Drosophila
Keywords: Comparative expression, genetic modification

overall_design:
Sxlfp7BO is a small deletion that removes the entire Sxl locus. The mutant Sxlfp7BO males were backcrossed to the wild-type Oregon-R flies for more than six generations to homogenize the genetic background. Two flanking phenotypic markers, cm and ct, on the proximal and distal sides of Sxlfp7BO, were used to trace the mutant allele during the backcross.
[cm Sxlfp7BO ct] and WT males were brothers from a single-pair cross: [cm Sxlfp7BO ct]/+  X   [cm Sxlfp7BO ct]/Y.
Male flies of 2-5 day old were decapitated on an ice pack; the heads were transferred directly into TRIzol before RNA extraction.  Three biological replicates were performed for each male type.

first: Thomas

last: Cline


title: Early responsive genes in human amnion epithelial FL cells induced by N-methyl-N’-nitro-N-nitrosoguanidine

status: Public on Jan 01 2009

city: Hangzhou

first: Hongjuan

last: Li

state: Zhejiang

address: 388 Yuhangtang Road

institute: Zhejiang University School of Medicine

summary:
The alkylating agent N-methyl-N’-nitro-N-nitrosoguanidine (MNNG) induces cellular DNA damages and other comprehensive alterations that lead to chromosomal aberrations, mutations, tumor initiations, and cell death. However, the molecular mechanism of MNNG-induced cellular stress remains unclear.We have genome-wide analyzed early transcriptional responses of human FL amnion epithelial cells after exposure to three relatively low doses of MNNG (0.2, 1.0, and 10.0µM),and differential gene expression profiles were obtained 4 h after exposure using oligonucleotide microarrays followed by validation with quantitative real-time RT-PCR.
The results demonstrate that the MNNG-responsive genes are involved in multiple cellular biological processes including transcription regulation, signal transduction, cell cycle regulation, cytoskeleton organization, protein synthesis, immune response, metabolism, etc. The possible roles of these genes and their related pathways in MNNG-induced cellular responses were discussed. This study helps to draw the whole picture how cells respond to environmental chemical exposure via transcriptional regulation.
Keywords: Dose response

overall_design: Human amnion epithelial FL cells were exposed to vehicle control (dimethyl sulfoxide) and increasing doses(0.2, 1.0, and 10.0µM)of N-methyl-N’-nitro-N-nitrosoguanidine(MNNG), respectively. The transcriptomes of the three treatments were compared to that of the control, respectively, to test the hypothesis that a characterized differential expression profile would be generated by exposure to various doses of this genotoxic agent.

first: Jimin

last: Shao

first: Xiangyun

last: Lu

first: Zhihua

last: Gao

first: Yingnian

last: Yu


title: SL2_expression

city: Kansas City

first: Marco

last: Blanchette

state: MO

institute: Stowers Institute for Medical Research

summary:
In this work, we have studied four closely related members of the five known Drosophila hnRNP A/B family, hrp36, hrp38, hrp40, and hrp48. Using splicing-sensitive microarrays, between 200 and 300 genes were detected as specifically regulated by each individual hrp protein following down regulation of the protein expression level by RNAi-mediated knock-down.

Keywords: Genetic modification

overall_design: This set of arrays evaluate the global expression level of all genes in wild-type SL2 cells.