title: Comparing active and repressed expression states of genes controlled by the Polycomb/Trithorax group proteins

city: Basel

first: Christian

last: Beisel

address: Mattenstrasse 26

department: Biosystems Science and Engineering

country: Switzerland

institute: ETH Zurich

summary:
Drosophila Polycomb group (PcG) and Trithorax group (TrxG) proteins are responsible for the maintenance of stable transcription patterns of many developmental regulators, such as the homeotic genes. We have used a ChIP-on- Chip approach to map the distribution of several PcG/TrxG proteins as well as histone modifications across the two homeotic complexes ANT-C and BX-C. Our data indicate the colocalization of the Polycomb repressive complex 1 (PRC1) with Trx and the DNA binding protein Pleiohomeotic (Pho) at discrete sequence elements as well as significant chromatin assembly differences in active and inactive regions. Trx binds to the promoters of active genes and noncoding transcripts. Most strikingly, in the active state Pho covers extended chromatin domains over many kilobases. The histone modifications investigated, seem to be mutually exclusive, with histone H3 trimethylated at lysine 27 covering inactive chromatin domains whereas hyperacetylated histone H4 is present only at active genes.
Keywords: ChIP-chip, cell line comparison

overall_design:
Currently it is unclear how the interplay between DNA elements, histones and their modifications and the PcG/TrxG-chromatin-associated proteins results in a stable gene expression state which is heritable through DNA replication and mitosis. A key to the understanding of the molecular mechanisms is to uncover the chromatin sites of action of the PcG/TrxG proteins and to correlate their binding patterns with the presence of histone modifications as well as with the expression level of their target genes.
Here, we generated a DNA tiling array that comprises the BX-C and the ANT-C as a series of overlapping 1 kb PCR fragments. Based on this tiling array we investigated the distribution of three core components of the PRC1 (PC, PH and PSC), the DNA binding protein PHO, TRX, hyperacetylated H4 and H3K27me3 in two different Drosophila embryonic cell lines that show different expression states for the AbdB and Dfd genes. The AbdB domain and the Dfd gene are highly active in SF4 and in Kc cells, respectively, whereas the other HOX genes are silent.
Two versions of the tiling array were used in the course of this study. Version 3 contained 10 and Version 4 contained 8 spotted replicates of each PCR fragment. With Version 3 arrays either three or four independent chromatin samples were taken for each antibody with dye swap, i.e. six or eight arrays. Normalization was based on spike-ins and all 10 spot replicates were averaged afterwards. The biological effect was estimated with a regularized linear model which contained a dye effect and treated the technical replicates like a random effect. With Version 4 arrays either four or six biological replicates were used where in every other hybridization Cy3 and Cy5 were interchanged between ChIP and input sample (four or six arrays total). Spike-ins served to determine the normalization parameters which were subsequently used to normalize all intensity measurements on every microarray. All 8 spot replicates were averaged. The statistical model applied to estimate the biological effect included a dye effect. Multiple testing correction was applied and false discovery rates (FDR) (as well as fold-changes) were reported and used for significance thresholding. All analyses were performed with R (www.rproject. org) and Bioconducter (www.bioconductor.org). The package arrayMagic was used for processing, the package vsn for normalisation and the package limma for statistical testing (Buness, A., Huber, W., Steiner, K., Sultmann, H., and Poustka, A. (2005). arrayMagic: two-colour cDNA microarray quality control and preprocessing. Bioinformatics 21, 554-556; Huber, W., von Heydebreck, A., Sultmann, H., Poustka, A., and Vingron, M. (2002). Variance stabilization applied to microarray data calibration and to the quantification of differential expression. Bioinformatics 18 Suppl 1, S96-104; Smyth, G. K., Michaud, J., and Scott, H. S. (2005). Use of within-array replicate spots for assessing differential expression in microarray experiments. Bioinformatics 21, 2067-2075).

first: Andreas

last: Buness

first: Britta

last: Koch

first: Stefan

middle: A

last: Haas

first: Marc

last: Hild

first: Ian

last: Roustan-Espinosa

first: Sabine

last: Schmitt

first: Renato

last: Paro


title: BW25113 ymgB and W/T in LBglu 24h biofilm cells

status: Public on Sep 21 2007

city: College Station

first: Jintae

last: Lee

state: TX

department: Chemical Engineering

country: USA

institute: Texas A&M University

summary:
For the microarray experiments, 10 g glass wool (Corning Glass Works, Corning, N.Y.) were used to form biofilms (30) in 250 mL in 1 L Erlenmeyer shake flasks which were inoculated with overnight cultures diluted that were 1:167.  The cells were shaken at 250 rpm and 37°C for 24 hours to form biofilms on the glass wool, and RNA was isolated from the suspension cells and the biofilm.
Keywords: E. coli biofilm

overall_design:
Strain: E. coli BW25113 wild type and ymgB mutant
Medium: LBglu
Cells: biofilm cells on glass wool
Time: 24h

first: Thomas

middle: K

last: Wood

first: Wolfgang

last: Peti

first: Rodolfo

middle: G

last: Contreras

first: Xue-Song

last: Zhang


title: Calcium-induced transcript variation in neuronal cells analyzed with exon microarrays.

status: Public on Sep 24 2007

city: PROVIDENCE

first: Alexander

last: Brodsky

state: RI

address: 593 Eddy St.

department: MCB

institute: Rhode Island Hospital/Brown University

summary:
Background:
Neuronal cells respond to changes in intracellular Ca2+ ([Ca2+]i) by affecting both the abundance and the architecture of specific mRNAs.  While Ca2+-induced transcription and transcript variation have both been recognized as important sources of gene regulation, the interplay of these two phenomena has not been evaluated together on a genome scale.
Results:
Here, we show that exon-centric microarrays can be used to resolve the Ca2+-modulated gene expression response into transcript-level and exon-level regulation. Global assessments of affected transcripts reveal modulation within distinct functional gene categories. We find that transcripts containing Ca2+-modulated exons show enrichment for Ca2+-ion binding, calmodulin-binding, plasma membrane-associated, and metabolic proteins. Additionally, we uncover instances of regulated exon use in potassium channels, neuroendocrine secretory proteins, and in metabolic enzymes, and demonstrate that regulated changes in exon expression give rise to distinct transcript isoforms.
Conclusions:
Our findings connect extracellular stimuli to specific exon behavior and suggest that changes in transcript and exon abundance are reflective of a coordinated gene expression response to elevated [Ca2+]i. The technology we describe here lends itself readily to the resolution of stimulus-induced gene expression at both the transcript and exon levels.
Keywords: Ca2+-modulated exons and transcripts

overall_design: Biological triplicate RNA samples are from untreated (ZERO) Human neuroblastoma IMR-32 cells (ATCC) and IMR-32 cells treated with 50 mM KCl for 0.5, 1.5, 3, 6, 12, or 24 hours, and exposed to 5µM thapsigargin (Tocris) for 24 hours (THAP).

first: Adrienne

middle: E

last: McKee

first: Nicola

last: Neretti

first: Luis

last: Carvalho

first: Richard

last: Park

first: Edward

last: Fox

middle: S

first: Pamela

last: Silver


title: Gene expression profiling of human adipose tissue of obese subjects in relation to bariatric surgery

status: Public on Sep 19 2007

city: Paris

first: Corneliu

last: Henegar

state: Ile de France

address: 15 Rue de l'Ecole de Medecine

country: France

institute: Institut national de la santé et de la recherche médicale

summary:
In human obesity, the stroma vascular fraction (SVF) of white adipose tissue (WAT) is enriched in macrophages. These cells may contribute to low-grade inflammation and to its metabolic complications. Little is known about the effect of weight loss on macrophages and genes involved in macrophage attraction. We examined subcutaneous WAT (scWAT) of 7 lean and 17 morbidly obese subjects before and 3 months after bypass surgery. Immunomorphological changes of the number of scWAT-infiltrating macrophages were evaluated, along with concomitant changes in expression of SVF-overexpressed genes. The number of scWAT-infiltrating macrophages before surgery was higher in obese than in lean subjects (HAM56+/CD68+; 22.6 +/- 4.3 vs. 1.4 +/- 0.6%, P < 0.001). Typical "crowns" of macrophages were observed around adipocytes. Drastic weight loss resulted in a significant decrease in macrophage number (-11.63 +/- 2.3%, P < 0.001), and remaining macrophages stained positive for the anti-inflammatory protein interleukin 10. Genes involved in macrophage attraction (monocyte chemotactic protein [MCP]-1, plasminogen activator urokinase receptor [PLAUR], and colony-stimulating factor [CSF]-3) and hypoxia (hypoxia-inducible factor-1alpha [HIF-1alpha]), expression of which increases in obesity and decreases after surgery, were predominantly expressed in the SVF. We show that improvement of the inflammatory profile after weight loss is related to a reduced number of macrophages in scWAT. MCP-1, PLAUR, CSF-3, and HIF-1alpha may play roles in the attraction of macrophages in scWAT.
Keywords: disease state analysis

overall_design:
Seventeen Caucasian morbidly obese women undergoing laparoscopic Roux-en-Y bypass were enrolled at Hôtel Dieu Hospital, Paris, France.  ScWAT biopsies were performed by a surgeon after local anesthesia (1% xylocaine) of the periumbilical area. A portion of each scWAT biopsy was  immediately frozen for RNA extraction and analysis. To study the effects of weight loss on scWAT gene expression, we processed 10 high-quality RNAs from scWAT biopsies collected before/after bypass by microarray technique.
Total RNA was prepared using the RNeasy total RNA Mini kit (Qiagen). RNA concentration and integrity were assessed using the Agilent 2100 Bioanalyzer (Agilent Technologies, Massy, France). For microarray experiments, 1 µg of total RNA from each total RNA sample preparation was amplified by MessageAmp RNA Kit (Ambion, Austin, TX, USA) and 3 µg of amplified RNA was labeled with cyanin dyes (Cy) using the CyScribe First-Strand cDNA labeling kit (Amersham Biosciences, Orsay, France). (available at http://cmgm.stanford.edu/pbrown/protocols/index). aRNA extracted from each scWAT sample before surgery was labeled with Cy3 dye, while the aRNA from each scWAT sample obtained 3 months after surgery was labeled with Cy5 dye. Several quality cross-checks (for total RNA quality, aRNA quality, dye incorporation efficiency, etc.) and microarray "dye swap" experiments were performed.
The labeled cDNA mixtures were hybridized according to the protocol described at http://cmgm.stanford.edu/pbrown/protocols/index

first: Karine

last: Clement

first: Raffaella

last: Cancello

first: Veronique

last: Pelloux

first: Christine

last: Rouault

first: Nathalie

last: Viguerie


title: Hepatectomy sham surgery 1h

status: Public on Sep 26 2007

city: Philadelphia

first: Egle

last: Juskeviciute

state: PA

address: 1020 Locust st. JAH 527

department: Pathology

institute: Thomas Jefferson University

summary:
4 Adult male Sprague-Dawley rats (275-350 g) were anesthetized and subjected to hepatectomy sham surgery (abdominal cavity was opened, liver   was handled, but no tissue resection was made). 1 hour after the surgery rats were killed and liver samples were harvested.
This study was conducted to analyzes the effects of surgical stress on gene expression levels in rat liver. It provides additional data to 1-6 h partial hepatectomy study (Series GSE7415).
Keywords: 1h hepatectomy sham surgery

sample_organism:
Rattus norvegicus
synthetic construct

overall_design: Reference Design with 4 biological repeats


title: AtNF-YB1 and CBF4 Overexpression Study in Arabidopsis

city: Hayward

first: Don

middle: R

last: Maszle

state: CA

address: 3935 Point Eden Way

institute: Mendel Biotechnology, Inc.

summary:
The baseline transcriptional profiles of Arabidopsis seedling shoots overexpressing CBF4 or AtNF-YB1 were compared with wild-type controls.
Keywords: overexpression_study

overall_design: Two independent homozygous 35S::AtNF-YB1 lines (#3 and #6), two independent 35S::CBF4 lines (#10 and #19, 23), and wild type Col-0 Arabidopsis were grown axenically on vertically-oriented tissue culture plates under 24 h light on standard MS media. Seedling shoot tissue (separated from root tissue) was harvested at 10 am after 14 days of growth and processed according to standard protocols for Affymetrix GeneChips. Two biological replicates for each transgenic line or three replicates for the single wild-type lines, each containing 20-24 seedlings, were hybridized to microarrays.


title: Comparative Genomics of Bordetella pertussis Reveals Progressive Gene Loss in Finnish Strains

status: Public on Sep 20 2007

city: Turku

first: Teemu

last: Kallonen

address: Kiinamyllynkatu 13

department: Derpartment of Infectious Disease Surveillance and Control

country: Finland

institute: National Institute for Health and Welfare

summary:
Background
Bordetella pertussis is a Gram-negative bacterium that infects the human respiratory tract and causes pertussis or whooping cough. The disease has resurged in many countries including Finland where the whole-cell pertussis vaccine has been used for more than 50 years. Antigenic divergence has been observed between vaccine strains and clinical isolates in Finland. To better understand genome evolution in B. pertussis circulating in the immunized population, we developed an oligonucleotide-based microarray for comparative genomic analysis of Finnish strains isolated during the period of 50 years.

Methodology/Principal Findings
The microarray consisted of 3,582 oligonucleotides (70-mer) and covered 94% of the genome of Tohama I, the strain of which the genome has been sequenced [21]. Twenty isolates from 1953 to 2004 were studied together with two Finnish vaccine strains and two international reference strains. The isolates were selected according to their characteristics, e.g. the year and place of isolation and pulsed-field gel electrophoresis profiles. Genomic DNA of the tested strains, along with reference DNA of Tohama I strain, was labelled and hybridized. The absence of genes as established with microarrays, was confirmed by PCR. Compared to the Tohama I strain, Finnish isolates lost 7 (8.6 kb) to 49 (55.3 kb) genes, clustered in one to four distinct loci. The number of lost genes increased with time, and one third of lost genes had functions related to ion transport, metabolism, or energy production and conversion. All four loci of lost genes were flanked by the insertion sequence element IS481.
Conclusion/Significance
Our results showed that the progressive gene loss occurred in Finnish B. pertussis strains isolated during a period of 50 years and confirmed that B. pertussis is dynamic and is continuously evolving, suggesting that the bacterium may use gene loss as one strategy to adapt to highly immunized populations.
Keywords: comparetive genomic hybridisation

overall_design: All together 26 samples were analysed. Tohama I was always the reference. GSM200718 and GSM200719 are replicates. GSM200712 and GSM200714 are dye swaps.

first: Eriikka

last: Heikkinen

first: Lilli

last: Saarinen

first: Rolf

last: Sara

first: Juhani

middle: T

last: Soini

first: Jussi

last: Mertsola

first: Qiushui

last: He


title: Identification of dASF/SF2 potential mRNA targets

status: Public on Sep 18 2007

city: Montpellier

first: johann

last: Soret

address: 1919, route de Mende

department: Institut de Genetique Moleculaire

institute: CNRS

summary:
Deciphering the role of alternative splicing in developmental processes relies on the identification of key genes whose expression is controlled by splicing regulators throughout growth of a whole organism. Targeting expression of five SR proteins in the developing eye of Drosophila allowed us to show that these splicing factors induce various phenotypic alterations concerning eye organogenesis and viability. Although both dASF/SF2 and B52 caused defects in ommatidia structure, only B52 impairs normal photoreceptor axons projection and neurogenesis in visual ganglia. Consistently, microarray analyses revealed that many of the B52 targets are involved in brain organogenesis and we show that their splicing profile is altered both in B52 loss and gain of function. Conversely, a large proportion of dASF/SF2 targets are involved in eye development. This differential effect argues that SR proteins confer accuracy to developmental gene-expression programs, thus ensuring tissue identity and supporting cell-lineage decisions.
Keywords: genetic modification

overall_design:
Experiment aimed at identifying dASF/SF2 potential mRNA targets in the Drosophila developing eye
Control: GMR X GFP-NLS trangenic larvae
The 2 IP- GFP cy3 vs ASF cy5 rep.1 and IP- GFP cy5 vs ASF cy3 rep.1 samples correspond to dye-swap experiments.
Two replicates (rep1 and rep2) are included.

first: Mathieu

last: Gabut

first: Jerome

last: Dejardin

first: Jamal

last: Tazi

first: Johann


title: shld1 effect on 3T3 cell

status: Public on Sep 27 2007

city: Stanford

first: Ling-Chun

last: Chen

address: 318 Campus Drive

department: Molecular Pharmacology

institute: Stanford University

summary:
NIH3T3 cells were cultured in medium containing 1 uM, 100 nM, 10 nM and 0 nM (serve as control) shld1  for 24 hours. Three independent cultures were obtained for each drug concentration.  Total RNA from experimental samples were extracted using RNeasy Kit (Qiagen). Mouse RNA pooled from 11 cell lines (Stratagene, La Jolla, CA) was used as reference. Cy5-dUTP and Cy3-dUTP, were used to label the experimental cDNA and the reference samples, respectively, and hybridized to the Mouse Exonic Evidence Based Oligonucleotide (MEEBO) arrays (http://www.microarray.org./sfgf/meebo). The hybridizations were performed by Stanford Functional Genome Facility using its standard protocol (Oligo Array Hybridization protocol, http://www.microarray.org./sfgf/docView.do?type=2).
A compound treatment design type is where the response to administration of a compound or chemical (including biological compounds such as hormones) is assayed.
Keywords: compound_treatment_design

overall_design: Computed


title: Mouse CIA Cytokine Array

city: Macclesfield

first: stephanie

middle: ann

last: roberts

state: Cheshire

address: Alderley Park

department: RIRA

country: United Kingdom

institute: astrazeneca

summary:
The molecular basis to autoimmune arthritis is unclear. Collagen Induced Arthritis (CIA) in mice, is a model that has many features that resemble Rheumatoid Arthritis (RA), although it does not perfectly duplicate RA. The study of CIA has provided insight into relevant pathogenesis and has aided in the identification of potential therapeutic targets. In this study we used Mouse Cytokine Expression Arrays to examine gene expression levels in joints at early, peak and decline stages during disease in DBA/1 mice. The aim of the study was to identify candidate molecules that may be involved in the development and progression of collagen-induced arthritis (CIA).
Keywords: Disease State Analysis

overall_design:
18 samples were analysed. 9 samples were from mice with CIA at early rising, peak and declining phases of the disease (3 mice at each time point). A further 9 samples were from normal joint tissue obtained from naïve DBA/1 mice at the same ages as the mice with CIA (also 3 mice each group).
Prepared samples were hybridized to R&D Systems Mouse Cytokine Arrays. These arrays were spotted with murine cDNAs representing 514 immuno-modulatory factors including 46 chemokines and receptors, 20 proteases, 17 integrins and 8 house keeping genes.

first: Shuhua

last: Han

first: George

last: Booth

first: Rachel

last: Barlow

first: Stephanie

last: Raines

first: Peter

last: Newham

first: Biao

last: Zheng


title: Increased dASF/SF2 expression does not affect global gene expression in Drosophila.

overall_design:
Experiment aimed at determining whether increased SR protein dASF/SF2 expression can affect global gene expression in Drosophila.
Control: GMR X GFP-NLS trangenic larvae
The 2 samples correspond to dye-swap experiments.


title: Increased B52 expression does not affect affect global gene expression in Drosophila

overall_design:
Experiment aimed at determining whether increased SR protein B52 expression can affect global gene expression in Drosophila.
Control: GMR X GFP-NLS trangenic larvae
The 2 samples correspond to dye-swap experiments.


title: Identification of B52 potential mRNA targets

overall_design:
Experiment aimed at identifying B52 potential mRNA targets in the Drosophila developing eye
Control: GMR X GFP-NLS trangenic larvae
The 2 IP- GFP cy3 vs B52 cy5 rep.1 and IP- GFP cy5 vs B52 cy3 rep.1 samples correspond to dye-swap experiments.
Two replicates (rep1 and rep2) are included.


title: Comprehensive gene expression analysis of Bruce-4 embryonic stem cells by SAGE

status: Public on Sep 22 2007

city: Bergisch Gladbach

first: Sandra

middle: Juliane

last: Pennartz

address: Friedrich-Ebert-Strasse 68

department: Research & Development

country: Germany

institute: Miltenyi Biotec GmbH

summary:
Mouse embryonic stem cells lines derived from different mouse strains show considerable variation concerning their culture conditions, gene targeting efficiency and germ line transmission capacity. To gain insight into the basis for this phenomenon, we generated a SAGE library of the C57BL/6 derived ES cell line Bruce-4 and compared their gene expression to 129 strain-derived cells, which in general show the best performance in gene targeting experiments.
Gene Ontology (GO) analyses for functional groups of genes demonstrated considerably higher expression of clusters indicating cellular differentiation as well as cell-cell and cell-matrix adhesion in R1 than in Bruce-4 cells. At the single gene level, almost 90% of the most strongly expressed transcripts in each library were significantly differential. Regarding transcripts that have been linked to stem cell state, seven of the 20 selected genes were expressed in both cell lines, eleven showed stronger expression in Bruce-4 ES cells and only one in R1 cells. These results suggest that expression of “stemness” genes cannot be used as a sole criterion for efficiency of ES cells in gene targeting experiments.
Keywords: Embryonic stem cells, SAGE, gene expression profile

overall_design: We generated a SAGE library of the C57BL/6-derived ES cell line Bruce-4 and compared their gene expression to an already available SAGE library of 129-derived R1 ES cells generated by Anisimov et al (GSM72878).

first: Harold

last: Cremer

first: Andreas

last: Bosio

first: Kay

last: Hofmann


title: Gene expression profiling in Bclaf1 deficient versus wild type post natal day1 lung

status: Public on Sep 15 2007

city: Toronto

first: Solmaz

last: Alizadeh-Azami

state: ON

address: 1 King's college circle

department: Pharmacology

country: Canada

institute: University of Toronto

summary:
In this study we compared the gene expression pattern in Bclaf1-deficient vs wild type lung tissue
Keywords: Gene expression profiling

overall_design:
RNA was extracted from dissected post natal day1 Bclaf1-deficient mice.
Labeling, hybridization and data acquisition was conducted at the University Health Network microarray centre.
http://www.microarrays.ca

first: Haya

last: Sarras

first: Laura

last: Tamblyn

last: Mcpherson


title: UV irradiation of human melanocytes

city: Zürich

first: Keith

last: Hoek

address: private

institute: Currently unaffilliated

summary:
Target genes of ultraviolet stress response in cutaneous melanocytes, potentially associated with solar induced melanocarcinogenesis, were characterized by cDNA microarray technology.
Keywords: stress response

overall_design: In cultured normal human melanocytes, 198 genes out of »9000 arrayed were found modulated > 1.9 times following artificial ultraviolet (mainly ultraviolet-B) irradiation (100 mJ per cm2).

first: Christine

last: Valery

first: Jean-Jacques

last: Grob

first: Patrick

last: Verrando


title: 7.5uM CG-1521 and 5uM TSA treatment of LNCaP Prostate Cancer cells

city: Notre Dame

first: Randy

last: Jeffrey

state: IN

address: Rm 232, Galvin Life Sciences Bldg

department: Biological Sciences

institute: University of Notre Dame

summary:
Genome wide expression changes following treatment with the HDACs (Histone Deacetylase Inhibitor) CG-1521 (7.5uM) or TSA (Trichostatin A) were investigated to determine regulatory targets and patterns of the HDAC Inhibitors.
Keywords: Expression response to treatment, data was used for a comparison of gene expression and regulation between CG-1521 and TSA in LNCaP Cells

overall_design: LNCaP Prostate Cancer cells were treated for a period of 24h with either CG-1521 (7.5uM) or TSA (5uM) following a 24h seeding period. At the selected time point, total RNA was harvested from the cells for hybridization and analysis by Nimblgen Systems Inc using the homo sapiens gene expression array.

first: Somdutta

last: Roy

first: Martin

last: Tenniswood


title: 50uM Casodex treatment of PC-346C Prostate Cancer cells

summary:
Genome wide expression changes following 50uM Casodex treatment were investigated to determine regulatory targets commonly overlooked in gene function specific microarrays and for comparison of the effects the wild-type AR expressing PC-346C cells against the mutant androgen receptor (AR) LNCaP cell line.
Keywords: Expression response to treatment, will be used with another array dataset for cell type comparison.

overall_design: PC-346C Prostate Cancer cells were treated for a period of 48h with or without 50uM Casodex following a 36h seeding period. At the selected time point, total RNA was harvested from the cells for hybridization and analysis by Nimblgen Systems Inc using the homo sapiens gene expression array.


title: 50uM Casodex treatment of LNCaP Prostate Cancer cells

summary:
Genome wide expression changes following 50uM Casodex treatment were investigated to determine regulatory targets commonly overlooked in gene function specific microarrays and for comparison of the effects of the mutant androgen receptor (AR) LNCaP cell line against the wild-type AR expressing PC-346C cells.
Keywords: Expression response to treatment, will be used with another array dataset for cell type comparison.

overall_design: LNCaP Prostate Cancer cells were treated for a period of 48h with or without 50uM Casodex following a 24h seeding period. At the selected time point, total RNA was harvested from the cells for hybridization and analysis by Nimblgen Systems Inc using the homo sapiens gene expression array.

first: Winnie

last: Wang


title: Expression data of HUVEC cells after TNF-alpha treatment

city: Komaba

first: Shuichi

last: Tsutsumi

country: Japan

institute: The University of Tokyo

type: Expression profiling by genome tiling array

summary:
Tumor necrosis factor a (TNF-aplha) is a potent cytokine that orchestrates the inflammatory response by sequentially activating the expression of many genes in cultured human umbilical vein cells (HUVECs).
We apply samples collected every 7.5 min to arrays and monitor the growth of several long transcripts.
Keywords: time course

overall_design: Before stimulation, HUVECs were serum-starved in EBM-2 (Clonetics) containing 0.5% FBS for 18 h, and then treated with 10 ng/ml TNF (Peprotec) and samples collected every 7.5 min for 3 h.

first: Tatsuhiko

last: Kodama

first: Hiroyuki

last: Aburatani

first: Sigeo

last: Ihara


title: SMAUG Is a Major Regulator of Maternal mRNA Destabilization in Drosophila

status: Public on Sep 14 2007

city: Mississauga

first: Zak

last: Razak

state: Ontario

address: 3359 Mississauga Rd

institute: Canadian Drosophila Microarray Centre

summary:
In animals, egg activation triggers a cascade of posttranscriptional events that act on maternally synthesized RNAs. We show that, in Drosophila, the PAN GU (PNG) kinase sits near the top of this cascade, triggering translation of SMAUG (SMG), a multifunctional posttranscriptional regulator conserved from yeast to humans. Although PNG is required for cytoplasmic polyadenylation of smg mRNA, it regulates translation via mechanisms that are independent of its effects on the poly(A) tail. Analyses of mutants suggest that PNG relieves translational repression by PUMILIO (PUM) and one or more additional factors, which act in parallel through the smg mRNA's 3' untranslated region (UTR). Microarray-based gene expression profiling shows that SMG is a major regulator of maternal transcript destabilization. SMG-dependent mRNAs are enriched for gene ontology annotations for function in the cell cycle, suggesting a possible causal relationship between failure to eliminate these transcripts and the cell cycle defects in smg mutants.
Keywords:  Identification of Maternal mRNAs

overall_design:
RNA was extracted from staged unfertilized eggs and stage 14 oocytes using a modification of the TRIzol (Invitrogen) method (Neal et al., 2003). Sample quality was evaluated by probing northern blots for known stable (rpA1) and unstable (Hsp83) transcripts. Total RNA was reverse transcribed as previously described (Neal et al., 2003), except that priming was carried out using random primers rather than oligo-dT in order not to bias the labeling toward mRNAs with long poly(A) tails. The fluorescently labeled cDNA probes were hybridized to 12Kv1 microarray slides obtained from the Canadian Drosophila Microarray Centre (http://www.flyarrays.com). These represent 10,500 distinct protein-coding genes, or 77% of the Drosophila protein-coding genome. Hybridization and scanning was also as previously described (Neal et al., 2003), using a PerkinElmer/GSI ScanArray 4000 scanner and the ScanArray software. The 16 bit TIFF image files were quantified using QuantArray Version 3 (PerkinElmer) using the adaptive quantification algorithm and analyzed using GeneTraffic Duo 3.2 (Iobion Informatics/Stratagene) after normalization to the known stable transcripts rpA1 and rp49.
Of the 10,500 distinct protein-coding genes on the microarray, 9,257 were analyzed for maternal expression. The averages of all of the raw values for 21 hybridizations of wild-type stage 14 oocyte RNA (three replicates of seven experiments) were sorted from highest to lowest. Maternal expression was assessed at different levels in the list by scanning the Berkeley Drosophila Genome Project in situ database (http://www.fruitfly.org/cgi-bin/ex/insitu.pl). RNA from the genes at the top of the table is maternally loaded (90.4% of transcripts with an average raw intensity > 20,000 are maternal; n = 73), whereas RNA from genes at the bottom is absent from early embryos (only 8.2% of those with an average intensity of < 2,000 are maternal; n = 73). Hsp83 appeared near the top of the list (sixth out of 9,257). Transcripts with average values between 3,500 and 5,000 were mostly maternal (75%; n = 76). As the values decreased, there was a decreasing frequency of maternal transcripts (58.7% between 3,000 and 3,500, n = 172; 46.9% between 2,800 and 3,000, n = 160; 26.2% between 2,500 and 2,800, n = 80; 24.7% between 2,000 and 2,500, n = 81). Using a cutoff value of 3,000, we calculate that 55% of all protein-coding genes are represented in stage 14 oocytes

first: Wael

last: Tadros

first: Aaron

middle: L

last: Goldman

first: Tomas

last: Babak

first: Fiona

last: Menzies

first: Leah

last: Vardy

first: Terry

last: Orr-Weaver

first: Timothy

last: Hughes

first: J

last: Westwood

first: Craig

last: Simbert

first: Howard

middle: D

last: Lipshitz


title: Whole genome ChIP/chip tiling of human ES cells for H3K4me3 and H3K27me3

city: Madison

first: Victor

last: Ruotti

state: WI

address: 425 Henry Mall

department: Biotechnology

institute: UW Madison

type: Genome binding/occupancy profiling by genome tiling array

summary:
Chromatin immunoprecipitation was performed using monoclonal antibodiesd. DNA was amplified, fluorescently labeled and hybridized to a series of 38 arrays containing a total of 14.6 million 50-mer oligonucleotides
Keywords: ChIP-chip hES whole genome

overall_design: Chromatin immunoprecipitated DNA from H3K4me

first: Guangjin

last: Pan

first: Shulan

last: Tian

first: Jeff

last: Nie

first: Chuhu

last: Yang

first: Hairong

last: Wei

first: Gudrun

last: Jonsdottir

first: Ron

last: Stewart

first: James

last: Thomson


title: A Marfan syndrome gene expression phenotype in cultured skin fibroblasts

city: Seattle

first: Walter

middle: L.

last: Ruzzo

state: WA

department: CSE

institute: University of Washington

summary:
Background:  Marfan syndrome (MFS) is a heritable connective tissue disorder caused by mutations in the fibrillin-1 gene. This syndrome constitutes a significant identifiable subtype of aortic aneurysmal disease, accounting for over 5% of ascending and thoracic aortic aneurysms.  Results:  We used spotted membrane DNA macroarrays to identify genes whose altered expression levels may contribute to the phenotype of the disease.  Our analysis of 4132 genes identified a subset with significant expression differences between skin fibroblast cultures from unaffected controls versus cultures from affected individuals with known fibrillin-1 mutations.  Subsequently, 10 genes were chosen for validation by quantitative RT-PCR. Conclusions:  Differential expression of many of the validated genes was associated with MFS samples when an additional group of unaffected and MFS affected subjects were analyzed (p-value < 3 x 10-6 under the null hypothesis that expression levels in cultured fibroblasts are unaffected by MFS status).   An unexpected observation was the range of individual gene expression. In unaffected control subjects, expression ranges exceeding 10 fold were seen in many of the genes selected for qRT-PCR validation. The variation in expression in the MFS affected subjects was even greater.
Keywords: disease state comparison; Marfan syndrome; cultured skin fibroblasts

overall_design: Using GF211 spotted cDNA arrays, we compared expression levels of 4132 genes in cultured skin fibroblasts from 17 Marfan subjects and  19 unaffected controls of similar age an sex distribution.  On average, each RNA sample was hybridized 2.5 times.  Ten selected genes were validated by quantitative real-time PCR in these subjects plus an additional 16 Marfan subjects and 16 controls.

first: Zizhen

last: Yao

first: Jochen

middle: C

last: Jaeger

first: Cecile

middle: Z

last: Morales

first: Mary

last: Emond

first: Uta

last: Francke

first: Dianna

middle: M

last: Milewicz

first: Stephen

last: Schwartz

first: Eileen

last: Mulvihill


title: Gene expression in the human VTA of cases with alcohol abuse

city: Brisbane

first: Traute

last: Flatscher-Bader

state: Queensland

address: Queensland Biosciences Precinct, Level 5, CSIRO LI, 306 Carmody Road

department: Faculty of Natural Resources, Agriculture and Veterinary Science

country: Australia

institute: University of Queensland

summary:
Drugs of abuse including nicotine and alcohol elicit their effect by stimulating the mesocorticolimbic dopaminergic system.  There is a high incidence of nicotine dependence in alcoholics.  To date only limited data is available on the molecular mechanism underlying the action of alcohol and nicotine in the human brain.  This study utilised gene expression screening to identify genes sensitive to chronic alcohol abuse within the ventral tegmental area of the human brain.
Keywords: gene expression, brain, alcohol abuse, human, ventral tegmental area

overall_design:
RNA was extracted individually from postmortem tissue samples of human ventral tegmental area (VTA) of  6 cases with and 6 cases without chronic alcohol abuse.
RNA was also extracted from various other brain region (nucleus accumbens, prefronatl cortex). All samples were linearly amplified individually as described.
Reference sample: Part of the aRNA from the VTA and those of the other brain regions was pooled.
Cy3 dCTP labelling of each individual VTA aRNA sample.
Cy5 dCTP labelling of reference pool.
One labelled VTA aRNA sample was hybridised together with labelled reference aRNA.
Samples of the same pathology (alcoholics or control cases) were treated as biological replicates.
Student's t-test was used to reveal differentially expressed genes.

first: Naomi

last: Landis

first: Peter

last: Wilce


title: Retionic acid gene expression time series in human embryonic stem cells

first: Victor

last: Ruotti

summary:
Retinoic acid RA time course experiment: H1 cells(WiCell Research Institute, WA01, Passage40) maintained in TeSR1 were treated with 1uM retinoic acid (RA) in a detailed time course:day0,day0.5,day1,day1.5,day2,day3, day4 and day5 . Total RNA was prepared by usage of the Qiagen RNA extraction kit.  mRNA was purified and then labeled and hybridized to HG17 NimbleGen human expression chips to evaluate changes in gene expression during RA-induced differentiation.
Keywords: Retionic acid gene expression time series human embryonic stem cells

overall_design: Samples from eight time points  were taken after RA treatment  (0,0.5,1,1.5,2,3,4,5 days) of H1 hES cells in TeSR1 medium. In addition, the experiment includes biological replicates at days 0 and 5.

first: Guangjin

last: Pan

first: Shulan

last: Tian

first: Jeff

last: Nie

first: Chuhu

last: Yang


title: Gene expression profiling of rheumatoid arthritis synovial tissues

status: Public on Sep 13 2007

last: Stanford Microarray Database

address: 300 Pasteur Drive

department: Stanford University, School of Medicine

institute: Stanford Microarray Database (SMD)

summary:
Spots were selected that had an expression level higher than 1.6 the local background. The use of a common reference sample allows the comparison of the relative expression levels across the tissue samples. All genes were expressed relative to their median expression level across the arrays of the same microarray platform, allowing us to combine the data from the 2 platforms. Data from Lymphochip and Stanford Human cDNA arrays was combined per patient. Only genes with reliable datapoints in at least 80% of the samples were used. Subsequently, genes with the same unigene identifier were averaged in SMD. For all further analysis genes were selected that showed at least a two-fold difference in expression level in at least one array.
A disease state experiment design type is where the state of some disease such as infection, pathology, syndrome, etc is studied.
Keywords: disease_state_design

overall_design: Using regression correlation

first: Cornelis

last: Verweij


title: Genome-wide analysis of melanoma cell lines using high-density single-nucleotide polymorphism arrays.

status: Public on Sep 12 2007

first: Mitchell

last: Stark

address: 300 Herston Rd Herston

institute: QIMR

type:
Genome variation profiling by SNP array
SNP genotyping by SNP array

summary:
Although a number of genes related to melanoma development have been identified through candidate gene screening approaches, few studies have attempted to conduct such analyses on a genome-wide scale. Here we use Illumina 317K whole-genome single-nucleotide polymorphism arrays to define a comprehensive allelotype of melanoma based on loss of heterozygosity (LOH) and copy number changes in a panel of 76 melanoma cell lines. In keeping with previous reports, we found frequent LOH on chromosome arms 9p (72%), 10p (55%), 10q (55%), 9q (49%), 6q (43%), 11q (43%), and 17p (41%). Tumor suppressor genes (TSGs) can be identified through homozygous deletion (HD). We detected 174 HDs, the most common of which targeted CDKN2A (n = 33). The second highest frequency of HD occurred in PTEN (n = 8), another well known melanoma TSG. HDs were also common for PTPRD (n = 7) and HDAC4 (n = 3), TSGs recently found to be mutated or deleted in other cancer types. Analysis of other HDs and regions of LOH that we have identified might lead to the characterization of further melanoma TSGs. We noted 197 regional amplifications, including some centered on the melanoma oncogenes MITF (n = 9), NRAS (n = 3), BRAF (n = 3), and CCND1 (n = 3). Other amplifications potentially target novel oncogenes important in the development of a subset of melanomas. The numerous focal amplifications and HDs we have documented here are the first step toward identifying a comprehensive catalog of genes involved in melanoma development, some of which may be useful prognostic markers or targets for therapies to treat this disease.
Keywords: High Density SNP array

first: Nicholas

last: Hayward


title: Salmonella Typhimurium transcription profiles in space flight

city: Irvine

first: Michael

last: McClelland

address: 132 Med Surge I

department: Microbiology & Molecular Genetics

institute: University of California, Irvine

summary:
Salmonella transcription profiles were obtained from samples flown on space shuttle mission STS-115, and compared to profiles from Salmonella grown under identical conditions on the ground.
Keywords: stress response, transcriptional profile

overall_design: Triplicate experimental samples were hybridized to slides that contain three identical Salmonella ORF arrays. Each hybridization was performed with Cy3 labeled total RNA  and Cy5 labeled gDNA as control.

first: James

last: Wilson

first: Steffen

last: Porwollik

first: Pui

last: Cheng

first: Cheryl

last: Nickerson


title: aCGH data from age-dichotomized ER+ breast tumors

city: Novato

first: Christina

last: Yau

address: 8001 Redwood Blvd

institute: Buck Institute of Age Research

type: Genome variation profiling by genome tiling array

summary:
To investigate the biological basis between aging and sporadic breast cancer incidence and prognosis, DNA samples
from matched ER+ invasive breast cancers diagnosed in either young (<45) or old (>70) women were analyzed with aCGH
Keywords: biomarker identification

overall_design: ER+ breast cancers collected from either young or old women, well matched for other clinical parameters were analyzed using aCGH.  Age associated genomic differences were assayed.

first: Vita

last: Fedele

first: Ritu

last: Roydasgupta

first: Jane

last: Fridlyand

first: Alan

last: Hubbard

first: Joe

middle: W

last: Gray

first: Karen

last: Chew

first: Shanaz

middle: H

last: Dairkee

first: Dan

last: Moore

first: Stefania

last: Tommasi

first: Angelo

last: Paradiso

first: Donna

last: Albertson

first: Christopher

last: Benz


title: Chromosomal rearrangement interferes with meiotic X-chromosome inactivation

city: Basel

first: Robert

last: Ivanek

address: Hebelstrasse 20

department: Department of Biomedicine

institute: University of Basel

summary:
Expression profiling of isolated populations of prepachytene spermatocytes, pachytene spermatocytes and spermatids of males carrying a translocation T(16;17)43H was performed to study the influence of this translocation on gene expression from individual chromosomes.
To evaluate the transcriptional behavior of individual chromosomes in the sterile B10-T43/+ males and their fertile congenic B10-T43/T43 and B10 counterparts, we compared their transcriptomes in sorted populations of pre-pachytene primary spermatocytes, pachytene primary spermatocytes and spermatids.
Keywords: individual genetic characteristics design

overall_design: Populations of pre-pachytene spermatocytes, pachytene spermatocytes and spermatids were isolated from three lines of mice: B10, B10-T43H/T43H and B10-T43H/+. All populations were isolated in duplicates from two animals. Hence, RNA was prepared from 18 samples and was then subject to labeling and hybridization on microarray chips.

first: David

last: Homolka


title: Mating response - six alpha factor concentrations (0.06, 0.2, 0.6, 6, 60 and 600 nM)

status: Public on Sep 11 2007

city: Rehovot

first: Ofer

last: Sarig

address: Weizmann Institute of Science

department: Molecular Genetics

country: Israel

institute: Weizmann Institute of Science

summary:
We examined the mating response of W303 bar1delta a-type cells to six alpha-factor concentrations (0.06, 0.2, 0.6, 6, 60 and 600 nM). In each alpha factor concentration between five to seven time points were collected. The time points in all experiments (except for concentration 600 nM were time point 15 min was omitted) were 5, 15, 30, 60, and 90 min. In some of the concentrations also time points 120 and 150 min were considered. The expression of cells before and after addition of alpha-factor were compared using S. cerevisiae cDNA microarrays, in all experiments the same sample before adding alpha factor was used as a control.
Keywords: yeast mating response, comparison among alpha factor concentrations

overall_design: The six time courses were processed in two groups, in each group all samples were processed in parallel. The group were: 0.06, 0.6, 60 nM and 0.2, 6, 600 nM. In all microarrays the same control sample was used (cells before adding alpha factor). All samples were measured by hybridization to S. cerevisiae cDNA microarray.


title: MGAS5005 rivRX knockout vs. rivRX overexpressing strain

city: Atlanta

first: Samantha

middle: Ann

last: Roberts

state: GA

address: 1510 Clifton Road

department: Microbiology

institute: Emory University

summary:
JRS1616/pREG696 is our MGAS5005 covR rivRX double knockout carrying empty vector. JRS1616/pJRS1624 is our covR rivRX double knockout carrying a plasmid which overexpresses rivRX.
Keywords: comparitive transcriptome analysis

overall_design: Four separate RNA preparations from each strain (RNA #A-D) were hybridized to duplicate arrays. Reference RNA consisted of pooled RNA from each preparation, labelled with Cy5.

middle: A

first: June

last: Scott


title: Isw2 ChIP in S cerevisiae

status: Public on Sep 10 2007

first: Toshio

last: Tsukiyama

address: 1100 Fairview Ave N

institute: FHCRC

summary:
Study to detect to genome wide localization of the ATP dependent chromatin remodelling factor Isw2 using ChIP.
Keywords: ChIP chip

overall_design: Isw2 binding was determined using a catalytically inactive mutant (K215R) that gives specific binding across the genome. Binding of the K215R mutant was compared to that of the WT protein (that binds nonspecifically) and also to that of control "input" DNA samples.


title: Transcriptinal profile of responses to ER and osmotic stress

city: Tallahassee

first: Andre

middle: Southernman Teixeira

last: Irsigler

state: FL

address: Chieftan Way

department: Biology

institute: Florida State University

summary:
Despite the potential of the endoplasmic reticulum (ER) stress response to accommodate adaptive pathways, its integration with other environmental-induced responses is poorly understood in plants. Here, we performed global expression profiling on soybean leaves exposed to polyethylene glycol treatment or to unfolded protein response (UPR) inducers to identify integrated networks between osmotic and ER stress-induced adaptive responses. The results unmasked the major branches of the ER-stress response, which includes enhancing protein folding and degradation in the ER, as well as specific osmotically regulated changes linked to cellular responses induced by dehydration. However, a small proportion (5.5%) of total up-regulated genes represented a shared response that seemed to integrate the two signaling pathways. These co-regulated genes were considered downstream targets based on similar induction kinetics and a synergistic response to the combination of osmotic- and ER-stress-inducing treatments.  Genes in this integrated pathway with the strongest synergistic induction encoded proteins with diverse roles. Two of them contained a plant-specific development and cell death (DCD) domain while another had homology to proteins with an ubiquitin-associated (UBA) domain. A NAC domain-containing protein exhibited robust early kinetics of induction consistent with a role as a transfactor. This integrated pathway diverged further from characterized ER-specific branches of UPR as downstream targets were inversely regulated by osmotic stress. Collectively, our results describe a novel branch of the ER stress response that integrates the osmotic signal to potentiate transcription of shared target genes.
Keywords: stress response

overall_design: In all experiments two independent biological replicates were used. For each biological replicate, two hybridizations were performed, with dye-swap.

first: André

first: Ping

last: Zhang

first: Ralph

last: Dewey

first: Rebecca

last: Boston

first: Elizabeth

last: Fontes


title: Mapping chromatin remodelling in delta isw2 cells

summary:
To define chromatin structure changes along the yeast genome using microarrays. Nucleosomal DNA from WT and delta isw2 yeast were hybridized and differences in signals were calculated.
Keywords: Nucleosomal DNA hybridization

overall_design: Nucleosomal DNA was harvested from WT and delta isw2 yeast (2 biological replicates of each) then fragmented into ~50bp lengths and then hybridized onto the tilling microarrays. WT signal was the compared to the mutant signal to find regions of chromatin structure change.

first: Iestyn

last: Whitehouse


title: gene differentially expressed in androgen independent prostate cancer

status: Public on Sep 09 2007

city: Shanghai

first: xuping

last: fu

country: China

institute: Institute of Genetics, Fudan University

summary:
Progression to androgen independent is the main cause of death in prostate cancer, and the mechanism is still unclear. By reviewing the expression profiles of 26 prostate cancer samples in a holistic view, we found a group of genes differentially expressed in androgen independent compared with androgen dependent groups (p value< 0.01, t test). Focusing on apoptosis, proliferation, hormone and angiogenesis, we found a group of genes such as thioredoxin domain containing 5 (TXNDC5), tumor necrosis factor receptor superfamily, member 10a (TNFRSF10A), ribosomal protein S19 (RPS19) and Janus kinase 2 (JAK2) up-regulated in androgen independent prostate cancer, which could play important roles in the transition from androgen dependent to androgen independent and could be biomarkers of prognosis.
Keywords: cell type comparison

overall_design: The main aim was comparing the androgen dependent and androgen independent prostate cancer to identify differentially expressed genes. In addition, we added several normal prostate tissue sample for comparisons. Totally 29 experiments were performed without replicates. 3 for normal prostate tissue, 8 for androgen independent cancer and 18 for androgen dependent prostate cancer. In all experiments, the reference samples are common reference, a pool with unrelated fetal tissues.

first: Qing

last: Wei

first: Ming

last: Li

first: Xuping

last: Fu

first: Rong

last: Tang

first: Yanqun

first: Mei

last: Jiang

first: Yao


title: UTX_CHIP_FIBROBLASTS

status: Public on Sep 07 2007

city: Stanford

first: John

middle: Louis

last: Rinn

address: 2155 CCSR

department: Dermatology

institute: Stanford University

summary:
The recent discovery of a large number of histone demethylases suggests a central role for these enzymes in regulating histone methylation dynamics. Histone H3K27 trimethylation (H3K27me3) has been linked to Polycomb Group (PcG) protein-mediated suppression of Hox genes and animal body patterning, X-inactivation and possibly maintenance of embryonic stem (ES) cell identity. An imbalance of H3K27 methylation due to over-expression of the methylase EZH2 has been implicated in metastatic prostate and aggressive breast cancers. Here we show that the related JmjC domain-containing UTX and JMJD3 catalyze demethylation of H3K27me3. UTX is enriched around the transcription start sites of many Hox genes in primary human fibroblasts where Hox genes are differentially expressed, but is selectively excluded from the Hox locus in ES cells in which Hox genes are largely silent. Consistently, RNAi inhibition of UTX led to increased H3K27me3 levels at some Hox gene promoters. Importantly, Morpholino inhibition of a zebrafish UTX homolog resulted in mis-regulation of Hox genes and a striking posterior developmental defect, which was partially rescued by wildtype, but not catalytically inactive human UTX. Taken together, these findings identified a small family of H3K27 demethylases with important, evolutionarily conserved roles in H3K27 methylation regulation and in animal anterior-posterior development.
Keywords: organism_part_comparison_design

overall_design:
An organism part comparison experiment design type compares tissues, regions, organs within or between organisms.

Controls: UTX occupancy was mapped by chromatin immunoprecipitation using anti-UTX antibody. As control, we performed mock chromatin immunoprecipitation with isotype-matched antibody (IgG). Two color, competitive hybridization experiments were performed to compare genomic DNA enrichment in UTX vs. IgG pulldowns.

Experimental variable: We compared primary human fibroblasts from two different anatomic locations--foot and lung.

first: Janos

last: Demeter


title: The Arabidopsis chromodomain-containing LHP1 colocalizes with histone H3 lysine27 trimethylation

city: Los Angeles

first: Steve

middle: E

last: Jacobsen

address: 621 Charles E. Young Dr. South

department: Molecular Cell and Developmental Biology

institute: University of California, Los Angeles

summary:
The POLYCOMB proteins are required for maintenance of silent chromatin states mediated by H3K27 trimethylation in animals, but POLYCOMB homologues are not found in plant genomes. Using DamID-chip, we found that the Arabidopsis chromodomain-containing protein LHP1 localizes to chromatin associated with H3K27me3 genome-wide. Furthermore, the LHP1 chromodomain binds H3K27me3 with high affinity. These results suggest that LHP1 shares similar functions with POLYCOMB.
Keywords: DamID-Chip

overall_design: Four replicates of DamID-chip and four replicates of Dam-only control

first: Steven


title: Genome-Wide Expression Dynamics of a Marine Virus and its Host during Lytic Infection

city: Haifa

first: Debbie

last: Lindell

address: Technion City

department: Department of Biology

institute: Technion - Israel Institute of Technology

summary:
We used microarray analysis to investigate whole genome transcriptome dynamics of the marine cyanobacterium Prochlorococcus sp. strain MED4 and the T7-like podovirus P-SSP7 over a time course during the 8 hour latent period of lytic infection prior to cell lysis.

Manuscript Summary: Interactions between bacterial hosts and their viruses (phages) lead to reciprocal genome evolution through a dynamic co-evolutionary process1-5.  Phage-mediated transfer of host genes – often located in genome islands – has had a major impact on microbial evolution1, 4, 6.  Furthermore, phage genomes have clearly been shaped by the acquisition of genes from their hosts2, 3, 5.  Here we investigate whole-genome expression of a host and phage, the marine cyanobacterium Prochlorococcus and a T7-like cyanophage during lytic infection, to gain insight into these co-evolutionary processes.  While most of the phage genome was linearly transcribed over the course of infection, 4 phage-encoded bacterial metabolism genes were part of the same expression cluster, even though they are physically separated on the genome.  These genes — encoding photosystem II D1 (psbA), high-light inducible protein (hli), transaldolase (talC) and ribonucleotide reductase (nrd) — are transcribed together with phage DNA replication genes and appear to make up a functional unit involved in energy and deoxynucleotide production needed for phage replication in resource-poor oceans.  Also unique to this system was the upregulation of numerous genes in the host during infection.  These may be host stress response genes, and/or genes induced by the phage.  Many of these host genes are located in genome islands and have homologues in cyanophage genomes. We hypothesize that phage have evolved to utilize upregulated host genes, leading to their stable incorporation into phage genomes and their subsequent transfer back to hosts in genome islands. Thus activation of host genes during infection may be directing the co-evolution of gene content in both host and phage genomes.
Keywords: time course, viral infection, marine cyanobacteria, podovirus, bacteriophage, stress response

sample_organism: Prochlorococcus marinus subsp. pastoris str. CCMP1986

overall_design: Triplicate independent cultures of the cyanobacterium Prochlorococcus MED4 were grown as described in the protocols section below. Each of the three cultures was divided into two subcultures. One subculture for each culture was infected with phage and the other subculture served as the paired uninfected control. Infection Treatment: The T7-like podovirus P-SSP7 was added to Prochlorococcus MED4 at a ratio of 3 infective phage per 1 cyanobacterial cell. Samples were taken for transcriptome analysis every hour after addition of phage during the 8 h latent period of infection: at 0, 1, 2, 3, 4, 5, 6, 7, 8 h after infection for a total of 9 samples per infected biological replicate. Control Treatment: Paired control (uninfected) cells were amended with filter-sterilized spent medium and samples taken at 0, 1, 2, 3, 4, 8 h after addition for a total of 6 samples per “uninfected” biological replicate. A total of 45 samples were analyzed - 27 infected samples from 3 biological replicates and 18 uninfected (control) samples from 3 biological replicates. Series 1, 2, 3 - infected cells; Series 4, 5, 6 - uninfected control cells. Paired infected and uninfected treatments: The paired treatments are: Series 1 and 4; 2 and 5; 3 and 6.

first: Maureen

last: Coleman

first: Matthias

last: Futschik

first: Trent

last: Rector

first: Gregory

last: Kettler

first: Robert

last: Steen

first: Sallie

last: Chisholm


title: Ovarian gene expression in the absence of FIGLA, an oocyte specific transcription factor

status: Public on Sep 08 2007

city: bethesda

first: saurabh

last: joshi

state: MD

address: 9000 rockville pk

department: national institute of diabestes digestive and kidney diseases

institute: national institutes of health

summary:
Ovarian folliculogenesis in mammals is a complex process involving cross talk between germ and somatic cells. Carefully orchestrated expression of transcription factors, cell adhesion molecules and growth factors are required for success. We have identified a germ-cell specific, basic helix-loop-helix transcription factor, FIGLA (Factor In the GermLine, Alpha) and demonstrated its involvement in two independent developmental processes: formation of the primordial follicle and coordinate expression of zona pellucida genes. Taking advantage of Figla null mouse lines, we have used a combined approach of microarray and Serial Analysis of Gene Expression (SAGE) to identify potential downstream targets genes. Using high stringent cutoffs, we find that FIGLA functions as a key regulatory molecule in coordinating expression of the NALP family of genes, genes of known oocyte-specific expression and a set of functionally un-annotated genes.
Keywords: Serial Analysis of Gene Expression

overall_design: The libraries were constructed using Sau3AI as the anchoring enzyme as described in the SAGE protocol (Cheval et al., 2002). Total RNA (~50 µg) was obtained from newborn normal and Figla null ovaries with an RNeasy mini kit (Qiagen, Valencia, CA, USA). The presence of MSY2 and the absence of ZP2 transcripts as detected by one-step qRT-PCR (Qiagen, Valencia, CA, USA) confirmed the identity of Figla null ovaries. Poly(A)+ RNA was isolated with oligo d(T)-conjugated magnetic beads (Dynal-Invitrogen, Calsbad, CA, USA). SAGE libraries are constructed as per the protocol described by Virlon B and group (Virlon B et al, 1999).Briefly, cDNA was synthesized from 5 μg of poly(A) RNAs with the cDNA synthesis system kit. Double stranded cDNA was synthesized using the Invitrogen SuperscriptII kit. The double-strand cDNA was digested with Sau3AI, and the 3′ end was isolated through binding to 1 mg of paramagnetic streptavidin beads (Dynal, Invitrogen, Calsbad, CA, USA), The streptavidin-bound cDNA was divided into two fractions. Each fraction was ligated to 10 pmol of either linker 1 or linker 2, then digested with BsmFI. The released cDNA tags were blunt-ended for 10 min at 42°C in a 50-μl reaction volume containing 20 mM Tris HCl (pH 7.5), 10 mM MgCl2, 25 mM NaCl, 10 mM DTT, 400 μM dNTP, and 10 units of T7 DNA polymerase (Pharmacia Biotech). The two fractions then were ligated to each other by using T4 DNA ligase. PCR (95°C, 30 sec; 58°C, 30 sec; 70°C, 45 sec: 26–28 cycles) was carried out on 1% of ligation reaction in a 100-μl reaction volume containing 20 mM Tris HCl (pH 8.3), 50 mM KCl, 2.5 mM MgCl2, 4 mM DTT, 100 μM dNTP, 50 pmol of primers 1 and 2, and 5 units of Taq polymerase. The products from 10–12 reactions were pooled. The 110-bp DNA fragment was purified by agarose gel electrophoresis and submitted to preparative PCR (120–150 reactions performed as described above, except that the amount of primers was reduced to 25 pmol and the number of cycles was 12). The PCR sample was digested with Sau3AI, and ditags were purified and concatenated as described (Velculescu et al, 1995). Concatenated ditags were cloned into Bluescript pKS (Stratagene, La Jolla, CA, USA) using the BamHI site and blue-white screening, prior to sequence, identified colonies with inserts. Plating, picking, DNA preparation and sequencing were performed at the the NIH Intramural Sequencing Center (NISC). Low quality sequence data were eliminated by Phred analysis.

first: Saurabh

last: Joshi

first: Holly

last: Davies

first: Jurrien

last: Dean


title: Germline and Somatic Disruption of DLG2 in Neuroblastoma

city: Philadelphia

first: Jayanti

last: Jagannathan

address: 34th & Civic center blvd

department: Oncology

institute: Childrens Hospital of Philadelphia

summary:
Data suggest that DLG2 functions as a human neuroblastoma suppressor gene
Keywords: aCGH

overall_design: To discover genetic events that might be involved in the initiation of neuroblastoma tumorigenesis, we performed a genome-wide scan for DNA copy number abnormalities in the germline and tumor DNAs from ten patients with neuroblastoma and complex congenital abnormalities using a customized high-resolution BAC array-based comparative genomic hybridization (aCGH) platform.


title: Gene expression profiling of sporadic neuroblastoma

status: Public on Sep 06 2007

city: Chiba

first: Akira

last: Nakagawara

address: 666-2 Nitona, Chuo-ku

institute: Chiba Cancer Center Research Institute

summary:
Integrated analysis of array-based comparative genome hybridization (aCGH) and gene expression profiling was conducted by using primary neuroblastomas. This submission corresponds the latter gene expression data.
Keywords: other

overall_design: Thirty mass screening-detected and 40 sporadic neuroblastomas were analyzed by both aCGH and gene expression profiling. A UCSF BAC array carrying 2,464 clones was used for the former experiment. For the latter experiment, an in-house cDNA microarray, carrying 5,340 cDNAs obtained from the oligo-capping cDNA libraries generated from anonymous neuroblastoma tissues, were used. Preparation of RNA, hybridization, reading of spots, and statistical analyses were conducted as reported previously [Ohira M, et al. Cancer Cell 2005;7:337-50.]. This series corresponds expression profiles for 40 sporadic neuroblastomas.


title: Human colon cancer cell lines: NT(untreated) vs EGF, Cetuximab and Gefitinib treatments

city: Bologna

first: rossella

last: solmi

address: Via Belmeloro 8

country: Italy

institute: University of Bologna

summary:
Gene Expression Profiling of HT-29 and Caco-2 colon cancer cell lines untreated compared with EGF, Cetuximab, Gefitinib,EGF plus Cetuximab and EGF plus gefitinib treatments.
Keywords: Gene Expression Profiling

overall_design: Five experimental conditions: NTvsEGF, NTvsCetuximab, NTvsGefitinib, NTvsEGFplusGefitinib, NTvsEGFplusCetuximab, for two human colon cancer cell lines (HT-29 and Caco-2).


title: Rapid, transcript-specific changes in splicing in response to environmental stress

status: Public on Sep 17 2007

city: Lexington

first: Gregg

middle: Brooks

last: Whitworth

state: VA

address: 204 West Washington St

institute: Washington and Lee Unversity

summary:
While the core splicing machinery is highly conserved between budding yeast and mammals, the absence of alternative splicing in Saccharomyces cerevisiae raises the fundamental question of why introns have been retained in ~5% of the 6,000 genes. Because Ribosomal Protein-encoding Genes (RPGs) are highly over-represented in the set of intron-containing genes, we tested the hypothesis that splicing of these transcripts would be regulated under conditions where translation is impaired. Using a microarraybased strategy, we find that within minutes after the induction of amino acid starvation the splicing of the majority of RPGs is specifically inhibited. In response to an unrelated stress, exposure to toxic levels of ethanol, splicing of a different group of transcripts is inhibited, while the splicing of a third set is actually improved. We propose that regulation of splicing, like transcription, can afford rapid and specific changes in gene expression in response to the environment.
Keywords: stress, environmental stress, time course, splicing, splicing-specific microarray

overall_design: Splicing-specific microarrays were used to assay changes to splicing induced in Saccharomyces cerevisiae in response to amino-acid starvation (50mM 3-AT), Ethanol toxicity (10% EtOH) and the deletion of several non-essential factors involved in mRNA decay.  The data includes time courses of shifts into each environmental stress as well as dye-flipped technical replicates of each time point.

first: Jeffrey

last: Pleiss

middle: B

first: Megan

last: Bergkessel

first: Christine

last: Guthrie


title: Overexpression of the HRD gene in Arabidopsis

status: Public on Sep 05 2007

city: Blacksburg

first: Andy

last: Pereira

address: Washington Street MC0477

department: Virginia Bioinformatics Institute

institute: Virginia Tech

summary:
Freshwater is a limited and dwindling global resource; therefore,
efficient water use is required for food crops that have high water
demands, such as rice, or for the production of sustainable energy
biomass. We show here that expression of the Arabidopsis HARDY
(HRD) gene in rice improves water use efficiency, the ratio of
biomass produced to the water used, by enhancing photosynthetic
assimilation and reducing transpiration. These drought tolerant
low-water-consuming rice plants exhibit increased shoot biomass
under well irrigated conditions and an adaptive increase in root
biomass under drought stress. The HRD gene, an AP2/ERF-like
transcription factor, identified by a gain-of-function Arabidopsis
mutant hrd-D having roots with enhanced strength, branching,
and cortical cells, exhibits drought resistance and salt tolerance,
accompanied by an enhancement in the expression of abiotic stress
associated genes. Although HRD overexpression in Arabidopsis
produces thicker leaves with more chloroplast-bearing mesophyll
cells, in rice there is an increase in leaf biomass and bundle sheath
cells that probably contribute to the enhanced photosynthesis
assimilation and efficiency. HRD overexpression was also studied for clues of molecular mechanisms involved using microarrays. The results exemplify application of a gene identified from the model plant Arabidopsis for the improvement of water use efficiency coincident with drought resistance in
the crop plant rice.
Keywords: Genetic modification transcription factor overexpression mutant

overall_design: Design includes the use of RNA from the hrd-D mutant overexpressing the HRD gene compared against that from WT Ws (control), both under well watered conditions, in two biological replications.

first: Aarati

last: Karaba

first: Shital

last: Dixit

first: Raffaella

last: Greco

first: Asaph

last: Aharoni

first: Kurniawan

middle: R

last: Trijatmiko

first: Nayelli

last: Marsch-Martinez

first: Arjun

last: Krishnan

first: Karaba

middle: N

last: Nataraja

first: Makarla

last: Udayakumar


title: NF-κB target gene microarray in HAECs in response to Pseudomonas aeruginosa

first: zhe

last: zhang

address: 401 Johnson Pavilion, 36th and Hamilton Walk

institute: University of Pennsylvania

summary:
An NF-κB target gene array (Panomics, Inc., Redwood City, CA) was performed to profile the PAO1-dependent expression pattern of 111 NF-kB-regulated genes in primary human airway epithelial cells (HAECs). A small scale microarray was carried out using NF-κB target gene array kit (Panomics, Inc., Redwood City, CA).  A long sense-strand oligonucleotide for each of the 111 human genes that have been previously shown to be regulated by NF-KB signaling pathway was spotted in duplicate on the nitrocellulose membrane. Biotinylated DNA was spotted along the right and bottom sides of the array membrane as control. The arrays were performed according to the manufacturer’s instructions.
Keywords: cellular response to bacteria

overall_design: In brief, HAECs were stimulated for 18 hours with the heat-killed bacteria PAO1 (multiplicity of infection: 100:1). Control experiment used PBS instead of bacteria. mRNA of HAECs was isolated using biotin-labeled oligo(dT)20 and streptavidin-conjugated magnetic particles (Roche, Germany). 500 ng mRNA was used to prepare biotin-labeled cDNA probes through incorporation of biotin-dUTP into cDNA via reverse transcription reactions. Each biotin-labeled cDNA probe was hybridized to an array membrane at 42°C overnight in a hybridization incubator (Fisher Scientific, Pittsburgh, PA). The membrane was washed and subjected to further incubations with blocking buffer and subsequent streptavidin-conjugated HRP. The membrane was developed through chemiluminescence reactions and detected immediately by 1 min exposure to Hyperfilm ECL X-ray films (Amersham, Piscataway, NJ). The dots on the array were quantified through densitometric analysis using GeneTools (Syngene, Frederick, MD) and the value of each dot was determined as D = (signal value – background value)/mean value of biotinylated DNA dots. Ratio (Dbacteria: Dcontrol) was used to determine the effect of bacteria stimulation on the gene regulation. Ratio>1 indicated an up-regulation of the gene. The experiments include two samples, one PBS control sample and one PAO1-stimulated sample.

first: Zhe

last: Zhang

first: William

last: Reenstra

first: Daniel

middle: J

last: Weiner

first: Jean-Pierre

last: Louboutin

first: James

last: Wilson


title: Microarray Analysis on Salt Stress Affected Genes in Tomato Seedlings

city: Nashville

first: Suping

last: Zhou

state: TN

address: 3500 John A Merritt Blvd

department: IAGER

institute: Tennessee State University

summary:
Large-scale gene expression affected by salt stress was analyzed with  tomato seedlings (Lycoperson esculentum Mill cv. Money Maker) by a cDNA microarray (Tom1).  The significantly differentially expressed genes (5% Benjamini-Hochberg false discovery rate) consisted of 1757 sequences in the analyzed tissues (cotyledons + shoot tip). Genes with over 2 fold difference were selected from the list and further categorized into different function and cellular processes. Tomato homologous genes for the chaperone proteins, antioxidant enzymes (catalase and peroxidase), and ion transporters (Na+-driven multidrug efflux pump, vacuolar ATPase, and others) were induced. The ACC oxidase and ethylene-responsive gene tomato homologs had higher transcript level after salt treatment. Multiple members with different expression patterns were identified for the  bZIP, WRKY, and MADS-box transcription regulator. Different genes in the signal transduction pathway, such as the protein kinases (Shaggy kinase, mitogen-activated protein kinase, ethylene receptor neverripe, and others), protein phosphatases, calmodulin, G-protein, and the N- myristoyltransferase, were regulated by salt stress. Most of the protease and the inhibitor homologs were suppressed by salt stress. In addition, different isoforms of cytochrome P450, genes for polyamine biosynthesis (putrescine and proline) and detoxification compounds (glutathione and thioredoxin), several key enzyme genes in the metabolic pathways of carbohydrates, amino acids, and fatty acids, were also affected by salt treatment. This study has provided a set of candidate genes, especially those in the regulatory machinery that can be further investigated to define salt stress in tomato and other plant species.
Keywords: treatment response

overall_design: Effect of 75mM NaCl on gene’s expression in tomato seedlings, Lycoperscon esculentum Mill. cv. Money Maker after 17 d of treatment. Each sample represents a pool of same tissue 20 seedlings grown in two flasks.  Experiment was done in triplicate.

first: Shu

last: Wei

first: Braden

last: Boone

first: Shawn

last: Levy


title: Identification of genes involved in macrophage activation and effector functions against Listeria monocytogenes.

city: Cape Town

first: Anita

last: Schwegmann

state: Western Cape

address: Institute for Infectious Diseases and Molecular Medicine, UCT medical School, Anzio Rd, Observatory

department: Division of Infectious Immunology

country: South Africa

institute: Institute for Infectious Diseases and Molecular Medicine, University of Cape Town

summary:
Listeria monocytogenes (LM) is a Gram-positive, facultative intracellular bacterium responsible for disseminated infections in immunocompromised individuals which can result in septicaemia and meningitis (1). Effective control of listeriosis requires both innate and adaptive immune responses with the principal mediators of bacterial killing being neutrophils and macrophages (2). Key cytokines driving the innate immune response are IFN-γ (3) and TNF (4), which promote macrophage activation and drive the production of anti-microbial mediators such as reactive oxygen intermediates (ROI) and reactive nitrogen intermediates (RNI). Several studies using gene deficient mice demonstrated that ROI and RNI are required for optimal pathogen killing (5-8), but that macrophages also utilise an alternative, unknown mechanism besides ROI and RNI to efficiently control bacterial infection. Mice deficient for the transcription factor NF-IL6 are highly susceptible to a number of intracellular bacterial and fungal infections, including LM (9-12). NF-IL6-deficient mice  infected with LM were unable to prevent bacterial dissemination, despite production of nitric oxide (NO), IFN-γ and TNF being equivalent to WT controls (12). From these results, we hypothesized that NF-IL6 acts downstream from IFN-γ and TNF during the activation of macrophage effector functions against LM (2). To facilitate the identification of putative effector genes in this unknown pathway, we compared gene expression profiles of WT and NF-IL6 deficient LM-infected macrophages by differential microarray.

REFERENCES
1. Gellin, B. G. & Broome, C. V. (1989) Jama 261, 1313-1320.
2. Brombacher, F. & Kopf, M. (1996) Res Immunol 147, 505-511.
3. Buchmeier, N. A. & Schreiber, R. D. (1985) Proc Natl Acad Sci U S A 82, 7404-7408.
4. Havell, E. A. (1989) J Immunol 143, 2894-2899.
5. Dinauer, M. C., Deck, M. B., & Unanue, E. R. (1997) J Immunol 158, 5581-5583.
6. Endres, R., Luz, A., Schulze, H., Neubauer, H., Futterer, A., Holland, S. M., Wagner, H., & Pfeffer, K. (1997) Immunity 7, 419-432.
7. MacMicking, J. D., Nathan, C., Hom, G., Chartrain, N., Fletcher, D. S., Trumbauer, M., Stevens, K., Xie, Q. W., Sokol, K., Hutchinson, N., et al. (1995) Cell 81, 641-650.
8. Shiloh, M. U., MacMicking, J. D., Nicholson, S., Brause, J. E., Potter, S., Marino, M., Fang, F., Dinauer, M., & Nathan, C. (1999) Immunity 10, 29-38.
9. Pizarro-Cerda, J., Desjardins, M., Moreno, E., Akira, S., & Gorvel, J. P. (1999) J Immunol 162, 3519-3526.
10. Screpanti, I., Romani, L., Musiani, P., Modesti, A., Fattori, E., Lazzaro, D., Sellitto, C., Scarpa, S., Bellavia, D., Lattanzio, G., et al. (1995) Embo J 14, 1932-1941.
11. Sugawara, I., Mizuno, S., Yamada, H., Matsumoto, M., & Akira, S. (2001) Am J Pathol 158, 361-366.
12. Tanaka, T., Akira, S., Yoshida, K., Umemoto, M., Yoneda, Y., Shirafuji, N., Fujiwara, H., Suematsu, S., Yoshida, N., & Kishimoto, T. (1995) Cell 80, 353-361.
Keywords: bone marrow derived macrophages, interferon-gamma, macrophage activation, macrophage effector activity, NF-IL6, gene-deficient mouse model, Listeria monocytogenes, intracellular pathogen

overall_design:
A 2 x 2 factorial experimental design was used to compare the gene expression patterns of bone marrow derived macrophages from WT and NF-IL6-/- mice that were untreated or were simultaneously activated with IFN-γ and infected with L. monocytogenes ("LM + IFN-γ"). Total RNA was isolated from untreated and "LM + IFN-γ" samples at 4 hours post-infection from 4 repeat infection experiments using TriReagent (Molecular Research Company, Cincinnati, USA). The total RNA was DNAseI treated and cleaned up using the RNeasy Mini Kit (Qiagen, Valencia, CA, USA) as per the manufacturer’s instructions. The mRNA was linearly amplified and labelled using the Amino Allyl MessageAmp™ II CyDye aRNA Amplification (Ambion, Austin, Texas, USA) as per the manufacturer’s instructions. The labelled cDNA was purified and hybridized to Mouse Exonic Evidence Based Oligonucleotide (Illumina, San Diego, CA, USA) microarrays printed at the Capar Microarray Facility (University of Cape Town, South Africa). For each infection experiment, labelled probes from the untreated and "LM + IFN-γ" samples for the same mouse group was hybridized to the same microarray. Image and data analysis was done using Limma (46) and TIGR MeV (47). Data was normalized using print-tip loess and quantile normalization. DE genes were identified using a paired t-test with unequal variance, pairing the relative gene expression ratio of "LM + IFN-γ" vs. untreated macrophages from WT and NF-IL6-/- from the same experiment. Top 10% of genes with the smallest p-values were selected further analysis (Table 2) and were functionally clustered.

Due to budget limitations, biological replicates were done instead of technical repeats and a “balanced dye” approach used instead of dye-swaps to compensate for dye-labelling bias. “Dye balance” in this context specifies that each condition is measured equally often with the Cy3 dye as with the Cy5 dye. Therefore, the "LM + IFN-γ" samples for each biological repeat was labelled with the opposite Cy-dye than in the previous biological repeat. Table 1 lists the "dye balance" labelling and hybridization reactions.The same principle was applied to “untreated” controls. There were 4 biological repeat experiments which were analysed over 8 individual arrays. None of the samples from the biological repeat experiments were pooled. Each biological infection experiment was performed independently of each other.

first: Reto

last: Guler

first: Antony

last: Cutler

first: Berenice

last: Arendse

first: Cathal

last: Seioghe

middle: G

last: Horsnell

first: Alexandra

last: Flemming

first: Michael

last: Leitges

first: Andreas

last: Kottmann

first: Gregory

last: Ryan

first: Winston

last: Hide

first: Frank

last: Brombacher


title: GAF competence

city: Nijmegen

first: Peter

last: Burghout

address: Kapittelweg 29

department: Laboratory of Pediatric Infectious Diseases

country: Netherlands

institute: Radboud University Nijmegen Medical Centre

summary:
We applied a novel negative selection strategy called genomic array footprinting (GAF) to identify genes required for genetic transformation of the gram-positive bacterium Streptococcus pneumoniae. Genome-wide mariner-transposon mutant libraries in S. pneumoniae strain R6 were challenged by transformation with an antibiotic resistance cassette and growth in the presence of the corresponding antibiotic. The GAF screen identified the enrichment of mutants in two genes, i.e., hexA and hexB, and the counter-selection of mutants in 21 different genes during the challenge. Eight of the counter-selected genes were known to be essential for pneumococcal transformation. Four other genes, i.e., radA, comGF, parB, and spr2011, have previously been linked to the competence regulon, and one, spr2014, was located adjacent to the essential competence gene comFA. Directed mutants in which seven of the eight remaining genes, i.e., spr0459/spr0460, spr0777, spr0838, spr1259/spr1260, and spr1357, were deleted, displayed reduced, albeit modest, transformation rates. No connection to pneumococcal transformation could be made for the eighth gene, which encodes the response regulator RR03. We further demonstrated that the gene encoding the putative DNA repair protein RadA is required for efficient transformation of chromosomal markers, whereas transformation with replicating plasmid DNA was not significantly affected. The radA mutant also displayed an increased sensitivity to treatment with the DNA-damaging agent methyl methanesulfonate. Hence, RadA is considered to have a role in recombination of donor DNA and DNA damage repair in S. pneumoniae.
Keywords: GAF competence

overall_design:
Selection for genes essential for transformation.
Aliquots of a marinerT7 transposon library generated in S. pneumoniae R6 containing approximately 40,000 independent mutants were transformed with an antibiotic resistance marker and grown without (initial library) or with (challenged library) antibiotics for 25 generation until DNA extraction.

first: Hester

last: Bootsma

first: Tomas

last: Kloosterman

first: Jetta

last: Bijlsma

first: Christa

last: de Jongh

first: Oscar

middle: P

last: Kuipers

last: Hermans


title: The effect of heat stress on the expression profile of potato tuber periderm genes

status: Public on Sep 01 2007

city: Rockville

first: Jia

last: Liu

address: 9712 Medical Center Drive

institute: Plant Genomics

summary:
The goal of the current research is to identify factors that involved with heat induced russeting of the potato tuber skin. Potato plants of the variety Desirèe were grown in pots filled with perlite, in a greenhouse under natural winter conditions (Nov 2005- Jan 2006, average temperatures range of 10-18°C). For the exposure of tubers to heat stress (H) hot water (33-35°C) was circulated in tubes lined at the internal side of the pots. The heat was applied one week before tubers harvest. Tubers were harvested at two time points: 8 weeks post sprouting and a week post mechanical vine killing. The skin (S) of young tubers was peeled by hand, as the remaining phelloderm (PH) layers, the periderm of young tubers (P) and the periderm of mature tubers (ST) were peeled using a scalpel blade. Leaves (L) and tuber flesh (TF) samples were collected as well. For each RNA sample 4 biological replicates were prepared; each one represents pooled tissues from 4-6 plants grown at different location in the greenhouse. RNA was extracted using CTAB protocol, and was further purified by RNeasy Mini Kit (Qiagen) using the On-Column DNase Digestion protocol.
Keywords: Loop design

overall_design: 28 hybs total

first: Ginzberg

last: Idit


title: Microarray analysis of ischemic lesions of WT and TNFRI KO mice

city: Athens

first: Era

last: Taoufik

state: Attiki

address: 127 Vas. Sofias Avenue

country: Greece

institute: Hellenic Pasteur Institute

summary:
TNF/TNFRI signaling has previously been shown to exert neuroprotective effects after brain ischemic injury. However, the detailed pathways involved remain unidentified. This study aims to identify key neuroprotective molecules activated by TNFRI after permanent ischemic injury in mice. Microarray analysis of the ipsilateral hemispheres of WT and TNFRI knockout at early time points (3 and 6 hours) after the middle cerebral artery occlusion mice were chosen to study the CNS response to injury without significant immune infiltration. Using the ATLAS mouse cDNA arrays that include known genes representative of multiple pathways, we identify novel protective pathways triggered by TNFRI signaling after ischemia.
Keywords: brain, ischemic lesion, comparison between WT and TNFRI knockout mice, cDNA array, two different time points (3 and 6 h)

overall_design: RNA was extracted from WT (n=2) and TNFRI KO (n=2) mice at each time point studied (3 and 6 h after ischemia). For each individual sample two hybridisations were performed.

middle: L

last: Roberts

first: Vivian

last: Tseveleki

first: Samuel

last: Valable

first: Didier

last: Divoux

first: Manuela

last: Mengozzi

first: Pietro

last: Ghezzi

last: Brines

first: Anthony

last: Cerami

first: Lesley

last: Probert


title: Gene expression profiles in the peel and pericarp of pepper fruit

summary:
The quality of the pepper fruit is significantly influenced by the properties of its surface such as color, glossiness and texture. The fruit surface is composed of a peel containing several layers including the cuticle, epidermis and the hypodermis. The peel acts as a protective barrier against biotic and abiotic stresses and is the most critical tissue affecting water loss during post harvest storage. The peel is composed of an outer epidermis with thick waxy (lipid) cuticle and few cell layers of thick-walled hypodermal cells. Despite its agronomic importance and due to the fact that the majority of studies in fruits have been conducted using flesh and peel tissues as a whole, the biochemical and genetic bases of variation in peel properties are largely unknown. In this proposal we aim to determine peel-specific gene expression in pepper by micro array hybridizations of peel and flesh RNA extracted at different developmental stages of the fruit.  The cultivar Celica (Capsicum annuum) that has a large blocky fruit will be used for studying gene expression in the peel and flesh. Plants were grown in the greenhouse during the spring of 2006. Fruits were harvested at three developmental stages: young- 10 days after anthesis, mature green- 30 days after anthesis and ripe red- 45 days after anthesis. These stages were chosen because each represents a distinct phase in fruit development. At each stage, a biological replicate consists of bulked tissue from 3 fruits from each of 3 plants (a total of 9 fruits). We have a total of 4 biological replicates. For each fruit, the peel was separated from the flesh by manual dissection using thin forceps and scalpel blade. Peel and flesh samples were immediately frozen in liquid nitrogen and stored at -800C until RNA extraction. Total RNA was extracted using the GenElute Mammalian Total RNA Miniprep kit (Sigma).
Keywords: Reference design

overall_design: 12 hybs total

first: Paran

last: Ilan


title: Functional genomic analysis of cold acclimation in the genus Solanum

summary:
Healthy five-weeks old S. tuberosum (Red Pontiac) and S. commersonii (Oka 5040) were subjected to cold-acclimating temperature (2oC) in the growth chamber. Control plants at identical developmental stage were also grown in growth chamber at optimum conditions for growth (28oC, 14/10, light/dark, 65% RH). Leaf tissues were collected from the control (reference) and cold-acclimating (experimental treatment, 2oC) plants for each species at 2, 4, 7, 10 and 14 days after the initiation of the control and low temperature treatments. A total of two plants for each species were used for both the control and treatment experiments and tissue samples from each plant (within replicate) were pooled for RNA isolation. These experiments were performed twice in order to produce two sets of biological replicates for each species for all sampling time-points.
Keywords: Reference design

overall_design: 17 hybs total

last: Benildo


title: Rat Brain-Heart-Liver Control Mix Self-Self Hybridization

first: Egle

last: Juskeviciute

address: 1020 Locust st. JAH 527

department: Pathology

institute: Thomas Jefferson University

summary:
Eight “self-self” hybridizations were prepared as follows. Total RNA was isolated from each of three rat livers and brains, and equal mass amounts of each of these six samples were combined to create a mixed RNA population. Cy3- and Cy5-labeled cDNA was generated from this RNA mixture using an aminoallyl nucleoside analog indirect labeling protocol. Specifically, eight separate 20 microgram aliquots of the total RNA mixture were reverse transcribed into cDNA in the presence of aminoallyl-dUTP and the resulting cDNA was then coupled to either Cy3 or Cy5 mono NHS ester (four reactions with each label).
Keywords: Self-Self Hybridization

overall_design: Two Color Experiment Dye-Swap Design with  identical biological sample and 4 technical replicates.


title: Comparative expression analysis between sigE (TTHB211) deletion mutant and wild-type of T. thermophilus HB8

status: Public on Sep 04 2007

city: Hyogo

first: Akeo

last: Shinkai

address: 1-1-1, Kouto, Sayo

department: SPring-8 Center

institute: RIKEN Harima Institute

summary:
We observed the expression profile of the total mRNA of sigE (TTHB211) deletion mutant of Thermus thermophilus HB8 strain grown in a rich (TT) medium at 70oC compared with that of wild type.
Keywords: cell type comparison

overall_design: Three wild-type and mutant T. thermophilus HB8 strains were each pre-cultured at 70oC for 16 h in 3 ml of TT medium containing 0.8% polypeptone, 0.4% yeast extract, 0.2% NaCl, 0.4 mM CaCl2, and 0.4 mM MgCl2, which was adjusted to pH 7.2 with NaOH.  The cells (2 ml) were inoculated into 1 liter of the same medium and then cultivated at 70oC.  Cells were collected after 8 h, and then crude RNA was extracted.  In order to compare mRNA expression in the sigE (TTHB211) deletion mutant with that of wild type, the expression of each mRNA at the time point was analyzed on a GeneChip as described under Sample Description Sheet of each sample in this website.

first: Aiko

last: Kashihara

first: Seiki

last: Kuramitsu

first: Shigeyuki

last: Yokoyama


title: Gene expression profiles of presenilin deficient DP thymocytes

city: Cologne

first: Bernhard

last: Gerstmayer

address: Stockheimer Weg 1

institute: Miltenyi Biotec GmbH

summary:
Notch1 signaling is an important regulator of cell fate in  early stages of T cell development.  Because Notch receptors can function redundantly, we sought an approach for inhibiting all endogenous Notch signaling in thymocytes.  Upon ligand engagement, Notch receptors undergo two successive proteolytic cleavages, the second involving a Presenilin (PS) containing complex with -secretase activity that releases a small intracellular fragment of Notch.  This activated form of Notch (NotchIC) translocates from the membrane to the nucleus, where  it acts as a transcription factor, inducing the expression of multiple target genes .  Since Presenilins are required for the activation of all four mammalian forms of Notch, we generated mice with deletions of both Presenilin1 and Presenilin2 genes, the only genes encoding Presenilin in the mouse genome.  To target Notch inactivation specifically to developing T cells, we introduced Cd4-Cre to mediate Presenilin gene deletion in a tissue- and stage-specific manner.  Direct target genes of Notch signaling are largely unknown, and are likely to be cell lineage and stage specific.  Therefore in order to identify potential Notch target genes , we compared RNA of thymocytes at the CD4+CD8+ stage of development from H-2b transgenic 5CC7 TCR/ RAG2-deficient / Presenilin1flox/flox /PS2-/- mice, with and without Cd4-Cre.
Keywords: presenilin, thymocytes, mouse

overall_design: Thymocyte single cell suspensions were generated using 100 µm nylon mesh (PGC Scientifics). DP thymocytes were isolated from RAG2° control or PS1/2° 5CC7 TCR H2bb mice by magnetic bead separation on anti-CD4 coated microbeads (Miltenyi Biotec.). Total RNA was isolated using TRIzol according to manufacturer's instructions (Invitrogen). After quantification and checking for integrity, 1 µg of RNA was subjected to linear amplification. The resulting aRNA was labeled by reverse transcription via Cy5-dCTP incorporation and Cy3-dCTP incorporation. The fluorescent labeled probes were subsequently combined and hybridized on topic-defined PIQORTM Immunology Microarrays Mouse Antisense.

first: K

last: Laky

first: B

last: Fowlkes


title: Effect of TIF on gene expression under inductive and non-inductive photoperiods

first: Jia

last: Liu

summary:
The primary objective of this project is to determine the physiological and developmental differences between photoperiod and root induced tuberization responses in potato using genomic techniques. These experiments will establish baseline genomic and proteomic response associated with a partially characterized root-produced compound (TIF) that either substitutes for, or over-rides, the primary leaf perceived photoperiod induction signal in potato. The proposed work will answer the following questions. Are photoperiod and root induced tuberization responses the same? Do photoperiod and root induced tuberization trigger the same developmental processes in the stolon? How are photoperiod and root induced signals expressed in the genome of the leaf, stolon and roots? The data from the proposed experiments will prove invaluable in identifying key pathways in specific tissues to target in subsequent experiments in order to determine identify of, and establish the mode of action of the TIF compound.
Keywords: Direct comparison

overall_design: 35 hybs total

first: Stutte

last: Gary


title: Synergistic Toxicity by Cadmium and Thiuram

city: Tsukuba

first: Emiko

last: Kitagawa

state: Ibaraki

address: Onogawa 16-1

institute: National Institute of Advanced Industrial Science and Technology

summary:
Bioassay is a system for monitoring toxicity of chemicals in the environment via the biological responses of experimental organisms.  These responses can be detected by analysis of genome-wide changes in mRNA expression levels using DNA microarray.  We applied this system for evaluation of synergistic toxicity by cadmium and thiuram, as this combination showed mutual growth inhibition in yeast.  Hierarchical cluster analysis using the mRNA expression profiles suggested the response of yeast to this combination is similar to that seen with cadmium treatment.  Functional characterization of induced genes by this combination treatment also suggests the enhanced toxicity of cadmium.  This toxicity was observed as the damage to mitochondrial functions which were not observed with either cadmium or thiuram treatments alone.  The potential toxicity to mitochondria by this combinational treatment was confirmed as the result of mitochondrial curing.  We could evaluate the synergistic toxicity by cadmium and thiuram and show the possible use of transcriptome bioassay for synergistic toxicity.
Keywords: stress response

overall_design: This series containes 3 treatment conditions with triplicate experiments from independent biological samples, respectively.

first: Hitoshi

last: Iwahashi

first: Emi

middle: I

last: Kojima

first: Yuko

last: Momose


title: Divergence of Transcription Factor Binding Sites in Related Yeast Species

city: Urrbrae

first: Anthony

last: Borneman

state: South Australia

address: Paratoo Rd

institute: The Australian Wine Research Institute

summary:
A fundamental problem in biology is the molecular basis for divergence among related organisms. We have investigated the level of divergence of transcription factor binding sites for two key factors that regulate developmental processes in the budding yeasts. The genomic binding locations for the Ste12 and Tec1 transcription factors in S. cerevisiae, S. mikatae and S. bayanus were mapped by chromatin immunoprecipitation combined with microarrays (chIP chip)1, 2 and compared to one another. While there was a large core network which was conserved in all three species, there were many instances of binding events whose relative levels differ significantly quantitatively in one species relative to another and as well as species-specific binding events. One interesting class of genes were identified that were bound only in S. mikatae and S. bayanus; many of these genes are targets of Ste12 in haploid strains of S. cerevisiae, suggesting that S. cerevisiae has uniquely acquired the ability to differentially regulate these genes in haploid and diploid cells in these species.  To extend these studies, the transcriptional network for the Ste12 homologue (Cph1) in Candida albicans was also mapped and compared to the Saccharomyces species. Again, there were several genes bound by Cph1 which are involved in mating in S. cerevisiae, suggesting that the precise delineation between many mating and pseudohyphal targets by Ste12 may be specific to S. cerevisiae. Overall our results demonstrate that transcription binding sites differ faster than gene content indicating that gene regulation at the level of transcription factor binding is likely to be a major mode of evolutionary divergence between related species.  We expect that this divergence is essential for the distinct ecological niches inhabited by these organisms.
Keywords: chIP-chip

sample_organism:
Saccharomyces bayanus
Saccharomyces cerevisiae
Candida albicans
Saccharomyces mikatae

overall_design: ChIP-chip was performed on Ste12 and Tec1 from S. cerevisiae, S. mikatae and S. bayanus in addition to Cph1 from S. cerevisiae. Three biological replicates were performed for each factor in each species with one replicate representing a dye swap.

first: Tara

last: Gianoulis

first: Zhengdong

middle: D

last: Zhang

first: Joel

last: Rozowsky

first: Michael

last: Seringhaus

first: Haiyuan

last: Yu

first: LuYang

last: Wong

first: Mark

last: Gerstein

last: Snyder


title: Mapping Nucleosome positions in WT and delta isw2 cells

city: Seattle

first: Toshio

last: Tsukiyama

address: 1100 Fairview Ave N

institute: FHCRC

summary:
To map nucleosome positions in WT and delta isw2 cells
Keywords: Nucleosomal DNA hybridization

overall_design: All samples are biological replicates. Nucleosome positions were defined by comparing the hybridization signal of full length (FL) nucleosomal DNA to that of fragmented (Frag) nucleosomal DNA.

first: Iestyn

last: Whitehouse