title: Identification of differentially regulated genes in Populus deltoides subjected to copper stress using 4.6K cDNA arrays

city: Talca

first: Fernando

last: Guerra

state: Talca

address: 2 Norte 685

department: Instituto de Biologia Vegetal y Biotecnologia

country: Chile

institute: Universidad de Talca

summary: cDNA macroarray expression profiling was carried out in poplar roots in order to identify genes regulated in response to exposition to copper stress. For this purpose, plants of a Populus deltoides clone grown in a hydroponic system during four  weeks were incubated in a nutrient solution (Hoagland's modiefied salt, ¼ strength) supplemented with copper (0 µM (control), 30 µM and 60 µM). Roots were sampled at 12 and 24 h after exposition in a time-course experiment.

overall_design: Three biological replicates named Rep1, Rep2 and Rep3 were obtained for each copper treatment (i.e., control, Cu30 and Cu60) at each time point (i.e., 12, 24). At each time point, two comparisons were performed: control treatment versus Cu30 and control treatment versus Cu60. For each comparison, biological triplicates were used to identify transcripts differentially regulated at the considered timepoint using a statistical analysis approach based on linear mixed models by the SAS system.

first: Sébastien

last: Duplessis

first: Annegret

last: Kohler

first: Francis

last: Martin

first: Jaime

last: Tapia

first: Pablo

last: Lebed

first: Francisco

last: Zamudio

first: Enrique

last: González


title: Heterosis-associated gene expression in maize (Zea mays L.) embryo and endosperm 6 days after pollination

status: Public on Aug 28 2009

city: Hamburg

first: Stefan

last: Scholten

address: Ohnhorststrasse 18

department: Biocenter Klein Flottbek

country: Germany

institute: University of Hamburg

summary: We demonstrated the manifestation of heterosis in hybrid maize embryo and endosperm tissue six days after fertilization in crosses of several inbred lines. Here we analyzed heterosis-associated gene expression pattern in these tissues of reciprocal crosses of two european maize inbred line combinations. Differences in gene expression were analyzed with custom microarrays by a combined approach of suppression subtractive hybridization and microarray hybridizations

overall_design: Analysis used isolated embryos and endosperm samples for a total of 48 microarray hybridizations of hybrid genotypes generated by reciprocal crosses of UH301/UH005 and UH250/UH301 and the corresponding inbred lines. The investigations are based on two biological replicates of the UH250/UH301 genotype combination and four biological replicates of the UH301/UH005 genotype combination. The hybridization experiments employed a loop design.

first: Stephanie

last: Meyer

first: Barbara

last: Sarholz

first: Alexander

last: Thiemann

first: Vera

last: Kühr

first: José

middle: F

last: Gutiérrez-Marcos

first: Hartwig

middle: H

last: Geiger

first: Hans-Peter

last: Piepho


title: The adult boar testicular and epididymal transcriptomes

status: Public on Sep 01 2009

city: NOUZILLY

first: JEAN-LUC

last: GATTI

address: Unité Mixte de Recherche :  INRA - CNRS (UMR 6175) -  Université François Rabelais de Tours - Haras nationaux

department: PHASE

country: France

institute: INRA

summary:
Background - Mammalians gamete production takes place in the testis but when they exit this organ, although spermatozoa have a specialized and distinct morphology, they are immotile and infertile. It is only after their travel in the epididymis that sperm acquire their motility and fertility. Epididymis can be divided in three gross morphological regions, head (caput), body (corpus) and tail (cauda), containing a long and unique convoluted tubule connecting the testis to the vas deferens. Results - In this study, the testis, the efferent ducts (vas efferens, VE), nine distinct successive epididymal segments and the deferent duct (vas deferens, VD) of four adult boars of known fertility were isolated and their mRNA extracted. The gene expression of each of these samples was analyzed using a pig generic 9K nylon microarray (AGENAE program; GEO accession number: GPL3729) spotted with 8931 clones derived from normalized cDNA banks from different pig tissues including testis and epididymis. Differentially expressed transcripts were obtained with moderated t-tests and F-tests and two data clustering algorithms based either on partitioning around medoïds (top down PAM) or hierarchical clustering (bottom up HCL) were combined for class discovery and gene expression analysis. Tissue samples analysis defined seven transcriptomic units: testis, vas efferens and five epididymal transcriptomic units. Meanwhile transcripts formed only four clusters related to the tissues. We have then used a specific statistical method to sort out genes specifically overexpressed (markers) in testis, VE or in each of the five transcriptomic units of the epididymis (including VD). Among these markers some well-known epididymal genes were retrieved while some were new genes or genes not yet reported in these boar tissues. The specific regional expression of some of these genes was further validated by PCR and Q-PCR. We also searched for specific pathways and functions using available gene ontology information. Conclusions - This study fulfilled the gap between those done in rodents and human, and provides tools that will be useful for further studies on the biochemical processes responsible for the formation and maintain of the epididymal regionalization and the development of a fertile spermatozoa.

Keywords: tissue type comparaison

overall_design: 96 samples - 12 tissue samples from 4 boars

first: Benoît

last: GUYONNET

first: Guillemette

last: MAROT

first: Jean-Louis

last: DACHEUX

first: Marie-José

last: MERCAT

first: Sandrine

last: SCHWOB

first: Florence

last: JAFFREZIC

first: Jean-Luc


title: Tiling microarray-based identification of small regulatory RNAs in the group A Streptococcus (GAS)

status: Public on Aug 25 2009

city: Reno

first: Paul

last: Sumby

state: NV

address: 1554 N. Virginia St.

department: Microbiology & Immunology

country: USA

institute: University of Nevada

type: Expression profiling by genome tiling array

type: Non-coding RNA profiling by genome tiling array

summary: Small regulatory RNAs (sRNAs) represent a major mechanism of virulence regulation in several pathogens.  To date, only three sRNAs have been described in GAS in any detail.  To identify whether sRNAs are common within the GAS genome, we created a custom Affymetrix tiling microarray in which all intergenic regions (>49 bp) of the MGAS2221 genome were tiled at high density.  Triplicate cultures of strain MGAS2221 in THY broth were grown to the exponential phase (O.D.600 of 0.5) of growth and RNA was isolated, converted to cDNA, fragmented, labeled, and hybridized to our custom array (one array per RNA sample [ hence three arrays total]).  Our data identified the presence of 40 candidate sRNAs within the MGAS2221 genome.

overall_design:
Three cultures of MGAS2221 were grown in THY broth to the exponential (O.D. 0.5) phase of growth.  Two volumes of RNA protect were added to the samples, the samples incubated at room temperature for 5 minutes, and the bacteria collected through centrifugation.  Total RNA was isolated via a mechanical disruption method, converted to cDNA, fragmented, labeled, and hybridized to our Affymetrix microarray.  Data was analyzed using tiling analysis software to combine the data from the three arrays and generate signal intensities.

The supplementary files 'GSE17789_2221sRNA_signal.bar' and 'GSE17789_2221sRNA_pvalue.bar' contain the signals and p-values, respectively, for Samples GSM444010-GSM444012. The BAR files are generated by TAS software using PM/MM signal intensities.  Signal intensity data is reported in a linear scale; the p-value is reported in -10log10 scale.  Probe analysis used a bandwidth of 40.  Interval analysis used a threshold of 0, a maximum gap of 20, and a minimum run of 60.


title: Gene Expression Data from Human Mesenchymal Stem Cells with and without TGF-B vs Human Annulus Disc Cells

city: Charlotte

first: Helen

last: Gruber

state: NC

address: 1542 Graden Terrace

department: Orthopaedic Surgery

institute: Carolinas HealthCare System

summary:
Autologous stem cell therapy has potential for biologic treatment of disc degeneration.  Due to ease of harvest and abundance, adipose-derived mesenchymal stem cells (AD-MSC) are readily available.  Our objectives were: 1) To develop/validate methods to harvest AD-MSC and direct them to a disc-like phenotype by three-dimensional (3D) culture and TGF-ß3 exposure; 2) To perform gene expression profiling for human AD-MSC and annulus cells in 3D culture; 3) To test whether disc cell-AD-MSC co-culture could augment proteoglycan production.
Stem cell plasticity offers potential for future biologic therapies for disc degeneration.  Data indicated that human AD-MSC can successfully be manipulated in 3D culture to express gene products important in the disc ECM (types I and II collagen, chondroitin sulfate, keratin sulfate, decorin), and that co-culture of annulus cells with AD-MSC enhances proteoglycan production.  Studies defined gene expression patterns of AD-MSC and human annulus cells in 3D culture, important as we explore the potential of MSC in biologic therapies for disc degeneration.

overall_design: AD-MSC were extracted from human adipose tissue, and characterized as stem cells using accepted criteria (direction into osteoblasts or chondrocytes; and cell surface marker criteria).  Three AD-MSC cultures were grown in 3D with or without TGF-ß3 for 2-3 weeks. Disc Tissue samples were obtained from surgical disc procedures performed on patients with herniated discs and degenerative disc disease. Seven disc cultures were also grown in 3-D for 2 weeks.  RNA was harvested according to instructions with the Trizol isolation method, checked for quality using the 2100 Bioanalyzer (Agilent Technologies, Inc., Santa Clara, CA USA), reverse-transcribed to double-stranded cDNA, subjected to two rounds of transcription, and hybridized to the DNA microarray in the Affymetrix Fluidics Station 400. Affymetrix human U133 X3P arrays were used. Using Genesifter, gene expression in AD-MSC with and witout TGF-B3 was compared to annulus cells. Gene expression of stem cells with TGF-ß3 was also compared to stem cells grown in the absence of TGF-ß3.

middle: E

first: Ray

last: Deepe

first: Gretchen

middle: L

last: Hoelscher

first: Jane

middle: A

last: Ingram

first: H

middle: J

last: Norton

first: Brian

last: Scannell

first: Bryan

last: Loeffler

first: Natalia

last: Zinchenko

first: Edward

middle: N

last: Hanley Jr

first: Hazel

last: Tapp


title: MicroRNA expression in rat soleus muscle during atrophy induced by hindlimb suspension

city: Lexington

first: Esther

middle: E.

last: Dupont-Versteegden

state: KY

address: 900 S. Limestone

department: Rehabilitation Sciences

institute: University of Kentucky

summary: The objective of the study was to identify microRNAs expressed in the rat soleus muscle and determine if their expression was changed in response to hindlimb suspension.

platform_organism:
Rattus norvegicus
synthetic construct

overall_design: The microRNA expression profile (Sanger miRBase 9.0) of the rat soleus muscle was determined in the following three groups: control (C), hindlimb suspended for 2 days (HS-2) or 7 days (HS-7).  Three replicates were performed for each group.

first: John

last: McCarthy

first: Karyn

last: Esser

first: A

last: Peterson


title: Transcriptome comparison of Streptococcus pyogenes strain MGAS2221 with that of an isogenic PEL mutant

summary: PEL is a small regulatory RNA that is present within the S. pyogenes genome.  In some strains of S. pyogenes, the PEL sRNA regulates many of the known virulence factors produced by this pathogen.  A genome-wide analysis of PEL-regulated genes had not been performed previously so we analyzed this by comparing the parental M1T1 isolate MGAS2221 with an isogenic PEL mutant.  Comparing gene transcript levels between these two strains at the exponential and stationary phases of growth, we identified that PEL has no regulatory function in MGAS2221.

overall_design: Three cultures of each GAS strain were grown in THY broth to exponential (O.D. 0.5) and stationary (O.D. 1.7) phases of growth.  Two volumes of RNA protect were added to samples recovered at these O.D.'s, the samples incubated at room temperature for 5 minutes, and the bacteria collected through centrifugation.  Total RNA was isolated via a mechanical disruption method, converted to cDNA, fragmented, labeled, and hybridized to our Affymetrix microarray.  Estimates of gene expression were calculated using GCOS software v1.4.


title: Regulation of dendritic cells and macrophages by an anti-apoptotic cell natural Ab that inhibits inflammatory arthritis

city: La Jolla

first: Gregg

last: Silverman

state: CA

address: 9500 Gilman Dr

department: Medicine

institute: UC San Diego School of Medicine

summary:
Although natural antibodies (NAbs) are present from birth, little is known about what drives their selection, and whether they have housekeeping functions. We now show that the prototypic T15-NAb, first identified because of its protective role in infection, is representative of a previously unknown type of NAb response that specifically recognizes and forms complexes with apoptotic cells, and which promotes cell-corpse engulfment by phagocytes. This T15-NAb-mediated process is dependent on the recruitment of C1q and mannose-binding lectin (MBL), which have known immune modulatory activities that also provide “eat me” signals for phagocytic clearance. Further investigation revealed that, the addition of T15-NAb significantly suppressed in vitro macrophage LPS-induced TNF-alpha and IL-6 secretion, as well as in vitro Toll-like receptor (TLR)-induced dendritic cell maturation and secretion of pro-inflammatory cytokines and chemokines. Significantly, high doses of this B-1 cell produced NAb also inhibited in vivo TLR–induced pro-inflammatory responses, and could suppress autoimmune inflammatory arthritis. These studies identify and characterize a previously unknown regulatory circuit by which a NAb product of innate-like B cells aids homeostasis by control of fundamental inflammatory pathways.

Keywords: Dendritic cells, Macrophages, Natural Antibodies, Apoptosis, Inflammation, Arthritis

sample_organism:
Mus musculus
unidentified

platform_organism:
Homo sapiens
Mus musculus

overall_design: In the study presented here, a total of 15 custom-spotted protein slides were hybridized with a T15 IgM monoclonal antibody at three different concentrations, an isotype control IgM, 2 slides were hybridized with serum from naїve mice, 2 slides incubated with serum from mice immunized with apoptotic cells, 2 slides were incubated with serum from mice immunized with saline, 2 slides were incubated with serum from mice injected with T15 IgM monoclonal antibody, 2 slides were incubated with serum from a lupus-prone mouse disease model and a negative control slide with no antibody hybridized

first: Yifang

last: Chen

first: Sahil

last: Khanna

first: Carl

middle: S

last: Goodyear

first: Yong-Beom

last: Park

first: Eyal

last: Raz

first: Steffen

last: Thiel

first: David

last: Boyle

first: Maripat

last: Corr

first: Dwight

last: Kono


title: Renal gene expression analysis: transcript-level data

city: Edinburgh

first: Donald

last: Dunbar

address: 47 Little France Crescent

department: Centre for Cardiovascular Science

country: United Kingdom

institute: University of Edinburgh

summary: This is a SubSeries of GSE12672

overall_design: 48 samples: 4 strains (F, L, FL, LF) x 3 time points (0, 7, 10 day induction) x 4 biological replicates each


title: Transcriptome analysis of Escherichia coli K-12 and ada mutant

city: Bethesda

first: Jonghwan

last: Baek

state: MD

address: 37 Convent Dr

department: NCI

institute: NIH

summary: Alkylation damage to DNA occurs when cells encounter alkylating agents in the environment or when active alkylators are generated by nitrosation of amino acids in metabolic pathways. To cope with DNA alkylation damage, cells have evolved genes that encode proteins with alkylation-specific DNA repair activities. It is notable that these repair systems are conserved from bacteria to humans.  In Escherichia coli, cells exposed to a low concentration of an alkylating agent, such as N-methyl-N’-nitro-N-nitrosoguanidine (MNNG) or methyl methanesulfonate (MMS), show a remarkable increase in resistance to both the lethal and mutagenic effects of subsequent high-level challenge treatments with the same or other alkylating agents. This increased resistance has been known as “adaptive response” to alkylation damage in DNA. To date, four genes have been identified as components of this response, ada, alkA, alkB and aidB. The ada gene encodes the Ada protein, which has the dual function of a transcriptional regulator for the genes involved in the adaptive response, and a methyltransferase that demethylates two methylated bases (O6meG and O4meT) and methylphosphotriesters produced by methylating agents in the sugar phosphate backbone. The differences between the wild-type and mutant strains were characterized at transcriptome levels. In addition, the global changes in gene expressions in response to alkylating agents (MMS), in E. coli K-12 W3110 and ada mutant strains were also analyzed. The analysis of time- and strain-dependent adaptive responses revealed the regulatory and physiological characteristics of the Ada-dependent adaptive response in E. coli.

overall_design: In order to examine the intracellular changes that are induced by the ada gene deletion in the MMS-untreated, normal growth condition, the expression levels of genes of ada mutant cells were compared with those of wild-type cells at the mid-log growth phase (at 0.5 h sampling point). Cells were cultivated at 37oC and 250 rpm in 100 mL of Luria-Bertani (LB) medium (10 g/L tryptone, 5 g/L yeast extract, and 5 g/L NaCl) in 250-mL Erlenmeyer flasks. Transcriptome analysis were also performed for the samples (E. coli wildtype and ada mutant strains) taken at 0.5, 1.5 and 3.9 h following MMS treatment for both MMS-treated and -untreated control cultures, and the expression levels were compared.

first: JongHwan

first: SangYup

last: Lee


title: Novel endogenous antisense transcripts (data for the mouse mammary tumors)

status: Public on Aug 24 2009

city: Tsukuba

first: Koji

last: Numata

address: 3-1-1 Koyadai

country: Japan

institute: RIKEN

summary: Recent transcriptomic analyses have uncovered widespread occurrence of natural antisense transcripts (NATs), which are transcribed from the opposite strand of another discrete transcribed region. We describe a novel probe-design technique for DNA microarray analysis that specifically targets the complementary strand of annotated genes, and show that this technique can successfully identify novel NAT expression in normal and tumor samples. Northern and in situ hybridization analyses of selected examples (Acaa1, Aard, and Thbd) confirmed their transcription and dynamic expression in a tissue- and cell-type-specific manner. Our data highlight the importance of NAT expression in the regulation of cellular processes and in oncogenesis. The fundamental basis of the technology described in this paper is applicable to all genes and sample types for the identification of NAT expression that is not possible by conventional cDNA/EST/CAGE information.

overall_design:
Total RNA for the GRS mammary tumor microarray experiments was isolated from normal and cancerous mammary glands of dissected GRS/A mice, by using Trizol reagent (Invitrogen Corporation).

Unpublished data has been masked in the supplementary feature extraction files.

first: Yuko

last: Osada

first: Yuki

last: Okada

first: Rintaro

last: Saito

first: Noriko

last: Hiraiwa

first: Hajime

last: Nakaoka

first: Naoyuki

last: Yamamoto

first: Kazufumi

last: Watanabe

first: Kazue

last: Okubo

first: Akio

last: Kanai

first: Kuniya

last: Abe

first: Hidenori

last: Kiyosawa


title: microRNA profiles of meningiomas

city: Charlestown

first: Okay

last: Saydam

address: Building 149, 13th st. 6th Floor, Room:6101

department: Neurology

institute: MGH

summary: Meningiomas, one of the most common human brain tumors, are derived from arachnoidal cells associated with brain meninges, usually benign and frequently associated with neurofibromatosis type 2. Here we define a human meningioma-typical miRNA profile and characterize effects of one down-regulated miRNA, miR-200a, on tumor growth.  Elevated levels of miR-200a inhibited meningioma cell growth in culture and in a tumor model in vivo.  Up-regulation of miR-200a decreased expression of transcription factors, ZEB1 and SIP1, with consequently increased expression of E-cadherin, an adhesion protein associated with cell differentiation. Down-regulation of miR-200a in meningiomas and arachnoidal cells resulted in increased expression of β -catenin and cyclin D1 involved in cell proliferation. miR-200a was found to directly target β -catenin mRNA, thereby inhibiting its translation and blocking β -catenin-Wnt signaling, frequently involved in cancer. A direct correlation was found between down-regulation of miR-200a and up-regulation of β-catenin in human meningioma samples. Thus, miR-200a appears to act as a multi-functional tumor suppressor miRNA in meningiomas through effects on the E-cadherin and β -catenin-Wnt signaling pathways. This reveals a previously unrecognized signaling cascade involved in meningioma tumor development and highlights a novel molecular interaction between miR-200a and Wnt signaling, thereby providing insights into novel therapies for meningiomas.

platform_organism:
Homo sapiens
Mus musculus
Rattus norvegicus

overall_design:
14 meningioma tumor samples were compared to 3 arachnoidal tissue control samples. Two technical replicates were performed for each sample. Normalization and processing included combining the data from technical replicates.

The below table contains quantile-normalized data.


title: Analysis of Spt16 depletion

city: Burjassot

first: Jose

last: Perez-Ortin

state: Valencia

address: Dr. Moliner 50

department: Bioquimica y Biologia Molecular

country: Spain

institute: Universitat de Valencia

summary:
Transcription rate analysis in Yeast by means of GRO, mRNA amount and ChIP-on-chip during the depletion of Spt16p.

Keywords: Genomic run-on GRO ChIP-chip

overall_design: Transcription rate analysis by means of GRO and mRNA amount (RA)  of three independent replicates during the depletion of Spt16p (Control and 5 & 7 hours after the depletion). Each time point replicate has been hybridized on a different macroarray (F14-F16). ChIP-on-chip analysis of Spt16 were done during exponential grow in YPD.

first: Silvia

last: Jimeno-Gonzalez

first: Vicent

last: Pelechano

last: Garcia-Martinez

first: Sebastian

last: Chavez


title: Mycobacterium tuberculosis TraSH experiment: Growth in wild type C57BL/6J versus iNOS-/- mice

status: Public on Aug 20 2009

first: Jeffrey

middle: P.

last: Murry

address: 10010 N. Torrey Pines Road

department: IMPL-Y

institute: Salk Institute

summary: A M. tuberculosis transposon library was used to infect WT and iNOS-/- mice.  Surviving mutants were recovered from spleens, genomic DNA was extracted, and labeled probes were synthesized from transposon ends.  Probes from each WT mouse were hybridized with probes from a similar iNOS-/- mouse.

overall_design: Two-condition experiment, Growth in WT vs. iNOS-/- mice.  Biological replicates: TraSH probe made from 5 wild type and 5 iNOS-/- mice after 3 weeks of infection and from 8 wild type and 8 iNOS-/- mice after 4 weeks of infection.  Technical replicates: TraSH probe was synthesized twice from each mouse and dyes were swapped.  One array contains probe from one iNOS-/- and one WT mouse.  Each biological replicate has two arrays, representing technical replicate dye swaps from one iNOS-/- and one WT mouse.

middle: P

first: Amit

middle: K

last: Pandey

first: Christopher

middle: M

last: Sassetti

first: Eric

last: Rubin


title: Novel endogenous antisense transcripts (data for the mouse normal tissues)

first: Koji

last: Numata

overall_design:
Total RNA for the mouse (C57BL/6J) microarray experiments was isolated from SL10 cells (fibroblast cell line), brain, heart, intestine, kidney, liver, lung, placenta (d.p.c. 10.5 and 13.5), spleen, stomach, testis, and thymus. Testis was from C57BL/6J males (8 to 10 weeks), placenta was from pregnant mice, and the other tissue was from both male and female mice.

Unpublished data has been masked in the supplementary feature extraction files.

first: Yuko

last: Osada

first: Yuki

last: Okada

first: Rintaro

last: Saito

first: Noriko

last: Hiraiwa

last: Nakaoka


title: Analysis of Mouse Dendritic and Antigen presenting cell and Mouse autoimmune and Inflammatory Response

status: Public on Aug 19 2009

city: Dallas

first: Kiyoshi

last: Ariizumi

state: TX

address: 5323 Harry Hines Blvd

department: Dermatology

institute: UT Southwestern Medical Center

summary: Antigen presenting cells (APC) express receptors recognizing microbes and regulating immune responses by binding to corresponding ligands on immune cells.  Having discovered a novel inhibitory pathway triggered by ligation of DC-HIL on APC to a heparin/heparan sulfate-like saccharide of syndecan-4 on activated T cells, we posited DC-HIL can recognize microbial pathogens in a similar manner.  We showed soluble recombinant DC-HIL to bind the dermatophytes Trichophyton rubrum and Microsporum audouinii, but not several bacteria nor Candida albicans.  Dermatophyte binding was inhibited completely by the addition of heparin.  Because DC-HIL contains an immunoreceptor tyrosine-based activation motif (ITAM)-like intracellular sequence, we questioned whether its binding to dermatophytes can induce tyrosine phosphorylation in dendritic cells (DC).  Culturing DC with T. rubrum (but not with C. albicans pseudohyphae) induced phosphorylation of DC-HIL, but not when the tyrosine residue of the ITAM-like sequence was mutated to phenylalanine.  To examine the functional significance of such signaling on DC, we cross-linked DC-HIL with mAb (surrogate ligand), which not only induced tyrosine phosphorylation but also upregulated expression of 23 genes among 662 genes analyzed by gene-array, including genes for profilin-1, Marcksl-1, C/EBP, LOX-1, IL-beta and TNF-alpha.  This cross-linking also upregulated expression of the activation markers CD80/CD86 and heightened APC capacity of DC to activate syngeneic T cells.  Our findings support a dual role for DC-HIL; inhibition of adaptive immunity following ligation of syndecan-4 on activated T cells, and induction of innate immunity against dermatophytic fungi.

overall_design: Gene expression analysis in mouse dendritic cells following stimulation of DC-HIL receptor

first: Jin-Sung

last: Chung


title: Microarray analysis of stalk-eyed fly lines selected for divergent eyespan

status: Public on Aug 22 2009

city: College Park

first: Gerald

last: Wilkinson

address: Bldg 144, 4094 Campus Dr

department: Biology

institute: University of Maryland

summary: Eye-stalks of some flies exhibit extreme sexual dimorphism, but little is known about the genetic mechanisms producing variation in these ornamental traits. Therefore, we constructed an EST database of genes expressed in the developing eye-antennal imaginal disc of the highly dimorphic species, Teleopsis dalmanni, and used this set of genes to construct high density oligoarrays and compare patterns of gene expression between lines of flies selected for divergent eyespan. Microarray experiments using eye-antennal disc tissue identified over 350 genes with significant differential expression between male flies from lines selected for high and low relative eyespan but did not reveal any primary biological process or pathway that is driving the expression differences.

overall_design: Custom 4x44K Agilent arrays were designed for approximately 3600 unique genes.  Each gene was represented by three 60 bp oligo probes and each probe was printed in triplicate on each array.  Each oligoarray was hybridized with two biological samples labeled either with cy3 or cy5.  Each sample came from 25 sets of eye-antennal imaginal discs that were dissected from gut-purged male larvae.  Samples were from lines of flies that had been under directional selection on males to either increase or decrease relative eyespan for 65 generations.  Dye labeling was switched between lines for every other array.  Ratios of median intensities were averaged across probes for each gene and then analyzed by SAM to identify genes with differential expression between lines.

first: Richard

last: Baker

last: Boore

first: Xianhui

last: Wang


title: Comparative Genomic Analysis of PhoP Regulon in S. Typhi and S. Typhimurium

city: Boston

first: Richelle

middle: C.

last: Charles

state: MA

address: 55 Fruit Street, Gray 504

department: Infectious Diseases

institute: Massachussetts General Hospital

summary: Transcriptional profiling of three sequenced S. enterica strains: S. Typhimurium LT2, S. Typhi CT18, and S. Typhi Ty2 in PhoP-inducing and non-inducing conditions in vitro, and compared these results to profiles of phoP-/Q- mutants derived from S. Typhimurium LT2 and S. Typhi Ty2.

sample_organism: Salmonella enterica subsp. enterica serovar Typhimurium str. LT2

sample_organism: Salmonella enterica subsp. enterica serovar Typhi str. Ty2

sample_organism: Salmonella enterica subsp. enterica serovar Typhi str. CT18

overall_design: Two-condition experiment:  Each strain above was grown in PhoP-inducing (Low Magnesium concentration) and PhoP non-inducing conditions (High Magnesium concentration) with 1 dye reversal.

first: Jason

middle: B

last: Harris

first: Alaullah

last: Sheikh

first: Lauren

last: Lebrun

first: Abdullah

last: Tarique

first: Regina

last: LaRocque

first: Tanya

last: Logvinenko

first: Firdausi

last: Qadri

first: Stephen

last: Calderwood

first: Edward

middle: T

last: Ryan


title: Inflammatory gene expression in response to sub-lethal ricin exposure in Balb/c mice

city: Salisbury

first: Jonathan

last: David

address: Porton Down

department: Biomedical Sciences

institute: Dstl

summary: The toxin ricin has been shown to cause inflammatory lung damage, leading to pulmonary oedema and, at higher doses, mortality. In order to understand the genetic basis of this inflammatory cascade a custom microarray platform (1509 genes) directed towards immune and inflammatory markers was used to investigate the temporal expression profiles of genes in a Balb/c mouse model of inhalational ricin exposure. To facilitate examination of those genes involved in both inflammatory cascades and wound repair the dose which was investigated was sub-lethal across a 96 hour time course. Histopathology of the lung was mapped across the time course and genetic responses considered in the context of overall lung pathology. 685 genes were found to be statistically significantly different compared to controls, across the time course and these genes have been investigated in the context of their biological function in ricin poisoning. As well as confirming key inflammatory markers associated with ricin intoxication (TNFa and Il1b) several pathways that are altered in expression were identified following pulmonary exposure to ricin. These genes included those involved in cytokine-cytokine signalling cascades (IL1, IL1r, IL1r2, Ccl 4, 6, 10), focal adhesion (Fn1, ICAM1) and tissue remodelling (VEGF, Pim1). Furthermore, the observed alteration in expression of matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) indicates a key role in membrane integrity and cellular adhesion in ricin poisoning. Data captured using this transcriptomic approach could be used to develop a specific approach to the treatment of inhalational ricin exposure. This work was conducted as part of a wider programme of work to compare a number of militarily relevant lung damaging agents, with a view to establishing a rational basis for the identification of more generic medical countermeasures.

overall_design: Four groups of six mice were exposed to ricin and a further groups exposed to vehicle only control (PBS). Following exposure mice were culled at 6 timepoints (1, 4, 7, 24, 48 and 96h). RNA was extracted and run on the custom array.

first: Lucy

first: Gareth

middle: D

last: Griffiths


title: The transcriptomic response of E. coli to triclosan exposure

city: Birmingham

first: Chrystala

last: Constantinidou

department: School of Biosciences

institute: University of Birmingham

summary:
The gene expression profile of E. coli K-12 MG1655 grown in minimal medium treated with 0.12 mg/L of the biocide triclosan has been analysed using whole genome oligonucleotide microarrays. "Control" RNA was isolated from three independently grown 50ml MOPS minimal media cultures of E. coli K-12 MG1655. “Test” RNA was isolated from three independently grown 50ml MOPS minimal cultures of E. coli K-12 MG1655, to which was added 0.12 mg/L of triclosan after reaching mid-logarithmic growth phase (OD600 ~ 0.7 +/- 0.02).
Keywords: dose response

overall_design: Cy5-labelled cDNA probes were synthesised from three independent biological extractions of RNA from E. coli MG1655 unexposed to triclosan. Cy3-labelled cDNA probes were synthesised from three independent biological extractions of RNA from E. coli MG1655 exposed to 0.12 mg/L of triclosan. A dye-swap control was also included, whereupon the control cDNA probes were labelled with Cy3 and the test cDNA probes labelled with Cy5. Probes were mixed to allow comparison of the control and test expression profiles and hybridised onto an E. coli spotted oligonucleotide array.

first: Andrew

last: Bailey

last: Constantinodou

first: Al

last: Ivens

first: Mark

middle: I

last: Garvey

last: Webber

first: Nick

last: Coldham

first: Jon

last: Hobman

first: John

last: Wain

first: Martin

last: Woodward

first: Laura

last: Piddock


title: A genome-wide expression comparison of productive and unproductive airway epithelial repair

status: Public on Aug 18 2009

city: Durham

first: Barry

middle: R

last: Stripp

address: 106 Research Drive, DUMC Box 103000

institute: Duke University

summary:
Defective epithelial repair in the setting of chronic lung disease has been suggested to contribute to uncontrolled extracellular matrix (ECM) deposition and development of fibrosis. We sought to directly test this hypothesis through gene expression profiling of total lung RNA isolated from mouse models of selective epithelial cell injury that are associated with either productive or abortive repair. Analysis of gene expression in repairing lungs of naphthalene-exposed mice revealed prominent clusters of up-regulated genes with putative roles in regulation of the extracellular matrix and cellular proliferation. Further analysis of tenascin C (Tnc), a representative matrix protein, in total lung RNA revealed a transient 4.5-fold increase in mRNA abundance 1 day after injury and a return to steady-state levels by Recovery Day 3. Tnc was deposited by the peribronchiolar mesenchyme immediately after injury and was remodeled to basement membrane subtending the bronchiolar epithelium during epithelial repair. Epithelial restitution was accompanied by a decrease in Tnc mRNA and protein expression to steady-state levels. In contrast, abortive repair using a transgenic model allowing ablation of all reparative cells led to a progressive increase in Tnc mRNA within lung tissue and accumulation of its gene product within the subepithelial mesenchyme of both conducting airways and alveoli. These data demonstrate that the ECM is dynamically remodeled in response to selective epithelial cell injury and that this process is activated without resolution in the setting of defective airway epithelial repair.
Bronchiolar Clara cells undergo phenotypic changes during development and in disease. These changes are poorly described due to a paucity of molecular markers. We used chemical and transgenic approaches to ablate Clara cells, allowing identification of their unique gene expression profile. Flavin monooxygenase 3 (Fmo3), paraoxonase 1 (Pon1), aldehyde oxidase 3 (Aox3), and claudin 10 (Cldn10) were identified as novel Clara cell markers. New and existing Clara cell marker genes were categorized into three classes based on their unique developmental expression pattern. Cldn10 was uniformly expressed in the epithelium at Embryonic Day (E)14.5 and became restricted to secretory cells at E18.5. This transition was defined by induction of Clara Cell Secretory Protein (CCSP). Maturation of secretory cells was associated with progressive increases in the expression of Fmo3, Pon1, Aox3, and Cyp2f2 between late embryonic and postnatal periods. Messenger RNA abundance of all categories of genes was dramatically decreased after naphthalene-induced airway injury, and displayed a sequence of temporal induction during repair that suggested sequential secretory cell maturation. We have defined a broader repertoire of Clara cell-specific genes that allows staging of epithelial maturation during development and repair.

overall_design: 32 samples were isolated and used to screen UniSet Mouse 20K I Bioarrays (total lung RNA Mus musculus). Control RNA was isolated from uninjured mice (N = 4 mice), and RNA was also isolated from mice exposed to naphthalene and recovered for 1, 2, 3, and 6 days (N = 4 mice per timepoint). Additionally, transgenic mice expressing HSV-tk under control of the mouse CCSP promoter were exposed to Ganciclovir (GCV) and recovered for 3, 6, and 9 days (N = 4 mice per timepoint).

first: Joshua

middle: C

last: Snyder

first: Anna

last: Zemke


title: Gallus gallus bursal FAE and IFE

city: Tsuruoka

first: Shinji

last: Fukuda

state: Yamagata

address: 246-2 Mizukami, Kakuganji

department: Institute for Advanced Biosciences

institute: Keio University

summary: A minor population of M cells within the follicle-associated epithelium (FAE) of intestinal Peyer’s patches (PP) serves as a major portal for entry of exogenous antigens. Characterization of the mammalian M cells, including identification of M-cell surface molecules used for bacterial uptake, has been hampered by their relative rarity.  In contrast, M cells constitute virtually all of the FAE cells in the avian bursa of Fabricius.  We therefore performed comparative gene expression profiling of chicken and murine FAE to identify commonly expressed genes by M cells in both species. The comprehensive transcriptome analysis revealed that 28 genes were commonly up-regulated in FAE from both species. In situ hybridization (ISH) revealed that annexin A10 (Anxa10) mRNA was scattered in FAE, and co-localized with Ulex europaeus agglutinin-1(UEA-1) that binds to M cells. Whole-mount immunostaining also revealed that cellular prion protein (PrPC) was expressed on the luminal side of the apical plasma membrane of M cells, and co-localized with grycoprotein2 (GP2) that recognizes only M cells in murine PP. Taken together, we found new M-cell-specific molecules by using comprehensive transcriptome analysis. These molecules conserved in M cells from both species might play critical roles in M-cell function and/or differentiation.

overall_design: Bursa of Fabricius were dissected from the rectum and soaked in Hank's Buffered Salt Solution (HBSS) (GIBCO) containing black ink for 5 min at room temperature to visualize FAE. After incubation, the tissue was treated with 40 mM EDTA/25 mM Hepes (pH 7.4) in HBSS for 5min at room temperature to collect bursal FAE or 30 min at 37ºC for bursal IFE. Bursal FAE and IFE were collected with a fine 29G needle under stereomicroscope monitoring (n=2)

first: Gaku

last: Nakato

first: Koji

last: Hase

first: Hiroshi

last: Ohno


title: UV treated Escherichia coli reveals a high number of differentially expressed regions and many novel transcripts

city: Oslo

first: Gard

last: Thomassen

state: Oslo

address: Rikshospitalet University Hospital

department: Molecular microbiology

country: Norway

institute: Centre for Molecular Biology and Neuroscience (CMBN) and Institute of Medical Microbiology

summary:
The transcriptional changes in Escherichia coli upon induction of the SOS response are investigated by utilizing custom designed oligonucleotide microarrays.

Keywords: Gene expression during the SOS response in Escherichia coli

sample_organism: Escherichia coli str. K-12 substr. MG1655

overall_design:
Escherichia coli K-12 MG1655 single colony in five parallells was grown to mid-log phase and exposed to UV to induce the SOS response. Total RNA was extracted from induced and uninduced cells and cDNA was prepared, fragmented and labelled prior to hybridizing to arrays.
The arrays was designed to maximize the genomic coverage whilst simultaneous including only probes estimated to give an approximately uniform binding affinity. Regions coding for non-hypothetical proteins or RNAs where covered less densely than the intergenic parts.

middle: O

first: Ragnhild

last: Weel-Sneve

first: James

last: Booth

first: Jessica

last: Lindvall

first: Karin

last: Lagesen

first: Knut

last: Kristiansen

first: Magnar

last: Bjørås

first: Torbjørn

last: Rognes

first: Alexander

last: Rowe


title: Bioactive Pyridine-N-oxide Disulfides from Allium stipitatum

status: Public on Aug 15 2009

city: Rockville

first: Helena

last: Boshoff

address: 12441 Parklawn Drive

department: LIG

institute: NIAID/ NIH

summary:
In this study, the researchers isolated a colorless waxy solid from the chloroform extract of Allium stipitatum as Compound 1. They demonstrated compound 1 is a potent bactericidal agent against nonreplicating Mycobacterium tuberculosis. Using microarray technology the authors report gene expression profiles in cells treated with either 2, 5, or 10 ug/ml of compound 1 or an equivalent amount of DMSO as control for 6 h. Gene expression studies revealed that transcriptional profiles elicited in response to compound 1 were similar to the profiles generated during treatment of cells with compounds such as menadione and 8-quinolinol that result in oxidative stress. They included the thioredoxin system components encoded by trxB2 and trxC as well as several genes associated with the heat shock response such as clpB, sigH, dnaJ, dnaK, hsp, Rv0331, Rv3463, Rv3054c, and Rv1334-Rv1335. These results suggest that compound 1 possibly generates damaged proteins and other oxidative stress signals as part of its mechanism of action.   The following is the full abstract of this published study.  O'Donnell G, et al. (2009) J Nat Prod 72(3):360-365  From Allium stipitatum, three pyridine-N-oxide alkaloids (1-3) possessing disulfide functional groups were isolated. The structures of these natural products were elucidated by spectroscopic means as 2-(methyldithio)pyridine-N-oxide (1), 2-[(methylthiomethyl)dithio]pyridine-N-oxide (2), and 2,2'-dithio-bis-pyridine-N-oxide (3). The proposed structure of 1 was confirmed by synthetic S-methylthiolation of commercial 2-thiopyridine-N-oxide. Compounds 1 and 2 are new natural products, and 3 is reported for the first time from an Allium species. All compounds were evaluated for activity against fast-growing species of Mycobacterium, methicillin-resistant Staphylococcus aureus, and a multidrug-resistant (MDR) variants of S. aureus. Compounds 1 and 2 exhibited minimum inhibitory concentrations (MICs) of 0.5-8 mug/ml against these strains. A small series of analogues of 1 were synthesized in an attempt to optimize antibacterial activity, although the natural product had the most potent in vitro activity. In a whole-cell assay at 30 mug/ml, 1 was shown to give complete inhibition of the incorporation of (14)C-labeled acetate into soluble fatty acids, indicating that it is potentially an inhibitor of fatty acid biosynthesis. In a human cancer cell line antiproliferative assay, 1 and 2 displayed IC(50) values ranging from 0.3 to 1.8 muM with a selectivity index of 2.3 when compared to a human somatic cell line. Compound 1 was evaluated in a microarray analysis that indicated a similar mode of action to menadione and 8-quinolinol by interfering with the thioredoxin system and up-regulating the production of various heat shock proteins. This compound was also assessed in a mouse model for in vivo toxicity.
A dose response design type examines the relationship between the size of the administered dose and the extent of the response of the organism(s).
Compound Based Treatment: Allium stipitatum (garlic) extract


title: EFTF vs GFP Experiment

city: Syracuse

first: Michael

middle: E

last: Zuber

state: NY

address: 766 Irving Ave

department: Ophthalmology

institute: SUNY Upstate Medical University

summary:
Xenopus laevis embryos were injected with mRNA for EFTFs at 2-cell stage. Animal caps collected at stage 9, cultured to the equivalent of stage 15 and RNA extracted. Four biological replicates of the EFTF-injected and GFP-injected (control) caps were used to profile transcript expression patterns using Affymetrix Xenopus Laevis GeneChip microarrays.
Keywords: Two way comparison

first: Andrea

last: Viczian

first: Yung

last: Lyou


title: Transcriptional analysis of Leishmania infantum antimony resistant strains using full-genome DNA microarrays

city: Québec

first: Philippe

last: Leprohon

state: Québec

address: 2705 boulevard Laurier

department: Centre hospitalier de l'Université Laval

country: Canada

institute: Université Laval

summary:
The aim of the experiment is to detect any gene expression modulation in an antimony resistant L.infantum Sb2000.1 strain compared to an antimony sensitive L.infantum WT strain. We want to identify the modulated genes associated with an antimony resistant phenotype. The Sb2000.1 resistant strain used is about 1000 times more resistant to SbIII than the WT sensitive strain.
The full-genome DNA microarrays includes one 70-oligonucleotide probe for each gene of L. infantum.

Keywords: drug resistance

overall_design: Two-condition experiment, resistant strain Sb2000.1 vs wild-type. Five biological replicates for each strain, independently grown and harvested. One replicate per array


title: Genomic DNA 1:1 mixtures in Allele Specific Expression study

city: Chicago

first: Justin

last: Borevitz

state: IL

department: Ecology and Evolution

institute: University of Chicago

type: Genome variation profiling by genome tiling array

summary:
Genomic DNA of Col-0 and Van-0 were mixed at 1:1 and labeled using Bioprime randon labeling kit. A total of 16ug labelled products were hybridized to AtSNPtile1.
Used as the reference of equal allele expression.

overall_design: Col and Van genomic DNA were mixed at 1:1 with four biological replicates. Single chanel. Total 4 samples.

first: Xu

last: Zhang


title: mRNA samples in Allele Specific Expression study

summary:
Col-0 and Van-0 total RNA were mixed at 1:1. Total RNA also extracted from F1 hybrid samples from Col x Van and Van x Col. PolyA RNA were purified and double-strand cDNA were synthesized, followed by bioprime random labeling. A total of 16ug labelled products were hybridized to AtSNPtile1.
cis regulatory effect was identified as allele specific expression in F1 hybrids. Composite trans regulatory effect was detected as deviation between parental expression and F1 hybrids expression.

overall_design: Each sample type has four replicates. Single chanel. Three sample types (Col and Van RNA 1:1 mixture, Col x Van F1 hybrids, Van x Col F1 hybrids). Total 12 samples.


title: Effect of sulfide on growth physiology and gene expression of Desulfovibrio vulgaris Hildenborough

city: Calgary

first: Sean

middle: Michael

last: Caffrey

state: Alberta

address: 2500 University Dr NW

department: Biological Scineces

institute: University of Calgary

summary: The effect of sulfide stress on Desulfovibrio vulgaris Hildenborough (DvH) gene expression was determined by comparing the gene expression profiles of DvH  under conditions in which sulfide was allowed to accumulate (high sulfide, average concentration 10 mM) against DvH cells grown under conditions in which sulfide was removed by continuous gassing (low sulfide, average concentration 1 mM). High sulfide significantly decreased the instantaneous growth rate constant and final cell density of the culture indicating a decreased bioenergetic fitness. Changes in gene expression caused by exposure to high sulfide were determined using full-genome DvH microarrays. The transcription of ribosomal protein-encoding genes was decreased, in agreement with the lower growth rate of DvH under high sulfide conditions. Interestingly, expression of the gene for DsrD, located downstream of the genes for dissimilatory sulfite reductase (DsrAB) was also strongly down-regulated. In contrast, the expression of many genes involved in iron accumulation, stress response and proteolysis, and chemotaxis were increased. This indicates that high sulfide represents a significant stress condition, in which the bioavailability of metals like iron may be lowered and in which proteins (e.g. metalloenzymes) may need to be refolded, or proteolytically degraded. Overall this leads to a reduced growth rate and less efficient biomass production with available resources.

sample_organism: Nitratidesulfovibrio vulgaris str. Hildenborough

overall_design: For each condition 2 unique biological samples were hybridized to 4 arrays that each contained duplicate spots. Genomic DNA was used as universal reference.

first: Gerrit

last: Voordouw


title: miRNA arrays for LPS-activated Akt1+/+ and Akt1-/- macrophages

status: Public on Aug 21 2009

city: Heraklion

first: Christos

last: Tsatsanis

state: Crete

address: Voutes

department: School of Medicine

country: Greece

institute: University of Crete

summary: MicroRNAs regulated by lipopolysaccharide (LPS) target genes that contribute to the inflammatory phenotype. Here we showed that the protein kinase Akt1, which is activated by LPS, positively regulated miRNAs let-7e, miR-181c but negatively regulated miR-155 and miR-125b. In silico analyses and transfection studies revealed that let-7e repressed Toll-like receptor 4 (TLR4) whereas miR-155 repressed SOCS1, two proteins critical for LPS-driven TLR signalling, which regulate endotoxin sensitivity and tolerance. As a result, Akt1-/- macrophages exhibited increased responsiveness to LPS in culture and Akt1-/- mice did not develop endotoxin tolerance in vivo. Overexpression of let-7e and suppression of miR-155 in Akt1-/- macrophages restored sensitivity and tolerance to LPS in culture and in animals. These results indicate that Akt1 regulates the response of macrophages to LPS by controlling miRNA expression. The data deposited here contain the entire analysis of miRNA profile of Akt1+/+ and Akt1-/- thioglycollate elicited peritoneal macrophages following stimulation with LPS for 3 hours in culture.

overall_design: Thioglycollate elicited macrophages were cultured in complete DMEM medium, stimulated with LPS for 3 hours and RNA was extracted. Samples were analyzed using Taq-man PCR miRNA arrays (Dana Farber microarray Facility).

first: D

last: Iliopoulos

first: P

middle: N

last: Tsichlis

first: A

last: Androulidaki

first: C


title: ChIP-on-chip from acute myeloid leukemia cell lines and clinical samples for H3K4me3, H3K27me3, and EZH2

city: Bethesda

first: Thomas

middle: A

last: Paul

address: 37 Convent Dr.

department: Laboratory of Cellular Oncology

institute: NCI-CCR

type: Genome binding/occupancy profiling by genome tiling array

summary:
Histone modifcations at the p15INK4b gene were compared in sample with p15INK4b DNA methylation vs. samples with no DNA methylation
AML clinical samples without DNA methylation exhibit bivalent histone modifications at p15INK4b, while clinical samples with DNA methylation display lower H3K4me3 and retain H3K27me3

overall_design: Comparison of AML cell lines and clinical samples with p15INK4b DNA methylation to those free of DNA methylation. AML cell lines KG-1, KG-1a Kasumi-1, AML-193 have p15INK4b DNA methylation. AML patient samples AML6, AML7, AML8 have p15INK4b DNA methylation.

first: Linda

last: Wolff


title: Age and albumin D site-binding protein control tissue plasminogen activator levels: neurotoxic impact.

city: Caen

first: Benoit

middle: Denis

last: Roussel

address: Bd Becquerel

department: GIP Cyceron

institute: INSERM

summary: Recombinant tissue-type plasminogen activator (tPA) is the fibrinolytic drug of choice to treat stroke patients. However, a growing body of evidence indicates that besides its beneficial thrombolytic role, tPA can also have a deleterious effect on the ischaemic brain. Although ageing influences stroke incidence, complications and outcome, age-dependent relationships between endogenous tPA and stroke injuries have not been investigated yet. Here, we report that ageing is associated with a selective lowering of brain tPA expression in the murine brain. Moreover, our results show that albumin D site-binding protein (DBP) as a key age-associated regulator of the neuronal transcription of tPA. Additionally, inhibition of DBP-mediated tPA expression confers in vitro neuroprotection. Accordingly, reduced levels of tPA in old mice are associated with smaller excitotoxic/ischaemic injuries and protection of the permeability of the neurovascular unit during cerebral ischaemia. Likewise, we provide neuroradiological evidence indicating the existence of an inverse relationship between age and the volume of the ischaemic lesion in patients with acute ischaemic stroke. Together, these results indicate that the relationship among DBP, tPA and ageing play an important role in the outcome of cerebral ischaemia.

overall_design:
For each stage (4 months and 21 months), samples were prepared by pooling an equal amount of the 3 individual RNA. Five µg of total RNA from each pool were reverse-transcribed in the presence of 7.5µM random hexamers (GE Healthcare), 75µM aminoallyl-dUTP (Sigma-Aldrich) and 100U Rtases Reverse-iT Blend (Thermo scientific) overnight at 37°C. Aminoallyl cDNAs were then labelled with Cy5 or Cy3 mono-Reactive Dye (GE Healthcare) according to the manusfacturer’s instructions. Labelled cDNAs were then purified on Qiaquick PCR prurification kit (Qiagen) and hybridized to the RNG-MRC_MM25k_EVRY microarrays (Le brigand et al., NAR, 2006) according to the RNG procedures (http://www.microarray.fr:8080/merge/index). Each pool was hybridized in duplicate dye-swap independent experiments.
After hybridization, median signal and median background intensities were extracted using Genepix 4.1 software (Axon Instruments). We have first evaluated the level of expression of each gene by calculating the mean ratio “signal/background” for each corresponding spot from the 2 dye-swap experiments. Genes were considered to be expressed if their mean ratio was higher or equal to 1.2. Genes with mean ratio < 1.2 in the 2 conditions were not used for further analysis. These filtered data were then submitted to VARAN (http//:www. bionet. espci.fr) for normalization by lowess fit and differential expression analysis (Golfier et al., 2004). Normalized log2ratio 21 months/4 months were then used for further statistical analysis by SAM software (Tusher et al., 2001) and t-test with a p-value equal to 5% using TIGR Multiexperiment Viewer (TM4:MeV, http://www.tm4.org/mev.html). The final list of differentially expressed genes was established by comparing the results obtained with the 3 methods.

first: Richard

last: Macrez

first: Amandine

last: Jullienne

first: Veronique

last: Agin

first: Eric

last: Maubert

first: Luce

last: Dauphinot

first: Marie-Claude

last: Potier

first: Laurent

last: Plawinski

first: Herve

last: Castel

first: Yannick

last: Hommet

first: Josep

last: Munuera

first: Joan

last: Montaner

first: Manuel

last: Yepes

first: Carine

last: Ali

first: Denis

last: Vivien


title: Genomic DNA concentration series in Allele Specific Expression study

first: Justin

last: Borevitz

summary:
Genomic DNA of COl-0 and Van-0 were labeled using Bioprime randon labeling kit. Labelled products were hybridized to AtSNPtile1 at a total amount of 6ug, 9ug, 12ug, 15ug, and 18ug.
This experiment is to test whether the overall target amount affect the log allele intensity ratio.

overall_design:
Each genotype has 5 hybridizations each at 6ug, 9ug, 12ug, 15ug and 18ug.
Single chanel.
Total two genotypes (Col, Van). Total 10 samples.

first: Xu

last: Zhang


title: A sentinel function for teat tissues in dairy cows: dominant innate immune response elements define response to mastitis

city: Beltsville

first: Anthony

middle: V.

last: Capuco

address: 10300 Baltimore Ave., Bldg 200, Rm 14

department: Beltsville Agricultural Research Center

institute: USDA-ARS

summary: Mastitis remains one of the most prevalent and costly diseases impacting the dairy industry worldwide. Escherichia coli is an environmental bacterium that frequently causes intramammary infections, the outcome of which depends on the capacity of the host to recognize and clear the bacterial pathogen. E. coli intramammary infection elicits localized and systemic responses, some of which have been characterized in mammary secretory tissue.  However, nothing is known about early transcriptome-wide responses to infection that occur within regions of the teat and mammary parenchyma.  Therefore, the objective of the current study was to use microarray analysis to characterize gene expression patterns and process networks that become activated in different regions of the mammary gland during the acute phase of experimentally induced intramammary infection with E. coli.  Tissues evaluated were from Furstenberg’s rosette, teat cistern, gland cistern, and lobulo-alveolar regions of control and infected mammary glands, 12 and 24 h after bacterial (or control) infusions.  For selected genes, quantitative RT-PCR was performed to confirm the microarray findings (Toll-like receptors; TLR1, TLR2, TLR4, TLR6) and evaluate the correspondence between mRNA level in tissues and protein level in milk (interleukin 8, tumor necrosis factor-alpha, haptoglobin, lipopolysaccharide-binding protein). The main networks activated by E. coli infection pertained to immune and inflammatory response, with marked induction of genes encoding proteins that function in chemotaxis, leukocyte activation and signaling. Genomic response was greatest in tissues of the teat and gland cisterns at 12 h post-infection, while tissue of the lobulo-alveolar region responded only, and most strongly of the regions, 24 h following the infection. Up regulation of TLR2, TLR4, IL8, TNF-alpha and bactericidal/permeability-increasing protein peaked at 12 h post infection in tissues of the teat and in the gland cistern, while the highest level of lipopolysaccharide-binding protein was reached at 24 h in the lobular alveoli region of infected quarters. Very few genes were down–regulated within the timeframe of this study. Similar genetic networks were impacted in all regions during early phases of E. coli intramammary infection, although regional differences throughout the gland were noted. Importantly, the tissues of the teat, which are first to encounter invading bacteria during a natural infection, responded rapidly and intensely, suggesting an important sentinel function for this region. Both resident mammary cells and infiltrating immune cells likely play an important role in recognition of pathogens and production of inflammatory mediators during mastitis.

overall_design: Three healthy lactating Holstein cows were used in this study.  All mammary glands were bacteria-free.  Immediately following the morning milking, 4 mL of E. coli suspension (100 colony forming units/mL) were infused into one mammary gland of the udder and the contralateral gland was infused with PBS (control gland).  After the PM milking 12 h later, the remaining ipsilateral quarter was infused with E. coli.  Cows were euthanized 24 h after initial infusions and tissues were collected from 4 regions of each mammary gland:  Furstenburg’s rosette, teat cistern, gland cistern and lobuloalveolar regions.  Tissues were snap frozen in liquid nitrogen, stored at -80C until processing.

first: Manuela

last: Rinaldi

first: Robert

middle: W

last: Li

first: Douglas

last: Bannerman

first: Kristy

last: Daniels

first: Christina

last: Clover

first: Marcos

middle: V

last: Silva

first: Max

middle: J

last: Paape

first: Bernadette

last: Van Ryssen

first: Christian

last: Burvenich


title: Eye primordia vs. posterior neural plate vs. lateral endoderm normalized the whole embryos

first: Michael

last: Zuber

summary:
Tissues from the eye primordia, lateral endoderm, and posterior
neural plate of stage 15 Xenopus laevis embryos were isolated and normalized to stage 15 whole embryos. Three biological replicates were prepared for each tissue and the expression patterns were profiled using Affymetrix Xenopus Laevis GeneChip microarrays.
Keywords: Three way comparison of Embryonic Tissue Isolation

first: Andrea

middle: S

last: Viczian

first: Yung

last: Lyou


title: Genomic DNA mixture series in Allele Specific Expression study

summary:
Genomic DNA of Col-0, Van-0 and their mixture at different proportion were labeled using Bioprime randon labeling kit. A total of 16ug labelled products were hybridized to AtSNPtile1.
This experiment is to test whether the log allele intensity ratio efficiently detects the allelic variation within targets.

overall_design: Templates include four Col genomic DNA replicates, four Van genomic DNA replicates, three Col and Van 1:1 mixture replicates, and one of each 1:2, 1:3, 1:5, 2:1, 3:1, 5:1 Col and Van mixture. Single chanel. Total 17 samples.


title: Dlx3 Regulates Lifr and Chrdl1 Expression in Mouse Embryonic Stem Cell Differentiation

city: Los Angeles

first: Nyam-Osor

last: Chimge

address: 2250 Alcazar street

department: IGM

institute: USC

summary:
Dlx3 over-expression in mouse embryonic fibroblasts changed the expression level of numerous genes involved in osteogenesis and embryonic stem cell-related pathways as revealed by microarray analysis.  From the list of Dlx3 modulated genes we focused our attention on the study of two candidates, Lifr and Chrdl1.  Chromatin immunoprecipitation demonstrated the recruitment of Dlx3 transcription factor to the promoters of Lifr and Chrdl1, and luciferase assays confirmed the role of Dlx3 in the regulation of Lifr expression.  Over-expression of Dlx3 in mouse embryonic stem cells stimulated Lifr and Chrdl1 expression and inhibited expression of Id proteins and Bmp4.  We show that Dlx3 increases the expression of both soluble and transmembrane forms of Lifr.  Soluble Lifr may regulate extracellular Lif levels via solution binding, while transmembrane Lifr mediates the signal transduction pathway. The data suggests that Dlx3 may be involved in stem cell differentiation in a dosage dependent way through its interaction with Lifr and with Chrdl1, a known antagonist of Bmp4.  We speculate that Dlx3 may also be involved in osteoblast differentiation through interactions involving Lifr, Bmp, and Id proteins and signaling via the JAK/STAT and MAPK pathways.  In summary, our data suggests that Dlx3 proteins play a significant role in a highly tuned network in early embryogenesis.

Keywords: treated vs.untreated

overall_design: Stable and Transient transfection of Dlx3 in MEF. Total of 4 hybridizations including biological replicates.


title: Combinatorial pharmacological approaches target EZH2-mediated gene repression in breast cancer cells

status: Public on Aug 12 2009

city: SINGAPORE

first: qiang

last: yu

address: BIOPOLIS STREET

country: Singapore

institute: GIS

summary:
Polycomb protein EZH2-mediated gene silencing is implicated in breast tumorigenesis through methylation of histone H3 on Lysine 27 (H3K27). We have previously shown that S-adenosylhomocysteine hydrolase (SAHH) inhibitor 3-Deazaneplanocin A (DZNep) can modulate histone methylation and disrupt EZH2 complex. Here, we used DZNep, together with other chromatin remodeling agents, as well as RNA interference-mediated EZH2 depletion, to probe the role of EZH2 in coordination with other epigenetic components in gene regulation in breast cancer cells. Through genome-wide gene expression analysis, coupled with extensive chromatin immunoprecipitation analysis of histone modifications, we have identified a variety of gene sets that are regulated either by EZH2 alone or through the coordinated action of EZH2 with HDAC and/or DNA methylation. We further found that tumor antigen GAGEs were regulated by distinct epigenetic mechanisms in a cell context-dependent manner, possibly reflecting mechanistic heterogeneity in breast cancer. Intriguingly, we found that EZH2 regulates a remarkable cohort of genes whose functions are highly enriched in immunoresponse and autocrine inflammation network, and their transcriptional activation upon EZH2 perturbation is cancer-specific, revealing a potential novel role of EZH2 in regulating cancer immunity. These findings demonstrate the complexity and diversity of epigenetic regulation in human cancer and underscore the importance for developing combinatorial pharmacologic approaches for effective epigenetic gene reactivation.

Keywords: Total RNA

overall_design: Cells were treated with 2.5 uM DZNep or 5uM AZA for 72 h, and TSA at 100 nM for 24 hours. For AZA treatment, the medium was replaced with freshly added AZA for every 24 h. For co-treatment of cells with DZNep and TSA, DZNep was added for 48 h followed by TSA for an additional 24 h. In total, there were seven different drug treatments (AZA, TSA, DZNep, AZA+TSA, DZNep+AZA, DZNep+TSA, DZNep+AZA+TSA). One group of untreated MCF-7 cells served as the control. Each treatment has 3 technical replicates.

first: Feng

last: Sun

first: Zhenlong

last: Wu

first: Qiang

last: Yu


title: Bemisia tabaci B and Q biotypes thermotolerance

status: Public on Aug 13 2009

city: Bet Dagan

first: Murad

last: Ghanim

address: Derech Hamakabim

department: Entomology

country: Israel

institute: Volcani Center

summary: The whitefly Bemisia tabaci (Gennadius) causes tremendous losses to agriculture by direct feeding on plants and by vectoring several families of plant viruses. The B. tabaci species complex comprises over 10 genetic groups (biotypes) that are well defined by DNA markers and biological characteristics. B and Q are amongst the most dominant and damaging biotypes, differing considerably in fecundity, host range, insecticide resistance, virus vectoriality, and the symbiotic bacteria they harbor. We used a spotted B. tabaci cDNA microarray to compare the expression patterns of 6,000 ESTs of B and Q biotypes under standard 25°C regime and heat stress at 40°C. Overall, the number of genes affected by increasing temperature in the two biotypes was similar. Gene expression under 25ºC normal rearing temperature showed clear differences between the two biotypes: B exhibited higher expression of mitochondrial genes, and lower cytoskeleton, heat-shock and stress-related genes, compared to Q. Exposing B-biotype whiteflies to heat stress was accompanied by rapid alteration of gene expression. For the first time, the results here present differences in gene expression between very closely related and sympatric B. tabaci biotypes, and suggest that these clear-cut differences are due to better adaptation of one biotype over another and might eventually lead to changes in the local and global distribution of both biotypes.

overall_design: 3 replicates comparing Q biotype under 40 degrees celsius (C) with three replicates under 25 C. The same number of replicates comparing B biotype under 40 C and 25 C, and three replicates comparing B and Q biotypes under 25 C.

first: Assaf

last: Mahadav

first: Svetlana

last: Kontsedalov

first: Henryk

last: Czosnek


title: Methylation profiling of differentially expressed regions between C57BL/6 and BALB/c in bone marrow derived macrophages

status: Public on Aug 10 2009

city: Regensburg

first: Elmar

last: Schilling

address: Franz-Josef-Strauss Allee 11

institute: University Hospital Regensburg

type: Methylation profiling by genome tiling array

summary:
DNA-methylation is a vital epigenetic mark that participates in establishing and maintaining chromatin structures and in regulating gene transcription during mammalian development and cellular differentiation. Inter-individual differences in methylation patterns may represent a major source of phenotypic variation, however, the determinants, inheritance, extent, and consequences of such differences are poorly understood. Here we have analysed methylation profiles of immune cells from two inbreed mouse strains (C57BL/6 & BALB/c) that represent prototypic models for Th1- or Th2-dominated immune responses. Using a methyl-CpG immunoprecipitation approach, genomes were fractionated into methylated and unmethylated genome pools and separately analysed at 180 genomic regions (covering 28 Mb of the mouse genome) that were selected based on differential gene expression between both mouse strains. Differentially methylated regions are detected by analyzing array probes for diametrically opposed enrichment behaviour between both hybridizations (e.g. a region that is relatively enriched in the unmethylated pool of BALB/c and shows reverse enrichment behaviour in the methylated pool is considered hypomethylated in BALB/c). The integrated analysis of hypo and hypermethylation profiles allowed the identification of several hundred differentially methylated regions, but also uncovered regions that were duplicated in one strain, contained single nucleotide polymorphisms or micro- and macro-deletions.

Keywords: MCIp-on-Chip; comparative genomic hybridization

overall_design: Methylation profiles of regions, harbouring differentially expressed genes in BMM, were compared between C57BL/6 and BALB/c. Two independently grown, harvested and MCIp treated gDNA preparations were used for each mouse strain (Two biological replicates; One replicate per array). Hypo- and hypermethylated pools serve as diametrically oposed technical replicates (mirror approach).

last: Rehli


title: Kawasaki Disease and Other Febrile Illnesses

city: Stanford

first: Stephen

last: Popper

department: Division of Infectious Diseases, Dept. of Medicine

institute: Stanford University

summary:
ABSTRACT  Background. Acute Kawasaki disease (KD) is difficult to distinguish from other acute rash/fever illnesses, in part because the etiologic agent(s) and pathophysiology remain poorly characterized. As a result, diagnosis and critical therapies may be delayed.  Methods.  We used DNA microarrays to identify possible diagnostic features of KD.  We compared gene expression patterns in the blood of 23 children with acute KD and 18 age-matched febrile children with three illnesses that resemble KD.  Results.  Genes associated with platelet and neutrophil activation were expressed at higher levels in KD patients than in patients with acute adenovirus infections or systemic adverse drug reactions but not in patients with scarlet fever; genes associated with B cell activation were also expressed at higher levels in KD patients than in controls.  A striking absence of interferon-stimulated gene expression in the KD patients was confirmed in an independent cohort of KD subjects.  We successfully predicted the diagnosis in 21 of 23 KD patients and 7 of 8 adenovirus patients using a set of 38 gene transcripts.   Conclusions.  These findings provide insight into the molecular features that distinguish KD from other febrile illnesses, and support the feasibility of developing novel diagnostic reagents for KD based on the host response.
A disease state experiment design type is where the state of some disease such as infection, pathology, syndrome, etc is studied.
Disease State: One of Kawasaki Disease (KD) or control (C) of Scarlet fever (C-sf), adenovirus infection (C-ai) or drug reaction (C-dr)


title: Campylobacter jejuni murine adaptation: unadapted v. adapted

status: Public on Aug 08 2009

city: East Lansing

first: Linda

middle: S.

last: Mansfield

state: MI

address: 181 Food Safety Bldg.

department: Microbiology and Molecular Genetics

institute: Michigan State University

summary: Expression arrays comparing Campylobacter jejuni 11168 before and after serial passage in C57 BL/6 IL-10 deficient mice.  Gene expression was compared during exponential growth in Bolton broth.

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

overall_design: Unpassaged (p0) C. jejuni 11168 was compared to the same strain after 1 passage (p1) and after three passages (p3) through the mice.  Biological replicates: 4 for p0 v. p1, 3 for p0 v. p3

first: John

middle: P

last: Jerome

first: Anne

last: Plovanich-Jones

first: Julia

last: Bell


title: A Saccharomyces cerevisiae strain widely used in bioethanol production

city: Durham

first: Juan Lucas

last: Argueso

state: SP

address: 371 CARL Building, Box 3054

department: Molecular Genetics and Microbiology

institute: Duke University Medical Center

summary: In the search for renewable sources of energy, bioethanol stands out as a benchmark biofuel because its production is based on a proven technological platform.  Bioethanol is produced mainly from the fermentation of carbohydrates derived from agricultural feedstocks by the yeast Saccharomyces cerevisiae.  One of the most widely adopted strains is PE-2, a heterothallic diploid naturally adapted to the sugar cane fermentation process used in Brazil.  Here we report the molecular genetic analysis of a PE-2 derived diploid (JAY270), and the complete genome sequence of a haploid derivative (JAY291).  The JAY270 genome is highly heterozygous (~2 SNPs per kilobase), and has several structural polymorphisms between homologous chromosomes.  These chromosomal rearrangements are confined to the peripheral regions of the chromosomes, and appear to reflect ectopic homologous recombination between repetitive DNA sequences.  Despite the complex karyotype of JAY270, this diploid, when sporulated, had a high frequency of viable spores (~93%).  Crosses of haploids derived from JAY270 to a haploid of the unrelated laboratory strain S288c also resulted in diploids that had good spore viability (75-95%).  Thus, the rearrangements that exist near the ends of chromosomes do not impair meiosis and spore viability, as they do not span regions that contain essential genes.  This observation is consistent with a model in which the peripheral regions of chromosomes represent plastic domains of the genome that are free to recombine ectopically and experiment with alternative structures that may be associated with a fitness benefit.  We also explore features of the JAY270 and JAY291 genomes that help explain their high adaptation to industrial environments, exhibiting desirable phenotypes such as high cell mass production and fermentation kinetics, high temperature growth and oxidative stress tolerance.  The genomic manipulation of such strains could enable the creation a new generation of industrial organisms, ideally suited for use as delivery vehicles for future bioenergy technologies.

overall_design: This microarray experiment was used to compare the relative gene expression levels between two unrelated S. cerevisiae strain backgrounds: JAY270 and JAY309.  Total RNA from each strain was prepared and used to synthesize differentially labeled cDNAs (Cy5 and C3 respectively). A positive Log2 (Red/Green) ratio indicates transcripts more abundant in JAY270, while a negative Log2 (Red/Green) ration indicates transcripts more abundant in JAY309.

first: Juan

middle: L


title: Purple sea urchin larvae alter their transcriptome in response to ocean acidification

status: Public on Aug 14 2009

city: San Francisco

last: Todgham

address: 1600 Holloway Ave

department: Biology

institute: San Francisco State University

summary: In this research we present a transcriptomics analysis of the physiological response of a marine calcifier, Strongylocentrotus purpuratus, to ocean acidification, a decline in ocean pH that results from the absorption of anthropogenic carbon dioxide (CO2). Larvae were raised from fertilization to prism stage in seawater with elevated CO2 conditions based upon IPCC emissions scenario B1 (540ppm CO2) and A1FI (1020ppm CO2).

overall_design:
Adult S. purpuratus were collected using SCUBA from Goleta Pier (Goleta,CA, USA) and maintained in flowing seawater tables at 15-16°C. Spawning was induced by coelomic injection of 0.5M KCl following standard methods (Strathmann 1987). Eggs were collected from 3 females and separately fertilized by sperm from a single male (i.e. all larvae were full or half-siblings). Each of the three replicate cultures were divided in three (nine cultures total) and held in 10L flow-through containers filled with seawater aerated through a side arm with commercially manufactured air containing three different pCO2: 380ppm CO2 (present day atmospheric CO2 level, pH 8.01 ± 0.01), 540ppm CO2, an optimistic atmospheric CO2 concentration predicted by the Intergovernmental Panel on Climate Change for 2100 (B1 scenario, pH 7.96 ± 0.01) and 1020ppm CO2 (A1FI scenario, pH 7.88 ± 0.01). Larvae were cultured at 15°C until prism stage (40h post-fertilization).
At the 40 hour time point, a sample of 60,000 larvae was removed from each of the 9 cultures in the 380, 540 and 1020 ppm treatments and stored in 1 mL Trizol at -80 ºC for later RNA analysis.
The larval cDNA sample from each replicate 540ppm CO2 culture was competitively hybridized against the 380ppm CO2 control culture from the same replicate female. Similarly, the larval cDNA sample from each replicate 1020ppm CO2 culture was competitively hybridized against the 380ppm CO2 control culture from the same replicate female.  Using dye swaps of technical replicates, treatment effects could be estimated independently of dye effects.

first: Gretchen

last: Hofmann


title: Comparison between caruncular and intercaruncular areas in the endometrium of cyclic and pregnant cows

city: Jouy-en-Josas

first: Olivier

last: Sandra

address: Domaine de Vilvert

department: UMR BDR

institute: INRA

summary:
At implantation the endometrium undergoes dramatic modifications necessary for its physical interactions with the trophoblast as well as the development of the conceptus. We aim to identify endometrial factors and pathways essential for a successful implantation in the caruncular [C] and the intercaruncular [IC] areas in cattle. The [C versus IC] direct comparison revealed 1177 and 453 DEG in cyclic and pregnant animals respectively, with a major impact of the conceptus on metabolism and cell adhesion.

Keywords: Microarray fluorescence

overall_design: 10 Samples comparing caruncle and intercaruncle in pregnant cows, 10 Samples comparing caruncle and intercaruncle in cyclic cows.

first: Nadéra

last: Mansouri-Attia

first: Julie

last: Aubert

first: Pierrette

last: Reinaud

first: Corinne

last: Giraud-Delville

first: Géraldine

last: Taghouti

first: Laurent

last: Galio

first: Robin

last: Everts

first: Séverine

last: Degrelle

first: Christophe

last: Richard

first: Isabelle

last: Hue

first: XCindy

last: Tian

first: Harris

last: Lewin

first: Jean-Paul

last: Renard


title: The RpoN Regulon of Escherichia coli O157:H7

city: Tampa

first: James

middle: Timothy

last: Riordan

state: FL

address: 4202 E Fowler Ave BSF 218

department: CMMB

institute: University of South Florida

summary:
There is increasing evidence to support a role for sigma factor 54 (RpoN) in the regulation of stress resistance factors and protein secretion systems important to bacterial transmission and pathogenesis.  In enterohemorrhagic E. coli O157:H7, acid resistance and type III secretion are essential determinants of gastric passage and colonization.  This study thus described the transcriptome of an rpoN null strain of E. coli O157:H7
(EcJR-8) to determine the influence of RpoN on virulence and stress resistance gene regulation, and further explored its contribution to glutamate-dependent acid resistance (GDAR).  Inactivation of rpoN resulted in the growth phase-dependent, differential expression of 104 genes.  This included type III secretion structural and regulatory genes encoded on the locus of enterocyte effacement (LEE), as well as GDAR genes gadA, gadBC and gadE.  Upregulation of gad transcript levels in EcJR-8 during logarithmic growth correlated with increased GDAR and survival in a model stomach.  Acid susceptibility was reconstituted in EcJR-8 complemented in trans with wild-type rpoN.  Acid resistance in EcJR-8 was dependent on exogenous glutamate, gadE and rpoS, but was independent of hns.  Results also suggest that GDAR may be controlled by RpoN at multiple regulatory levels.  This study supports the hypothesis that RpoN is an important regulator of virulence and stress resistance factors in E. coli O157:H7, and is the first to examine the mechanism by which it represses GDAR.

overall_design: Hybridizations measured transcriptional differences between an rpoN null and wild-type (WT) strain of E. coli O157:H7 Sakai at logarithmic and transition phase.  Image files (TIFF) of hybridized microarray slides were generated using an Axon 4000B scanner (Molecular Devices), and analyzed using GenePix Pro software (Molecular Devices, ver. 6.0).  The resulting microarray intensity data was log2-transformed, and normalized using the LOWESS algorithm in MAANOVA ver. 0.98-8 (R ver. 2.2.1).

middle: T

first: Jillian

last: Tietjen

first: Coilin

last: Walsh

first: Thomas

last: Whittam


title: Cycle versus Pregnancy

first: Olivier

last: Sandra

summary:
At implantation the endometrium undergoes dramatic modifications necessary for its physical interactions with the trophoblast as well as the development of the conceptus. We aim to identify endometrial factors and pathways essential for a successful implantation in the caruncular [C] and the intercaruncular [IC] areas in cattle. Using a bovine oligo-array, expression profiles were established at day 20 of the estrous cycle or pregnancy (implantation) showing 446 and 1295 differentially expressed genes (DEG) in [C] and [IC] areas respectively. The impact of the conceptus was higher on the immune response function in [C] but more prominent on the regulation of metabolism function in [IC].

Keywords: Fluorescence Microarray

overall_design: 18 intercaruncular samples, 18 caruncular Samples. Comparisons of cycle (d20) and pregnancy (d20).

first: Nadéra

last: Mansouri-Attia


title: Transforming growth factor beta signaling in hypertensive remodeling of porcine aorta

city: College Station

first: Natasa

last: Popovic

address: 355 Reynolds Medical Bldg.

institute: Texas A&M University

summary: A porcine aortic coarctation model was used to examine regulation of gene expression in early hypertensive vascular remodeling. Aortic segments were collected proximal (high pressure) and distal (low pressure) to the coarctation after 2 weeks of sustained hypertension (mean arterial pressure>150mmHg). Porcine 10K oligoarrays used for gene expression profiling of the two regions of aorta revealed downregulation of cytoskeletal and upregulation of extracellular region genes relative to the whole genome. A genomic database search for transforming growth factor-β (TGFβ) control elements (TCE) showed that 19% of the genes that changed expression due to hypertension contained putative TCEs. Real time PCR and microarray analysis showed no change in expression of TGFβ1, TGFβ2, TGFβ3, or bone morphogenetc proteins (BMP)2 and 4, yet immunohistochemical staining for phosphorylated SMAD2, an indicator of TGFβ signaling, and for phosphorylated SMAD1/5/8, an indicator of signaling through the BMPs, showed the highest percentage of positively stained cells in the proximal aortic segments of occluded animals. For TGFβ signaling, this increase was significantly different from sham-operated controls. Western blot analysis showed no difference in total TGFβ1 protein levels with respect to treatment or aortic segment. Immunohistochemistry showed that the protein levels of latency associated peptide, was decreased in proximal segments of occluded animals. Collectively, these results suggest that activation of TGFβ, but not altered expression, may be a major mechanism regulating early hypertensive vascular remodeling.

overall_design:
Dye-coupled RNA samples isolated from the same animal were paired according to the balanced block experimental design with a complete dye swap.
Quality control was done by combining 3 flagging systems: F635-B635>10; 40% of pixels>B635 +2SD; F635/B635>2.5. If the gene failed only one of these criteria, it was flagged and was not included in further analysis. Same criteria were used to flag measurements in 532 channel.
Filtering: A gene had to have absent flags in at least 4 out of 6 samples in order to be included in the analysis. Genes with at least 1.5 times upregulated or 1.5 downregulated gene expression and T-test p<0.05 were considered to be significant.

first: Eric

last: Bridenbaugh

first: Jin-Jia

last: Hu

first: Theresa

last: Fossum

first: Jay

middle: D

last: Humphrey

first: Emily

last: Wilson

first: Marina

last: Vannucci

first: Qianxing

last: Mo

first: Matthew

last: Miller

first: Jerome

last: Trzeciakowski

first: Jessemy

last: Neiger


title: Transcriptional response of Escherichia coli O157 TUV93-0 to small molecule inhibitors of TTS

status: Public on Aug 11 2009

city: Sydney

first: Jai

middle: Justin

last: Tree

state: NSW

address: Rm s110 Bldg F25, UNSW, Gate 11 Botany St

department: School of Biotechnology and Biomolecular Sciences

country: Australia

institute: University of New South Wales

summary:
The transcriptional profile of Escherichia coli O157 treated with small molecule inhibitors of type III secretion was determined. Four variations of the small molecule inhibitor were assessed for global changes in transcription by treating cells with 20uM of inhibitor or an equivalent volume of DMSO (inhibitor solvent).
Keywords: treatment, dose, Cy3, Cy5, 2-colour

overall_design: For each inhibitor, biological tripicates of treated (20uM inhibitor in DMSO) and untreated (DMSO only) cultures were grown in 25ml of MEM-HEPES supplemented with 0.1% Glucose and 250nM Fe(NO3)2. RNA was extracted from 15ml of culture at OD 0.7 and labeled with Cy5 (treated) and Cy3 (untreated) (excepting ME0055 that was performed as a dye swap). Labeled untreated RNA was pooled prior to hybridisation. Treated RNA (biological replicates) and pooled untreated RNA was hybridised to microarray slides. Slides were scanned using an Axon 4100A scanner and data processed using Genespring GX.

first: David

middle: G

last: Gally

first: Andrew

last: Roe


title: Early Transcriptomic response of coral larvae to hyperthermal stress

city: Lafayette

first: Mauricio

last: Rodriguez-Lanetty

state: LA

address: 300 E St Mary blvd

institute: University of Louisiana at Lafayette

summary: In this study, we examined the very early transcriptional response of aposymbiotic coral larval host (still not engaged in symbiosis) to hyperthermal stress.  This experimental setting provided a scenario and opportunity to study the direct effect of environmental stressors on the host cell per se.  Using a cDNA microarray constructed for Acropora millepora and Q-RT-PCR assays, we identified a number of genes that were significantly up- and down-regulated with increase of seawater temperature.  Down-regulation of several key component of DNA/RNA metabolism was detected implying inhibition of this cellular metabolic process, however the down-regulation of overall protein synthesis was not simple and random, which suggest that the response to stress is a more complicated adjustment to the metabolic needs of the cell.  We identified four significant outcomes during the very early hours of the transcriptional response to hyperthermal stress in coral larvae. First, molecular chaperones responded to hyperthermal stress by increasing their expression as expected, but the response was immediate and extremely rapid during the first 3 hours of heat exposure. Secondly, elevated temperature triggers down-regulation of a fluorescent protein homolog, DsRed-type FP, suggesting that this gene might be used as a potential molecular marker for monitoring hyperthermal stress in nature. Thirdly, the downregulation of a coral mannose-binding lectin under hyperthermal stress might compromise the coral immune defense and bring about susceptibility to pathogenic diseases. And lastly, an absence in the response of oxidative stress genes in aposymbiotic coral larvae during the early hours to hyperthermal stress suggest that the up-regulation of cnidarian host oxidative stress genes reported during thermal stress in algal/host symbiosis might be triggered directly by ROS generated by photosynthetic-dysfunctionally algal endosymbionts that diffuse into host cells, as very little ROS seems to be produced by the host cells from thermal-associated host cellular damage.

overall_design:
We applied a reference microarray design for the multi-factorial experiment outlined in the study, including two factors: Temperature (3 levels: 24 ˚C, 28 ˚C, 31 ˚C) and Time (2 levels: 3h and 10h).  Samples from time zero were used to generate the reference sample for the microarray hybridization experiments.  A total of 18 microarrays were used in the entire experiment. Reference samples in each array was labeled with Cy3, and the actual experimenatl samples with Cy5.
Ratio-Intensity plots were constructed for each array data to explore whether or not intensity dependence of log ratios, which appears as curvature, was present.   Because curvatures were detected in a few of the arrays, an rLowess curve fitting transformation was applied to the data.  The transformation was applied to all the arrays to keep consistence in the whole data.  Quantile normalisation was also applied to mean log-intensities in order to make the distributions essentially the same across arrays.  To detect differentially expressed genes among treatment through the course time of the experiment, a 2-way ANOVA model was fitted to the log transformed intensity data using the microarray analysis software GeneSpring (Agilent Technology). To correct for Type I error derived from multiple testing, the Benjamini and Hochberg method was applied as a false discovery rate (FDR) approach.

first: Saki

last: Harii

first: Ove

last: Hoegh-Guldberg


title: CGH analysis of adrenocortical tumors

status: Public on Aug 06 2009

city: Budapest

first: Viola

last: Tamasi

address: Nagyvarad sqr 4

department: Genetic, Cell- and Immunobiology

country: Hungary

institute: Semmelweis University

summary:
Adrenocortical tumors are common; their prevalence may reach up to 5-7% in pathological series. Most of them are benign and hormonally inactive, however, rare hormone-secreting (aldosterone and cortisol) and malignant forms are associated with significant morbidity and mortality. The prognosis of adrenocortical cancer (ACC) is poor with an overall five-year survival below 30 %.
In this study, CGH analysis was performed on 4 ACC (adrenocortical carcinoma), 4 IA (hormonally inactive adrenocortical adenoma) and 3 CPA (cortisol producing adrenocortical adenoma) samples. Tissue digestion, labeling, hybridization and data analysis of genomic DNA were performed according to the Agilent Technologies (Santa Clara, CA) protocol version 2.0 for 105 K arrays.
As expected, many of the observed aberrations were generally consistent with those of other preciously published data and will provide the basis for determination how genomic diversity impacts biological function and human diseases, such as cancer.

overall_design: In this study, biopsies from adrenocortical tumors (4 ACC, 4 IA  and 3 CPA samples) were analysed with CGH. Tissue digestion, labeling, hybridization, and data analysis of genomic DNA were performed according to the Agilent Technologies (Santa Clara, CA) protocol version 2.0 for 105 K arrays. Slides were scanned with Agilent Microarray scanner and data were extracted with Feature Extraction software version 9.5.1.1. DNA Analytics software was used (version 4.0.85, Agilent Technologies, Santa Clara, CA) for data analysis. The starting and ending points of the aberrations were confirmed by the ADM-2 algorithm with 6.0 threshold.

first: Péter

middle: M

last: Szabó

first: Viktor

last: Molnár

first: Márton

last: Andrásfalvy

first: Zsófia

last: Tömböl

first: Roland

last: Farkas

first: Katalin

last: Kövesdi

first: Attila

last: Patócs

first: András

last: Falus

first: Károly

last: Rácz

last: Igaz


title: Gene Expression in NS0 cells under ammonium stress

status: Public on Aug 05 2009

city: Clemson

first: Sarah

last: Harcum

state: SC

address: 105 Collings Street

department: Bioengineering

institute: Clemson University

summary:
Ammonium is a waste product that inhibits cell growth, recombinant protein production, and protein glycosylation in mammalian cell culture.  Recent studies have demonstrated that ammonium adversely affects glycosylation-related gene expression in Chinese hamster ovary (CHO) cells.  However, since the CHO cell line species has not been fully sequenced, a glycosylation transcriptome analysis is not possible in this cell line.  Therefore, to further understand the effects of ammonium on glycosylation-related gene expression, NS0 cells, a mouse myeloma cell line, were cultured under elevated ammonium.  NS0 cells are similar to CHO cells, in that the NS0 cells are anchorage-independent and commonly used to commercially produce recombinant proteins. Additionally, DNA microarrays containing all known mouse glycosylation-related genes were available to be used to examine gene expression.
NS0 cells were cultured under normal (control), elevated ammonium, elevated salt, and elevated ammonium with proline.  It was observed that the control and treatments culture growth rates were not significantly different; however, the final cell densities were significantly different.  The DNA microarray data was analyzed using a Welch ANOVA test with a Benjamini and Hochberg false discovery rate correction for the multiple comparisons of the glycosylation-genes.  No significant difference in gene expression levels between the four conditions examined were observed. The results of this study demonstrated that NS0 cells, at the gene expression level, are insensitive to ammonium. Thus, the decreased glycosylation observed in NS0 cell cultures at elevated ammonium is likely due to changes in synthesis and degradation enzyme activity.  In contrast, CHO cells have decreased glycosylation levels due to decreased sialytransferase gene expression and not increased degradation enzyme activity. Therefore, even though NS0 and CHO cells are both commonly used recombinant hosts for glycoprotein synthesis, it appears that NS0 and CHO cells had different control mechanisms respect to glycosylation-related gene expression under elevated ammonium.

overall_design: NS0 cells (ECACC#85110503), originally from the European Collection of Cell Culture, were donated to Clemson University by Merck, Inc. NS0 cells are a mouse myeloma cell line with lymphoblast morphology, non-secreting clone, and cholesterol auxotroph.  NS0 cells cultured under four conditions were examined:  Control (C), Ammonium-Stressed (A), Salt-Stressed (S), and Ammonium-Stressed with Proline added to mitigate the negative effects of ammonium (P).  Triplicates of each condition were used.  The cultures were be monitored during the normal batch growth phase.  To identify genes sensitive to ammonium in growing cultures, the 90-h time point was selected for RNA isolation and gene expression analysis.  Other culture parameters that were monitored include:  Cell density, viability, and glucose.

first: Mary

last: Caldwell


title: Genome-wide DNA methylation in MDS/secondary AML and de novo AML

city: Miami

first: Maria

middle: Eugenia

last: Figueroa

address: 1501 NW 10th Ave, BRB 709A, Locator code C227

department: Human Genetics

institute: University of Miiami

summary:
An increasing body of work reveals aberrant hypermethylation of genes occurring in and potentially contributing to the pathogenesis of myeloid malignancies.  Several of these diseases, such as myelodysplastic syndromes (MDS), are responsive to DNA methyltransferase inhibitors. In order to determine the extent of promoter hypermethylation in such tumors we compared the distribution of DNA methylation of 14,000 promoters in MDS and secondary AML patients enrolled in a phase I trial of 5-azacytidine and the histone deacetylase inhibitor entinostat against de novo AML patients and normal CD34+ bone marrow cells. The MDS and secondary AML patients displayed more extensive aberrant DNA methylation involving thousands of genes than did the normal CD34+ bone marrow cells or de novo AML blasts.  Aberrant methylation in MDS and secondary AML tended to affect particular chromosomal regions, occurred more frequently in Alu poor genes, and included prominent involvement of genes involved in the WNT and MAPK signaling pathways.   DNA methylation was also measured at days 15 and 29 after the first treatment cycle.  DNA methylation was reversed at day 15 in a uniform manner throughout the genome, and this effect persisted through day 29, even without continuous administration of the study drugs.

Keywords: DNA methylation profiling

overall_design: Direct comparison of DNA methylation in bone marrow samples from  patients with Myelodysplastic syndrome or secondary Acute Myeloid Leukemia (AML) at baseline and after in vivo treatment with 5-azacytidine + etinostat. A comparison to de novo normal karyotype AML was also performed.  Two control groups were included: one consisting of 8 CD34+ bone marrow samples from healthy donors and a second one consisting of matched CD34+ and CD34- fractions from the bone marrows of 4 healthy donors.

middle: E

first: Ari

last: Melnick