title: Differential expression analysis of genes from the mantle tissue of pearl oyster: Pinctada maxima

status: Public on Feb 05 2010

city: Pacific Grove

first: Luke

last: Gardner

state: California

address: 120 Ocean View Blvd, Hopkins Marine Station

institute: Stanford University

summary:
The mantle is a thin tissue from which proteins are secreted dictating the mollusk shell construction. As a conserved organ involved in shell formation throughout mollusks, the mantle is an excellent foundation from which to study biomineralization. A P. maxima mantle tissue specific cDNA microarray, termed PmaxArray 1.0, has been developed comprising 5000 cDNA transcripts derived from the mantle tissue of P. maxima. This tool has been used to investigate the spatial functional dynamics of the mantle tissue identifying over 2000 PmaxArray 1.0 spots as differentially expressed spatially within this organ. Gene expression profiles observed for these transcripts indicated 5 major spatial functions for the mantle, 3 of which have been putatively attributed to shell formation roles associated with nacre microstructure, calcite prismatic microstructure and periostracum. These transcripts are further examined with in situ expression localization and comparative sequence analyses in reference to potential shell formation roles. This spatial investigation has expedited the elucidation of functions within the dynamic mantle organ, paying particular attention to of shell biomineralization.

Keywords: Spatial expression profiling by array

overall_design: The mantle tissue from 9 animals was dissected into 5 separate sections: outer fold (OF), middle fold (MF), inner fold (IF), ventral mantle tissue (VM) and dorsal mantle tissue (DM). Total RNA was extracted from these tissues and pooled across subjects in order to reduce the effect of biological variation; such that 3 individuals were pooled together totaling 3 pooled replicate samples for each tissue. All the biologically pooled tissue types were compared against a common reference in which total RNA from all tissues types and all nine animals was equally pooled. A total of 30 dual channel microarrays hybridizations were performed and analyzed.

first: Abigail

last: Elizur


title: MicroRNAs in the HIV and Major Depressive Brain

status: Public on Feb 01 2010

city: La Jolla

first: Erick

middle: T

last: Tatro

state: CA

address: 9500 Gilman Dr

department: Psychiatry

country: USA

institute: University of California San Diego

type:
Expression profiling by RT-PCR
Non-coding RNA profiling by array
Other

summary:
Micro-RNAs (miRNAs) are small, non-coding RNAs that regulate gene networks, helping to control cell function and phenotype.  MiRNAs in the central nervous system (CNS) are considered to be involved in development, function, and homeostasis.  Past genomic studies focused on messenger-RNA (mRNA) expression using large-scale hybridization-based arrays for elucidating gene expression in the brain during HIV-infection.  Because miRNAs affect the downstream functions of mRNAs, it is desirable to understand both mRNA and miRNA perturbations, together in the same clinical specimens.  HIV infection affects the CNS through inflammation and glial cell activation leading to long-term changes in cellular function of neurons and glia.  One mechanism by which this may occur is through changes in the miRNA repertoire.  The goals of this study were to:  1. Identify changes in miRNA expression that occurred in HIV; 2. Determine whether detectable miRNA expression of the frontal cortex (FC) could differentiate HIV from HIV/Major Depressive Disorder (MDD, neurobehavioral diagnostic category); 3. Adapt a method to meaningfully integrate gene expression data and miRNA expression data in clinical samples.
The present study was performed by pooling RNA isolated from the FC of 3 patients from each of three groups: Control, HIV+, and HIV/MDD, and assessing miRNA expression profile using Applied Biosystems Taqman MicroRNA Array v2.0.  A larger sample size of 4 HIV+ patients and 4 HIV/MDD patients were used and non-pooled for gene expression analysis on Affymetrix U133 Plus 2.0 array.  We present a method for integrating the two datasets in a Target Bias Analysis. Micro-RNAs were clustered into three types: A) Those with many dysregulated mRNA targets of less stringent significance; B) Fewer dysregulated target-genes of highly stringent significance; C) Spectrum from non-bias to combinations of A and B.  In HIV/MDD, more miRNAs were down-regulated than in HIV alone. The dysregulated miRNAs clustered on Chromosomes 14, 17, 19, and X.  A small subset of dysregulated genes had many 3’UTR target-sites for dysregulated miRNAs. Specific miRNA families at targeted chromosomal loci are dysregulated. Certain miRNAs serve as key circuits in gene regulatory networks in the adult human brain, and this is affected by HIV, and may be implicated in neurobehavioral disorder. Some genes may serve as hubs of miRNA activity.

overall_design: Retrospective microRNA expression analysis of autopsy brain tissue.  Three HIV/MDD and three HIV-only subjects without neuropsychiatric conditions are identified and three age-matched Control subjects are compared. Technical triplicates are performed.

middle: R

last: Scott

first: Timothy

middle: B

last: Nguyen

first: Eliezer

last: Masliah

first: Cristian

middle: L

last: Achim

first: Ian

middle: P

last: Everall


title: Infection with Salmonella or Pseudomonas or E. coli - Interactions between plants and human pathogenic bacteria.

status: Public on Mar 25 2010

city: BEAUCOUZE

first: Sandra

last: Pelletier

address: 42, rue Georges Morel - BP 60057

country: France

institute: INRA

summary:
au07-07_salmonella - infection with Salmonella or Pseudomonas or E. coli. Identification of genes involved in early Arabidopsis response to pathogenic and non-pathogenic bacteria. Arabidopsis thaliana Col-0 seedlings were infected for 2 hours with a) Salmonella typhimurium strain 14028s, b) Pseudomonas syringae DC3000 or c) Escherichia coli DH5A

Keywords: treated vs untreated comparison

overall_design: 6 dye-swap - CATMA arrays

first: Heribert

last: Hirt

first: Jean-Pierre

last: Renou

first: Marie-Laure

last: Martin-Magniette


title: Eyeless: Pitx3 mutant mouse line, Expression profiling of liver

status: Public on Dec 01 2009

city: Neuherberg

first: Martin

last: Irmler

state: Bayern

address: Ingolstaedter Landstrasse 1

department: Institute of Experimental Genetics

country: Germany

institute: Helmholtz Zentrum München GmbH

summary:
A new mouse mutant arose spontaneously at the 5th inbred generation in a congenic C3H/He line. Since the mice suffered from anophthalmia or microphthalmia, we referred to this new mutant as eyeless (provisional gene symbol: eyl). The mutation follows an autosomal recessive pattern of inheritance and was mapped to chromosome 19 close to Pitx3; complementation tests confirmed that eyl is an allelic variant of aphakia (gene symbol: ak). Sequence analysis of the Pitx3 gene identified an insertion of a G in exon 4 (after cDNA position 416: 416insG). The altered open reading frame is predicted to result in a hybrid protein still containing the Pitx3 homeobox, but 121 new amino acids instead of the OAR domain at its C-terminal part.
The new Pitx3eyl allele shares the ophthalmological and brain defects of the aphakia mouse including the loss of dopamine transporter 1 transcripts in the midbrain. Moreover, we observed in the mutants an increased extramedullary hematopoiesis in the spleen, and frequently also hepatosteatosis, combined with a reduced body weight. Fat mass, fat content and subcutaneous fat are significantly decreased in females, but not in males. Additionally, we observed a broad spectrum of behavioural changes in the homozygous mutants including reduced forelimb grip strength and increased nociception. In addition to these alterations being apparent in both sexes, we observed in female Pitx3eyl-/- mice increased anxiety, reduced locomotor activity, reduced object exploration and increased social contacts, but decreased anxiety and increased arousal in males.
Most of these defects identified in the new Pitx3 mutation can also be observed in Parkinson patients. Therefore, the Pitx3eyl mutant is a valuable new model, because it is the first mutant mouse line carrying a point mutation within the coding region of the Pitx3 gene.

overall_design: 5 male animals, 5 experiments including 2 dye swap hybridisations

first: Johannes

last: Beckers

first: Marion

last: Horsch

first: Jochen

last: Graw


title: Divergent gene-expression profiles between proepicardium and epicardium differentiation

status: Public on Dec 21 2010

city: Leiden

first: Henk

last: Buermans

state: ZH

address: Albinusdreef 2

department: Human Genetics

country: Netherlands

institute: Leiden University Medical Center

summary:
The HH16/17 chicken proepicardium (PE) gives rise to the embryonic epicardium (Epi) which significantly contributes formation of the coronary vasculature during cardiac development. In contrast to explanted Epi cells, explanted PE cells can undergo differentiation into a cardiac myocyte phenotype.
In order to assess which genes are associated with PE differentiation into distinc cellular lineages, two interconnented microarray gene-expression series were performed.
1: Gene-expression profiles at 12, 24, 36, 48, 60, 72 and 120 hours during cardiac myocyte differentiation from including the HH16/17 PE (t0h) were determined by hybridizing Cy3 and Cy5 labelled amplified RNA to the ArkGen 20K oligo arrays in a 2-color looped experiment design, i.e., hybridization of successive time-points per array, including dye swaps, resulting in four technical replicates for each time point.
2: Gene expression changes during normal embryonic Epi maturation were assess by hybridizing Epi from stages HH25, HH29, HH32 and HH37 in all possible pair-wise combinations, including dye-swaps, leading to 6 replicates per time point. To allow for valid comparisons between the Epi and untreated PE differentiation, these two array series were connected via hybridization of both Epi stage HH25 and HH29 with the PE explant at 48 hours in culture, with dye swaps.

Keywords: time course, cardiac myocyte differentiation, embryonic coronary vessel formation, cell type comparison, mRNA expression

overall_design: Proepicardium to cardiac myocyte differentiation: 16 arrays for 8 time points [t0h,t12h,t24h,t36h,t48h,t60h,t72h,t120h]; successive time points hybridized per array in a loop design with dye swaps + Embryonic epicardium dfferentiation: 12 arrays for 4 time points [HH25,HH29,HH32,HH37]; all possible sample combination were hybridized with dye swaps + Epicardium to Proepicardium data connection: 4 arrays; comparing HH25 and HH29 Epicardium to t48h Proepicardium with dye swaps.

first: Bram

last: van Wijk

first: Maurice

middle: J

last: van den Hoff

first: Peter-Bram

middle: A

last: 't Hoen


title: Roles of relSpn in Stringent Response, Global Regulation, and Virulence of Serotype 2 Streptococcus pneumoniae D39

status: Public on Jan 11 2010

city: Bloomington

first: Krystyna

middle: M

last: Kazmierczak

state: IN

address: 1001 E. Third St.

department: Biology

institute: Indiana University

summary: Transcriptome analysis of D39 rel+Spn and delta-relSpn strains treated with mupirocin revealed relSpn-independent (translation stress), relSpn-dependent (stringent response), and delta-relSpn-dependent changes suggesting that relSpn and (p)ppGpp amount play wide-ranging homeostatic roles in pneumococcal physiology, besides adjusting macromolecular synthesis and transport in response to nutrient availability.

overall_design:
Wildtype and rel deletion mutant (Delta rel) bacteria were grown statically in BHI broth at 37C in an atmosphere of 5% CO2. Cultures were divided in two upon reaching a density of OD620 ~0.1, and lithium mupirocin was added to one culture (t=0) to a final concentration of 100 ng per mL. Treated and untreated cultures were harvested after 20 min further incubation and RNA was extracted by a hot lysis-acid phenol protocol.

We performed the following comparisons: Wildtype + mupirocin versus Wildtype untreated (reference); Delta rel + mupirocin versus Delta rel untreated (reference); Delta rel untreated versus Wildtype untreated (reference); Delta rel + mupirocin versus Wildtype + mupirocin (reference).
For each comparison, microarray data were obtained from three independent biological replicates and included one dye swap.

first: Kyle

last: Wayne

first: Andreas

last: Rechtsteiner

first: Malcolm

middle: E

last: Winkler


title: Maize Comparative Genomic Hybridization (CGH) between B73, Mo17, M0022 (RIL), and M0023 (RIL)

status: Public on Nov 20 2009

city: Ames

first: Patrick

middle: S.

last: Schnable

state: IA

address: 2035B Roy J Carver Co-Lab

institute: Iowa State University

type: Genome variation profiling by genome tiling array

summary: Following the domestication of maize over the past 10,000 years, breeders have exploited the extensive genetic diversity of this species to mold its phenotype to meet human needs. The extent of structural variation, including copy number variation (CNV) and presence/absence variation (PAV), which are thought to contribute to the extraordinary phenotypic diversity and plasticity of this important crop, have not been elucidated. Whole-genome, array-based, comparative genomic hybridization (CGH) revealed a level of structural diversity between the inbred lines B73 and Mo17 that is unprecedented among higher eukaryotes. A detailed analysis of altered segments of DNA conservatively estimates that there are several hundred CNV sequences among the two genotypes, as well as several thousand PAV sequences that are present in B73 but not Mo17. Haplotype-specific PAVs contain hundreds of single-copy, expressed genes that may contribute to heterosis and to the extraordinary phenotypic diversity of this important crop.

overall_design:
In our experimental design we had seven replicates of B73 (one with Cy3 and six with Cy5) and seven replicates of Mo17 (six with Cy3 and one with Cy5).  Images were processed and spatial normalization of data within the array was conducted according to Nimblegen's standard protocol.
The RIL samples (M0022 and M0023) were in included because we used the probe's B73 and Mo17 signals of those samples during our analysis.

first: Nathan

last: Springer

first: Kai

last: Ying

first: Yan

last: Fu

first: Tieming

last: Ji

first: Cheng

last: Yeh

first: Yi

last: Jia

first: Wei

last: Wu

first: Todd

last: Richmond

first: Jacob

last: Kitzman

first: Heidi

last: Rosenbaum

first: A

last: Iniguez

first: W

last: Barbazuk

first: Jeffrey

last: Jeddeloh

first: Dan

last: Nettleton

middle: S


title: Normal expression of facial prominences in stage 18 chicken embros

status: Public on Oct 30 2009

city: Vancouver

first: Joy

last: Richman

state: BC

address: 2350 Health Sciences Mall

department: Oral Health Sciences

country: Canada

institute: University of British Columbia

summary:
The face is one of the three regions most frequently affected by congenital defects in humans. In order to understand the molecular mechanisms involved it is necessary to have a more complete picture of gene expression in the embryo. Here we use microarrays to profile expression in chicken facial prominences, post neural crest migration and prior to differentiation of mesenchymal cells. Chip-wide analysis revealed that maxillary and mandibular prominences had similar expression profiles while the frontonasal mass chips were distinct. Of the 3094 genes that were differentially expressed in one or more regions of the face, a group of 56 genes was subsequently validated with quantitative PCR and a subset examined with in situ hybridization. Microarrays trends were consistent with the QPCR data for the majority of genes (81%). On the basis of QPCR and microarray data, groups of genes that characterize each of the facial prominences can be determined.
We used microarrays to detail the global programme of gene expression underlying facial morphogensis
keyword: craniofacial

overall_design: Chicken embryos were selected at a stage after neural crest cells have ceased migration, when facial prominences have formed and prior to ctyodifferentiation of cartilage, bone, muscle. Microdissection was used to isolate facial prominences. Embryos were dissected on ice in Hanks Buffered Salt Solution and batches of 26-32 pieces were snap frozen in liquid Nitrogen. These pooled facial prominences comprised one biological replicate. A total of three biological replicates were generated for the frontonasal mass, maxillary and mandibular prominences.

first: Marcela

last: Buchtová


title: Down-regulation of miR-92 in human plasma is a novel marker for acute leukemia patients

status: Public on Dec 31 2009

city: Sinjuku-ku

first: Masami

last: Tanaka

state: Tokyo

address: Sinjuku 6-1-1

country: Japan

institute: Tokyo Medical University

summary: MicroRNAs are a family of 19- to 25-nucleotides noncoding small RNAs that primarily function as gene regulators.  Aberrant microRNA expression has been described for several human malignancies, and this new class of small regulatory RNAs has both oncogenic and tumor suppressor functions.  Despite this knowledge, there is little information regarding microRNAs in blood especially because microRNAs in blood, if exist, were thought to be digested by RNase.  Recent studies, however, have revealed that microRNAs exist and escape digestion in serum and plasma.  Thus, we performed microRNA microaray to obtain insight into microRNA deregulation in the plasma of a leukemia patient.  Here, we have revealed that microRNA-638 (miR-638) is stably present in human plasmas, and miR-92a dramatically decreased in the plasmas of acute leukemia patients.  Our data indicate that the ratio of miR-92a/miR-638 in plasma has strong potential for clinical application as a novel biomarker for detection of leukemia.

overall_design: Seven normal samples and two acute leukemia samples ware profiled and compared.

first: Kosuke

last: Oikawa

first: Masakatsu

last: Takanashi

first: Motoshige

last: Kudo

first: Junko

last: Ohyashiki

first: Kazuma

first: Masahiko

last: Kuroda


title: Specific and pleiotropic patterns of mRNA regulation by ArcZ

status: Public on Oct 19 2009

city: Norwich

first: Sacha

last: Lucchini

address: Colney Lane

department: Molecular Microbiology

country: United Kingdom

institute: Institute of Food Research

summary: The small RNA, ArcZ (previously RyhA/SraH), was discovered in several genome-wide screens in Escherichia coli and Salmonella. Its high degree of genomic conservation, its frequent recovery by shotgun sequencing, and its association with the RNA chaperone, Hfq, identified ArcZ as an abundant enterobacterial “core” small RNA of unknown function. Here, we report that ArcZ acts as a post-transcriptional regulator in Salmonella, repressing the mRNAs of the widely distributed sdaCB (serine uptake) and tpx (oxidative stress) genes, and of STM3216, a horizontally acquired methyl-accepting chemotaxis protein (MCP). Both sdaCB and STM3216 are regulated by sequestration of the ribosome binding site. In contrast, the tpx mRNA is targeted in the coding sequence (CDS), arguing that CDS targeting is more common than appreciated. Transcriptomic analysis of an arcZ deletion strain further argued for the existence of a distinct set of Salmonella loci specifically regulated by ArcZ. In contrast, increased expression of the sRNA altered the steady-state levels of >16% (≥750) of all Salmonella mRNAs, and rendered the bacteria non-motile. Deep sequencing detected a dramatically changed profile of Hfq-bound sRNAs and mRNAs suggesting that the unprecedented degree of pleiotropic regulation might in part be caused by titration of Hfq binding by ArcZ.

overall_design:
This study used three different approaches to identify target genes and biological role of the small RNA ArcZ in Salmonella Typhimurium.

Transcriptomic analysis of ArcZ overexpression: Strain JVS-0082 (ΔarcZ) was transformed with plasmids pJV300 (control) and pKP48-1 (parcZ), and grown in liquid culture (LB broth) inoculated 1:100 from an overnight culture for 6h after cells had reached an OD600=2.0 to attain late stationary phase.

Transcriptomic analysis of arcZ mutant strain: Strains JVS-007 (WT) and JVS-0082 (ΔarcZ) were transformed with plasmid pJV300 (control) and grown in liquid culture (LB broth) inoculated 1:100 from an overnight culture for 6h after cells had reached an OD600=2.0 to attain late stationary phase.

Transcriptional effects of ArcZ pulse expression: Strain SL1344 was transformed with plasmids pKP8-35 (pBAD-control) and pKP4-13 (pBAD-ArcZ), and grown in liquid culture (LB broth) inoculated 1:100 from an overnight culture to an OD600 of 1.5. Expression of the insert was induced with L-arabinose (0.2% final concentrations) for 10 min.

Three biological replicates were performed for each strain/condition.

first: Kai

last: Papenfort

first: Nelly

last: Said

first: Tim

last: Welsink

first: Jay

middle: C

last: Hinton

first: Joerg

last: Vogel


title: Transcriptional analysis of the effects of 089 on a panel of yeast strains

status: Public on Dec 11 2009

city: Torino

first: Irene

last: Stefanini

address: via Accademia Albertina

country: Italy

institute: University of Turin

summary:
The analysis allow to delve into the molecular activity of the molecule 089, wich induced a growth decrease on various yeast strains.
Yeast strains selected are knock-out mutants in genes involved in MDR (BY4742Δerg6, BY4742Δsnq2) and the parental strain (BY4742).

overall_design: Six samples analyzed, two biological replicatesfor each yeast strain, the referenze samples is the treatment with 089 of the parental strain, BY4742.

first: Andrea

last: Trabocchi

first: Emmanuela

last: Marchi

first: Antonio

last: Guarna

first: Duccio

last: Cavalieri


title: Involvement of the post-transcriptional regulator Hfq in Yersinia pestis virulence

status: Public on Sep 01 2009

city: Beijing

first: Ruifu

last: Yang

address: No.20, DOngdajie, Fengtai

country: China

institute: Institute of Microbiology and Epidemiology

summary:
Yersinia pestis is the causative agent of plague, which is transmitted primarily between fleas and mammals and is spread to humans through the bite of an infected flea or contact with afflicted animals. Hfq is proposed to be a global post-transcriptional regulator that acts by mediating interactions between many regulatory small RNAs (sRNAs) and their mRNA targets. Sequence comparisons revealed that Y. pestis appears to produce a functional homologue of E. coli Hfq.
Phenotype comparisons using in vitro assays demonstrated that Y. pestis Hfq was involved in resistance to H2O2, heat and polymyxin B and contributed to growth under nutrient-limiting conditions. The role of Hfq in Y. pestis virulence was also assessed using macrophage and mouse infection models, and the gene expression affected by Hfq was determined using microarray-based transcriptome and real time PCR analysis. The macrophage infection assay showed that the Y. pestis hfq deletion strain did not have any significant difference in its ability to associate with J774A.1 macrophage cells. However, hfq deletion appeared to significantly impair the ability of Y. pestis to resist phagocytosis and survive within macrophages at the initial stage of infection. Furthermore, the hfq deletion strain was highly attenuated in mice after subcutaneous or intravenous injection. Transcriptome analysis supported the results concerning the attenuated phenotype of the hfq mutant and showed that the deletion of the hfq gene resulted in significant alterations in mRNA abundance of 243 genes in more than 13 functional classes, about 23% of which are known or hypothesized to be involved in stress resistance and virulence.
Our results indicate that Hfq is a key regulator involved in Y. pestis stress resistance, intracellular survival and pathogenesis. It appears that Hfq acts by controlling the expression of many virulence- and stress-associated genes, probably in conjunction with small noncoding RNAs.

overall_design: Y. pestis WT strain 201 and the Δhfq::KmR mutant were grown at 26C in LB medium to middle exponential phase and then transferred to 37C for 3 hr. Two independent bacterial cultures of strain 201 (control condition) and the Δhfq::KmR mutant (test condition) were considered biological replicates for RNA isolation. Four separate labeled probes were made for each RNA preparation as technical replicates.

first: Jing

last: Geng

first: Lei

first: Yanfeng

last: Yan

first: Yanping

last: Han


title: A Dysregulated Proinflammatory Status in Preterm School-aged Children with PVL-induced Spastic Cerebral Palsy

status: Public on Sep 30 2009

city: Tainan

first: Chang-yi

last: Lin

address: No.1, University Road

department: institute of basic medical science

country: Taiwan

institute: National Cheng Kung University

summary: Injury occurring during critical periods of development may have long-term effects on inflammatory responses. Periventricular leukomalacia (PVL) is the most common cause of cerebral palsy (CP) in preterm infant. Activated leukocytes are the main source of inflammatory cytokines that give rise to white matter damage and CP in preterm infant. Here, we tested the hypothesis that inflammation profiles as pathogenic mediators for the occurrence of PVL in the neonatal period may persist in preterm children with CP at school age.

overall_design: Five preterm children with PVL-induced CP and gestational age-matched five preterm children with normal neurodevelopment were recruited from follow up clinics. Proinflammatory gene expression in the PBMCs from preterm children were determined by Superarray PCR study.

first: Chang-Yi

first: Ying-Chao

last: Chang

first: Shan-Tair

last: Wnag

first: Ting-Yang

last: Lee

first: Chiou-Feng

first: Chao-Ching

last: Huang


title: Comparison of gene expression in 23 Cryptococcus gattii isolates

status: Public on Aug 07 2009

city: Birmingham

first: Robin

last: May

address: Edgbaston

department: School of Biosciences

institute: University of Birmingham

type: Expression profiling by genome tiling array

summary:
An array analysis of C. gattii (C. bacillosporus), intended to identify loci associated with the hypervirulence of the Vancouver Island Outbreak (VIO).  23 C. gattii isolates, representing both VIO strains and control strains, were grown for 24 hours in mammalian macrophages.  RNA was isolated and gene expression for each strain quantified relative to pooled RNA from all 23 samples.  Linear regression was used to identify loci showing positive or negative correlation with "intracellular proliferation rate", a proxy measure for virulence.  Data from this analysis is included in Ma et al, 2009, PNAS 106(31) 12980-12985.  The abstract is included below.

In 1999, the population of Vancouver Island, Canada, began to experience an outbreak of a fatal fungal disease caused by a highly virulent lineage of Cryptococcus gattii. This organism has recently spread to the Canadian mainland and Pacific Northwest, but the molecular cause of the outbreak remains unknown. Here we show that the Vancouver Island outbreak (VIO) isolates have dramatically increased their ability to replicate within macrophages of the mammalian immune system in comparison with other C. gattii strains. We further demonstrate that such enhanced intracellular parasitism is directly linked to virulence in a murine model of cryptococcosis, suggesting that this phenotype may be the cause of the outbreak. Finally, microarray studies on 24 C. gattii strains reveals that the hypervirulence of the VIO isolates is characterized by the up-regulation of a large group of genes, many of which are encoded by mitochondrial genome or associated with mitochondrial activities. This expression profile correlates with an unusual mitochondrial morphology exhibited by the VIO strains after phagocytosis. Our data thus demonstrate that the intracellular parasitism of macrophages is a key driver of a human disease outbreak, a finding that has significant implications for a wide range of other human pathogens.

overall_design: Dual-colour hybridization (each sample was hybridized against pooled RNA), one array per isolate, 23 biological samples, no technical replicates.

first: Hansong

last: Ma


title: Expression profiles of four hematopoietic cell types isolated from the bone marrow of BXD recombinant inbred mice.

status: Public on Sep 12 2009

city: Haren

first: Bruno

last: Tesson

address: Kerklaan 30

department: Groningen Bioinformatics Center

institute: University of Groningen

summary: Genetical genomics is a strategy for mapping gene expression variation to expression quantitative trait loci (eQTLs). We performed a genetical genomics experiment in four functionally distinct but developmentally closely related hematopoietic cell populations isolated from the BXD panel of recombinant inbred mouse strains. This analysis allowed us to analyze eQTL robustness/sensitivity across different cellular differentiation states. Although we identified a large number (365) of “static” eQTLs that were consistently active in all four cell types, we found a much larger number (1283) of “dynamic” eQTLs showing cell-type-dependence. Of these, 140, 45, 531, and 295 were preferentially active in stem, progenitor, erythroid and myeloid cells, respectively. A detailed investigation of those dynamic eQTLs showed that in many cases the eQTL specificity was associated with expression changes in the target gene. We found no evidence for target genes that were regulated by distinct eQTLs in different cell types, suggesting that large-scale changes within functional regulatory networks are uncommon. Our results demonstrate that heritable differences in gene expression are highly sensitive to the developmental stage of the cell population under study. Therefore, future genetical genomics studies should aim at studying multiple well-defined and highly-purified cell types in order to construct as comprehensive a picture of the changing functional regulatory relationships as possible.

overall_design: These data consist of total mRNA obtained from purified Lin-Sca-1+c-Kit+ multi-lineage cells, committed Lin-Sca-1-c-Kit+ progenitor cells, erythroid TER-119+ cells, and myeloid Gr-1+ cells, isolated from the bone marrow of ~25 genetically related and fully genotyped BXD – C57BL/6 (B6) X DBA/2 (D2) – recombinant inbred mouse strains (female) in three batches. Samples were assayed in three batches which are labeled in the metadata provided here. Some BXD lines have biological replicates, they are designated with letters (e.g. BXD28a and BXD28b are biological replicates). For some mice, not all four cell populations could be assayed. There are also a few technical replicates (see description column).

first: Alice

last: Gerrits

first: Yang

last: Li

first: Leonid

middle: V

last: Bystrykh

first: Ellen

last: Weersing

first: Albertina

last: Ausema

first: Bert

last: Dontje

first: Xusheng

last: Wang

first: Rainer

last: Breitling

first: Ritsert

last: Jansen

first: Gerald

last: de Haan


title: Genome-Wide Transcriptional Response of Silurana (Xenopus) tropicalis to Infection with the Deadly Chytrid Fungus

status: Public on Aug 31 2009

city: Davis

first: Matt

last: Settles

address: 451 Health Sciences Sr

department: Genome Center

institute: University of California, Davis

summary: Emerging infectious diseases are of great concern for both wildlife and humans. Several highly virulent fungal pathogens have recently been discovered in natural populations, highlighting the need for a better understanding of fungal-vertebrate host-pathogen interactions. Because most fungal pathogens are not fatal in the absence of other predisposing conditions, host-pathogen dynamics for deadly fungal pathogens are of particular interest. The chytrid fungus Batrachochytrium dendrobatidis (hereafter Bd) infects hundreds of species of frogs in the wild. It is found worldwide and is a significant contributor to the current global amphibian decline. However, the mechanism by which Bd causes death in amphibians, and the response of the host to Bd infection, remain largely unknown. Here we use whole-genome microarrays to monitor the transcriptional responses to Bd infection in the model frog species, Silurana (Xenopus) tropicalis, which is susceptible to chytridiomycosis. To elucidate the immune response to Bd and evaluate the physiological effects of chytridiomycosis, we measured gene expression changes in several tissues (liver, skin, spleen) following exposure to Bd. We detected a strong transcriptional response for genes involved in physiological processes that can help explain some clinical symptoms of chytridiomycosis at the organismal level. However, we detected surprisingly little evidence of an immune response to Bd exposure, suggesting that this susceptible species may not be mounting efficient innate and adaptive immune responses against Bd. The weak immune response may be partially explained by the thermal conditions of the experiment, which were optimal for Bd growth. However, many immune genes exhibited decreased expression in Bd-exposed frogs compared to control frogs, suggesting a more complex effect of Bd on the immune system than simple temperature-mediated immune suppression. This study generates important baseline data for ongoing efforts to understand differences in response to Bd between susceptible and resistant frog species and the effects of chytridiomycosis in natural populations.

overall_design: For each microarray, a competitive hybridization was performed, such that two samples were labeled with different Cy dyes and hybridized to the same chip to provide a direct comparison between frogs. A total of 13 chips were analyzed from 26 samples comprising 2 conditions (control and infected), 3 tissues (Dorsal skin, liver and spleen) and 2 timepoints (early and late).  Three biological replicates were used for each condition and tissue at each time point.  A dye-balance scheme was conducted for condition within each tissue at each time point in order to account for the effects of dye bias.

first: Erica

last: Rosenblum


title: Expression profiling of asense mutant VNC cells in Drosophila embryos

status: Public on Oct 01 2009

city: London

first: Tony

middle: David

last: Southall

address: South Kensington Campus

department: Life Sciences

institute: Imperial College London

summary: The expression profile of asense mutant neuroblasts and GMCs were compared to wildtype in order to investigate the function of Asense in neural development. Abstract of paper: Neural stem cells must strike a balance between self-renewal and multipotency, and differentiation. Identification of the transcriptional networks regulating stem cell division is an essential step in understanding how this balance is achieved. We have shown that the homeodomain transcription factor, Prospero, acts to repress self-renewal and promote differentiation. Amongst its targets are three neural stem cell transcription factors, Asense, Deadpan and Snail, of which Asense and Deadpan are repressed by Prospero. Here we identify the targets of these three factors throughout the genome. We find a large overlap in their target genes, and indeed with the targets of Prospero, with 245 genomic loci bound by all factors. Many of the genes have been implicated in vertebrate stem cell self-renewal, suggesting that this core set of genes is crucial in the switch between self-renewal and differentiation. We also show that multiply bound loci are enriched for genes previously linked to nervous system phenotypes, thereby providing a short-cut to  identifying genes important for nervous system development.

overall_design: 8 asense mutant samples were directly compared to 8 wildtype samples. 3 out of the 8 were dye-swapped.

middle: D

first: Andrea

middle: H

last: Brand


title: Coding sequence targeting by MicC RNA reveals bacterial mRNA silencing downstream of translational initiation

status: Public on Jul 16 2009

first: Sacha

last: Lucchini

summary: It is well established that small noncoding RNAs (sRNAs) of bacteria recognize the 5’ regions of target mRNAs, generally at the ribosome binding site. Until now, the translated coding sequence (CDS) of mRNA appeared to be refractory to sRNA interactions. We have discovered that Salmonella MicC RNA silences ompD mRNA by targeting the CDS, at codons 23-26. Analyses of MicC-ompD RNA complexes in vitro, and of chimeric sRNAs and ompD reporter fusions in vivo, show that interactions are confined to the CDS, and that a ≤12 bp RNA duplex involving the conserved 5’ end of MicC is essential and sufficient for target repression. MicC cannot act as a translational repressor at this downstream position being unable to inhibit 30S or 70S ribosome activity on the ompD mRNA. Instead, MicC accelerates RNase E-dependent ompD mRNA decay. The degradosome contributes to target destabilization, and facilitates the efficient degradation of processed ompD mRNA. Our data show that bacterial gene silencing by sRNAs can take place downstream of the translational initiation site by triggering irreversible mRNA decay. This ability of sRNAs allows targets in the CDS to be silenced without interference from the strong RNA helicase activity of elongating ribosomes.

overall_design: To determine the targets of the small regulatory RNA MicC in S. Typhimurium, we looked at the effect of a short pulse of MicC over-expression on the Salmonella transcriptome. To achieve over-expression, the micC gene was cloned in the pBAD plasmid and induced with 0.2% L-arabinose for 10 min. We then extracted the total RNA for transcriptional profiling. A strain carrying the pBAD plasmid w/o insert was used as negative control. 3 biological replicates were performed. This sRNA target identification strategy has been described in Papenfort et al; Molecular Microbiology (2006) 62(6), 1674–1688.

first: Verena

last: Pfeiffer

first: Kai

last: Papenfort

first: Jay

last: Hinton

first: Joerg

last: Vogel


title: Identification and validation of novel ERBB2 (HER2, NEU) targets including genes

status: Public on Jun 10 2009

first: Martin

last: Irmler

summary:
V-erb-b2 erythroblastic leukemia viral oncogene homolog 2 (ERBB2;
synonyms HER2, NEU) encodes a transmembrane glycoprotein with tyrosine kinase-specific activity that acts as a major switch in different signal-transduction processes. ERBB2 amplification and overexpression have been found in a number of human cancers, including breast, ovary and kidney carcinoma. Our aim was to detect ERBB2- regulated target genes that contribute to its tumorigenic effect on a genomewide scale. The differential gene expression profile of ERBB2- transfected and wild-type mouse fibroblasts was monitored employing DNA microarrays. Regulated expression of selected genes was verified by RT-PCR and validated by Western blot analysis. Genomewide gene expression profiling identified (i) known targets of ERBB2 signaling, (ii) genes implicated in tumorigenesis but so far not associated with ERBB2 signaling as well as (iii) genes not yet associated with oncogenic transformation, including novel genes without functional annotation. We also found that at least a fraction of coexpressed genes are closely linked on the genome. ERBB2 overexpression suppresses the transcription of antiangiogenic factors (e.g., Sparc, Timp3, Serpinf1) but induces expression of angiogenic factors (e.g., Klf5, Tnfaip2, Sema3c). Profiling of ERBB2-dependent gene regulation revealed a compendium of potential diagnostic markers and putative therapeutic targets. Identification of coexpressed genes that colocalize in the genome may indicate gene regulatory mechanisms that require further study to evaluate functional coregulation.

overall_design: Erbb2 transfected NIH3T3 cells versus un-transfected cells, 4 technical replicates including two colour swap experiments

first: Beckers

last: Johannes

first: Seliger

last: Barbara

first: Rieger

last: Sandra

first: Horsch

last: Marion


title: FOXM1 in Early Human Squamous Cell Carcinoma Oncogenesis and it Is Enhanced by Nicotine during Malignant Transformation

status: Public on Jun 13 2009

first: Muy-Teck

last: Teh

state: England

address: Blizard Building, 4, Newark Street,

department: Centre for Clinical & Diagnostic Oral Sciences

institute: Queen Mary University of London

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

summary:
Genome-wide SNP profilling for loss of heterozygosity (LOH) during FOXM1B-induced malignant transformation in a human premalignant oral keratinocyte line SVpgC2a.

Keywords: oral cancer, keratinocytes, head and neck squamous cell carcinoma, FOXM1, genomic instability, SNP array, loss of heterozygosity, malignant transformation

overall_design: Eight FOXM1B-induced malignant clones (SVFN1-8) that grew in soft agar were grown as individual cell lines and gDNA harvested from each clone for SNP array analysis compared to wild-type non-transformed SVpgC2a as control to obtain loss of heterozygosity profiles.

first: M


title: Interferon regulatory factor 5 expression levels regulate murine lupus in a type-I interferon-independent manner

status: Public on Sep 09 2009

city: Stanford

first: Chih Long

last: Liu

address: 269 Campus Drive

department: Medicine - Imm. & Rheum

institute: Stanford University

summary: Polymorphisms in the transcription factor interferon regulatory factor 5 (IRF5) are strongly associated in human genetic studies with an increased risk of developing the autoimmune disease systemic lupus erythematosus.  However, the biological role of IRF5 in lupus pathogenesis, if any, is not known.  In this study we show that IRF5 is absolutely required for disease development in the FcgRIIB-/-Yaa and FcgRIIB-/- lupus models.  In contrast to IRF5-sufficient FcgRIIB-/-Yaa mice, IRF5-deficient FcgRIIB-/-Yaa mice do not develop lupus manifestations and have a phenotype comparable to wildtype mice.  Strikingly, full expression of IRF5 is required for the development of autoimmunity, as IRF5-heterozygotes had dramatically reduced disease.  One effect of IRF5 is to induce the production of the type I interferon IFN-gamma, a cytokine implicated in lupus pathogenesis.  To address the mechanism by which IRF5 promotes disease, we evaluated FcgRIIB-/-Yaa mice lacking the type I interferon receptor IFNAR1.  Unlike the IRF5-deficient and IRF5-heterozygous FcgRIIB-/-Yaa mice, IFNAR1-deficient FcgRIIB-/-Yaa mice maintained a substantial level of residual disease.  Furthermore, in FcgRIIB-/- mice lacking Yaa, IRF5-deficiency also markedly reduced disease manifestations, indicating that the beneficial effects of IRF5 deficiency in FcgRIIB-/-Yaa mice are not due only to inhibition of the enhanced TLR7 signaling associated with the Yaa mutation. Overall, we demonstrate that IRF5 plays an essential role in lupus pathogenesis in murine models and that this is mediated through pathways beyond that of type I interferon production.  The fact that even IRF5 heterozygous mice developed minimal disease makes IRF5 a particularly attractive therapeutic target.

platform_organism:
Escherichia coli
Gallus gallus
Homo sapiens
Bos taurus
Oryctolagus cuniculus
Mus musculus

overall_design:
Serum samples from a total of 70 mice were run on the Utz Lab Whole Protein Autoantigen Array V1.0 (a single-color platform) in order to profile their autoantibodies against a library of autoimmune antigens.  All samples were run once with no replicates.  The samples consisted of the following groups:

For data appearing in Figure 3D, illustrating that mice lacking IRF5 have their autoantibody levels significantly affected:

R2Yaa IRF5+/+:  12
R2Yaa IRF5+/-:  11
R2Yaa IRF5-/-:  14
C57BL/6 ("WT" control):  13

Total mice (arrays) for this group:  50

For data appearing in Figure 6D, illustrating that mice lacking Ifnar1 do not have their autoantibody levels significantly affected:

R2Yaa Ifnar+/+:  10
R2Yaa Ifnar-/-:  10

Total mice (arrays) for this group:  20

first: Christophe

last: Richez

first: Kei

last: Yasuda

first: Ramon

middle: G

last: Bonegio

first: Amanda

last: Watkins

first: Tamar

last: Aprahamian

first: Patricia

last: Busto

first: Rocco

last: Richards

first: Chih

first: Regina

last: Cheung

first: Paul

last: Utz

first: Ann

last: Marshak-Rothstein

first: Ian

last: Rifkin


title: Types I GT1, II Me49, III CTGara strains - tachyzoite and Compound 1

status: Public on May 12 2009

city: St Louis

first: Michael

middle: Sean

last: Behnke

state: MO

address: 660 S. Euclid Ave

institute: Washington University School of Medicine

summary: Previous studies suggested that specific bradyzoite genes were differentially expressed in the three major lineages common to North America and Europe (Radke et al., 2006). In order to determine if these differences extended to the global transcriptome, we characterized whole-cell mRNA levels in both tachyzoite and bradyzoite populations from three primary strain isolates differentiated in vitro. It is well understood in the field that long passage history is associated with the loss of developmental competence (Frenkel et al., 1976). Therefore, the Type I-GT1, Type II-Me49B7 and Type III-CTG strains studied here were maintained at < 20 cell culture passages from the oocyst stage and are capable of completing both intermediate and definitive host life cycles. These experiments used newly constructed Affymetrix GeneChips that include probe sets for ~8000 genes, and we assessed the induction of bradyzoite genes using a strong inducer of bradyzoite differentiation, Compound 1 (Radke et al., 2006).

overall_design: Both tachyzoite and induced (Compound 1/48hrs - bradyzoite) RNA for strains representing the three major lineages of Toxoplasma gondii were hybridized .

middle: W

last: White


title: Comparative analysis of gene expression in two yeast species and their interspecific hybrid

status: Public on Apr 01 2009

city: Rehovot

first: Itay

last: Tirosh

address: herzl

department: Molecular genetics

country: Israel

institute: Weizmann Institute of Science

summary:
We compared the genome-wide expression profiles of two yeast species (S. cerevisiae and S. paradoxus) using a two-species microarray that contain species-specific probes and can thus measure the expression levels of the two species simultaneosly. In Addition, we used the array to measure expression levels of the interspecific hybrid of these yeast species, while discriminating between the alleles that correspond to the two parental species. Comparison of the between-species differences and the within-hybrid allele differences allows us to separate cis from trans effects. Also, comparison of the overall expression in the hybrids (both alleles) with their parental species allows us to analyze hybrid over-expression and under-expression.

Keywords: comparative transcriptome analysis, hybrid gene expression

sample_organism: Saccharomyces cerevisiae x Saccharomyces paradoxus

platform_organism:
Saccharomyces cerevisiae
Saccharomyces paradoxus

overall_design: We analyzed four conditions (rich media, glycerol, heat shock and TSA). For each conditions, we hybridized the pooled sample of both species dyed with cy3/cy5 and a sample of the hybrid dyed with the alternate fluorescent color. Each experiment was done with 2 biological repeats, except for the rich media experiments done with 4 biological repeats.

first: Sharon

last: Reikhav


title: The host transcriptome remains unaltered during the establishment of coral-algal symbioses

city: Thuwal

first: Christian

middle: Robert

last: Voolstra

address: Red Sea Research Center

country: Saudi Arabia

institute: King Abdullah University of Science and Technology

summary: Coral reefs are based on the symbiotic relationship between corals and photosynthetic dinoflagellates of the genus Symbiodinium. We followed gene expression of coral larvae of Acropora palmata and Montastraea faveolata after exposure to Symbiodinium strains that differed in their ability to establish symbioses. We show that the coral host transcriptome remains almost unchanged during infection by competent symbionts, but is massively altered by symbionts that fail to establish symbioses. Our data suggest that successful coral-algal symbioses depend mainly on the symbionts' ability to enter the host in a stealth manner rather than a more active response from the coral host.

sample_organism:
Acropora palmata
Orbicella faveolata

platform_organism:
Acropora palmata
Orbicella faveolata

overall_design:
Acropora palmata Samples: Three biological replicates of pooled larvae from each species and condition (i.e. untreated control, inoculated with competent Symbiodinium strain, inoculated with incompetent Symbiodinium strain) for both time points were hybridized against a pooled reference. Pooled references were constructed by combining equal amounts of aRNA from all control samples from A. palmata. References were labeled with Cy3, samples with Cy5.

Montastraea faveolata Samples: Three biological replicates of pooled larvae from each species and condition (i.e. untreated control, inoculated with competent Symbiodinium strain, inoculated with incompetent Symbiodinium strain) for both time points were hybridized against a pooled reference. Pooled references were constructed by combining equal amounts of aRNA from all control samples from M. faveolata. References were labeled with Cy3, samples with Cy5.

Symbiodinium sp. CassKB8: competent strain
Symbiodinium sp. EL1: incompetent strain
Symbiodinium sp. Mf1.05b: competent strain

first: Chrisitan

middle: R

first: Jodi

last: Schwarz

first: Julia

last: Schnetzer

first: Shinichi

last: Sunagawa

middle: K

last: DeSalvo

first: Mary-Alice

last: Coffroth

first: Alina

last: Szmant

first: Monica

last: Medina


title: Expression Profiling of Dll1 mutant mouse line

status: Public on Mar 01 2009

city: Neuherberg

first: Martin

last: Irmler

address: Ingolstaedter Landstrasse 1

institute: Helmholtz Zentrum München GmbH

summary: BACKGROUND: The Notch signaling pathway is an evolutionary conserved signal transduction pathway involved in embryonic patterning and regulation of cell fates during development and self-renewal. Recent studies have demonstrated that this pathway is integral to a complex system of interactions, involving as well other signal transduction pathways, and implicated in distinct human diseases. Delta-like 1 (Dll1) is one of the known ligands of the Notch receptors. The role of the Notch ligands is less well understood. Loss-of-function of Dll1 leads to embryonic lethality, but reduction of Delta-like 1 protein levels has not been studied in adult stage. METHODOLOGY/PRINCIPAL FINDINGS: Here we present the haploinsufficient phenotype of Dll1 and a missense mutant Dll1 allele (Dll1(C413Y)). Haploinsufficiency leads to a complex phenotype with several biological processes altered. These alterations reveal the importance of Dll1 mainly in metabolism, energy balance and in immunology. The animals are smaller, lighter, with altered fat to lean ratio and have increased blood pressure and a slight bradycardia. The animals have reduced cholesterol and triglyceride levels in blood. At the immunological level a subtle phenotype is observed due to the effect and fine-tuning of the signaling network at the different levels of differentiation, proliferation and function of lymphocytes. Moreover, the importance of the proteolytic regulation of the Notch signaling network emphasized. CONCLUSIONS/SIGNIFICANCE: In conclusion, slight alterations in one player of Notch signaling alter the entire organism, emphasizing the fine-tuning character of this pathway in a high number of processes.

overall_design: Four organs (liver, spleen, thymus, brain) of the Dll1 mutant mouse line analysed by cDNA microarray technology. Experiments include four biological replicates for reference (wildtype) and mutant animals. Two technical replicates for each mutant mouse were performed. As reference pooled RNA of the same organ was used. 50% of the chip hybridisations are dye sway experiments.

first: Johannes

last: Beckers

first: Marion

last: Horsch


title: miRNA expression profile in subcutaneous adipose tissue from morbid obese patients

status: Public on May 10 2009

city: Baronissi (SA)

first: Rosanna

last: Martinelli

address: Via Allende

department: Medicine and Surgery

institute: Salerno University

summary:
To evaluate wether miRNA expression patterns contributes to obesity total RNA were purified from subcutaneous adipose tissue (SAT)and used in  miRNA microarrays. Platform contain LNA-modified probes for all human miRNAs present in release 8.2 of the miRBase microRNA Registry.  Two RNA pools from 3 non obese men (CM) and from 5 non obese women (CW) were used as controls.
Expression profiling revealed that a large set of miRNAs is expressed in SAT. Forty two miRNAs changed by at least 1.5 folds in 17/20 obese subjects versus non obese control pool. Particularly, 21/42 were up-regulated and 21/42 were down-regulated. Among the differentially expressed miRNA, miR-519d, miR-498 and miR-150 were up-regulated, miR-659 and miR-371-3p_MM2 were down-regulated consistently in 20/20 obese subjects.

overall_design: To search for miRNAs eventually associated with obesity we used a microarray to evaluate the expression of 1458 different miRNAs in 10 obese women (OBW) and 10 obese men (OBM). Two RNA pools from 3 non obese men (CM) and from 5 non obese women (CW) were used as controls.


title: Systematic identification of genes involved in chicken muscle development

status: Public on Jan 01 2009

city: Wilmington

first: Qi

last: Zheng

state: DE

address: 1701 Rockland Road

institute: Nemours Children's Health

summary:
The genetic closeness and divergent muscle growth rates of broilers and layers make them great models for myogenesis study. In order to discover the molecular mechanisms determining the divergent muscle growth rates and muscle fiber sizes in different chicken lines, we systematically identified differentially expressed genes between broilers and layers during muscle development (postnatal 1 day, 2 weeks, 4 weeks, 6 weeks and 8 weeks) by microarray hybridization experiment. Taken together, 543 differentially expressed probe sets were found between broilers and layers across different developmental stages, including genes related to muscle growth and hypertrophy, fatty acid transportation and metabolism, protein degradation, and several important signaling pathways. The expression profiles of a few differentially expressed genes were highly correlated with the growth rates of broilers and layers. We also identified SNPs within upstream transcription factor binding sites of a few differentially expressed genes, indicating that these SNPs might be the causing factor of the expression differences of these genes between broilers and layers. These studies provided new clues for deciphering mechanisms underlining muscle development and organ size control in different chicken lines, will shed light on the study of human muscle related disease as well.
Keywords: Time-course studies of two different intra-species breeds

overall_design: Pectoralis major muscles were sampled from broilers and layers at indicated developmental stage (postnatal 1 day, 2 weeks, 4 weeks, 6 weeks and 8 weeks). Trizol extraction of total RNA was performed according to the manufacturer's instructions. Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 2 ug total RNA. Affymetrix Gene Chip array hybridization was carried out according to Affymetrix Expression Analysis Technical Manual by GeneTech Biotechnology Limited Company (Shanghai, China). Following labeled cRNA fragmentation, 15 ug of cRNA were hybridized for 16 hr at 45℃ on Affymetrix Chicken Genome Array. After standard washing and staining, GeneChips were scanned using the Affymetrix GeneChip Scanner 7G.

first: Yong

last: Zhang

first: Ying

last: Chen

first: Ning

last: Yang

first: Xiujie

last: Wang

first: Dahai

last: Zhu


title: Mitochondrial dysfunction in Arabidopsis thaliana.

status: Public on Feb 15 2009

city: Rosario

first: Paula

last: Casati

state: Santa Fe

address: Suipacha 531

department: Biological Sciences

country: Argentina

institute: Universidad Nacional de Rosario

summary: We characterized the global response of plants carrying a mitochondrial dysfunction induced by the expression of the unedited form of the ATP synthase 9 subunit. The u-ATP9 transgene driven by A9 and Apetala3 promoters induce mitochondrial dysfunction revealed by a decrease in both oxygen uptake and ATP levels, with an increase in ROS and a concomitant oxidative stress response. The transcriptome analysis of young Arabidopsis flowers, validated by RT-PCR and enzymatic or functional tests, show dramatic changes in u-ATP9 plants. Both lines present a modification in the expression of several genes involved in carbon, lipid and cell wall metabolism, suggesting that an important metabolic readjustment occurs in plants with a mitochondrial dysfunction. Interestingly, transcript levels involved in mitochondrial biogenesis, protein synthesis, and degradation are affected. Moreover, several mRNA levels for transcription factors and DNA binding proteins were also changed. Some of them are involved in stress and hormone response, suggesting that several signaling pathways overlap. Indeed, the transcriptome data reveal that the mitochondrial dysfunction dramatically alters genes involved in signaling pathways, including those involved in ethylene, absicic acid and auxin signal transduction. Our data suggest that the mitochondrial dysfunction model used in this rapport may be useful to uncover the retrograde signaling mechanism between the nucleus and mitochondria in plant cells.

overall_design: Arabidopsis oligonucleotide microarrays fabricated by the University of Arizona contain 26,000 oligonucleotides (http://www.ag.arizona.edu/microarray/). RNA was isolated from 6-week-old flowers from u-ATP9 and wt plants. The experimental (mutant) and reference (wild type) RNA samples were reverse-transcribed and directly labeled with either Cy5-dUTP or Cy3-dUTP fluorescent dye (GE Healthcare), using random hexamer primers (Invitrogen). Excess nucleotides and primers were removed using QIAquick PCR Purification Kit (Qiagen). Labeled samples were mixed and then hybridized to microarray for 15 h at 60°C. The slides were washed at room temperature in three wash steps: 2 x SSC, 0.5% SDS; 0.5 x SSC; and 0.05 x SSC for 5 min each with gentle shaking. The slides were scanned with a GenePix 4000B Scanner (Axon Instruments Inc., Union City, CA). Normalization between the Cy3 and Cy5 fluorescent dye emission channels was achieved by adjusting the levels of both image intensities. The experiments were repeated four times with samples from different experiments, as biological replicates. In dye swapping experiments, the RNA samples from different experiments were reciprocally labeled, both as a biological and technical repetition for comparing the reproducibility of the experiments. Hybridization intensities for each microarray element were measured using ScanAlyze 4.24 (available at http://genome-www4.stanford.edu/MicroArray/SMD/restech.html). The two channels were normalized in log space using the z-score normalization on a 95% trimmed data set. We removed unreliable spots according to the following criteria: spots flagged as having false intensity caused by dust or background on the array were removed; and spots for which intensity was less than three fold above background were also eliminated. Data from multiple experiments were normalized (Bolstad, 2003) and signals from spots from different experiments were statistically analyzed using Significance Analysis of Microarrays using the one class response (SAM, http://www-stat.stanford.edu/~tibs/SAM/), cut at a false discovery rate < 10% (Tusher, 2001).

first: María

last: Busi

last: Gomez-Lobato

first: Sebastián

middle: P

last: Rius

first: Eduardo

middle: J

last: Zabaleta

first: Diego

middle: F

last: Gomez-Casati

first: Alejandro

last: Araya


title: Multiple functions for Eya3 in mammalian organism

status: Public on Dec 17 2008

last: Irmler

summary:
The important function of Eya (eyes absent) in eye development of  D.melanogaster is well known, the function of its mammalian orthologs Eya1-4 is only partially understood. Yet, no phenotype for Eya2 and Eya3 in humans or mouse have been described. In our study we analysed new Eya-deficient mouse model generated by insertional mutagenesis. Expression analysis of Eya3 by in-situ hybridizations and β-Gal-staining revealed a broad expression of the gene throughout development, e.g. in brain, eyes, branchial arches, somites and limbs. This diverse expression pattern suggests pleiotropic effects of Eya3 in mammalian organisms. In contrast to D. melanogaster, a close examination of the eye by various methods did not show any obvious ocular defects in mutant animals. A broad phenotypical characterization of homozygous mutants displayed decreased body weight and body length. Additionally, behavioral changes as a educed forward locomotor activity and altered exploratory behaviour were identified. A significant reduction of forelimb grip strength and an impairment of cardiac processes in Eya3-/- mice suggest a role for Eya3 in muscle development and heart function. Our results implicate that Eya3 plays various roles outside the eye with effects particularly on heart and muscle function in the mouse.
Keywords: Eya3-deficient mice, cDNA microarrays, brain, heart, muscle

overall_design: analysis of 4 Eya3 mutant mice versus a pool of 5 wildtype animals; brain, heart, and muscle; 8 hybridisations for each tissue: two technical replicates for each mutant animal including one dye-swap experiments

first: Jochen

last: Graw

first: Troben

last: Söker


title: Repeatability and comparability of microRNA microarray

status: Public on May 14 2009

city: Kyoto

first: Akira

last: Myomoto

address: Yoshida-Shimoadachi

institute: Kyoto Univ.

summary:
We compared repeatability and comparability of microRNA microarray using 5 different platforms. and compared microarray data generated from five different microRNA microarray platforms with quantitative RT-PCR. Our data suggested that the most accurate and repeatable methods for microRNA expression profiling are Agilent and Toray, and the numbers of detected microRNA at Toray are more than at Agilent.
Keywords: repeatability, comparability, microRNA, microarray

platform_organism:
Homo sapiens
Mus musculus
Rattus norvegicus

overall_design: We compared repeatability and comparability of microRNA microarray using 5 different platforms (Agilent, Ambion, Exiqon, Invitrogen, and Toray). In addition, we compared microarray data generated from five different microRNA microarray platforms with quantitative RT-PCR.

first: F

last: Sato

first: S

last: Tsuchiya

first: K

last: Terasawa

first: G

last: Tsujimoto


title: The global expression profile of Bacillus subtilis to the toxic carbonyls methylglyoxal (MG) and formaldehyde (FA)

status: Public on Dec 13 2008

city: Ithaca

first: John

middle: D.

last: Helmann

state: NY

address: 372 Wing Hall

department: Microbiology

institute: Cornell University

summary: Abstract of associated menuscript; Quinones and alpha,beta-unsaturated carbonyls are naturally occurring electrophiles that target cysteine residues via thiol-(S)-alkylation. We analyzed the global expression profile of Bacillus subtilis to the toxic carbonyls methylglyoxal (MG) and formaldehyde (FA). Both carbonyl compounds cause a stress response characteristic for thiol-reactive electrophiles as revealed by the induction of the Spx, CtsR, CymR, PerR, ArsR, CzrA, CsoR and SigmaD regulons. MG and FA triggered also a SOS response which indicates DNA damage. Protection against FA is mediated by both the hxlAB operon, encoding the ribulose monophosphate pathway for FA fixation, and a thiol-dependent formaldehyde dehydrogenase (AdhA) and DJ-1/PfpI-family cysteine proteinase (YraA). The adhA-yraA operon and the yraC gene, encoding gamma-carboxymuconolactone decarboxylase, are positively regulated by the MerR-family regulator, YraB(AdhR). AdhR binds specifically to its target promoters which contain a 7-4-7 inverted repeat (CTTAAAG-N4-CTTTAAG) between the -35 and -10 elements. Activation of adhA-yraA transcription by AdhR requires the conserved Cys52 residue in vivo. We speculate that AdhR is redox-regulated via thiol-(S)-alkylation by aldehydes and that AdhA and YraA are specifically involved in reduction of aldehydes and degradation or repair of damaged thiol-containing proteins, respectively.

overall_design: WT (-FA) vs. WT (+ 1 mM FA), WT (-MG) vs. WT (+ 2.8 mM MG), WT (-MG) vs. WT (+ 5.6 mM MG). Each experiement was conducted triplicates using three independent total RNA preparations. For all datasets used for final analysis, untreated samples were labeled with Cy3 and treated samples with Cy5.

first: Thi

last: Nguyen

first: Warawan

last: Eiamphungporn

first: Ulrike

last: Mäder

first: Manuel

last: Liebeke

first: Michael

last: Lalk

last: Hecker

first: Haike

last: Antelmann


title: An analysis of the impact of infection by Buchnera aphidicola APS on gene expression of Drosophila S2 cells.

status: Public on Dec 15 2008

city: Cambridge

first: Roslin

last: Russell

address: Downing Street

department: Department of Genetics

institute: University of Cambridge

summary:
An analysis of the impact of infection by Buchnera aphidicola APS (isolated from Acyrthosiphon pisum strain LL01) on gene expression of S2 cells.  All comparisons are made against a pool of RNA from S2 cells not exposed to B. aphidicola.  B. aphidicola freshly isolated from the aphids, and data are collected at 1, 6 and 24 hours after exposure of S2 cells to the B. aphidicola preparation.
Keywords: time course, Buchnera aphidicola APS, aphids, Drosophila melanogaster S2 cells


title: Generation of Induced Pluripotent Stem Cells from Adult Rhesus Monkey Fibroblasts

status: Public on Dec 07 2008

city: Beijing

first: hongkui

last: deng

address: Yiheyuan Road 5#, Haidian District

department: College of Life Sciences

institute: Peking University

summary:
Induced pluripotent stem (iPS) cells can be generated from somatic cells by transduction with several transcription factors in both mouse and human. However, direct reprogramming in other species has not been reported. Here, we established an efficient method to generate monkey iPS cells from fibroblasts by retrovirus-mediated introduction of the four monkey transcription factors OCT4 (POU5F1), SOX2, KLF4, and c-MYC. The monkey iPS cells displayed ES-like morphology, expressed ES cell-marker genes, shared similar global gene profiles and methylation status in the OCT4 promoter to those of monkey ES cells, and possessed the ability to differentiate into three germ layers in vitro and in vivo. Our results suggest that the mechanism of direct reprogramming is conserved among species. The efficient generation of monkey iPS cells will allow investigation of the feasibility of therapeutic cloning in primate model with various diseases.

Keywords: Induced pluripotent stem, iPS, Rhesus monkey

overall_design: We analysed each sample (Rhesus monkey fibroblast, embryonic stem cell (ES) and induced pluripotent stem cell (iPS)) for three replications and sought to see high similarty between iPS and ES.

first: Haisong

last: Liu

first: Hongkui

last: Deng


title: Monocular Visual Deprivation in Macaque Monkeys: A Profile in the Gene Expression of the LGN by LCM

status: Public on Dec 01 2008

city: Cleveland

first: Henry

middle: J.

last: Kaminski

state: OH

address: 11100 Euclid Avenue

department: Neurology

institute: Case Western Reserve University

summary:
Laser capture microdissection was used to obtain individual LGN layers for DNA microarray of Rhesus array in macaque monkeys that Monocular Visual Deprivation was generated by either opaque dark contact lens or tarsoraphoplasty at birth.
Keywords: Lateral Geniculate Nucleus, monocular deprivation, Rhesus array, Laser Capture Microdissection, Gene expression

overall_design: Lateral Geniculate Nucleus from two months and four months monocular deprived monkey were used. Each Lateral Geniculate Nucleus has deprived parvocellular layers, non-deprived parvocellular layers considered as a replicates.

first: Georgiana

last: Cheng

first: Bendi

last: Gong

first: Lan

last: Zhou

first: Denise

last: Hatala

first: Scott

last: Howell

first: Xiaohua

last: Mustari


title: Prostaglandin E2 regulates B cell proliferation through a candidate tumor suppressor, Ptger4

city: Evry

first: Olivier

last: ALIBERT

address: 2 rue Gaston Crémieux

department: IRCM

institute: CEA

summary:
Recognition of foreign antigens by B cell receptor (BCR) on mature B cells leads to their clonal expansion, which is critically important for the effective host defence of the organism. However, excessive antigenic responses or reaction of B cells against body’s own components frequently lead to diverse immune diseases, such as B-cell lymphoma or autoimmunity, that often affect humans. Identification of genes that restrain uncontrolled proliferation of B cells is therefore an important goal towards understanding the origin of such diseases. Here we identify Ptger4 as a negative feedback regulator of B-cell proliferation in response to BCR triggering, and show that its encoded EP4 receptor is a principal molecule conveying the growth-suppressive effect of prostaglandin E2 (PGE2). In controlled in vitro assays, Ptger4-/- B cells showed augmented proliferative response and increased expression of activating genes upon BCR stimulation. Stable knock-down of Ptger4 in B-cell lymphoma markedly accelerated tumour spread in mice, while Ptger4 overexpression yielded significant protection. Lack of Ptger4 rendered mouse B cells completely resistant to proliferation arrest signalled by PGE2, and we find by transcriptional profiling that intrinsic activity of Ptger4 and PGE2-EP4 signalling target a similar set of activating genes. We further show that Ptger4 inhibits mouse B-cell activation in vivo and find it significantly downregulated in human B-cell lymphoma. Our results demonstrate that Ptger4 functions in B cells as a candidate tumour suppressor whose activity is regulated by the presence of PGE2 in the microenvironment. These findings suggest that targeting EP4 receptor for prostaglandin may present a novel strategy for treatment of B-cell diseases.
Keywords: time course, cell type comparison, compound treatment design, microarray experiment record

overall_design: B cells were extracted from spleen of Ptger4+/+ and Ptger4-/- mice, and incubated in vitro for the indicated period with or without anti-IgM (Fab')2 antibody fragments and co-treated with or without PGE2. Total RNA was extracted, amplified, labelled with Cy3 or Cy5, and hybridized to mouse 24k oligo-arrays using a dye-swap strategy.

first: Jernej

last: Murn

first: Simon

last: Tendil

last: Alibert

first: Beverly

last: Koller

first: Xavier

last: Gidrol


title: mRNA profile in purified human peripheral blood eosinophil treated with retinoic acids

status: Public on Oct 25 2008

city: Akita

first: Shigeharu

last: Ueki

department: Department of Clinical and Laboratory Medicine

institute: Akita University School of Medicine

summary:
Objectives: Eosinophils cultured with all-trans RA (ATRA) and 9-cis RA showed dramatically induced cell survival, and the efficacy of RAs (10-6 M) was similar to that of IL-5 (1 ng/ml), the most critical cytokine for eosinophil activation.  Using a gene microarray, we studied mRNA profile regulated by RAs.
Methods: Peripheral venous blood was obtained from subjects with mild eosinophilia.  Eosinophils were isolated by sedimentation with 6% dextran followed by centrifugation on 1.088 Percoll (Pharmacia, Uppsala, Sweden) density gradients.  The cells were further purified by negative selection using anti-CD16 immunomagnetic beads and a MACS system (Miltenyi Biotec, Bergisch Gladbach, Germany).  Purified eosinophils were resuspended in RPMI 1640 medium (Gibco, Grand Island, NY) with 10% FCS, and incubated with vehicle, 10-6 M of 9-cis RA, or ATRA in 1% human serum albumin-coated plates, each 4 × 106 cells, for 4 h.  Then cells were lysed, and total RNA was isolated using Isogen (Nippon Gene, Tokyo, Japan) as per the manufacturer’s instructions.  RNA was repurified with phenol-chloroform extraction and ethanol precipitation.  For each cell culture condition, the total RNA from three donors was pooled, enabling us to acquire adequate amounts of RNA for microarray analysis.
Results: There was little variation between the samples treated by 9-cisRA and ATRA.  The comparison between vehicle control and 9-cisRA, or vehicle control and ATRA, identified an increase (greater than twofold change over control) in 19 and 11 transcripts, respectively.   The number of decreased transcripts (less than 0.5-fold) resulting fromby 9-cisRA or ATRA exposure wasere 19 and 8 transcripts, respectively.  In these transcripts, the expression changes relative to vehicle control were smaller in withthe ATRA compared with 9-cis RA, suggesting that 9-cis RA induced gene transcription more effectively rather than ATRA.  Of note, among the ten genes with the biggestmost decrease relative tofrom vehicle control, we found an apoptosis- related gene, caspase-3.
Conclusions: We screened several genes involved in RA-induced prolonged eosinophil survival.

overall_design: Purified peripheral blood eosinophils were incubated with vehicle, 10-6 M of 9-cis RA, or ATRA in 1% human serum albumin-coated plates, each 4 × 106 cells, for 4 h.  Then cells were lysed, and total RNA was isolated.  RNA was repurified with phenol-chloroform extraction and ethanol precipitation.  For each cell culture condition, the total RNA from three donors was pooled, enabling us to acquire adequate amounts of RNA for microarray analysis.

first: Gulixian

last: Mahemuti

first: Hajime

last: Oyamada

first: Hikari

last: Kato

first: Wataru

last: Ito

first: Masahide

last: Takeda

first: Hiroyuki

last: Kayaba

first: Junichi

last: Chihara


title: Diagnosis of Morphologically Challenging Cases of Renal Epithelial Neoplasms by Virtual Karyotypes with SNP Arrays

status: Public on Nov 01 2008

city: Houston

first: Federico

middle: A

last: Monzon

state: TX

address: 6565 Fannin St., MS205

department: Pathology

institute: The Methodist Hospital Research Institute

summary:
Renal tumors with complex morphology require extensive workup for accurate classification.  Chromosomal aberrations that define subtypes of renal epithelial neoplasms have been reported. We explored if whole-genome chromosome copy number and loss-of-heterozygosity analysis with single nucleotide polymorphism (SNP) arrays can be used to identify these aberrations in cases where morphology was unable to definitively classify these tumors.

Keywords: Chromosome copy number and LOH analysis (virtual karyotyping) with SNP Genotyping Arrays

Keywords: Genome variation profiling by SNP array

overall_design: We analyzed 50 paraffin-embedded tissues representing tumors with classic morphology from clear cell renal cell carcinoma (RCC), papillary RCC, chromophobe RCC, and oncocytoma with Affymetrix GeneChip 10K 2.0 Mapping arrays. We then analyzed 25 tumors that could not be classified by morphology and classified them based on the presence of defining chromosomal anomalies identified in the "classic" cohort.

first: Jill

middle: M

last: Hagenkord

first: Anil

last: Parwani


title: SAGE Libraries from Highly Purified Mouse Hematopoietic Stem Cells from the E14.5 Fetal Liver and Adult Bone Marrow

status: Public on Oct 18 2008

city: Vancouver

first: BCGSC

last: BC Cancer Agency

address: 675 West 10th Avenue

department: Gene Expression

institute: Canada's Michael Smith Genome Sciences Centre

summary: Recent improvements in the development of methods for isolating functionally validated populations of nearly pure (>20%) murine hematopoietic stem cells (HSCs) have made it possible to analyze the molecular basis of the properties of these cells with increased precision. One intriguing feature of HSCs is the change they undergo in many of their key properties during development a change that affects the control of their self-renewal, cycling status, differentiated progeny output and steel factor sensitivity. To investigate how these differences are mediated, we undertook a genome-wide analysis of the transcripts present in highly purified fetal and adult HSCs using an adaptation of the LongSAGE methodology that allows its application to small numbers of cells (10 ng of RNA) by inclusion of an initial PCR amplification step that preserves the transcript repertoire while excluding less than 0.25% of the transcripts. The LongSAGE methodology was adopted because it is sequence-based and thus quantitative and independent of prior knowledge of expressed genes or variations in their hybridization to matching or related cDNAs, ESTs or derived oligos. A suspension of 10,000 lin-Sca-1+CD43+Mac1+ fetal liver (FL) cells (20% pure HSCs as determined by 16-week limiting dilution and single cell transplantation experiments) was obtained from embryonic day 14.5 fetuses. From these cells, we generated a 160,000-tag LongSAGE library containing 7865 tags that map uniquely to the mouse genome (using the RefSeq database through DiscoverySpace; www.bcgsc.ca/DiscoverySpace). A suspension of 3700 CD45midlin-Rho-SP cells (30% pure HSCs) was isolated from adult mouse bone marrow (BM) and then used to generate a 37,000-tag LongSAGE library (956 uniquely mapped tags). Both of these libraries contained tags identifying transcripts that have been previously reported to be associated with HSCs from FL and/or adult BM, including c-kit, pbx-1, tgf-, cul-4a, PrP, c-myc, robo1, sox17, as well as a number of Smarc transcripts, ubiquitin ligase transcripts and TNF-related transcripts. As a first test of the utility of the libraries, we looked for differences in the expression of genes that have been broadly associated with differences in cellular proliferative activity. This comparison identified many such tags in the FL HSC library that were absent from the adult BM HSC library, including multiple cyclins (A2, B2, C, D1, D2, D3, E1, F, H, I, J L1, L2), cdc20, cdc5b, and plk, as well as 34 of the top 50 proliferation genes identified by Venezia et al (PLoS Biology, 2004) to be selectively expressed in adult BM HSCs that had been stimulated to proliferate. Other transcripts that were present at significantly higher levels in the FL HSC library (95% C.I. using Audic Claverie statistics) included msl2, rbx1, lmo2, pfn1, and 16 members of the tripartite motif protein (trim) family. Conversely, many transcripts for components of the proteosome, involved in nucleic acid binding, and transcripts coding for proteins with receptor activity were present at higher levels (or uniquely) in the adult BM HSC library. Taken together, these findings establish the validity and potential of these permanent HSC transcriptome resources for further investigation of mechanisms that determine the different biology of fetal and adult HSCs.

overall_design: RNA was collected from E14.5 fetal livers and 10-12 week old adult bone marrow with the phenotypes that isolate highly purified hematopoietic stem cells (Lin-/CD45+/SP/Rho- for bone marrow and Lin-/Mac1+/CD43+/Sca1+ for fetal liver)

first: David

last: Kent

first: Yun

last: Zhao

first: Michelle

last: Bowie

first: Brad

last: Dykstra

first: Jay

last: Cheyne

first: Yongjun

first: Allen

last: Delaney

first: Martin

last: Hirst

first: Marco

last: Marra

first: Connie

last: Eaves


title: miRNA profiles of HCC from Southeast China

status: Public on Sep 21 2008

city: Guangzhou

first: Jian-Rong

last: Yang

address: No. 74, Zhongshan 2nd Road

institute: Sun Yat-sen University

summary: We evaluated the expression of known human miRNAs in human hepatocellular carcinoma (HCC) and normal hepatic tissues from southeast China, and identified the differentially expressed miRNAs in HCC tissues.

overall_design: We use microRNA array platform from CapitalBio Corp. to access the miRNA expression profiles in HCC and non-tumor liver samples from Southeast China. There were 5 HCC samples and 3 non-tumor liver samples in our study. As the microarray platform we used was based on a older version of miRBase, we mapped the probe sequences to a newer version of miRBase before these data was applied to further analysis.

first: Hang

last: Su

first: Teng

last: Xu

first: Jun

last: Huang

first: Li

first: Yunfei

last: Yuan

first: Shi-Mei

last: Zhuang


title: Expression analysis of Bacteroides fragilis NCTC 9343 delta-ungD1 delta-ungD2 delta-PSH acapsular mutant

status: Public on Sep 02 2008

city: Boston

first: Michael

last: Coyne

state: MA

address: 181 Longwood Avenue

department: Medicine

institute: Brigham & Women's Hospital

summary:
Investigation of whole genome gene expression level changes in a Bacteroides fragilis NCTC 9343 delta-ungD1 delta-ungD2 delta-PSH triple mutant, compared to the wild-type strain.  The mutations engineered into this strain render it acapsular.

The mutants analyzed in this study are further described in Coyne, M. J., M. Chatzidaki-Livanis, L. C. Paoletti, and L. E. Comstock. 2008. Role of glycan synthesis in colonization of the mammalian gut by the bacterial symbiont Bacteroides fragilis. PNAS 105(35):13098-13103 (PID 18723678).

overall_design: A six chip study using total RNA recovered from three separate wild-type cultures of Bacteroides fragilis NCTC 9343 and three separate cultures of a triple mutant strain, Bacteroides fragilis NCTC 9343 delta-ungD1 delta-ungD2 delta-PSH, in which ungD1 (BF1706), ungD2 (BF2848), and six genes (BF3454 through BF3459) of the PSH capsular polysaccharide locus are truncated or deleted entirely.  Each chip measures the expression level of 4,302 genes from Bacteroides fragilis NCTC 9343 and the associated plasmid pBF9343 with fourteen 24-mer probe pairs (PM/MM) per gene, with three-fold technical redundancy.

first: Laurie

middle: E

last: Comstock

first: Maria

last: Chatzidaki-Livanis


title: Profiling differentially expressed earthworm genes in response to TNT, RDX, or TNT+RDX exposure.

status: Public on Aug 28 2008

city: Vicksburg

first: Ping

last: Gong

state: MS

address: 3909 Halls Ferry Road

department: Environmental Laboratory

institute: US Army ERDC

summary:
To understand molecular mechanisms of the joint effects of 2,4,6-trinitrotoluene (TNT) and hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX), both widely used ordnance compounds, we constructed a microarray consisting of 4,032 cDNA isolated from the earthworm Eisenia fetida using the suppressive subtractive hybridization technique. Worms were exposed to TNT-, RDX-, or TNT+RDX-spiked soil for 28 days (TNT 50 mg/kg, RDX 30 mg/kg).
Keywords: Combined toxicity of TNT and RDX to earthworm (Eisenia fetida)

overall_design: We analyzed 40 arrays for 4 treatments (control, TNT 50ppm, RDX 30ppm, TNT 50ppm + RDX 30ppm) with 5 biological replicates per treatment using an interwoven loop design.

first: Xin

last: Guan

first: Edward

last: Perkins


title: Isolation and transcriptional profiling of embryonic human neural crest cells

status: Public on Aug 07 2008

city: Marseille

first: Heather

middle: C

last: Etchevers

address: 27 boulevard Jean Moulin

department: INSERM U1251

institute: INSERM/Aix-Marseille Univ

summary:
To identify new candidate genes for neurocristopathies, a Long-SAGE library has been made from human neural crest cells (hNCC) derived from a normal embryo at Carnegie stage 13.  Data analysis indicates the expression of a number of genes in multiple GO functional classes that were anticipated based on animal studies and/or human disease. An average linkage hierarchical clustering was performed using SAGE data from hNCC and 14 other normal tissues and cell lines. This and other analyses indicate a high degree of molecular identity between hNCC and the two human embryonic stem cells (hES) lines included, compared to the other tissue or cell types included, including more tissue-restricted stem cells, such as mesenchymal stem cells, or hNCC derivatives such as Schwann cells.
Keywords: Cell type comparison

overall_design:
Total RNA was extracted from a confluent 10 cm dish of hNCC (line N5) from a female human embryo at 28-32 dpf (Carnegie stage 13). Half the RNA was used to construct a SAGE bank with long tags using NlaIII and MmeI. Data was then compared to currently publicly available SAGE data from fourteen banks made from normal human tissues for hierarchical clustering.
Third-party GEO accession numbers for human SAGE data:
GSM41360, hES3;
GSM41362, hES4;
GSM48250, sciatic nerve;
GSM48251, Schwann cells (in vitro);
GSM31931, brain substantia nigra;
GSM824, muscle;
GSM785, liver;
GSM708, kidney;
GSM676, brain white matter;
GSM761, brain cerebellum;
GSM762, lung.

Other third-party data:
Cancer Genome Anatomy Project http://cgap.nci.nih.gov/SAGE/SAGELibraryFinder: SAGE_Prostate_normal_B_2 (prostate)

Umbilical cord- and bone marrow-derived mesenchymal stem cell data had been downloaded from currently broken links http://bit.fmrp.usp.br/uc-msc_tags/ and http://bit.fmrp.usp.br/msc_tags/ as referenced in Panepucci et al. (2004) Stem Cells 22:1263-1278; DOI: 10.1634/stemcells.2004-0024 (UC-MSC) and Silva et al. (2003) Stem Cells 21:661-669.

first: Sophie

last: Thomas

first: Marie

first: Patrick

last: Wincker

first: Pu-Ting

first: Marcy

last: Speer

first: Arnold

last: Munnich

first: Stanislas

last: Lyonnet

first: Michel

last: Vekemans

first: Candice

last: Babarit


title: Expression data for a study on cross-species hybridization on single-species microarrays

status: Public on Sep 03 2008

city: Hamilton

first: Frederic

last: Chain

state: Ontario

address: 1280 Main Street West

department: Biology

institute: McMaster University

type:
Expression profiling by array
Genome variation profiling by array

summary:
Gene expression was examined in testis and brain tissue between two species (Xenopus laevis and Xenopus borealis) and their hybrid.

Genomic DNA hybridizations of the microarray target and non-target species were used to select probes that are unbiased between species.

sample_organism:
Xenopus borealis
Xenopus laevis
Xenopus muelleri
Xenopus laevis x Xenopus borealis

overall_design:
Gene expression:
3 biological replicates were used for each tissue type.

Genomic hybridization:
We used the ratio of probe gDNA hybridization intensities between the target and non-target species to create a probemask for expression data from these species.
Genomic DNA was extracted from blood samples of several Xenopus individuals.  Xenopus laevis individuals were lab bread, Xenopus borealis individuals were from Kenya, and Xenopus muelleri individuals were from Swaziland.

first: Frédéric

first: Dora

last: Ilieva

first: Ben

last: Evans

first: Darcy

middle: B

last: Kelley


title: Gene regulation in ki-ras transfected murine fibroblasts

status: Public on Aug 01 2008

city: Neuherberg

first: Martin

last: Irmler

address: Ingolstaedter Landstrasse 1

department: Institute of Experimental Genetics

institute: Helmholtz Zentrum München GmbH

summary:
Ki-ras gene mutations are involved in the carcinogenesis of acute myeloid leukemia, melanoma and different carcinomas. In order to define potential mutation-specific therapeutic targets, expression profiling analyses were performed using stable transfected NIH3T3 cells carrying different ki-ras gene mutations. Maybe, this analysis allows the discorvery of ras-associated cellular mechanisms, which might lead to the identification of physiological targets for pharmacological interventions of the treatment of Ki-ras-associated human tumors.
Keywords: cDNA microarray, murine fibroblast, ki-ras mutations, carcinogenesis

overall_design: 4 transfected murine fibroblast cell lines (mock, wt-kras, Asp, Val) were compared to non-transfected cell line. For each of the four comparison 4 chip experiments were performed including 2 dye swaps.

first: Johannes

last: Beckers

first: Marion

last: Horsch

first: Barbara

last: Seliger

first: Christian

last: Recktenwald


title: Identification of RNA molecules associated with the RNA-chaperone Hfq

status: Public on Aug 23 2008

city: Norwich

first: Sacha

last: Lucchini

address: Colney Lane

department: Molecular Microbiology

institute: Institute of Food Research

summary:
We used high-throughput methods to discover the Salmonella RNAs that are targeted by the bacterial Sm-like protein, Hfq. Our generic approach, using chromosomal epitope tagging and coIP-on-chip will also be useful to identify the post-transcriptional regulons of many other RNA-binding proteins.
Keywords: coIP-chip

platform_organism: Salmonella enterica subsp. enterica serovar Typhimurium

overall_design: To identify the direct Hfq RNA targets, we co-immunoprecipitated RNA from a strain expressing a 3xFLAG-tagged Hfq-protein using an anti-3xFLAG antibody. As a negative control, we used the same antibody (anti-3xFLAG) to co-immunoprecipitate RNA from an isogenic strain expressing the non-tagged Hfq-protein. Two independent biological replicates were generated.

first: Alexandra

last: Sittka

first: Kai

last: Papenfort

first: Cynthia

last: Sharma

first: Gary

last: Rowley

first: Katarzyna

last: Rolle


title: BW25113 hha mutant vs wild type biofilm cells 4hr LB and 4,15,24hr LB glu

status: Public on Jun 17 2008

city: State College

first: Thomas

middle: K.

last: Wood

state: PA

address: Room 161 Fenske Laboratory, University Park

department: Chemical Engineering & Biochemistry and Molecular Biology

institute: Pennsylvania State University

summary:
E. coli K-12 BW25113 mutant strain hha expression in biofilm cells relative to E. coli wild-type strain expression in biofilm cells. Samples were cultured in LB with glasswool at 37C for 4 hours and in LB glu with glasswool at 37C for 4, 15 and 24 hours.

Hha is a temperature- and osmolarity-dependent modulator of gene expression that is induced 30-fold in Escherichia coli biofilms. Here we show through whole-transcriptome analysis that Hha decreases biofilm formation in both LB and LB glu media by (i) repressing fliC encoding the main structural flagellar protein flagellin, (ii) by repressing fimA encoding the major structural subunit of type I fimbriae, (iii) by repressing ihfA encoding a subunit of the transcriptional regulator IHF that induces the transcription of type I fimbriae genes, (iv) by regulating tnaA encoding tryptophanase that inhibits biofilms, and (v) by repressing ybaJ that forms an operon with hha. Corroborating the microarray data, hha deletion increased motility 3.2 ± 0.1-fold, decreased extracellular indole concentrations 12 ± 2-fold, and decreased type I fimbriae (as measured by yeast agglutination). Biofilm tests using single and double mutants of fimA and ihfA and transcriptional studies of the fimA, ihfA and ybaJ-hha promoters confirmed that Hha represses biofilm by inhibiting type I fimbriae production and that it negatively regulates its own transcription and that of ybaJ. Nickel-enhanced DNA microarrays to determine in vivo Hha binding sites confirmed that Hha binds the ybaJ-hha promoter, that it binds fimZ, a positive regulator of fimA, and that it binds to the rare codon tRNAs argU, ileXY, and proL. Sequence analysis of fimZ, fimB, fimE, and the type I fimbriae gene cluster fimAICDFGH revealed a high bias for the rare codons of arginine, isoleucine, proline, leucine, and threonine, and overexpressing Hha leads to cell death.  Therefore, it appears Hha decreases biofilms by decreasing type I fimbriae production as a result of inhibiting synthesis of tRNAs for rare codons.
Keywords: effect of hha deletion in biofilm formation 4 hr LB and 4,15 and 24 hr LB glu

overall_design:
Strains: E.coli K-12 BW25113 wild-type, mutant hha
Medium:LB and LB glu
Biofilm grown on glass wool
Time: 4 hr for LB, 4,15 and 24 hr for LB glu
Cell type: biofilm

RNA isolation and DNA microarrays. Pre-cultures of E. coli BW25113 and BW25113 hha mutant were done for 15 h in LB and LB with kanamycin (50 μg/mL), respectively. Biofilm cells of each strain were prepared as described before (Zhang et al., 2007), and then the total RNA was isolated from biofilm cells as described in (Ren et al., 2004). The E. coli Genechip antisense genome array chips (P/N 900381, Affymetrix, Santa Clara, CA) c were used to analyze the complete E. coli transcriptome as described before (Zhang et al., 2007). All the procedures were done according to the Gene Expression Technical Manual (Affymetrix) and the DNA microarray data analysis was done using the GeneChip operating software (Affymetrix). The data quality was confirmed following the manufacturer's specifications (GeneChip Expression Analysis: Data Analysis Fundamentals; Affymetrix) and based on the expected signals of E. coli BW25113 and the hha mutant genotypes (e.g., signals of the deleted genes, araA and rhaA were low for both BW25113 and BW25113 hha mutant, while signal of hha were low for hha mutant). A gene was considered differentially expressed only when the P value was lower than 0.05 and the expression ratio was higher than: 2-fold for the 4 h LB and LB glu microarrays, higher than 4-fold for the LB 15 h microarray and higher than 2.46-fold for the 24 h LB glu microarray since the standard deviation for the expression ratio for all of the genes were: 1.4 for the 4 h LB, 1.38 for the 4 h LB glu, 3.29 for the 15 h LB glu and 1.63 for the 24 h LB glu microarray. The gene functions were obtained from the National Center for Biotechnology Information database (http://www.ncbi.nlm.nih.gov/) (Edgar et al., 2002), and the SRI International Institute for Genomic Research, University of California at San Diego, and UNAM database (http://ecocyc.org/) (Keseler et al., 2005).

first: Rodolfo

last: Garcia-Contreras

first: Xue-Song

last: Zhang

first: Younghoon

last: Kim


title: Control of RpoS in global gene expression of Escherichia coli in minimal media

status: Public on Oct 10 2008

first: Herb

last: Schellhorn

state: ON

address: 1280 Main St. W.,

summary: RpoS, an alternative sigma factor, is critical for stress response in Escherichia coli. The RpoS regulon expression has been well characterized in rich media that support fast growth and high growth yields. In contrast, though RpoS levels are high in minimal media, how RpoS functions under such conditions has not been clearly resolved. In this study, we compared the global transcriptional profiles of wild type and an rpoS mutant of E. coli grown in glucose minimal media using microarray analyses. The expression of over 200 genes was altered by loss of RpoS in exponential and stationary phases, with only 48 genes common to both conditions. The nature of the RpoS-controlled regulon in minimal media was substantially different from that expressed in rich media. Specifically, the expression of many genes encoding regulatory factors (e.g., hfq, csrA and rpoE) and genes in metabolic pathways (e.g., lysA, lysC and hisD) were regulated by RpoS in minimal media. In early exponential phase, protein levels of RpoS in minimal media were much higher than that in LB media, which may at least partly account for the observed difference in the expression of RpoS-controlled genes. Expression of genes required for flagellar function and chemotaxis was elevated in the rpoS mutant. Western blot analyses show that the flagella sigma factor FliA was expressed much higher in rpoS mutants than in WT in all phase of growth. Consistent with this, the motility of rpoS mutants was enhanced relative to WT. In conclusion, RpoS and its controlled regulators form a complex regulatory network that mediates the expression of a large regulon in minimal media.

overall_design: A precise rpoS deletion mutant of MG1655 was constructed using the red recombinase method. Wild type and rpoS mutants were inoculated in triplicate into M63 glucose (0.2%) minimal media at a starting OD of 0.0001 and grown aerobically at 37oC. Cultures were harvested at OD600= 0.3 in exponential phase and at OD600= 1.5 in stationary phase. For RNA extraction, cultures were mixed directly with a boiling lysis buffer containing SDS and EDTA followed by acidic hot phenol (65C) to minimize RNA degradation. RNA samples were hybridized to Affymetrix E. coli Antisense Genome Array according to Affymetrix's standard protocols.

first: Tao

last: Dong


title: Effect of hfq-deletion on Salmonella transcriptome

summary:
The expression profile of an S. Typhimurium hfq mutant-strain was compared to the parental strain under 2 different growth conditions; early stationary phase and SPI-1 (Salmonella pathogenicity island 1) inducing condition.
Keywords: Genetic modification

overall_design: This study comprises two separate experiments performed under different growth conditions, were the gene expression profile was compared to that of the parental strain. Three biological replicates were performed for each strain and condition. For this study, we used Salmonella genomic DNA as the comparator which also acted as the control for spot quality.

first: Jay

last: Hinton

first: Joerg

last: Vogel


title: Comparison of small medium and large healthy ovarian pig follicles

status: Public on Jun 12 2008

city: Castanet-Tolosan Cedex

first: Gwenola

last: Tosser-Klopp

address: Chemin de Borde-Rouge  BP 52627

institute: INRA

summary:
Three different stages of pig antral follicles have been studied in a granulosa-cell transcriptome analysis on nylon microarrays (1152 clones). The data have been generated from 7 RNA follicle pools and several technical replicates were made. Four Large, one Medium and two Small follicles pools were considered. For each follicle pool, 2 radioactive labellings were performed. Each membrane was exposed 16 hours (to avoid saturation of the signal of highly expressed genes) and 28 hours (to get some signal from lowly expressed genes). Each probe was hybridised on GPL3971 scag_scai Sus scrofa 1.2K mono array and on GPL3970 scag_scai Sus scrofa 4.6K triplicate array  (except for GFS2172 which was labelled only once and hybridised onto 2  GPL3971 scag_scai Sus scrofa 1.2K mono array membranes), so that 4 spots are available for each gene (and 2 spots for GFS2172), for a given RNA and a given radioactive labelling.
Keywords: granulosa-cell transcriptome analysis

overall_design: data consisted in (6 RNA x 2 labellings) + (GFS2172 RNA X 1 labelling)= 13 probes, 26 hybridisations, 52 images.