title: Gene expression signatures of HO-1 in BeWo cells

status: Public on Feb 15 2011

city: Vienna

first: Stefanie

last: Tauber

address: Dr. Bohr-Gasse 9

department: CIBIV

country: Austria

institute: MFPL

summary: Heme Oxygenase-1 (HO-1) is expressed in many cancers and  influences the growth, survivall and metastasis of tumors, however, the molecular mechanisms remains largely unknown. To identify a common mechanism of action of HO-1 in cancer, we studied the global effect of HO-1 on the transcriptome of multiple tumors. Genome-wide expression profiling of HO-1 expressing versus HO-1 silenced cancer cells and a further data mining analysis of the preexisting expression database of 190 human tumors of 14 cancer types led us to identify 14 genes, the expression of which correlated firmly and universally with that of HO-1 (P < 0.001). These genes included regulators of cell plasticity and extracellular matrix remodeling (MMP2, ADAM8, TGFβ1, BGN, COL21A1, PXDN), signaling (CRIP2, MICB), amino acid transport and glycosylation (SLC7A1 and ST3GAL2), estrogen and phospholipid biosynthesis (AGPAT2 and HSD17B1), protein stabilization (IFI30) and phosphorylation (ALPPL2). PXDN, one of the genes being co-expressed with HO-1, was selected for further analysis. Immunofluorescence and western blotting confirmed positive correlation of PXDN with HO-1 levels in BeWo cancer cells as well as co-localization in invasive extravillous trophoblast cells of first trimester placenta. Loss of HO-1 in BeWo cells correlated with reduced cell adhesion to Collagen type I, Fibronectin and Laminin. The adhesion-promoting effects of HO-1 were dependent on PXDN expression, as loss of PXDN in HO-1 expressing BeWo cells led to reduced cell attachment to Laminin and Fibronectin coated wells.

overall_design: We used gene expression profiling to determine the genome-wide effect of HO-1 on the transcriptome of BeWo trophoblast cells. We specifically selected BeWo  cells for our studies because these cells express HO-1 naturally. We knocked down endogenous HO-1 in BeWo cells using retroviral transduction with a micro-RNA adapted retroviral vector targeting human HO-1 sequence. RNA isolated from control (LMP) or miHO1 infected (miHO-1) cells was labeled and hybridized to human genome-wide gene level 1.0 ST arrays

first: Martin

last: Bilban


title: Prospective comparison of genome-wide aCGH platforms for the detection of CNVs in MR (NimbleGen)

status: Public on Feb 17 2011

city: Vancouver

first: Tracy

last: Tucker

address: Box 153 4480 Oak Street

country: Canada

institute: University of British Columbia

type: Genome variation profiling by genome tiling array

summary: Clinical laboratories are adopting array comparative genomic hybridization (AGH) as a standard clinical test. A number of whole genome AGH systems are available, but little is known about the comparative performance in a clinical context.  We prospectively studied 30 children with idiopathic MR and both unaffected parents of each child using Affymetrix 500K GeneChip SNP arrays, Agilent Human Genome 244K oligonucleotide arrays and NimbleGen 385K Whole-Genome oligonucleotide arrays. We determined whether CNVs called on these platforms were detected by Illumina Hap550 beadchips or SMRT 32K BAC whole genome tiling arrays and tested 15 of the 30 trios on Affymetrix 6.0 SNP array.  The Affymetrix 500K, Agilent and NimbleGen platforms identified 3061 autosomal and 117 X chromosome CNVs in 30 trios. 147 of these CNVs were de novo, but only 33 (22%) of the de novo CNVs were found on more than one platform. Performing genotype-phenotype correlations, we identified 7 pathogenic and 4 possibly pathogenic CNVs for MR. All 11 of these CNVs were detected by both the Agilent and NimbleGen arrays, 9 by the Affymetrix 500K and Illumina beadchips, and 5 by the SMRT BAC array. Two of the 4 pathogenic or possibly pathogenic CNVs present in the trios tested with the Affymetrix 6.0 array were identified.   Our findings demonstrate that different results are obtained with different AGH platforms and illustrate the trade-off that exists between sensitivity and specificity. The large number of apparently false positive CNV calls supports the need for validating clinically important findings with a different methodology.

overall_design: 30 trios were analysed consisting of child (proband) and both normal parents.

first: Alexandre

last: Montpetit

first: David

last: Chai

first: Susanna

last: Chan

first: Sébastien

last: Chénier

first: Bradley

middle: P

last: Coe

first: Allen

last: Delaney

first: Patrice

last: Eydoux

first: Wan

middle: L

last: Lam

first: Sylvie

last: Langlois

first: Emmanuelle

last: Lemyre

first: Marco

last: Marra

first: Hong

last: Qian

first: Guy

middle: A

last: Rouleau

last: Vincent

first: Jacques

last: Lmichaud

first: Jan

middle: M

last: Friedman


title: Prospective comparison of genome-wide aCGH platforms for the detection of CNVs in MR (Illumina HumanHap550)

last: Tucker

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

overall_design: 30 childern with MR


title: The stress response factors Yap6, Cin5, Phd1, and Skn7 direct targeting of the conserved co-repressor Tup1-Ssn6

status: Public on Mar 01 2011

city: Buffalo

first: Michael

last: Buck

state: NY

address: 701 Ellicott St

department: Biochemistry

country: USA

institute: SUNY at Buffalo

type: Genome binding/occupancy profiling by array

summary: The budding yeast transcriptional corepressor Tup1-Ssn6 is a model for studying similar repressosome complexes in multicellular eukaryotes. Tup1-Ssn6 does not bind DNA directly, but is directed to individual promoters by one or more DNA-binding proteins, referred to as Tup1 recruiters. We determined the genomic distribution of Tup1 and Ssn6 by ChIP-chip and found that most loci bound by Tup1-Ssn6 could not be explained by co-occupancy with a known recruiting cofactor. Furthermore, we found that individual deletions of known Tup1 recruiters did not significantly alter Tup1 binding profile. These two observations suggest that Tup1 recruitment typically depends on multiple recruiting cofactors, and that new Tup1 recruiting proteins remain to be discovered. To identify new recruiting proteins we computationally screened for factors with binding patterns similar to the observed Tup1-Ssn6 genomic distribution. Four top candidates, Cin5, Skn7, Phd1, and Yap6, all known to be associated with stress response gene regulation, were experimentally confirmed to physically interact with Tup1 and/or Ssn6.  Incorporating these new cofactors with previously characterized cofactors now accounts for the majority of Tup1 binding across the genome, and expands our understanding of the mechanism by which Tup1-Ssn6 is directed to its targets.

overall_design: ChIP-chip

middle: J

first: Sean

middle: E

last: Hanlon

first: Deirdre

middle: C

last: Tatomer

first: Jason

middle: D

last: Lieb

last: Rizzo


title: Structural variation among soybean cultivars

status: Public on May 03 2011

city: Saint Paul

first: Robert

last: Stupar

state: MN

address: 1991 Upper Buford Circle

department: Agronomy and Plant Genetics

institute: University of Minnesota

summary: CGH was used to compare structural variation among four soybean cultivars (Archer, Minsoy, Noir1 and Williams 82).  Four additional hybridizations were performed with these and other accessions (Kingwa, Williams, M92-220, Richland and Essex) to confirm the patterns observed.

overall_design: Three different soybean cultivars (Archer, Minsoy and Noir1) were hybridized against the common reference genotype Williams 82.  The soybean tiling array consists of 700k probes, spaced at approximately 1.1 kb intervals.  Four additional hybridizations were performed with these and other accessions (Kingwa, Williams, M92-220, Richland and Essex) to confirm the patterns observed.

first: William

last: Haun

first: Pudota

middle: B

last: Bhaskar


title: Microarray analysis of human neuroblastoma cells exposed to aluminum, B-amyloid or the B-amyloid aluminum complex

status: Public on Feb 10 2011

city: Padua

first: Karina

last: Fincati

address: V.le G. Colombo, 3

department: Department of Biology, University of Padua

country: Italy

institute: CNR-Institute for Biomedical Technologies

summary:
A typical pathological feature of Alzheimer’s disease (AD) is the appearance in the brain of senile plaques made up of B-amyloid (AB) and neurofibrillary tangles. Ad is also associated with an abnormal accumulation of some metal ions and we have recently shown that one of these, aluminum (Al), plays a relevant role in affecting AB aggregation and neurotoxicity.  In this study, employing a microarray analysis of 29,166 genes, we investigated the effects induced by the exposure to the AB-Al complex on the gene expression profile of a cell line of neuronal-like cells, the SH-SY5Y.
Results from the microarray essay indicate that, compared to AB or Al alone, exposure to AB-Al produced selective changes in gene expression. Some of the genes selectively over or under expressed are directly related to AD.

overall_design: Analysis used a neuronal cell lines, the SH-SY5Y, exposed to either AB,  the AB-Al complex or Al alone. The SH-SY5Y without treatment were used that a control. For the exposure to AB, six replicates were performed (three dye swap and three non-dye swap), while for exposure to AB-Al and Al  two replicates each were made (one dye swap and one non-dye swap).

first: Denise

last: Drago

first: Valentina

last: Gatta

first: Stefano

last: Sensi

first: Paolo

last: Zatta


title: Aedes aegypti circadian expression profiling

status: Public on Mar 17 2011

city: Fort Collins

first: Andrey

last: Ptitsyn

state: CO

address: 1682 Canpus Delivery

department: Center for Bioinformatics

institute: Colorado State University

summary: Background. Aedes aegypti is arguably the most studied of all mosquito species in the laboratory and is the primary vector of both Dengue and Yellow Fever flaviviruses in the field. A large number of transcriptional studies have been made in the species and these usually report transcript quantities observed at a certain age or stage of development.  However, circadian oscillation is an important characteristic of gene expression in many animals and plants, modulating both their physiology and behavior. Circadian gene expression in mosquito species has been previously reported but for only a few genes directly involved in the function of the molecular clock.    Results. Herein we analyze the transcription profiles of 21,494 messenger RNAs using an Ae. aegypti Agilent® microarray. Transcripts were quantified in adult female heads at 24 hours and then again at 72 hours and eight subsequent time points spaced four hours apart.  We document circadian rhythms in multiple molecular pathways essential for growth, development, immune response, detoxification/pesticide resistance.  Circadian rhythms were also noted in ribosomal protein genes used for normalization in reverse transcribed PCR (RT-PCR) to determine transcript abundance.  We report pervasive oscillations and intricate synchronization patterns relevant to all known biological pathways.   Conclusion. These results argue strongly that transcriptional analyses either need to be made over time periods rather than confining analyses to a single time point or development stage or exceptional care needs to be made to synchronize all mosquitoes to be analyzed and compared among treatment groups.

overall_design:
12 samples representing 48h timeline sampled every 4h. Each chip has one channel taken by sample at ZG24h and the other by ZG72h, ZG76h and so on. Each time point has two independent replicates from two independent pools of mosquitoes sampled on the same calendar day. These time series are concatenated to form one continuous 48h time series for each gene.

The circadian expression analysis presented in the associated paper encompasses a test timeframe of 72 – 92 hour (Cy3-labeled).

Supplementary file: The unabridged matrix of processed data (includes duplicated features) is linked below.

first: Guadalupe

last: Reyes-Solis

first: Karla

last: Saavedra-Rodriguez

first: Jonathan

last: Betz

first: Erica

last: Suchman

middle: O

last: Carlson

last: Black IV


title: Recruitment of BCL6 to target genes in Philadelphia chromosome positive acute lymphoblastic leukemia

city: San Francisco

first: Huimin

last: Geng

state: CA

address: 513 Parnassus Ave., MSB HSW 1201

department: Department of Laboratory Medicine

institute: UCSF

type: Genome binding/occupancy profiling by genome tiling array

summary: The Philadelphia chromosome (Ph) encodes the oncogenic BCR-ABL1 tyrosine kinase, which defines a subset of acute lymphoblastic leukemia (ALL) with a particularly unfavorable prognosis. Tyrosine kinase inhibitors (TKI) are widely used to treat patients with leukemia driven by BCR-ABL1 and other oncogenic tyrosine kinases. In response to TKI-treatment, BCR-ABL1 ALL cells upregulate BCL6 protein levels by ~90-fold, i.e. to similar levels as in diffuse large B cell lymphoma (DLBCL) with BCL6 translocations. In this study, we used genome tiling arrays to identify BCL6 target genes with specific recruitment of BCL6.

overall_design: Three Ph+ ALL cell lines (BV-173, NALM-1 and TOM-1) in duplicate were either treated with 10µM STI571 (Imatinib) for 24 hours or cultured in absence of STI571.

first: Cihangir

last: Duy

first: Christian

last: Hurtz

first: Seyedmehdi

last: Shojaee

first: Leandro

last: Cerchietti

first: Srividya

last: Swaminathan

first: Lars

last: Klemm

first: Soo-mi

last: Kweon

first: Rahul

last: Nahar

first: Melanie

last: Braig

first: Eugene

last: Park

first: Yong-mi

last: Kim

first: Wolf-Karsten

last: Hofmann

first: Sebastian

last: Herzog

first: Hassan

last: Jumaa

first: Phillip

last: Koeffler

first: Jessica

last: Yu

first: Nora

last: Heisterkamp

first: Thomas

middle: G

last: Graeber

first: Hong

last: Wu

first: Hilda

last: Ye

first: Ari

last: Melnick

first: Markus

last: Müschen


title: Stage-specific expression of TNFα regulates bad/bid-mediated apoptosis and RIP1/ROS-mediated secondary necrosis in Birnavirus-infected fish cells

status: Public on Feb 01 2011

city: Taipei

first: Wei-Lun

last: Wang

address: No 128, Sect. 2, Academia Rd, Nankang, Taipei

department: Institute of Cellular and Organismic Biology

country: Taiwan

institute: Academia Sinica

summary: Infectious pancreatic necrosis virus (IPNV) is a fish-derived pathogen and possess a bi-segmented, double-stranded RNA genome. By using zebrafish 14K oligo-microarray and quantitative RT-PCR, we identified differential expression of a defined subset of genes involved in apoptosis and immunity at 6-, 12- and 24- hour after IPNV infection. Transcripts divided into 11 functional categories were significantly modulated by IPNV, including immune response, apoptosis, transcription, signal transduction, lipid and cholesterol metabolism, carbohydrate metabolism, oxidative phosphorylation, cell cycle, protein degradation, protein folding and stress response, protein synthesis, nucleoside metabolism and synthesis. Most of pro-apoptotic bcl-2 family members were up-regulated after IPNV infection. Activation of pro-apoptotic members might disrupt potential of mitochondria and leaded to the mitochondria-mediated apoptosis in the late stage of IPNV infection. After treating the IPNV-infected cells with TNFα inhibitor AP126, expression of two bcl-2 family genes Bad and Bid and activation of caspase-8 and -3 had been inhibited significantly in early stage of IPNV infection. Expression of RIP-1 and Bmf-1 these two necroptosis-related genes and production of ROS were diminished in virus-infected cells which pre-treated with AP126. Our study shows the interactions between host cells and IPNV, the molecular mechanisms involved in IPNV-induced pathogenesis, and the variation of transcriptome through TNFα during infection which shows the important component of host defense. TNFα might lead to apoptosis in early stage and necrosis in late stage in ZF4 cells infected by IPNV. TNFα is crucial to apoptosis and ROS-mediated necrosis caused by IPNV in zebrafish cells.

overall_design:
Zebrafish ZF4 cell line was derived from 24h post-fertilization zebrafish embryos. Previous studies have been shown that adult zebrafish and zebrafish cell line could be infected with IPNV. In the present study, ZF4 cells were infected with IPNV, and total RNA was isolated from infected and uninfected control cells at 0 h, 6 h, 12 h, 24 h post-infection. Microarray analysis gene expression between IPNV-infected and uninfected cells relative to internal control on slides.
The zebrafish 14K oligo microarray we used comprise 1800 zebrafish gene sequences from the NCBI and a database of 12768 putative open reading frames using NCBI zebrafish EST sequence information. Data files were imported into GeneSpring GX 7.3 for further analysis. Expression data sets must pass all the following quality control categories before used for cluster analysis. Clustering analysis allowed us to observe differences in cellular gene expression. A similar expression pattern was seen when comparing differentially expressed host cell genes between 12 and 24 h post-infection. The expression profiles were significantly different between 6 and 12 h post-infection. Therefore, we conclude that the regulation of host gene expression was changed after 12 h post-infection, and progressed into the late stage.
To understand the interactions between host cells and IPNV and the molecular mechanisms involved in IPNV-induced pathogenesis, we used zebrafish oligo microarrays to investigate the gene expression profiles of IPNV-infected zebrafish embryonic cells. We also studied expression of specific genes related to immune response and apoptosis which were not present on the microarray we used by real-time quantitative RT-PCR. We used Pathway Studio’s software to analyze the altered genes in cellular pathway for IPNV, TNFα was shown to be crucial to these genes of host response. Our study proved the variation of transcriptome during infection, which shows the activation of important component of host defense, apoptosis and necrosis through TNFα mediate pathway.

first: Jiann-Ruey

last: Hong

first: Gen-Hwa

last: Lin

first: Hong-Yi

last: Gong

first: Ming-Wei

last: Lu

first: Wangta

last: Liu

first: Ching-Chun

first: Jen-Leih


title: Liver effects of Aflatoxin B1 (AFB1) in wild type (C57BL/6J) and hepatitis C virus-transgenic (HCV-Tg) mice

status: Public on Jan 25 2011

city: Chapel Hill

first: Ivan

last: Rusyn

state: NC

address: 0031 MHRC

institute: University of North Carolina

summary:
Viral hepatitis and aflatoxin B1 (AFB1) exposure are common risk factors for hepatocellular carcinoma (HCC). The incidence of HCC in individuals co-exposed to hepatitis C (HCV) or B virus and AFB1 is greater than could be explained by the additive effect, yet the mechanisms are poorly understood due to lack of an animal model. This study investigated the outcomes and mechanisms of combined exposure to HCV and AFB1. We hypothesized that HCV transgenic (HCV-Tg; expressing core, E1, E2, and p7, nucleotides 342-2771) mice will be prone to hepatocarcinogenesis when exposed to AFB1. Neonatal (7 days old) HCV-Tg or C57BL/6J wild-type mice were exposed to AFB1 (6 ug/g bw) or tricaprylin vehicle (15 ul/g bw) and male offspring were followed for up to 12 months. No liver lesions were observed in vehicle-treated wild type or HCV-Tg mice. Tumors (adenomas or carcinomas) and preneoplastic lesions (hyperplasia or foci) were observed in 22.5% (9 of 40) of AFB1-treated wild-type mice. In HCV-Tg, the incidence of tumorous or pre-tumorous lesions was significantly elevated (50%, 18 of 36), with the difference largely due to a 2.5-fold increase in the incidence of adenomas (30.5% vs 12.5%). While oxidative stress and steato-hepatisis were observed in both AFB1-treated groups, molecular changes indicative of the enhanced inflammatory response and altered lipid metabolism were more pronounced in HCV-Tg mice. In summary, HCV proteins core, E1, E2 and p7 are sufficient to reproduce the additive co-carcinogenic effect of HCV and AFB1 which is a known clinical phenomenon.
HCV transgenic mice (SL-139 strain, pAlbSVPA-HCV-S, containing the structural genes core, E1, E2, and p7, nucleotides 342-2771 of HCV genotype 1b, strain N, under the control of the murine albumin promoter/enhancer) on C57BL/6J (Jackson Laboratory, Bar Harbor, ME) background were previously reported in Lerat et al (Lerat et al., Gastroenterology 122, 352-365, 2002). Transgenic animals were identified after weaning as detailed in Korenaga et al (Korenaga et al., J Biol. Chem. 280, 37481-37488, 2005). Neonatal (7 days old) mice were administered a single dose of AFB1 (6 ug/g bw) or tricaprylin vehicle (15 ul/g bw) by intra-peritoneal injection. Male mice were maintained on the regular animal chow with free access to food and water for up to 12 months. All animal experiments were approved by the UNC Animal Care and Use Committee.

overall_design: There were 20 liver samples used for microarray analysis (12 month time point). All samples were run in one batch. There are 4 groups: WT/Control (4 samples - all biological replicates, i.e., different animals); WT/AFB1 (6 samples); HCV/Control (4 samples); HCV/AFB1 (6 samples). This was a 2 color design with a common reference mRNA. No dye swaps or replicate arrays were included.

first: Emmanuelle

last: Jeannot


title: Expression of microRNAs and their gene targets are dysregulated in pre-invasive breast cancer (microRNA)

city: Boston

first: Bethany

last: Hannafon

state: MA

address: 650 Albany Street

department: Medicine/Section of Hematology and Oncology

institute: Boston University School of Medicine

summary: Introduction: microRNAs (miRNAs) are short non-coding RNAs that negatively regulate gene expression and may play a causal role in invasive breast cancer. Since many genetic aberrations of invasive disease are detectable in earlier stages, we hypothesized that miRNA expression dysregulation and the predicted changes in gene expression would also be found in early breast neoplasias.   Methods: Expression profiling of 365 miRNAs by RT-qPCR was combined with laser-capture microdissection to obtain an epithelial specific miRNA expression signature of normal breast epithelium (n=9) and of paired samples of histologically normal epithelium (HN) and ductal carcinoma in situ (DCIS) (n=16). To determine how miRNAs may control the expression of co-dysregulated mRNAs we also performed gene expression microarray analysis in the same paired HN and DCIS samples and integrated this with miRNA-target prediction. We further validated several target pairs by modulating the expression levels of miRNAs in MCF7 cells and measured the expression of target mRNAs and proteins.  Results: Thirty-five miRNAs were aberrantly expressed between RM, HN and DCIS. Twenty-nine miRNAs and 420 mRNAs were aberrantly expressed between HN and DCIS. Combining these two datasets with miRNA-target prediction we identified two established target pairs (miR-195:CCND1 and miR-21:NFIB) and tested several novel miRNA:mRNA target pairs. Over-expression of the putative tumor-suppressor miR-125b, under-expressed in DCIS, repressed the expression of MEMO1, which is required for ErbB2-driven cell motility (also a target of miR-125b); and NRIP1/RIP140, which modulates the transcriptional activity of the estrogen receptor. Knockdown of the putative oncogenic miRNAs miR-182 and miR-183, both highly over-expressed in DCIS, increased the expression of CBX7 (which regulates E-cadherin expression), DOK4, NMT2, and EGR1. Augmentation of CBX7 by knockdown of miR-182 expression, in turn, positively regulated the expression of E-cadherin, a key protein involved in maintaining normal epithelial cell morphology which is commonly lost during neoplastic progression.   Conclusions: These data provide the first miRNA expression profile of normal breast epithelium and of pre-invasive breast carcinoma. Further, we demonstrate that altered miRNA expression can modulate gene expression changes that characterize these early cancers. We conclude that miRNA dysregulation likely plays a substantial role in early breast cancer development.

overall_design: The expression of 365 microRNAs were measured in 19 total samples via multiplex reverse transcription PCR using the TaqMan Human MicroRNA Low Density Array. Patients age ranged from 42-75. Equal amounts of total RNA from 9 reduction mammoplasty samples (age range 42-75) were combined into a pooled RM control (PRM), this sample was run in triplicate. Remaining 16 samples consist of matched samples of ductal carcinoma in situ and adjacent histologically normal breast epithelium, these are identified by case number and histologic lesion.

middle: N

first: Carol

last: Rosenberg


title: A combined genetic and genomic approach uncovers molecular basis of wine yeast fermentation traits

status: Public on Jan 19 2011

city: Montpellier

first: Bruno

last: Blondin

address: 2 place Pierre Viala

country: France

institute: INRA

summary: Industrial wine yeast strains possess specific abilities to ferment under stressing conditions and give a suitable aromatic outcome. Although the fermentations properties of Saccharomyces cervisiae wine yeasts are well documented little is known on the genetic basis underlying the fermentation traits. Besides, although strain differences in gene expression has been reported, their relationships with gene expression variations and fermentation phenotypic variations is unknown. To both identify the genetic basis of fermentation traits and get insight on their relationships with gene expression variations, we combined fermentation traits QTL mapping and expression profiling in a segregating population from a cross between a wine yeast derivative and a laboratory strain.

overall_design: 40 samples are analysed with 2 technical replicates, using a unique reference named pool of the 30 segregants. The transcriptome of each segregant is compared to the transcriptome of the pool. The transcriptome of 5 biologic replicates of each parental strain is also compared to this reference. An haploid  derivative of the commercialized wine yeast EC1118 which sequence is available (Novo et al. 2009. PNAS, 106:16333-16338) called 59A was used as industrial wine yeast. It is a prototroph strain and has a MATa sexual type. The haploid laboratory strain S288C (MATa) was used for crossing.

first: Chloé

last: Ambroset


title: Transcriptome response of high- and low-light-adapted Prochlorococcus strains to changing iron availability

status: Public on Jan 11 2011

city: Cambridge

first: Anne

last: Thompson

address: 15 Vassar Street

institute: MIT

summary: Prochlorococcus contributes significantly to ocean primary productivity. The link between primary productivity and iron in specific ocean regions is well established and iron limitation of Prochlorococcus cell division rates in these regions has been shown. However, the extent of ecotypic variation in iron metabolism among Prochlorococcus and the molecular basis for differences is not understood. Here, we examine the growth and transcriptional response of Prochlorococcus strains, MED4 and MIT9313, to changing iron concentrations. During steady state, MIT9313 sustains growth at an order-of-magnitude lower iron concentration than MED4. To explore this difference, we measured the whole-genome transcriptional response of each strain to abrupt iron starvation and rescue. Only four of the 1159 orthologs of MED4 and MIT9313 were differentially expressed in response to iron in both strains. However, in each strain, the expression of over a hundred additional genes changed, many of which are in labile genomic regions, suggesting a role for lateral gene transfer in establishing diversity of iron metabolism among Prochlorococcus. Furthermore, we found that MED4 lacks three genes near the iron-deficiency-induced gene (idiA) that are present and induced by iron stress in MIT9313. These genes are interesting targets for studying the adaptation of natural Prochlorococcus assemblages to local iron conditions as they show more diversity than other genomic regions in environmental metagenomic databases.

sample_organism: Prochlorococcus marinus subsp. pastoris str. CCMP1986

overall_design: We used microarrays to measure the whole-genome expression response of Prochlorococcus MED4 and MIT9313 to iron stress and recovery from iron stress.

middle: W

first: Katherine

middle: H

last: Huang

first: Mak

last: Saito

first: Sallie

last: Chisholm


title: Differential Expression of Genes Characterizing Muscle Fiber Phenotype in Turkeys

status: Public on Jan 13 2011

city: Raleigh

first: Lidia

middle: Sylwia

last: Nierobisz

address: 2711 Founders Dr.

institute: North Carolina State University

summary: Because of its high plasticity and rapid growth rate, turkey skeletal muscle is an important model for studying mechanisms responsible for vertebrate skeletal muscle development and function.   Skeletal muscle is composed of metabolically heterogeneous myofibers that exhibit high plasticity at both the morphological and transcriptional levels.  The objective of this study was to employ microarray analysis to elucidate the differential gene expression between the tonic- “red” Anterior latissimus dorsi (ALD) muscle, the phasic- “white” Posterior latissimus dorsi (PLD), and  “mixed”-phenotype Biceps femoris (BF)  in 1-week and 19-week old male turkeys.   A total of 170 differentially expressed genes were identified in the analyzed muscle samples (P<0.05). Gene Go analysis software was utilized to identify top gene networks and metabolic pathways involving differentially expressed genes.  The largest differences were observed between ALD and PLD muscles, where 32 genes were over-expressed and 82  genes were under-expressed in ALD1-PLD1 comparison; and 70 genes were over-expressed and 70 under-expressed in ALD19-PLD19 comparison.   The largest number of genes over-expressed in ALD muscles as compared to other muscles, code for extracellular matrix proteins such as dystroglycan and collagen. Furthermore, a number of genes involved in glycolytic metabolism were under-expressed in ALD muscles as compared to BF and PLD muscles.  The gene analysis revealed that phenotypically “red” BF muscle has high expression of glycolytic genes usually associated with “white” muscle phenotype. Muscle-specific differences were observed in expression levels of genes coding for proteins involved in mRNA processing and translation regulation, proteosomal degradation, apoptosis, and insulin resistance.

overall_design: The study utilized Nicholas turkey males at two different stages of growth (1 week old: high satellite cell mitotic activity; 19 weeks old: low satellite cell mitotic activity, hypertrophy, market age).  Six 1-week-old and six 19-week-old Nicholas turkey males (Meleagris gallopavo) were randomly selected from a single flock. The experiments were designed to compare genes characterizing muscle fiber phenotype at each time point, and to elucidate age-related differences in gene expression levels. Each analyzed cDNA sample was derived from six pooled RNA samples, resulting with six cDNA samples per muscle (one cDNA per bird).  To minimize dye bias, the cDNA samples were labeled either with Cy3 or Cy5.   A total of 17 Turkey Skeletal Muscle Long Oligo (TKSMLO) microarrays were utilized.  Birds were randomly assigned to each array and the cDNA hybridizations we performed at random. The array data was Log2 transformed and then normalized by using locally weighed regression and smoothing within each array and across all arrays.  Mixed model ANOVA with Bonferroni correction of P=0.05 for multiple testing was performed to increase the sensitivity of detected differences in gene expression levels.

middle: S

first: Kelly

middle: R

last: Sporer

first: Gale

last: Strasburg

first: Kent

last: Reed

first: Sandra

last: Velleman

first: Christopher

last: Ashwell

first: J

middle: V

last: Felts

first: Paul

last: Mozdziak


title: Canine normal tissue gene expression profiles

status: Public on Dec 31 2010

city: Gaithersburg

first: Joseph

last: Briggs

state: MD

address: 37 Convent Drive

department: Pediatric Oncology Branch

institute: National Cancer Institute

summary: Our understanding of disease is increasingly informed by changes in gene expression between normal and abnormal tissues.  The release of the canine genome sequence in 2005 provided an opportunity to understand health and disease in the dog using investigative techniques including gene expression profiling.  Accordingly, we now present a publicly accessible canine normal tissue gene expression database that will streamline the study of canine tissues and facilitate comparative genomic analysis with other mammals.  The Affymetrix platform was utilized to catalogue the genetic signatures of normal canine tissues including:  liver, kidney, heart, lung, cerebrum, lymph node, spleen, jejunum, pancreas and skeletal muscle.  The quality of the database was assessed in several ways.  Organ defining gene sets were identified for each tissue and functional enrichment analysis revealed themes consistent with known physio-anatomic functions for each organ.  In addition, a comparison of orthologous gene expression between matched canine and human normal tissues uncovered remarkable similarity. Public access and use of this data, using infrastructure identical to that currently in use for human normal tissues, has been established and allows for additional cross-species comparisons.  To demonstrate the utility of this dataset, novel canine gene annotations were established based on comparative analysis of dog and human tissue selective gene expression and manual curation of canine probeset mapping. It is expected that this dataset will contribute to more advanced study of disease in the dog and biologically robust biomedical studies that utilize the dog as a model for translational research.

overall_design: 39 pathologically normal organ samples were collected from four dogs, 10 organs from each dog.  Organ samples included the pancreas, kidney, liver, lung, heart, skeletal muscle, jejunum, cerebrum, spleen and peripheral lymph node.

first: Melissa

last: Paoloni


title: Interplay between manganese and zinc homeostasis in the human pathogen Streptococcus pneumoniae

status: Public on Feb 02 2011

city: Bloomington

first: Krystyna

last: Kazmierczak

state: IN

address: 1001 E. Third St.

department: Biology

institute: Indiana University

summary: ICP-MS analysis of Streptococcus pneumoniae D39 reveals a high cell-associated manganese (Mn(II)) concentration that is comparable to that of zinc (Zn(II)).  Stressing these cells with 100-200 µM Zn(II) leads to a slow-growth phenotype and a reduction in total Mn(II) concentration, with no decrease in the concentrations of other metal ions. Supplementation of the growth media with as little as 10 µM Mn(II) fully restores wildtype growth patterns and normal levels of cell-associated Mn(II).  DNA microarray analysis reveals that zinc stress induces the expected upregulation of czcD (encoding a zinc effluxer), but also a pleiotropic transcriptional response suggestive of mild cell wall stress. Genes encoding a nitric oxide detoxification system (nmlR) and the Mn(II) uptake system (psaABC) are also induced.  We conclude that Zn(II) toxicity results in a cytoplasmic Mn(II) deficiency, possibly caused by competition at the Mn(II) uptake transporter protein PsaA.

overall_design:
Bacteria were grown statically in Brain Heart Infusion media (Bacto BHI, Becton Dickinson) or BHI supplemented with metals at 37C in an atmosphere of 5% CO2 to a culture density of OD620~0.2 and were processed as described in the related Sample records. Samples were collected from three independent biological replicates, including one dye swap.
Data were analyzed using software from the TM4 Microarray Software Suite (www.tm4.org). GenePix results files (.gpr) were converted to TIGR MultiExperiment Viewer file format (.mev) using ExpressConverter 2.1 software and median spot intensities. Lowess (block) normalization was performed on non-background subtracted data using TIGR MIDAS 2.21 software. Spots flagged as bad (A=0 non-saturated pixels in spot; X= low spot intensity relative to background) were removed from the analysis. The cut-off for statistical significance of differential expression was set at Bayesian P =0.001 and an average up or down fold change of at least 1.8 fold.
The data were normalized both with and without removal of flagged spots and used to calculate expression ratios and Bayesian P values. Ratios and P values did not differ significantly with or without removal of flagged spots.

first: Faith

last: Jacobsen

first: John

middle: P

last: Lisher

first: Malcolm

last: Winkler

first: David

last: Giedroc


title: Transcriptome analysis of Bacillus cereus B3711 during silver ion stress condition

status: Public on Dec 14 2010

city: Bangalore

first: Genotypic

last: technology

state: Karnataka

address: 259, Apoorva 4th cross,80 feet Road,RMV 2ND STAGE

country: India

institute: Genotypic Technology

summary: We have employed a whole genome microarray for gene expression profilling to understand the global view of the cellular mechanism and metabolic response of B. cereus B3711 exposed to 1 mM silver nitrate. RNA was extracted from B. cereus grown under silver treated and untreated set of conditions in duplicates at two different time points (30 & 60 min). Comparing the transcript profile of the control and treated cells, several differentially expressed genes were identified. These includes genes involved in various metabolic pathways, such as arginine and alanine metabolism, membrane transport system and alternate respiratory proteins such as arsenate reductase, stress related proteins, proteins of transcriptional regulators, hypothetical proteins and protein of two-component systems.

overall_design: Agilent one-color experiment,Organism: Bacillus cereus  ,Agilent-023971 Genotypic designed Custom Bacillus cereus 8x15k  , Labeling kit: Agilent Quick-Amp labeling Kit (p/n5190-0442)

first: M

last: Ganesh Babu

first: P

last: Gunasekaran


title: MiRNAs expression data from Atm+/+ and Atm-/- mouse embryonic fibroblasts after DNA damage at various time point

status: Public on Dec 09 2010

city: Houston

first: Guohui

last: Wan

state: TX

address: 7777 Knight Road

department: Cancer Biology

institute: The University of Texas M.D. Anderson Cancer Center

summary: Atm+/+ and Atm-/- mouse embryonic fibroblasts were treated with DNA damaging agent neocarzinostatin (NCS), and cells were harvested at indicated time points for the microarray analyses of whole-genome miRNAs.

overall_design: To examine how miRNAs are regulated in the DNA damage response, we assessed the genome-wide mature miRNA expression in Atm+/+ and Atm-/- littermate mouse embryonic fibroblasts (MEFs).


title: Study of ecs mutation

status: Public on Jan 01 2011

city: Geneva

first: FRANCOIS

last: Patrice

state: Ge 4

address: Rue Gabrielle-Perret-Gentil, 4

department: Service of Infectious Diseases

country: Switzerland

institute: Genomic Research Laboratory

summary:
Ecs is an ATP-binding cassette (ABC) transporter present in aerobic and facultative anaerobic gram-positive Firmicutes. Inactivation of Bacillus subtilis Ecs causes pleiotropic changes in the bacterial phenotype including inhibition of intramembrane proteolysis. The molecule(s) transported by Ecs is (are) still unknown.
In this study we mutated the ecsAB operon in two Staphylococcus aureus strains, Newman and LS-1. Phenotypic and functional characterization of these Ecs deficient mutants revealed a defect in growth, increased autolysis and lysostaphin sensitivity, altered composition of cell wall proteins including the precursor form of staphylokinase and an altered bacterial surface texture. DNA microarray analysis indicated that the Ecs deficiency changed expression of the virulence factor regulator protein Rot accompanied by differential expression of membrane transport proteins, particularly ABC transporters and phosphate-specific transport systems, protein A, adhesins and capsular polysaccharide biosynthesis proteins. Virulence of the ecs mutants was studied in a mouse model of hematogenous S. aureus infection. Mice inoculated with the ecs mutant strains developed markedly milder infections than those inoculated with the wild-type strains and had consequently lower mortality, less weight loss, milder arthritis and decreased persistence of staphylococci in the kidneys. The ecs mutants had higher susceptibility to ribosomal antibiotics and plant alkaloids chelerythrine and sanguinarine.
Our results show that Ecs is essential for staphylococcal virulence and antimicrobial resistance probably since the transport function of Ecs is essential for the normal structure and function of the cell wall. Thus targeting Ecs may be a new approach in combating staphylococcal infection.

overall_design: WT and ecsA mutant strains were hybridized at 3 and 6 hours of growth in rich medium (4-replicates)

first: Jarmo

middle: T

last: Juuti

first: Manfred

last: Saller

first: Arnold

last: Driessen

first: Vesa

last: Kontinen


title: IFN-γ Upregulates Survivin and Ifi202 Expression to Induce Survival and Proliferation of Tumor-Specific T Cells

status: Public on Dec 03 2010

city: Augusta

first: Kebin

last: Liu

state: GA

address: 1410 Laney Walker Blvd

department: Biochemistry and Molecular Biology

institute: Medical College of Georgia

summary: Transcriptional profiling of mouse tumor-specific cytotoxic T lymphocytes (CTL) comparing activation-induced gene expression profiles induced by tumor-specific antigen (Ag) and CD3 mAb. CTLs were stimulated with either Ag or CD3 mAb for 3 and 24 h, respectively. The stimulated CTLs were compared to un-stimulated CTLs. The objective was to identify differential gene expression profiles induced by the two activation (Ag vs CD3 mAb) conditions.

overall_design: Two-condition experiments, un-stimulated vs stimulated CTLs. Biological replicates: 3 replicates for each comparison. Unstimulated CTLs were compared to stimulated CTLs (Ag and CD3 mAb, respectively).

first: Mary

last: Zimmerman

first: Dafeng

last: Yang

first: Xiaolin

last: Hu

first: Feiyan

first: Nagendra

last: Singh

first: Darren

last: Browning

first: Vadivel

last: Ganapathy

first: Divaker

last: Choubey

first: Phillip

last: Chandler

first: Scott

middle: I

last: Abrams


title: Microorganisms with novel dissimilatory (bi)sulfite reductase genes are widespread and part of the core microbiota in low-sulfate peatlands.

status: Public on Nov 26 2010

city: Vienna

first: Doris

last: Steger

state: Vienna

address: Althanstrasse 14

department: Department of Microbial Ecology

institute: University of Vienna

summary: Peatlands of the Lehstenbach catchment (Germany) house so far unidentified microorganisms with phylogenetically novel variants of the dissimilatory (bi)sulfite reductase genes dsrAB. These genes are characteristic for microorganisms that reduce sulfate, sulfite, or some organosulfonates for energy conservation, but can also be present in anaerobic syntrophs. However, nothing is currently known regarding the abundance, community dynamics, and biogeography of these dsrAB-carrying microorganisms in peatlands. To tackle these issues, soils from a Lehstenbach catchment site (Schlöppnerbrunnen II fen) from different depths were sampled at three time points over a six-year period to analyze the diversity and distribution of dsrAB-containing microorganisms by a newly developed functional gene microarray and quantitative PCR assays. Members of novel, uncultivated dsrAB lineages (approximately representing species-level groups) (i) dominated a temporally stable but spatially structured dsrAB community and (ii) represented ‘core’ members (up to 1-1.7% relative abundance) of the autochthonous microbial community in this fen. In addition, denaturing gradient gel electrophoresis (DGGE)- and clone library-based comparison of the dsrAB diversity in soils from a wet meadow, three bogs, and five fens of various geographic locations (distance ~1-400 km), identified one Syntrophobacter-related and nine novel dsrAB lineages to be widespread in low-sulfate peatlands. Signatures of biogeography in dsrB-DGGE data were not correlated with geographic distance but could largely be explained by soil pH and wetland type, implying that distribution of dsrAB-carrying microorganisms in wetlands on the scale of a few hundred kilometers is not limited by dispersal but determined by contemporary environmental conditions.

sample_organism:
uncultured sulfate-reducing bacterium
environmental samples
environmental samples

overall_design: 36 dsrAB clones for chip evaluation, 33 hybridizations of labeled dsrAB RNA from environmental peatsoil samples

first: Manuel

last: Hofer

first: Alexander

last: Loy


title: Differential gene expression in lactating rats compared against age matched virgin controls

status: Public on Nov 21 2010

city: Lexington

last: Vore

state: KY

address: 1095 Va Drive

department: Graduate Center of Toxicology

institute: University of Kentucky

summary:
Gene level analysis of lactating day 10-11 16h Sprague-Dawley dams compared against age matched virgins.
Lactation is a time of significantly increased energy demands imposed by the suckling young that requires a proportional adjustment in the ability of the lactating dam to absorb nutrients and to synthesize critical biomolecules to meet the dietary needs of both the offspring and the dam.  We have shown an increase in the size and hydrophobicity of the bile acid pool during lactation [1], implying an increased absorption and disposition of lipids, sterols, nutrients, and xenobiotics.   In order to investigate changes at the level of mRNA, we utilized an exon array and calculated gene level summaries to investigate changes in gene expression in the liver, duodenum, jejunum, and ileum of lactating dams when compared against age-matched virgin controls.

overall_design: Affymetrix Rat Exon 1.0ST arrays were used to determine expression of genes in the livers and small intestines of day 10-11 lactating Sprague Dawley rats and compared against virgins of comparable age.

first: Antony

last: Athippozhy

first: Liping

last: Huang

first: Clavia

last: Wooton-Kee

first: Tianyong

last: Zhao

first: Paiboon

last: Jungsuwadee

middle: J

last: Stromberg


title: Global transcriptome analysis indicates that depletion of rps13 and ribosomes leads to cell cycle arrest in G1

status: Public on Nov 29 2010

city: St Louis

first: Michael

middle: Sean

last: Behnke

state: MO

address: 660 S. Euclid Ave

department: Molecular Microbiology

institute: Washington University School of Medicine

summary: Affymetrix Toxoplasma microarray data was analyzed to determine whether there was an association with a G1 transcript profile. Four hundred seventy-five genes with significant gene expression differences (Welch T-test, P = 0.01) between ±ATc samples were identified. Genes were clustered into those increased (153) or decreased (322) -ATc. There are a number of genes with diminished expression and pronounced over expression -ATc including LDH1, 4 zinc finger proteins, leucine-rich protein, and ABC transporter-like molecules, myosin-related molecules with many fold increases and one AP2 protein with a small increase. These gene sets have a large overlap with G1 genes defined in an ongoing cell cycle expression study in T. gondii (Behnke, White). Over 60% of those genes that have diminished expression -ATc overlap with genes that are diminished at G1, and over 40% of genes with increased expression -ATc overlap with genes that are up regulated at G1. A hypergeometric distribution of this association suggests that the probability of this happening by chance is essentially zero. Thus, by both methods, FACS and arrays, parasites -ATc display a G1 profile.

overall_design: To determine the global transcriptional differences of the RSP13 conditional mutant with and without anhydrotetracycline (ATc). Four replicates each.

first: Rima

last: McLeod

last: White


title: Chemokine Transcripts as Targets of the RNA-Binding Protein HuR in Human Airway Epithelium

status: Public on Nov 10 2010

city: Baltimore

first: Jinshui

last: Fan

address: 5200 Eastern Avenue

department: Division of Allergy & Clinical Immunology

institute: Johns Hopkins University

summary:
HuR is a regulator of mRNA turnover or translation of inflammatory genes through binding to adenylate-uridylate-rich elements (ARE) and related motifs present in the 3’untranslated region (UTR) of mRNAs.
We aimed to identify HuR targets in the human airway epithelial cell line BEAS-2B challenged with TNFa plus IFNg, a strong stimulus for inflammatory epithelial responses.  Ribonucleoprotein (RNP)
complexes from resting and cytokine-treated cells were immunoprecipitated (IP) using anti-HuR and isotype-control antibody, and eluted mRNAs were reverse-transcribed and hybridized to an inflammatory-focused
gene array.  The chemokines CCL2, CCL8, CXCL1 and CXCL2 ranked highest among 27 signaling and inflammatory genes significantly enriched in the HuR RNP-IP from stimulated cells over the control IP.
Among these, 20 displayed published HuR binding motifs. Association of HuR with the four endogenous chemokine mRNAs was validated by single-gene RNP-IP, and shown to be 3’UTR-dependent by biotin
pull-down assay.  Cytokine treatment increased mRNA stability only for CCL2 and CCL8, and transient silencing and overexpression of HuR affected only CCL2 and CCL8 expression in primary and transformed
epithelial cells.  Cytokine-induced CCL2 mRNA was predominantly cytoplasmic; conversely, CXCL1 mRNA remained mostly nuclear and unaffected, as CXCL2, by changes in HuR levels.  Increase in cytoplasmic
HuR and HuR target expression partially relied on the inhibition of AMP-dependent kinase, a negative regulator of HuR nucleocytoplasmic shuttling.

overall_design:
We postulate that HuR critically regulates the epithelial response, by associating with multiple adenylate-uridylate-rich elements (ARE)-bearing, functionally related inflammatory transcripts.  On the basis of
genome-wide studies probing the relationship between RNA-binding proteins and the functional profile of their associated transcripts, we combined the specific HuR immunoprecipitation of RNPs and the
genome-scale microarray to profile the target mRNAs of HuR in the human airway epithelial cell line BEAS-2B challenged with very strong inflammatory stimulation, TNFa plus IFNG. The array platform
we used is the Human Autoimmune and Inflammatory Response Gene Array (SuperArray Bioscience, Frederick, MD). To validate the association of HuR and its target mRNAs, we planned to apply biotin pull-down
assay. HuR overexpression and knockdown assays were also planned to further verify the association of HuR and its target mRNAs. Overall, we did three biological replicates for the IP array experiments, which
include both IgG1 control IP and HuR IP arrays.

first: Faoud

last: Ishmael

first: Xi

last: Fang

first: Chris

last: Cheadle

first: Allen

last: Myers

first: Shau-Ku

first: Ulus

last: Atasoy

first: Myriam

last: Gorospe

first: Cristiana

last: Stellato


title: The zinc cluster transcription factor Ahr1p directs Mcm1p regulation of Candida albicans adhesion

city: Montreal

first: Adnane

last: Sellam

state: Quebec

address: 6100 Royalmount

department: Anatomy and Cell Biology

institute: Adnane Sellam

summary: Biofilm development by Candida albicans requires cell adhesion for the initial establishment of the biofilm and the continued stability after hyphal development occurs; however, the regulation of the process has not been fully established. Using chromatin immunoprecipitation coupled to microarray analysis (ChIP-chip) we have characterized a regulon containing the Mcm1p factor that is required for the initial surface adhesion during biofilm formation. In the yeast Saccharomyces cerevisiae several Mcm1p regulons have been characterized in which regulatory specificity is achieved through co-factors binding a sequence adjacent to the Mcm1p-binding site. This new Mcm1p regulon in C. albicans also requires a co-factor, which we identify as the transcription factor Ahr1p. However, in contrast to the other yeast regulons, Ahr1p alone binds the target promoters, which include several key adhesion genes, and recruits Mcm1p to these sites. Through transcription profiling and qPCR analysis, we demonstrate that this Ahr1p-Mcm1p complex directly activates these adhesion genes. When the regulon was disrupted by deleting AHR1, the strain displayed reduced adherence to a polystyrene surface. We also demonstrate a role for the regulon in hyphal growth and in virulence. Our work thus establishes a new mechanism of Mcm1p-directed regulation distinct from those observed for other Mcm1p co-regulators.

overall_design: We performed genome wide occupancy experiments (YPD, 30oC) with Ahr1p and Mcm1p to determine their binding sites. To confirm an interaction between the two factors we also performed genome wide occupancy with Mcm1p in an ahr1 deletion strain. To complement with genome wide occupancy experiments, we performed a transcription profile with an ahr1 deletion strain under yeast conditions (YPD, 30oC).

first: Christopher

last: Askew

first: Elias

last: Epp

first: Jaideep

last: Mallick

first: Herve

last: Hogues

first: Alaka

last: Mullick

first: Andre

last: Nantel

first: Malcolm

last: Whiteway


title: Molecular characterization of AML with ins(21;8)(q22;q22q22) reveals similarity to t(8;21) AML

status: Public on Oct 26 2010

city: Berlin

first: Lars

last: Bullinger

address: Augustenburger Platz 1

department: Hematology, Oncology and Tumorimmunology

country: Germany

institute: Charité

summary: In acute myeloid leukemia (AML) non-random clonal chromosome aberrations are detectable in ~55% of adults with AML. Translocation t(8;21)(q22;q22) resulting in the 5'RUNX1/3'RUNX1T1 fusion gene occurs in ~8% of acute myeloid leukemia (AML) cases. Also, insertions ins(8;21) and ins(21;8) have been described that show a broad heterogeneity at the molecular level with inserted fragment sizes ranging from 2.4 to 44 Mb. Microarray-based comparative genomic hybridization (arrayCGH) in 49 intermediate-risk AML and RT-PCR-based screening in 532 AML cases allowed the detection of ins(21;8)/ins(8;21) in three cases; arrayCGH and subsequent RT-PCR revealed an ~0.5 Mb sized inserted fragment generating the 5'RUNX1/3'RUNX1T1 fusion gene in one case with a submicroscopic ins(21;8)(q22;q22q22) whereas the other two cases were identified by banding analysis and RT-PCR, respectively. Gene expression profiling (GEP) and a detailed review of the literature highlighted similar biological features of AML cases with ins(21;8)/ins(8;21) and t(8;21)(q22;q22). Our study demonstrates the potential of high-resolution array-based analysis and GEP and provides further evidence that AML with insertions generating the 5'RUNX1/3'RUNX1T1 fusion not only biologically resemble the t(8;21)(q22;q22) AML subgroup, but might also share their prognostically favorable clinical behavior. Thus, similar treatment options should be considered in these patients.

overall_design: An all pairs experiment design type is where all labeled extracts are compared to every other labeled extract.


title: Stepwise histone replacement by SWR1 requires dual activation with histone H2A.Z and canonical nucleosome

status: Public on Oct 12 2010

city: Bethesda

first: Carl

last: Wu

address: 37 Convent Dr, 6068

department: LBMB

institute: NIH/NCI

summary: Histone variant H2A.Z-containing nucleosomes are incorporated at most eukaryotic promoters. This incorporation is mediated by the conserved SWR1 complex, which replaces histone H2A in canonical nucleosomes with H2A.Z in an ATP-dependent manner. Here, we show that promoter-proximal nucleosomes are highly heterogeneous for H2A.Z in Saccharomyces cerevisiae, with substantial representation of nucleosomes containing one, two, or no H2A.Z molecules. SWR1-catalyzed H2A.Z replacement in vitro occurs in a stepwise and unidirectional fashion, one H2A.Z-H2B dimer at a time, producing heterotypic nucleosomes as intermediates and homotypic H2A.Z nucleosomes as end products. The ATPase activity of SWR1 is specifically stimulated by H2A-containing nucleosomes without ensuing histone H2A eviction. Remarkably, further addition of free H2A.Z-H2B dimer leads to hyperstimulation of ATPase activity, eviction of nucleosomal H2A-H2B and deposition of H2A.Z-H2B. These results suggest that the combination of H2A-containing nucleosome and free H2A.Z-H2B dimer acting as both effector and substrate for SWR1 governs the specificity and outcome of the replacement reaction.

overall_design: Total nucleosomes from MNase-treated nuclear extracts were fractionated by sequential immunoprecipitation into homotypic H2A/H2A (AA), heterotypic H2A/H2A.Z (AZ), and homotypic H2A.Z/H2A.Z (ZZ) nucleosomes.

first: Ed

last: Luk


title: Chromatin regulators as capacitors of inter-species expression variations

status: Public on Nov 01 2010

city: Rehovot

first: Itay

last: Tirosh

address: herzl

department: Molecular genetics

country: Israel

institute: Weizmann Institute of Science

summary:
We analyzed the genome-wide expression profiles of mutant strains of two yeast species in which eight different chromatin regulators and one transcription factor were deleted (one gene deleted in each strain). Comparison of the inter-species expression differences between the wild-types of these species and their mutants showed that deletion of regulators tends to increase the amount of inter-species expression differences, suggestng that the regilators are normally buffering hidden variability. In addition, we measured allele-specific expresion levels of the interspecific hybrid formed by mating each of the two corresponding mutants (deleted for the same regulator).
Keywords: Comparative transcriptome analysis

sample_organism: Saccharomyces cerevisiae x Saccharomyces paradoxus

platform_organism:
Saccharomyces cerevisiae
Saccharomyces paradoxus

overall_design: Nine gene deletions were examined. For each gene deletion, there are four datasets - S.cerevisiae and S. paradoxus or the hybrid each with two biological repeats. These four datasets are contained within a single slide of our custom two-species microarray, as it includes two subarrays (blocks) and hybridization was performed with two dyes.


title: MicroRNA Expression in Multiple Myeloma is Associated with Genetic Subtype, Isotype and Prognostic Outcome

status: Public on Sep 29 2010

city: Oxford

first: charles

last: lawrie

address: John Radcliffe Hospital

department: NDCLS

country: United Kingdom

institute: University of Oxford

summary: MicroRNAs are small RNA species that regulate gene expression post-transcriptionally and are aberrantly expressed in many cancers including lymphoma. However, the role of microRNAs in the pathogenesis of multiple myeloma (MM) is poorly understood. We therefore elucidated the complete miRNome of purified tumor (CD138+) cells from 33 patients with MM, 5 patients with monoclonal gammopathy of undetermined significance (MGUS) and 9 disease controls. Unsupervised cluster analysis revealed that MM samples have a distinct microRNA expression profile. The majority of microRNAs aberrantly expressed in MM (109/129) were up-regulated. A comparison of these microRNAs with those aberrantly expressed in other B-cell and T-cell lymphomas revealed a surprising degree of similarity (~40%) suggesting the existence of a common lymphoma microRNA signature. We identified 39 microRNAs associated with the pre-malignant condition MGUS. Twenty-three (59%) of these were also aberrantly expressed in MM suggesting common microRNA expression events in MM progression. MM is characterized by multiple chromosomal abnormalities of varying prognostic significance. We identified specific microRNA signatures associated with the most common IgH translocations (t(4;14) and t(11;14)) and del(13q). Expression levels of these microRNAs were distinct between the genetic subtypes (by cluster analysis) and correctly predicted these abnormalities in >85% of cases using the support vector machine algorithm. Additionally, we identified microRNAs associated with light chain only myeloma, as well as IgG and IgA-type MM. Finally, we identified 32 microRNAs associated with event-free survival (EFS) in MM, ten of which were significant by univariate (logrank) survival analysis. In summary, this work has identified aberrantly expressed microRNAs associated with the diagnosis, pathogenesis and prognosis of MM, data which will prove an invaluable resource for understanding the role of microRNAs in this devastating disease.

platform_organism:
Gallus gallus
Homo sapiens
Mus musculus
Rattus norvegicus

overall_design: Plasma cells (CD138+) cells were purified from bone marrow samples of 33 patients with MM, 5 patients with monoclonal gammopathy of undetermined significance (MGUS) and 9 disease controls.

first: Charles

last: Lawrie

first: Erica

last: Ballabio

first: Xiao-He

last: Chen

first: Rajko

last: Kušec

first: Steve

last: Taylor

first: Debbie

last: Hay

first: Daniela

last: Tramonti

first: Nigel

last: Saunders

first: Tim

last: Littlewood

first: Francesco

last: Pezzella

first: Jackie

last: Boultwood

first: Jim

last: Wainscoat

first: Chris

last: Hatton

first: Jianxiang

last: Chi


title: Coordinated Progression Through Two Subtranscriptomes Underlies the Tachyzoite Cycle of Toxoplasma gondii

status: Public on Aug 27 2010

first: Michael

last: Behnke

summary: We describe the cell cycle transcriptome of the Toxoplasma gondii that has emerged as a major genetic model for the study of Apicomplexa parasites.  Two distinct transcriptional waves accompany the relatively simple binary replication of tachyzoite stage (endodyogeny) functionally separating conserved gene expression in the eukaryotic G1 phase from the lineage-specific expression that predominates in the parasite S phase and mitotic periods.  This division of transcriptional focus closely mirrors the intimate relationship that has evolved between mitosis and building of the daughter parasites and invasion organelles.  Promoter mechanisms appear to orchestrate the induction of gene expression in dividing tachyzoites with up to two dozen cell cycle AP2 factors likely acting within a transcriptional regulatory network to coordinate the parasite cell cycle transcriptome.

overall_design: To characterize the cell cycle transcriptome of Toxoplasma tachyzoites, we expanded a thymidine-synchrony model to isolate sufficient RNA for microarray expression studies.  The ToxoGeneChip microarray (http://ancillary.toxodb.org/docs/Array-Tutorial.html) was used to measure mRNA expression in 13 duplicate samples (R0 to R12) covering 12 hours post-synchronization and nearly two tachyzoite replication cycles.

first: John

middle: C

last: Wootton

first: Margaret

middle: M

last: Lehmann

first: Josh

last: Radke

first: Olivier

last: Lucas

first: Julie

last: Nawas

first: L

last: Sibley

last: White


title: Genome wide profiling of HrpL-regulated genes in the necrotrophic plant pathogen Dickeya dadantii 3937

status: Public on Oct 21 2010

city: Golden

first: Shihui

last: YANG

address: 1617 Cole Blvd

department: National Bioenergy Center

institute: National Renewable Energy Lab

summary:
Background
Dickeya dadantii is a necrotrophic pathogen causing disease in many plants. Previous studies have demonstrated that the type III secretion system (T3SS) of D. dadantii is required for full virulence. HrpL is an alternative sigma factor that binds to the hrp box promoter sequence of T3SS genes to up-regulate their expression.

Methodology/Principal Findings
To explore the inventory of HrpL-regulated genes of D. dadantii 3937 (3937), transcriptome profiles of wild-type 3937 and a hrpL mutant grown in a T3SS-inducing medium were examined. Using a cut-off value of 1.5, significant differential expression was observed in sixty-three genes, which are involved in various cellular functions such as type III secretion, chemotaxis, metabolism, regulation, and stress response. A hidden Markov model (HMM) was used to predict candidate hrp box binding sites in the intergenic regions of 3937, including the promoter regions of HrpL-regulated genes identified in the microarray assay. In contrast to biotrophic phytopathgens such as Pseudomonas syringae, among the HrpL up-regulated genes in 3937 only those within the T3SS were found to contain a hrp box sequence. Moreover, direct binding of purified HrpL protein to the hrp box was demonstrated for hrp box-containing DNA fragments of hrpA and hrpN using the electrophoretic mobility shift assay (EMSA). In this study, a putative T3SS effector DspA/E was also identified as a HrpL-upregulated gene, and shown to be translocated into plant cells in a T3SS-dependent manner.

Conclusion/Significances
We provide the genome-wide study of HrpL-regulated genes in a necrotrophic phytopathogen (D. dadantii 3937) through a combination of transcriptomics and bioinformatics, which led to identification of several effectors. Our study indicates the extent of differences for T3SS effector protein inventory requirements between necrotrophic and biotrophic pathogens, and may allow the development of different strategies for disease control for these different groups of pathogens.

overall_design: Analysis used aerobic steady state RNA from wild-type as control samples for comparison to the experimental samples of hrpL mutant taken 6 h post-inoculation in T3SS inducible minimum media. The microarray results are based on the geomean of ten normalized expression values derived from three biological replicates and two dye-swap experiments with a technology replicate for each array.

last: Yang

first: Quan

last: Peng

first: Qiu

last: Zhang

first: Lifang

last: Zou

first: Yan

last: Li

first: Christelle

last: Robert

first: Leighton

last: Pritchard

first: Hui

last: Liu

first: Raymond

last: Hovey

first: Qi

last: Wang

first: Paul

last: Birch

first: Ian

middle: K

last: Toth

first: Ching-Hong


title: New insights on resistance mechanisms to antimicrobial compounds in a plant pathogen biofilm

status: Public on Aug 31 2010

city: Cordeiropolis

first: Alessandra

middle: Alves

last: De Souza

state: Sao Paulo

address: Rod Anhanguera KM 158

department: Centro de Citricultura Sylvio Moreira

country: Brazil

institute: Instituto Agronomico de Campinas

summary: Investigation of whole genome gene expression level changes in Xylella fastidiosa 9a5c biofilm, submitted to treatments with sub inhibitory and inhibitory concentrations of copper and tetracycline.

overall_design: A study of Xylella fastidiosa 9a5c was done using total RNA recovered from biofilm bacterial cells submitted to 3 or 7mM of CuSO4 or 100 or 800 µg/ml of tetracycline.  Each chip measures the expression level of 2832 genes from Xylella fastidiosa 9a5c with thirteen 60-mer probe pairs (PM/MM) per gene, with five-fold technical redundancy.

first: Ligia

last: Muranaka

first: Marco

middle: A

last: Takita

first: Jacqueline

last: Olivato

first: Luciano

last: Kishi

first: Marcos

last: Machado

last: Souza


title: Sex-biased expression in heads of three Drosophila species

status: Public on Jul 30 2010

first: Brian

last: Oliver

address: 50 South Drive

department: LBG

institute: NIDDK, NIH

summary: Investigation of sex-biased expression across species have relied on measurements from whole flies which sample the extensive expression differences found in the germline and gonads of females and males. We wanted to examine genes with sex-biased expression in a somatic tissue to analyze patterns of genes with sex-biased expression in the context of a tissue more phenotypically similar between females and males. We used a Nimblegen microarray designed for Drosophila melanogaster and two custom micoarrays designed for D. pseudoobscura and D. mojavensis to survey gene expression differences in heads of females and males.

sample_organism:
Drosophila melanogaster
Drosophila mojavensis
Drosophila pseudoobscura

overall_design: PolyA+ mRNA was isolated in quadruplicate from dissected heads of 8 day old adult females and males. Female and male mRNA were labeled with Cy3 or Cy5 dyes separately and co-hybridized to species-specific microarrays. We analyzed a total of 24 channels of data.

first: John

last: Malone


title: Expression Profile of Chicken Primitive Streak (HH4) Tissues

status: Public on Jul 29 2010

city: Kobe

first: Guojun

last: Sheng

state: Hyogo

address: 2-2-3, Minatojima-minamimachi, Chuo-ku

department: Center for Developmental Biology

country: Japan

institute: RIKEN

summary: In birds and mammals, all mesoderm cells are generated from the primitive streak. Nascent mesoderm cells contain unique dorso-ventral (D/V) identities depending on their relative ingression position along the streak. Molecular mechanisms controlling this initial phase of mesoderm diversification are not well-understood. Using chick model, we generated high-quality transcriptomic datasets of different streak regions and analyzed their molecular heterogeneity.

overall_design: Streak tissues were dissected from stage-matched HH4 chick embryos and were further divided into four equal pieces along its anterio-posterior length (termed “A”, “B”, “C” and “D”, with “A” representing most anterior and therefore most dorsal and “D” most posterior and therefore most ventral). Each of the four regions was represented by three independent samples (n=3), with each sample containing 5-7 μg total RNA derived from 85-93 pooled pieces. 5 μg of RNA from each sample were used to screen Affymetrix Chicken Genome Array without an amplification step.

first: Cantas

last: Alev

first: Yuping

last: Wu

first: Takeya

last: Kasukawa

first: Lars

last: Jakt

first: Hiroki

last: Ueda


title: Regional Immune Response to Trichostrongylis colubriformis - Gene Expression during the Development of Immunity

status: Public on Sep 01 2010

city: Puddle Alley

first: Alan

middle: Francis

last: McCulloch

state: Mosgiel

address: Invermay

country: New Zealand

institute: AgResearch NZ

summary: Afferent Lymph was collected over a 8 week period from up to 5 pen raised, nematode naïve, outbreed Romney sheep (Tag No’s  1001, 1002, 1003, 1004, 1006). During this time a Pre-Infection sample was collected, the sheep were immunised by 3 truncated (2 week) T. colubiformis infections (40,000 Tc L3, Immunising Infections 1-3). The treatment regime (shown below) will enable us to determine changes in gene expression during the development of immunity

overall_design: The overall design is summarised in the below table.

first: Jacqui

last: Knight

first: Tony

last: Pernthaner

first: Wayne

last: Hein


title: Lhx1 is required for specification of renal progenitor cell field

city: Hong Kong

first: Hui

last: Zhao

address: The Chinese University of Hong Kong

department: School of Biomedical Sciences

country: Hong Kong

institute: The Chinese University of Hong Kong

summary: The goal of this study is to investigate the molecular mechanism of lhx1 on regulation of pronephros formation during the early embryonic development. In the vertebrate embryo the kidney is derived from the intermediate mesoderm. The LIM-class homeobox transcription factor lhx1 is expressed early in the intermediate mesoderm and is one of the first genes to be expressed in the nephric mesenchyme. The animal cap cells can be induced by treatment of activin and retinoic acid to differentiate into pronephros tissue. In this study we investigated the role of Lhx1 in differentiation of pronephros by depleting lhx1 in the organ culture system. We generated the gene expression profile of early pronephros tissue, and demonstrated that expression of genes from all the kidney domains is affected by the absence of lhx1. Taken together our results highlight an essential role for Lhx1 in pronephros formation.

overall_design:
lhx1 is involved in driving specification of intermediate mesoderm into nephrogenic mesenchyme. Lhx1 is initially expressed throughout the entire intermediate mesoderm. To determine the role of lhx1 pronephros formation, we performed a microarray analysis using an explant culture system. Xenopus tissue explants can be surgically isolated and cultured under specific conditions to be driven towards many distinct tissue types. Formation of pronephric cell fates is induced by culturing isolated explants in the presence of Activin and RA (AcRA). Treatment of dissected explants of stage 9 blastulae embryos with 10ng/ml Activin and 1x10-4 M retinoic acid can induce differentiation of the pluripotent ectoderm into pan-kidney tissue. For this experiment, both blastomeres of 2-cell embryos were injected with a total of 800pg lhx1 DEED-AS. Explants were dissected and treated with AcRA and expression of pax8 at stage 15 (based on timing of paired control whole embryos) was analyzed. We observed a lack of induction of pax8 expression in lhx1-depleted explants under AcRA treatment conditions in which expression of this gene is normally induced. Based on this observation, microarray analysis was carried out to identify genes whose expression is affected by the absence of lhx1.
Explants of injected embryos with 800pg of lhx1 DEED-AS were dissected, treated with pronephric tissue inductive conditions (AcRA) and harvested after 24 hours incubation at 14C (Fig. S5B). The sibling control embryos reached stage 12.5. Explants from uninjected embryos +AcRA and -AcRA as well as explants from DEED injected embryos -AcRA were also harvested. Approximately 12 caps were pooled for each RNA preparation and the analysis was performed using triplicates.

first: Cecilia

last: Cirio

first: Caroline

last: Haldin

first: Carsten

last: Stuckenholz

first: Xiongfong

last: Chen

first: Sung-Kook

last: Hong

first: Igor

middle: B

last: Dawid

first: Neil

last: Hukriede


title: Upregulation of FOXM1 Induces Genomic Instability in Human Epidermal Keratinocytes

status: Public on Jun 28 2010

city: London

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

summary:
Genome-wide 10k SNP profiling of FOXM1B-transduced N/TERT and primary normal human epidermal keratinocytes (NHEK). The aim of this study was to study the cancer initiation role of UVB and FOXM1B upregulation in NHEK. Upregulation of FOXM1B alone (without UVB) was found to directly induce genomic instability in the form of copy number aberration (CNA) and low levels of loss of heterozygosity (LOH) in primary NHEK. The FOXM1B-induced CNA was found to be retained and accumulated in subsequent cell culture passages. UVB-exposure resulted in significant chromosomal LOH and CNA in N/TERT cells expressing FOXM1B but not in EGFP-expressing cells. This indicates that UVB corroborated with FOXM1B to recruit LOH and CNA which may predispose cell to malignant transformation. Collectively, these results indicate that aberrant upregulation of FOXM1B in skin keratinocytes following UVB exposure may be an early mechanism whereby cells acquire genomic changes required for oncogenesis.

Keywords: Genome-wide SNP profiling for loss of heterozygosity (LOH) and copy number aberration (CNA), FOXM1, UVB, Keratinocytes, Basal cell carcinoma, genomic instability, carcinogenesis, squamous cell carcinoma.

overall_design: 1) Three individual normal primary NHEK cultures were retrovirally transduced (with either EGFP or EGFP-FOXM1B) and left to grow for 4 days prior to SNP array analysis. 2) EGFP or EGFP-FOXM1B-tansduced NHEK cells were harvested at passage 1, 2 and 3 for SNP array analysis to investigate if genomic instability is maintained and accumulated in subsequent passages. 3) N/TERT cells transduced with either EGFP or EFOX were UVB-irradiated and left to grow for 50 days in culture prior to SNP array analysis.


title: Mouse Soleus Muscle Chronic Electrical Stimulation Study

status: Public on Jun 01 2010

city: Pittsburgh

first: John

middle: Michael

last: Krill-Burger

state: PA

address: Rm. WG21.3 Shadyside Hospital

institute: University of Pittsburgh Medical Center

summary: Stimulation of the mouse hindlimb via the sciatic nerve was used to induce contractions for 4 hours to investigate acute muscle gene activation in a model of muscle phenotype conversion. Initial force production (1.6 + 0.1 g/g body weight) declined 45% within 10 min and was maintained for the remainder of the experiment. Force returned to initial levels upon completion of the study. An immediate-early growth response was present in the EDL (FOS, JUN, ATF3, MAFK) with a similar but attenuated pattern in the soleus. Transcript profiles showed decreased fast fiber specific mRNA (myosin heavy chains 2A, 2B; troponins T3, I; alpha-tropomyosin, m-creatine kinase) and increased slow transcripts (myosin heavy chain slow/1beta, troponin C, tropomyosin 3gamma) in the EDL. Histological analysis of the EDL revealed glycogen depletion without inflammatory cell infiltration or myofiber damage in stimulated vs. control muscles. Several fiber type specific transcription factors (EYA1, TEAD1, NFATc1 and c4, PPARG, PPARGC1alpha and beta, BHLHB2) increased in the EDL along with transcription factors characteristic of embryogenesis (KLF4, SOX17, TCF15, PKNOX1, ELAV). No established in vivo satellite cell markers or the genes activated during our parallel studies of satellite cell proliferation in vitro (CYCLINS A2, B2, C, E1, MyoD) increased in the stimulated muscles. These data indicated that onset of fast to slow phenotype conversion occurred in the EDL within 4 hours of stimulation without satellite cell recruitment or muscle injury but was driven by phenotype specific transcription factors from resident fiber myonuclei including activation of nascent developmental transcriptional programs.

overall_design: Adult male Swiss Webster mice (30-35 g) were anesthetized, a bipolar electrode was implanted adjacent to the sciatic nerve and the hindlimb immobilized. The voltage-force relation was determined to establish supramaximal stimulation conditions and the length-tension relation was determined to set the resting length for maximum twitch tension. Contractions were induced by sciatic nerve stimulation (0.5 msec duration, 2-5 volts). The muscles were allowed to rest 15 minutes for full metabolic recovery at physiologic temperatures. Supramaximal stimulation was applied at a rate of 10 Hz for 4 hours. At the end of each experiment the soleus muscles were carefully dissected and flash frozen in liquid nitrogen for analysis of mRNA expression via microarray analysis. The contralateral, unstimulated Soleus provided a genetically matched, paired control for each specimen.

first: William

last: LaFramboise

first: Robert

middle: W

last: Wiseman


title: Transcriptomic analysis comparing the spontaneous acapsular strain AL1114 to the parent capsulated strain AL1115

status: Public on May 24 2010

city: St Lucia

first: Jason

last: Steen

address: Carmody Road

department: IMB

country: Australia

institute: University of Queensland

summary: P. multocida is the causative agent of a wide range of diseases of animals, including fowl cholera in birds. Fowl cholera isolates of P. multocida generally express a capsular polysaccharide composed of hyaluronic acid. There have been reports of spontaneous capsule loss in P. multocida fowl cholera-causing strains but the mechanism by which this occurs has not been determined. In this study, we identified three independent strains that had spontaneously lost the ability to produce capsular polysaccharide.  Quantitative RT-PCR showed that these strains had significantly reduced transcription of the capsule biosynthetic genes, but DNA sequence analysis identified no mutations within the cap biosynthetic locus. However, whole genome sequencing of paired capsulated and acapsular strains identified a single nucleotide polymorphism within fis that was present only in the acapsular strain. Sequencing of fis from two independently derived spontaneous acapsular strains showed that each contained a mutation within fis. Complementation of these strains with an intact copy of fis returned normal capsule expression to all strains. Therefore, expression of a functional Fis protein is absolutely required for normal capsule expression in P. multocida.DNA microarray studies comparing one of the acapsular pairs (AL114 to AL1115) identified approximately 30 genes as down-regulated in the mutant; including pfh_B2 which encodes the filamentous hemagglutinin, a known P. multocida virulence factor and the cross protective surface antigen plpE.

platform_organism: Pasteurella multocida subsp. multocida str. Pm70

overall_design: Biological triplicates of each strain were analysed in a single colour experimental design

middle: D

last: Boyce


title: Adult Male Swiss Webster Hind Limb Muscle Satellite Cells

summary: Stimulation of the mouse hindlimb via the sciatic nerve was used to induce contractions for 4 hours to investigate acute muscle gene activation in a model of muscle phenotype conversion. Initial force production (1.6 + 0.1 g/g body weight) declined 45% within 10 min and was maintained for the remainder of the experiment. Force returned to initial levels upon completion of the study.  An immediate-early growth response was present in the EDL (FOS, JUN, ATF3, MAFK) with a similar but attenuated pattern in the soleus.  Transcript profiles showed decreased fast fiber specific mRNA (myosin heavy chains 2A, 2B; troponins T3, I; alpha-tropomyosin, m-creatine kinase) and increased slow transcripts (myosin heavy chain slow/1beta, troponin C, tropomyosin 3gamma) in the EDL.  Histological analysis of the EDL revealed glycogen depletion without inflammatory cell infiltration or myofiber damage in stimulated vs. control muscles. Several fiber type specific transcription factors (EYA1, TEAD1, NFATc1 and c4, PPARG, PPARGC1alpha and beta, BHLHB2) increased in the EDL along with transcription factors characteristic of embryogenesis (KLF4, SOX17, TCF15, PKNOX1, ELAV). No established in vivo satellite cell markers or the genes activated during our parallel studies of satellite cell proliferation in vitro (CYCLINS A2, B2, C, E1, MyoD) increased in the stimulated muscles. These data indicated that onset of fast to slow phenotype conversion occurred in the EDL within 4 hours of stimulation without satellite cell recruitment or muscle injury but was driven by phenotype specific transcription factors from resident fiber myonuclei including activation of nascent developmental transcriptional programs.

overall_design: Hind limb muscle satellite cells were pooled in each of three adult male Swiss Webster mice. Culture conditions were used to seperate a proliferating and differentiated group of satellite cells from each of the three mice.


title: Regional Immune Response to Trichostrongylis colubriformis - Gene Expression during Challenge of Immunized Sheep

first: Alan

last: McCulloch

summary: Afferent Lymph was collected from up to 5 pen raised, nematode naïve, outbreed Romney sheep (Tag No’s 1001, 1002, 1003, 1004, 1006) over a 12 week period. During this time a Pre-Infection sample was collected, the sheep were immunised by 3 truncated (2 week) T. colubiformis infections (40,000 Tc) followed by a Challenge Infection with 40,000 T. colubiformis L3 (Challenge Infection weeks 1-3). The Challenge Infection will enable us to determine changes in gene expression when the sheep gut rejects the parasitic nematodes.

first: Jaqui

last: Knight


title: Mouse EDL Muscle Chronic Electrical Stimulation Study

summary: Adult male Swiss Webster mice (30-35 g) were anesthetized, a bipolar electrode was implanted adjacent to the sciatic nerve and the hindlimb immobilized. The voltage-force relation was determined to establish supramaximal stimulation conditions and the length-tension relation was determined to set the resting length for maximum twitch tension. Contractions were induced by sciatic nerve stimulation (0.5 msec duration, 2-5 volts). The muscles were allowed to rest 15 minutes for full metabolic recovery at physiologic temperatures. Supramaximal stimulation was applied at a rate of 10 Hz for 4 hours. At the end of each experiment the extensor digitorum longus (EDL) muscles were carefully dissected and flash frozen in liquid nitrogen for analysis of mRNA expression via microarray analysis. The contralateral, unstimulated EDL provided a genetically matched, paired control for each specimen.

overall_design: Stimulation of the mouse hindlimb via the sciatic nerve was used to induce contractions for 4 hours to investigate acute muscle gene activation in a model of muscle phenotype conversion. Initial force production (1.6 + 0.1 g/g body weight) declined 45% within 10 min and was maintained for the remainder of the experiment. Force returned to initial levels upon completion of the study.  An immediate-early growth response was present in the EDL (FOS, JUN, ATF3, MAFK) with a similar but attenuated pattern in the soleus.  Transcript profiles showed decreased fast fiber specific mRNA (myosin heavy chains 2A, 2B; troponins T3, alpha-tropomyosin, m-creatine kinase) and increased slow transcripts (myosin heavy chain slow/1beta, troponin C, tropomyosin 3gamma) in the EDL.  Histological analysis of the EDL revealed glycogen depletion without inflammatory cell infiltration or myofiber damage in stimulated vs. control muscles. Several fiber type specific transcription factors (EYA1, TEAD1, NFATc1 and c4, PPARG, PPARGC1alpha and beta, BHLHB2) increased in the EDL along with transcription factors characteristic of embryogenesis (KLF4, SOX17, TCF15, PKNOX1, ELAV). No established in vivo satellite cell markers or the genes activated during our parallel studies of satellite cell proliferation in vitro (CYCLINS A2, B2, C, E1, MyoD) increased in the stimulated muscles. These data indicated that onset of fast to slow phenotype conversion occurred in the EDL within 4 hours of stimulation without satellite cell recruitment or muscle injury but was driven by phenotype specific transcription factors from resident fiber myonuclei including activation of nascent developmental transcriptional programs.

first: Kelly

middle: L

last: Bombach

first: David

last: Ankrapp

first: Roop

last: Jayaraman

first: Patti

last: Petrosko


title: Large-Scale Expression Analysis Reveals Distinct microRNA Profiles at Different Stages of Human Neurodevelopment

status: Public on May 27 2010

city: Ottawa

first: Brandon

last: Smith

state: Ontario

address: 1200 Montreal Road

department: Institute for Biological Sciences

institute: National Research Council Canada

summary:
Background: MicroRNAs (miRNAs) are short non-coding RNAs predicted to regulate one third of protein-coding genes via mRNA targeting. In conjunction with key transcription factors, such as the repressor REST (RE1 silencing transcription factor), miRNAs play crucial roles in neurogenesis, which requires a highly orchestrated program of gene expression to ensure the appropriate development and function of diverse neural cell types. Whilst previous studies have highlighted select groups of miRNAs during neural development, there remains a need for amenable models in which miRNA expression and function can be analyzed over the duration of neurogenesis.

Principal Findings: We performed large-scale expression profiling of miRNAs in human NTera2/D1 (NT2) cells during retinoic acid (RA)-induced transition from progenitors to fully differentiated neural phenotypes. Our results revealed dynamic changes of miRNA patterns, resulting in distinct miRNA subsets that could be linked to specific neurodevelopmental stages. Moreover, the cell-type specific miRNA subsets were very similar in NT2-derived differentiated cells and human primary neurons and astrocytes.  Further analysis identified miRNAs as putative regulators of REST, as well as candidate miRNAs targeted by REST. Finally, we confirmed the existence of two predicted miRNAs; pred-MIR191 and pred-MIR222 associated with SLAIN1 and FOXP2, respectively, and provided some evidence of their potential co-regulation.

Conclusions: In the present study, we demonstrate that regulation of miRNAs occurs in precise patterns indicative of their roles in cell fate commitment, progenitor expansion and differentiation into neurons and glia. Furthermore, the similarity between our NT2 system and primary human cells suggests their roles in molecular pathways critical for human in vivo neurogenesis.

platform_organism:
Homo sapiens
Mus musculus

overall_design:
The experiment consists of a total of 51 arrays: 29 retinoic acid time series arrays (0,2,4,6,8,12,14,21 and 28 days), 2 each of NT2-derived neurons and astrocytes, 12 primary human fetal astrocytes, 3 primary human embryonic astrocytes and 3 primary human neurons.  Each condition has a minimum of 2 biological replicates.  The samples were compared as single channel experiments.

NOTE: The raw data files were submitted as generated in Quantarray with 2 channels, but due to issues with the control sample dye (Cy5 on Channel 1), only the Channel 2 (Cy3) data was analysed.

first: Julie

last: Treadwell

first: Dongling

last: Zhang

first: Iain

last: McKinnell

first: Dao

last: Ly

first: P

middle: R

last: Walker

first: Marianna

last: Sikorska


title: Lactate modulates gene expression in human mesenchymal stem cells

status: Public on May 16 2010

city: Tübingen

first: Kay

last: Nieselt

state: DE

address: Sand 14

department: Department for Computer Science

institute: Institute for Bioinformatics and Medical Informatics

summary:
Wound healing is associated with high rates of cell replication and lactate accumulation even under normoxic and hyperoxic conditions. Lactate accounts for various effects in tissue regeneration, such as collagen synthesis, angiogenesis, modulation of cytokine patterns and as recently shown for stem cell homing. Its influence on genes involved in cell replication has not been shown yet. Therefore, the effect of lactate considering genes involved in different cellular processes was investigated. Human umbilical vein endothelial cells (HUVEC) were cultured and incubated with lactate for different periods of time. Gene expression analysis was performed using custom-designed oligonucleotide microarrays.
Keywords: Human gene expression study

overall_design: Reference design with Cy3 labeled uniRNA and Cy5 labeled sample RNA.

first: Derek

last: Zieker

first: Katharina

last: Buck


title: Functional Genomics Analysis of the Yeast Iron Responsive Transcription Factor Aft1 Reveals Iron-Independent Functions

status: Public on May 01 2010

first: Linda

last: Harris

address: 960 Carling Ave.

department: Eastern Cereal & Oilseed Research Centre

institute: Agriculture & Agri-Food Canada (AAFC)

summary: The Saccharomyces cerevisiae transcription factor Aft1 is activated in iron-deficient cells to  induce the expression of iron regulon genes, which coordinate the increase of iron uptake and  remodel cellular metabolism to survive low iron conditions. In addition, Aft1 has been  implicated in numerous cellular processes including cell cycle progression and chromosome  stability; however it is unclear if all cellular effects of Aft1 are mediated through iron  homeostasis. To further investigate the cellular processes impacted by Aft1 using genome-wide  synthetic lethal and synthetic dosage lethal screens, we identified greater than 70 deletion mutants  that are sensitive to perturbations in AFT1 levels. Our genetic network reveals that Aft1 impacts  a diverse range of cellular processes, including the RIM101 pH pathway, cell wall stability, DNA  damage, protein transport, chromosome stability and mitochondrial function. Surprisingly, only  a subset of mutants identified are sensitive to iron fluctuations or display genetic interactions  with mutants of iron regulon genes AFT2 or FET3. We demonstrate that Aft1 works in parallel  with the RIM101 pH pathway and the role of Aft1 in cell wall structure and DNA damage repair  is mediated by iron. In contrast, we show that the role of Aft1 in chromosome maintenance is  independent of its iron regulatory role and that alterations in iron levels do not impact chromosome  loss rates. Our study clearly demonstrates a novel iron-independent role for Aft1.

overall_design: For this experiment 4 samples (2 reference and 2 test samples), consisting of 3 biological reps were hybridized. Cy3 one-color labelling was used for each sample.

first: Sharon

last: Berthelet

first: Jane

last: Usher

first: Kristian

last: Shulist

first: Nancy

last: Maltez

first: Ying

last: Fong

first: Kristin

last: Baetz

first: Akil

last: Hamza

first: Anne

last: Johnston


title: Elevated Myocardial Na+/H+ Exchanger Elicits the Expression of Genes that Lead to Cardiac Hypertrophy

status: Public on May 17 2010

city: La Jolla

first: Dan

last: Zhou

state: CA

address: CMG 103, 9500 Gilman Drive, MC0735

department: Pediatrics

institute: University of California, San Diego, School of Medicine

summary: In myocardial disease, elevated expression and activity of Na+/H+ exchanger isoform 1 (NHE1) is detrimental.  To better understand the involvement of NHE1, transgenic mice with elevated heart-specific NHE1 expression were studied.   N-line mice expressed wild-type NHE1 and K-line mice expressed activated NHE1.  Cardiac morphology, interstitial fibrosis and cardiac function were examined by histological staining and echocardiography.  Differences in gene expression between the N- or K-line and non-transgenic littermates were probed using genechips.  We found that NHE1 K-line hearts (but not N-line) developed hypertrophy, including elevated heart weight to body weight and increased cross sectional area of the cardiomyocytes, interstitial fibrosis, as well as depressed cardiac function.  N-line transgenic hearts had modest changes in gene expression (50 up-regulations and 99 down-regulations, P<0.05), whereas K-line hearts had a very strong transcriptional response (640 up-regulations and 677 down-regulations, P<0.05).  In addition, the magnitude of expression alterations was much higher in K-line than in N-line transgenic mice.  The most significant changes in gene expression were involved in cardiac hypertrophy, cardiac necrosis/cell death and cardiac infarction.  Secreted phosphoprotein 1 signaling pathways were up-regulated while peroxisome proliferative activated receptor gamma signaling was down-regulated in K-line mice.  Our study shows that expression of activated NHE1 elicits specific pathways of gene activation in the myocardium that lead to cardiac hypertrophy, cell death and infarction.

overall_design: Alterations on gene expression profiles were determined in the hearts of transgenic mice carrying either a wild-type NHE1 gene (N-line) or a constitutively active NHE1 mutant gene (K-line).

first: Jin

last: Xue

first: Gabriel

middle: G

last: Haddad


title: Assessment of basal cell carcinoma and rhabdomyosarcoma tumorigenesis in Blm-deficient Ptch1+/- mice

status: Public on Jan 14 2010

city: San Francisco

first: Bing

last: Huey

address: 1450 Third Street

department: Helen Diller Comprehensive Cancer Center

institute: University of California at San Francisco

summary: Basal cell carcinomas (BCCs) have relative genomic stability and relatively benign clinical behavior but whether these two are related causally is unknown.  To investigate the effects of introducing genomic instability into murine BCCs, we have compared ionizing radiation-induced tumorigenesis in Ptch1+/- mice vs. that in Ptch1+/- mice carrying mutant Blm alleles. We found that BCCs in Ptch1+/- Blmtm3Brd/tm3Brd mice had a trend towards greater genomic instability as measured by array CGH and that these mice developed significantly more microscopic BCCs than did Ptch1+/- Blm+/tm3Brd or Ptch1+/- Blm+/+ mice. The mutant Blm alleles also markedly enhanced the formation of rhabdomyosarcomas (RMS), another cancer to which Ptch1+/- mice and PTCH1+/- (basal cell nevus syndrome) patients are susceptible. Highly recurrent but different copy number changes were associated with the two tumor types and included losses of chromosomes 4 and 10 in all BCCs and gain of chromosome 10 in 80% of RMSs. Loss of chromosome 11 and 13, including the Trp53 and Ptch1 loci respectively, occurred frequently in BCCs, suggesting tissue-specific selection for genes or pathways that collaborate with Ptch deficiency in tumorigenesis. Despite the quantitative differences, there was no dramatic qualitative difference in the BCC or RMS tumors associated with the mutant Blm genotype.

overall_design: We investigated the effect of Blm deficiency on ionizing radiation-induced basal cell carcinoma and rhabdomyosarcoma tumorigenesis in Ptch1+/- mice. Six BCC and five RMS samples were obtained from separate mice. Liver tissue from each mouse was used as the normal reference.

first: Parastoo

last: Davari

first: Jeniffer

last: Hebert

first: Donna

last: Albertson

first: Ritu

last: Roy

first: Maria

last: Mancianti

first: Andrew

middle: E

last: Horvai

first: Lisa

last: Mcdaniel

first: Roger

last: Schultz

first: Ervin

middle: H

last: Epstein Jr.