title: New rapid scheme for distinguishing subspecies of the Mycobacterium abscessus group and identification of Mycobacterium massiliense with inducible clarithromycin resistance

city: Bethesda

first: Timothy

middle: G

last: Myers

state: MD

address: 50 South Drive, Room 5509

department: Research Technologies Branch

institute: National Institute of Allergy and Infectious Diseases

summary: Mycobacterium abscessus [M. abscessus (sensu lato) or M. abscessus group] comprises three closely related taxa with taxonomic status under revision: M. abscessus sensu stricto, M. bolletii and M. massiliense. We describe here a simple, robust and cost effective PCR-based method for distinguishing among M. abscessus, M. massiliense and bolletii. Based on the M. abscessus ATCC 19977T genome, discriminatory regions were identified between M. abscessus and M. massiliense from array-based comparative genomic hybridization. A typing scheme using PCR primers designed for four of these locations was applied to 46 well-characterized clinical isolates comprising 29 M. abscessus, 15 M. massiliense and 2 M. bolletii previously identified by multi-target sequencing. Interestingly, 2 isolates unequivocally identified as M. massiliense were shown to have a full length erm(41) instead of the expected gene deletion and showed inducible clarithromycin resistance after 14 days. We propose using this PCR-based typing scheme combined with erm(41) PCR for a straightforward identification of M. abscessus, M. massiliense and M. bolletii and assessment of inducible clarithromycin resistance. This method can be easily implemented into a routine workflow providing subspecies level identification within 24 hours of isolation of M. abscessus group.

sample_organism: Mycobacteroides abscessus subsp. bolletii

overall_design: Two-color CGH with 4 independent Mycobacterium clinical isolates and the M massiliense type strain (CCUG 48898) labeled with Cy3 were cohybridized with the M abscessus type strain (ATCC 19977) labeled with Cy5 on a tiling array designed against the  M abscessus type strain

first: Shamira

last: Shallom

first: Paul

last: Gardina

first: Yinong

last: Sebastian

first: Patricia

last: Conville

first: Hervé

last: Tettelin

first: Kenneth

middle: N

last: Olivier

first: Gulbu

last: Uzel

first: Elizabeth

last: Sampaio

first: Steven

last: Holland

first: Adrian

last: Zelazny


title: MicroRNA profiling of brain metastasis competent cell-derived exosomes

status: Public on Jul 16 2013

city: Houston

first: Laura

last: Camacho

address: One Baylor Plaza

country: USA

institute: Baylor College of Medicine

summary: Objective of this work was to characterize the miRNA profile of exosomes isolated from brain-homing cell lines (MDA-MB-231BR, CTC1BMSM, and 70W) with their respective parental non-brain metastatic cell lines (MDA-MB-231P, CTC1P and MeWo).

overall_design: Exosomes derived from the six cell lines were isolated. Brain metastatic cell-derived exosomal miRNAs were compared with non brain metastatic cell lines (MDA-MB-231BR versus MDA-MB-231P, CTC1BMSM versus CTC1P, and 70W versus MeWo).

first: Dario

last: Marchetti


title: Transcriptomics of lateral root initiation in Maize

city: Gent

first: boris

last: parizot

address: Technologiepark 927

country: Belgium

institute: VIB Department of Plant Systems Biology

summary: We developed a method to synchronize the induction of lateral roots in primary and adventitious roots of Zea mays, and used it to perform a genome-wide transcriptome analysis of the pericycle cells in front of the phloem poles during lateral root initiation.

overall_design: Lateral roots were induced in primary and adventitious roots of Maize. For the primary root, plants were germinated and grown 64 hours in NPA 50 µM, and then transfered to NAA 50 µM. For the adventitious roots, plants were germinated and grown in water for 6 days, then tranfered 4 days in NPA 25 µM, and finally transfered to NAA 25 µM. For all these roots, pericycle cells located in front of the phloem poles in segments of roots located between 5 and 10 mm distance from the root tip were isolated using laser capture microdissection after cryosection. Material was sampled at 0 hours (NPA) and after 2, 3 and 4 hours of NAA treatment, for both the primary and adventitious roots and also after 6 hours and 9 hours of NAA treatment for the adventitious roots.

first: Leentje

last: Jansen

first: Boris

last: Parizot


title: Arabidopsis thaliana seed germination timecourse

city: Sutton Bonington

first: Simon

middle: Peter

last: Pearce

state: Leicestershire

address: Sutton Bonington Campus

department: Plant and Crop Sciences

country: United Kingdom

institute: University of Nottingham

summary: This series analyses germinating Arabidopsis seeds with both temporal and spatial detail, revealing two transcriptional phases that are separated with respect to testa rupture. Performed as part of the ERA-NET Plant Genomics grant vSEED.

overall_design: Arabidopsis seeds were dissected into four tissues at nine time-points during seed germination. The tissues were the combined micropylar and chalazal endosperm (MCE), the remaining endosperm (PE), the radicle and embryonic axis (RAD) and the cotyledons (COT). At testa and endosperm rupture the seeds were sampled in separate pre- and post-ruptured populations.

first: Bas

middle: J

last: Dekkers

middle: P

first: Marieke

last: van Bolderen-Veldkamp

first: Alex

last: Marshall

first: George

middle: W

last: Bassel

first: John

middle: R

last: King

first: Andrew

middle: T

last: Wood

first: Kerstin

last: Müller

first: Paweł

last: Widera

first: James

last: Gilbert

first: Natalio

last: Krasnogor

first: Gerhard

last: Leubner-Metzger

first: Michael

last: Holdsworth

first: Leonie

last: Bentsink


title: Differential gene expression between Salmonella Typhimurium biofilms on glass vs cholesterol surfaces

status: Public on Jul 09 2013

city: Boston

first: Geoffrey

last: Gonzalez

address: 55 Fruit St

department: Infectious Diseases

institute: Massachusetts General Hospital

summary:
Salmonella spp. biofilms have been implicated in persistence in the environment and plant surfaces. In addition, Salmonella is able to form biofilms on the surface on cholesterol gallstones.
The ability of Salmonella spp. on these surfaces is superior to biofilm formation on surfaces on glass or plastic. Thus, we hypothesized that Salmonella gene expression is specific during biofilm development on cholesterol surfaces.

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

overall_design:
Flow through assays were performed whereby S. Typhimurium was inoculated into chambers coated with glass or cholesterol. At 24h post-inoculation, planktonic (from the flow through), biofilms (from glass or cholesterol) were collected.
Thus we had 4 samples: Planktonic (2) and Biofilms (2), each with 2 biological replicates

last: Gonzalez-Escobedo

middle: S

last: Gunn


title: Liver transcriptome profile in pigs with extreme phenotypes of intramuscular fatty acid composition

city: Bellaterra

first: Yuliaxis

last: Ramayo-Caldas

state: Barcelona

address: Centre for Research in Agricultural Genomics

department: Animal Genetics

country: Spain

summary: The liver transcriptomes of two female groups (High and Low) with phenotypically extreme intramuscular fatty acid composition were sequenced using RNA-Seq [accn: SRA053452, subid: 86092,  Bioproject: PRJNA168072]. A total of 146 and 180 unannotated protein-coding genes were identified in intergenic regions for the L and H groups, respectively. In addition, a range of 5.8 to 7.3% of repetitive elements was found, with  SINEs being the  most abundant elements. The expression in liver of 186 (L) and 270 (H) lncRNAs was also detected. The higher reproducibility of the RNA-Seq data was validated by RT-qPCR and porcine expression microarrays, therefore showing a strong correlation between RT-qPCR and RNA-Seq data (ranking from 0.79 to 0.96), as well as between microarrays and RNA-Seq (r=0.72). A differential expression analysis between H and L animals identified 55 genes differentially-expressed between groups. Pathways analysis revealed that these genes belong to biological functions, canonical pathways and three gene networks related to lipid and fatty acid metabolism. In concordance with the phenotypic classification, the pathways analysis inferred that linolenic and arachidonic acids metabolism was altered between extreme individuals. In addition, a connection was observed among the top three networks, hence suggesting that these genes are interconnected and play an important role in lipid and fatty acid metabolism.

overall_design: According to the score information for the first principal component the animals were ranked in two groups and ten females (5 per group) were assayed with high-density oligonucleotide microarray chips (GeneChip® Porcine) from Affymetrix.

first: Nuria

last: Mach

first: Anna

last: Esteve-Codina

first: Jordi

last: Corominas

last: Castelló

first: Maria

last: Ballester

last: Estellé

first: Noelia

last: Ibáñez-Escriche

first: Ana

middle: I

last: Fernández

first: Miguel

last: Pérez-Enciso

first: Josep

middle: M

last: Folch


title: Murine Bone Marrow Macrophages: Control vs. RANKL Stimulated

status: Public on Jul 10 2013

city: Tokyo

first: Hitoshi

last: Kohsaka

address: 1-5-45 Yushima Bunkyo-ku

department: Department of Medicine and Rheumatology

country: Japan

institute: Tokyo Medical and Dental University

summary: To identify the microRNAs that are involved in osteoclastogenesis, microRNA expression profiles in mouse bone marrow macrophages (BMMs) stimulated with RANKL (BMOc) were compared with that of control untreated BMMs. These results provide insights into the mechanisms to regulate osteoclastogenesis and  bone resorption activities in osteoclasts by microRNA.

platform_organism:
Homo sapiens
Mus musculus
Rattus norvegicus

overall_design: BMMs were cultured with 20 ng/ml M-CSF in the presence or absence of 50 ng/ml RANKL for 24 hours. Cells were collected for total RNA isolation, and were subjected to microRNA array analysis.

first: Fumitaka

last: Mizoguchi

first: Yousuke

last: Murakami


title: Toxicity and transcriptomic analysis in Hyalella azteca suggests increased exposure and susceptibility of epibenthic organisms to Zinc Oxide Nanoparticles

status: Public on Jul 24 2013

first: Helen

middle: C

last: Poynton

state: MA

address: 100 Morrissey Blvd.

department: School for the Environment

institute: UMass Boston

summary: Abstract:  Nanoparticles (NPs) are expected to make their way into the aquatic environment where sedimentation of particles will likely occur, putting benthic organisms at particular risk.  Therefore, organisms such as Hyalella azteca, an epibenthic crustacean which forages at the sediment surface, is likely to have a high potential exposure.  Here we show that Zinc Oxide  (ZnO) NPs are more toxic to H. azteca compared with the corresponding metal ion, Zn2+.  Dissolution of ZnO NPs contributes about 50% of the Zn measured in the ZnO NP suspensions, and cannot account for the toxicity of these particles to H. azteca.  However, gene expression analysis is unable to distinguish between the ZnO NP exposures and Zinc Sulfate (ZnSO4) exposures at equitoxic concentrations.  These results lead us to hypothesize that ZnO NPs provide and an enhanced exposure route for Zn2+ uptake into H. azteca, and possibly other sediment dwelling organisms.  Our study supports the prediction that sediment dwelling organisms are highly susceptible to the effects of ZnO NPs and should be considered in the risk assessment of these nanomaterials.

overall_design: This experiment included four different treatments and an untreated control.  Each treatment or control, consisted of ten independent replicates of twenty Hyalella azteca.  Of these, six were randomly chosen to be used for the microarray analysis.

first: James

last: Lazorchak

first: Bonnie

last: Blalock


title: Identification of WRKY22 direct targets under submergence in Arabidopsis with ChIP

status: Public on Jul 03 2013

city: Taipei

first: Fu-Chiun

last: Hsu

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

department: Agricultural Biotechnology Research Center

country: Taiwan

institute: Academia Sinica

type: Genome binding/occupancy profiling by genome tiling array

summary: To identify direct targets of WRKY22, we created a transgenic Arabidopsis line that expresses a c-myc epitope-tagged WRKY22 and used ChIP followed by microarray hybridization (ChIP-chip) to screen for candidates and validate the in vivo protein-DNA interactions with ChIP followed by quantitative PCR (ChIP-Q-PCR).  The WRKY22 and c-myc epitope tag fusion construct was generated and transformed into wrky22-ko2 plants.  The resulting transgenic lines should have better ChIP efficiency than the wild-type background, due to the reduced competition for WRKY22 binding sites from endogenous WRKY22.  ChIP-enriched DNA fragments were identified using criteria of a window of +300 to －1200 of a gene for a promoter, a width of 4 probes or more, and a false discovery rate (FDR) < 0.1.  The ChIP-chip experiments were repeated six times, i.e., six biological replicas.  Candidates were defined by the presence of the promoter in three out of six biological replicas.  Candidates were then classified based on their hypoxic responsiveness with a positive response defined as gene expression levels exhibiting > 2 or < 0.5-fold induction in any time point under submergence treatments in expression array data.

overall_design: Comparison of c-myc tagged WRKY22 transgenic plants vs wild-type (Columbia) plants.  Both materials were submergence treated for 3 hours.

first: Hsu

last: Fu-Chiun

first: Chou

last: Mei-Yi

first: Peng

last: Hsiao-Ping

last: Shu-Jen

first: Li

last: Ya-Ru

first: Shih

last: Ming-Che


title: Cold-induced changes in gene expression in brown adipose tissue, white adipose tissue and liver

status: Public on Jul 08 2013

city: Loughborough

first: Neil

last: Graham

summary: Cold exposure imposes a metabolic challenge to mammals that must be met by a coordinated repsonse in different tissues to prevent hypothermia. This study reports analysis of transcriptome profiles in brown adipose tissue, liver, white adipose of mice in repsonse to 24 hour cold exposure

overall_design: Total RNA was extracted from brown adipose, whie adipose and liver from cold treated and control plants. 3 replicates of brown adipose and white adipose and 2 replicates of liver samples.

first: Andrew

last: Shore

first: Karamitri

last: Angeliki

first: Paul

last: Kemp

first: John

last: Speakman

first: Michael

middle: A

last: Lomax


title: Identification of microRNAs dysregulated in cellular senescence driven by endogenous genotoxic stress

status: Public on Jun 29 2013

city: Pittsburgh

first: Saleem

last: Khan

state: PA

address: 450 Technology Drive

department: Microbiology and Molecular Genetics

institute: University of Pittsburgh

summary: We compared microRNA (miRNA) expression in Ercc1-/- primary mouse embryonic fibroblasts (MEFs) and wild-type (WT) MEFs in 20% O2 and 3% O2 growth conditions to identify miRNAs that drive cellular senescence.

overall_design: MicroRNA expression in WT and Erccc1-/- MEFs was measured at passage 3 and passage 7 after growth in 20% and 3% O2. Two independent experiments were performed for each growth condition.

first: Lolita

last: Nidadavolu


title: Molecular Insight into the Association between Cartilage Regeneration and Ear Wound Healing in Genetic Mouse Models: Targeting New Genes in Regeneration

status: Public on Jul 04 2013

city: St. Louis

first: Muhammad Farooq

last: Rai

state: MO

address: 660 S. Euclid Ave.

department: Orthopaedic Surgery

institute: Washington University School of Medicine

summary:
Certain mouse strains have the ability to heal both ear wounds and knee articular cartilage injuries.  A positive genetic correlation between cartilage healing and protection from osteoarthritis exists, which suggests that a common core of genes may operate in healing process.
Here, we analyzed knee joint tissues from healer and non-healer mice and found that four genes were common to both ear wound and knee cartilage healing.  Specific sequence differences between haler and non-healer parental strains were identified as potential causal polymorphisms. We suggest a common molecular and genetic basis of tissue healing.

overall_design:
We collected tissue lysates from the formalin-fixed paraffin-embedded sections from mouse knee joints and analyzed the expression of several genes by Affymetrix QunatiGene Plex assay.
Please note that 8-9 replicates were performed per each sample. The averaged (across the replicate samples) data was provided for each sample record and non-averaged data (from each replicate) is provided as a Series supplementary file (*setI.txt contains data from Panel 321347 and *setII.txt from panel 321389).

first: Muhammad

middle: F

first: Eric

last: Schmidt

first: Audrey

last: McAlinden

last: Cheverud

first: Linda

last: Sandell


title: Identification of GATA1 transcriptional regulation in the megakaryoblastic cell line Meg01

status: Public on Jul 01 2013

city: Detroit

first: Yubin

last: Ge

state: MI

address: 110 East Warren Ave.

institute: Karmanos Cancer Institute

summary: chIP-on-chip analysis of GATA1 in the megakaryoblastic cell line Meg01.

overall_design: The IP was obtained with a GATA1 C-terminus antibody.  The control sample was a normal goat IgG pull down done side by side with the GATA1 (C-20) antibody using the same soluble chromatin sample from non-treated Meg-01 cells. A 2-chip microarray set (Agilent 240K) was used for the chIP-on-chip analysis.

first: Alan

last: Dombkowski

first: Jeffrey

last: Taub


title: The expression profile of SLC transporter genes in resectable pancreatic cancer

status: Public on Jun 30 2013

city: Prague 10

first: Pavel

last: Soucek

address: Srobarova 48

country: Czech Republic

institute: National Institute of Public Health

summary: Objectives: The aim of this study was to investigate the prognostic significance of thirteen anticancer drug-relevant solute carrier transporters (SLCs) in pancreatic cancer in the context of clinical-pathological characteristics and the KRAS mutation status of tumors. Methods: Tumors and non-neoplastic pancreatic tissues were obtained from 32 histologically verified patients with pancreatic ductal adenocarcinoma. The transcript profile of SLCs was assessed using quantitative real-time PCR. KRAS mutations in exon 2 were assessed by high resolution melting analysis and confirmed by sequencing. Results: SLC22A3 was upregulated and SLC22A1, SLC22A2, SLC22A11, SLC28A1, SLC28A3 and SLC29A1 were downregulated when compared with non-neoplastic pancreatic tissues. Moreover, significantly lower levels of SLC22A1, SLC22A11 and SLC29A1 were found in tumors with angioinvasion. There was also a significantly higher transcript level of SLC28A1 in tumors with regional lymph nodes affected by metastasis. The study found that a high expression of SLC22A1 or SLC28A1was significantly associated with poor overall survival in unselected patients. In contrast, a high expression of SLC22A3 or SLC29A3 was significantly associated with longer overall survival in patients treated with nucleoside analogs. Finally, SLC levels were not found to be associated with KRAS mutation status in exon 2. Conclusions: This study identified a number of associations of SLCs with prognosis of pancreatic cancer patients.

overall_design: Transcript levels of thirteen anticancer drug-relevant solute carrier transporters (SLCs) were determined by qPCR in pancreatic ductal adenocarcinomas and non-neoplastic tissue samples from 32 pancreatic cancer patients. MRPL19, ELF1 and POLR2A  were used as reference genes for data normalization.

first: Beatrice

last: Mohelnikova-Duchonova

first: Veronika

last: Brynychova


title: Sensitivity of cholangiocarcinoma cells to gemcitabine is affected by microRNA expression

status: Public on Jun 02 2013

city: Yonago-city

first: Kinya

last: Okamoto

state: Tottori

address: 36-1, Nishi-cho

department: Second Dept. Internal Medicine

institute: Tottori University

summary: We examined the relationship between miRNA expression and the sensitivity of CCA cells to Gem. Two intrahepatic CCA cell lines, HuH28 and HuCCT1 were used. HuCCT1 cells were more sensitive to Gem than were HuH28 cells. The miRNA expression profiles of HuH28 and HuCCT1 were determined by microarray analysis. Eighteen miRNAs were differentially expressed whose ratios over ± 2log2 between HuH28 and HuCCT1. Among these 18 miRNAs, ectopic overexpression of each of three downregulated miRNAs in HuH28 (miR-29b, miR-205, miR-221) restored Gem sensitivity to HuH28. Suppression of one upregulated miRNA in HuH28, miR-125a-5p, inhibited HuH28 cell proliferation independently to Gem treatment.

platform_organism:
Homo sapiens
Mus musculus
Rattus norvegicus
Human alphaherpesvirus 1
Human alphaherpesvirus 2
Human betaherpesvirus 5
human gammaherpesvirus 4
JC polyomavirus
Human immunodeficiency virus 1
Murid gammaherpesvirus 4
Human gammaherpesvirus 8
Merkel cell polyomavirus
Mus musculus cytomegalovirus 2
Betapolyomavirus hominis
Betapolyomavirus macacae

overall_design: Cell lines and cultures  Two human intrahepatic CCA cell lines, HuCCT1 and HuH28, were purchased from Japan Health Science Research Resources Bank (Osaka, Japan). Each cell line was cultured in RPMI-1640 medium (Invitrogen, Life Technologies Corp., CA, USA) that contained 10% fetal bovine serum (Nichirei Bioscience, Tokyo, Japan) and in humidified conditions at 37 ˚C and 5% CO2. Antibiotics were not added to the culture medium when cells were prepared for transfection with miRNA mimics or oligonucleotides.   Gem treatment  Gem hydrochloride was purchased from Wako (Osaka, Japan). A stock solution was prepared at 1 mmol/L (1 x 10-3 M) and was further diluted to anyone of several different final working concentrations from 1 x 10-4 to 1 x 10-7 M with cell culture medium that lacked antibiotics. Transfection of miRNA mimics, antisense oligonucleotides, or siRNA for miRNA target genes were performed 24 hr before the Gem treatment. All assays were conducted 72 hr after Gem treatment.

first: Kenichi

last: Miyoshi

first: Yoshikazu

last: Murawaki


title: Developmental exposure to valproic acid alters the expression of microRNAs involved in neurodevelopment in zebrafish

status: Public on Jun 19 2013

city: Woods Hole

first: Neelakanteswar

last: Aluru

address: 45 Water Street

department: Biology

institute: Woods Hole Oceanographic Institution

summary: Congenital malformations are a prevalent cause of infant mortality in the United States and their induction has been linked to a variety of factors, including exposure to teratogens. However, the molecular mechanisms of teratogenicity are not fully understood. MicroRNAs are an important group of small, non-coding RNAs that regulate mRNA expression. MicroRNA roles in early embryonic development are well established, and their disruption during development can cause abnormalities. We hypothesized that developmental exposure to teratogens such as valproic acid alters microRNA expression profiles in developing embryos. Valproic acid is an anticonvulsant and mood-stabilizing drug used to treat epilepsy, bipolar disorder and migraines. To examine the effects of valproic acid on microRNA expression during development, we used zebrafish embryos as a model vertebrate developmental system. Zebrafish embryos were continuously exposed to valproic acid (1 mM) or vehicle control (ethanol) starting from 4 hours post-fertilization (hpf) and sampled at 48 and 96 hpf to determine the miRNA expression profiles prior to and after the onset of developmental defects. At 96 hpf, 95% of the larvae showed skeletal deformities, abnormal swimming behavior, and pericardial effusion. Microarray expression profiling was done using Agilent zebrafish miRNA microarrays. Microarray results revealed changes in miRNA expression at both the time points. Thirteen miRNAs were differentially expressed at 48 hpf and 22 miRNAs were altered at 96 hpf. Among them, six miRNAs (miR-16a, 18c, 122, 132, 457b, and 724) were common to both time points. Bioinformatic target prediction and examination of published literature revealed that these miRNAs target several genes involved in the normal functioning of the central nervous system. These results suggest that the teratogenic effects of valproic acid could involve altered miRNA expression.

overall_design: Small RNA profiles were deteremined in valproic acid exposed zebrafish embryos using Agilent miRNA microarrays

first: Kristina

last: Deak

first: Matthew

last: Jenny

first: M

middle: E

last: Hahn


title: Mycobacterium tuberculosis ESX-3 conditional mutant expression

status: Public on Jun 12 2013

city: Padua

first: Agnese

last: Serafini

address: Via Gabelli 63

department: Molecular Medicine

country: Italy

institute: University of Padua

summary: Transcriptional profile of esx-3 conditional mutant strain (TB79) vs its parental strain (TB38) grown in 7H9 in presence of 100ng/ml Atc for 48h

overall_design: One experimental condition. 2 independent biological replicates. One replicate per array.

first: Riccardo

last: Manganelli


title: Expression data from human OC+BAP+ double positive versus OC-BAP- double negative cells

status: Public on May 17 2013

city: Padova

first: Nicol

last: Poncina

address: via Giustiniani, 2

department: Department of Medicine

institute: University of Padova

summary:
Subpopulation of circulating monocyte/macrophage lineage cells show a calcifying ability in vivo and in vitro. These cells may contribute to intimal calcification within atherosclerotic lesions and may have pathophysiological clinical and therapeutic implication in vascular disorders. Such cells express Osteocalcin (OC) and Bone Alkaline Phosphatase (BAP).
We analysed the expression profile of human OC+BAP+ cells versus OC-BAP- cells

overall_design: We compared three biological repilcates of OC+BAP+ versus OC-BAP- (each population coming from the same donor), each of the three coming from different healthy non diabetic donors.

first: Lisa

last: Menegazzo

first: Mattia

last: Albiero

first: Kenneth

last: Dyar

first: Angelo

last: Avogaro

first: Gian-Paolo

last: Fadini


title: Pseudonocardia dioxanivorans strain CB1190 during growth on THF or succinate

status: Public on Jul 12 2013

city: Philadelphia

first: Christopher

middle: Michael

last: Sales

address: 3141 Chestnut St, Curtis 251

department: Civil, Architectural & Environmental Engineering

institute: Drexel University

type:
Expression profiling by array
Third-party reanalysis

summary: Analysis of gene expression in Pseudonocardia dioxanivorans strain CB1190 during growth on THF (tetradhydrofuran) or succinate, and re-analysis of GSE33197 published expression data.

overall_design:
Two new treatments, based on carbon/energy source used for growth: THF and succinate. Three previously published treatments (pyruvate, dioxane, and glycolate) from GEO Series GSE33197 were also reanalyzed. Triplicate microarrays (biological replicates) were prepared for each treatment. Gene expression changes were compared directly between the new treatments (THF vs. succinate) and to previously published data during growth on pyruvate (GEO accession numbers GSM821862-GSM821864) in order to yield THF vs. pyruvate and succinate vs. pyruvate contrasts. Included as part of the re-analysis of GSE33197 were data from dioxane and glycolate treatments (GSM821865-GSM821870).

The complete dataset representing: (1) the THF- and succinate-treated CB1190 Samples and (2) the pyruvate-, dioxane- and glycolate-treated CB1190  Samples from Series GSE33197 (re-processed using RMA), is linked below as a supplementary file (GSE48814_complete_data.txt).

first: C

first: L

last: Alvarez-Cohen


title: Overexpression of ATF4 in BEAS2B cells

status: Public on May 01 2013

first: Katrina

last: Steiling

address: 72 East Concord Street, E-631

department: Medicine

institute: Boston Univeristy School of Medicine

summary: To determine gene expression changes induced by ATF4 overexpression, RNA was isolated from BEAS2B cells after overexpression of ATF4 or negative control.  mRNA expression was profiled using Affymetrix Human Gene 1.0 ST Arrays.

overall_design: RNA isolated from BEAS2B cells after overexpression of ATF4 or empty vector was processed and hybridized to Affymetrix Human Gene 1.0 ST Arrays.  Experiments were performed in triplicate.  Data from the 6 microarrays were used for RMA normalization.  RMA normalization was performed in the R statsitical environment using the affy package.  A t-test was used to determine the association of gene expression with ATF4 overexpression.

first: Avrum

last: Spira


title: Differential microRNA expression profiles of mouse lungs infected with 2009 pandemic H1N1 influenza virus and seasonal H1N1 virus

status: Public on Apr 17 2013

city: Beijing

first: Peng

last: Li

address: Dongda street 20# Fengtai District

department: Department of Infectious Disease Control

country: China

institute: Institute of Disease Control and Prevention

summary: To further understand the molecular pathogenesis of the 2009 pandemic H1N1 influenza virus infection, we profiled cellular miRNAs of lung tissue from BALB/c mice infected with influenza virus BJ501 and a mouse-adapted influenza virus A/Puerto Rico/8/34 (H1N1)(PR8) as a comparison.

overall_design: Five groups of mice were selected, and three of each group were used to profile the miRNA, two were in case for unqualified RNA extraction. Whole lungs from mice infected by BJ501 or PR8  were harvested on 2,5 days post infection (dpi), and compared with lung samples from 5 uninfected mice.

first: Zhihao

last: Wu

first: Chuanfu

last: Zhang

first: Xiaoai

first: Hongbin

last: Song


title: Mouse strain-specific responses to commensal gut bacteria at the mucosal gene expression level

status: Public on Apr 09 2013

city: Nuthetal

first: Gunnar

last: Loh

address: Arthur-Scheunert-Allee 114-116

department: Gastrointestinal Microbiology

country: Germany

institute: German Institute of Human Nutrition

summary: The host genotype has been proposed to contribute to individually composed bacterial communities in the gut. To provide deeper insight into interactions between gut bacteria and their host, we associated germ-free C3H and C57BL/10 mice with intestinal bacteria from a C57BL/10 donor mouse. Analysis of microbiota similarity between the experimental animals with denaturing gradient gel electrophoresis (DGGE) 13 weeks after association revealed the development of a mouse strain specific microbiota. Gene expression in the colonic mucosa was analyzed with a microarray approach taking advantage of a modified Affymetrix mouse genome chip. We detected 202 genes whose expression differed significantly by a factor of < 2. Application of bioinformatics tools demonstrated that functional terms including signaling/secretion, lipid degradation/catabolism, guanine nucleotide/guanylate binding and immune response were significantly enriched in differentially expressed genes. We had a closer look at the 56 genes with expression differences of < 4 and observed a higher expression in C57BL/10 mice of the genes coding for toll-like receptor 1 (4-fold) and angiogenin 4 (33-fold) which are involved in the recognition and response to gut bacteria. In contrast, a 70-fold higher expression of the phospholipase A2, group IIA-encoding gene (Pla2g2a) was detected in C3H mice. In addition, a number of interferon-inducible genes were higher expressed in C3H than in C57BL/10 mice including Gbp1 (18-fold), Mal (7-fold), Oasl2 (7-fold), Ifi202b (7-fold), Rtp4 (6-fold), Ly6g6c (5-fold), Ifi27l2a (5-fold), Usp18 (5-fold), Ifit1 (5-fold), Ifi44 (4-fold), and Ly6g (4-fold) indicating that these cytokines play an essential role in microbiota regulation. However, genes coding for interferons, their receptors or factors involved in interferon signaling pathways were not differentially expressed between the two mouse strains. Taken together, our study confirms that the host genotype is involved in the establishment of host-specific bacterial communities in the gut. Based on expression differences after colonization with the same bacterial inoculum, we propose that Pla2g2a and interferon-dependent genes may contribute to this phenomenon.

overall_design: Total RNA was extracted from the colonic mucosa  and hybridization was performed using 12.5 – 20 µg of cDNA on a customized Affymetrix nugomm 1a520177 chip.


title: Expression profile of Yeast Frataxin Mutants

status: Public on Mar 06 2013

city: Lleida

first: Jordi

last: Tamarit

address: Av Rovira Roure 80

department: Ciències Mèdiques Bàsiques

institute: Universitat de Lleida

summary: Yeast Frataxin Homologue 1 has been involved in oxidative stress and iron-sulfur biogenesis within the mitochondria. We have investigated the expression profile of conditional Yfh1 mutants. Yfh1 depletion leads to activation of iron uptake and repression

overall_design: Two strains were used: WT (strain W303-1A) and tetYFH1 (YFH1 gene under the control of a tet promoter regulatable with doxycycline). We compared the expression profile of tetYFH1 vs tetYFH1 + 10h doxycycline. As control we also compared W303-1A vs W303-1A

first: Armando

last: Moreno-Cermeño

first: David

last: Alsina

first: Elisa

last: Cabiscol

first: Joaquim

last: Ros


title: MCL1 is deregulated in subgroups of diffuse large B-cell lymphoma

status: Public on Jan 04 2013

city: Münster

first: Michael

last: Grau

state: Nordrhein-Westfalen

address: Albert-Schweitzer-Campus 1

department: Faculty of Medicine

institute: University of Münster

type: Genome variation profiling by genome tiling array

summary:
MCL1 is an anti-apoptotic member of the BCL2 family that is deregulated in various solid and hematological malignancies. However, its role in the molecular pathogenesis of diffuse large B-cell lymphoma (DLBCL) is unclear. We analyzed gene expression profiling data from 350 DLBCL patient samples and detected that activated B-cell-like (ABC) DLBCLs express MCL1 at significantly higher levels compared to germinal center B-cell-like (GCB) DLBCL patient samples (p=2.7 x 10(-10)) [PMID 23257783]. Immunohistochemistry confirmed high MCL1 protein expression predominantly in ABC DLBCL in an independent patient cohort (n=249; p=0.001).
To elucidate molecular mechanisms leading to aberrant MCL1 expression, we analyzed array comparative genomic hybridization (aCGH) data of 203 DLBCL samples [GSE11318] and identified recurrent chromosomal gains/amplifications of the MCL1 locus that occurred in 26% of ABC DLBCLs. In addition, aberrant STAT3 signaling contributed to high MCL1 expression in this subtype. Knockdown of MCL1 as well as treatment with the BH3-mimetic obatoclax induced apoptotic cell death in MCL1 positive DLBCL cell lines. In summary, MCL1 is deregulated in a significant fraction of ABC DLBCLs and contributes to therapy resistance. These data suggest that specific inhibition of MCL1 might be utilized therapeutically in a subset of DLBCLs.

overall_design: This GEO dataset is comprised of aCGH measurements for DLBCL cell lines, which are used in the above-mentioned paper. Cell lines were measured against the DNA of a healthy male donor who in turn was measured against a pool of healthy DNAs to correct for individual CNVs of the donor.

first: Georg

last: Lenz


title: Transcriptomic response of the polycyclic aromatic hydrocarbon-degrading Sphingomonas sp. LH128 upon short-term starvation stress

status: Public on Feb 14 2013

city: Heverle

first: Tekle Tafese

last: Fida

address: Kastelpark Arenberg 220

department: Earth and environmental Science

institute: KU Leuven

summary: This study examines genome-wide expression of the phenanthrene-degrading Sphingomonas sp. LH128 as a response to short-term starvation stress. For this purpose, the strain was subjected to complete nutrient starvation for 4h after growth on a rich medium. Survival was monitored by plating and transcriptomic response was determined by whole-genome microarray analysis. The data showed no major differences were obsrved in gene expression and the viability of the cells were not affected during short-term incubation time

overall_design: Transcriptomic response of phenanthrene degrading Sphingomonas sp. LH128 starved for 4h in isotonic solution of 0.01 mM MgS04 was studied using genome-wide gene expression analysis. For this purpose, the strain was pregrown in minimal medium to an OD600 of 0.5, washed twice with 0.01 mM MgS04 and resuspended in the same solution to an OD of 0.5. RNA was extracted both from starved cells and from the initial culture (non-starved cells) and cDNA was synthesized and labeled with Cy3. Transcriptomic response of three replicates were analyzed and compared with the initial inoculum

first: TEKLE

last: FIDA

first: SLVIA

last: MORENO

first: HERMANN

last: HEIPIEPER

first: JAN-ROELOF

last: VAN DER MEER

first: DIRK

last: SPRINGAEL


title: Transcriptomic response of the polycyclic aromatic hydrocarbon-degrading Sphingomonas sp. LH128 upon long-term starvation stress

status: Public on Oct 25 2012

summary: This study examines genome-wide expression of the phenanthrene-degrading Sphingomonas sp. LH128 as a response to long-term starvation stress. For this purpose, the strain was subjected to complete nutrient starvation for 6 months after growth on a rich medium. Survival was monitored by plating, physiological response was examined by flow cytometry and FAME analysis, and this response was related to transcriptomic response as determined by whole-genome microarray analysis. The data showed that decreased gene functions involved in ribosomal proteins biosynthesis, decreased chromosomal replication, increased gene functions involved in stringent regulation of gene expression, increased gene functions involved in genetic exchange and recombination, increased efflux systems, increased degradation of rRNA, and increased β-oxidation of fatty acids to access stored nutrients. Genes involved in PAH degradations appears to be with decreased expression.

overall_design: Transcriptomic response of phenanthrene degrading Sphingomonas sp. LH128 starved for 6 month in isotonic solution of 0.01 mM MgS04 was studied using genome-wide gene expression analysis. For this purpose, the strain was pregrown in minimal medium to an OD600 of 0.5, washed twice with 0.01 mM MgS04 and resuspended in the same solution to an OD of 0.5. RNA was extracted both from starved cells and from the initial culture (non-starved cells) and cDNA was synthesized and labeled with Cy3. Transcriptomic response of three replicates were analyzed and compared with the initial inoculum

first: JAN


title: Different responses of murine and human macrophages to leptospiral infection revealed by comparative array analysis

status: Public on Mar 15 2013

first: Feng

last: Xue

state: Beijing

address: No. 95, Yong'an Road

department: Department of Microbiology and Parasitology

institute: Beijing Tropical Medicine Research Institute, Beijing Friendship Hospital, Capital Medical University

summary: Leptospirosis is a re-emerging tropical infectious disease caused by the pathogenic Leptospira spp. The different host innate immune responses were partially related to the different severity of leptospirosis. In this study, we employed transcriptomics and cytokine array to comparatively calculate the responses of murine peritoneal macrophages (MPM) and human peripheral blood monocytes (HBM) to leptospiral infection, and uncover a series of different expression regulations of these two immune cells. The regulation percentages in several biological processes of MPM, such as the antigen process and presentation, the regulation of membrane potential, and the innate immune response, etc., were much greater than those of HBM (>2 folds). In HBM and MPM, the CASP8 and FADD-like apoptosis regulator genes were significantly up-regulated, which supported previous results that the caspase-8/3 pathway plays an important role in macrophage apoptosis during leptospiral infection. The key component of complement pathway, C3, was only up-regulated in MPM. Several cytokines (e.g. IL-10, TNF-alpha) were differently expressed at both mRNA and protein level in MPM and HBM. The differences in the transcriptomics and the cytokine expression revealed in this study were partially consistent with the different outcomes of the chronic and acute leptospirosis; thus, these findings facilitated further molecular study on the innate immune response to leptospiral infection.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Murine peritoneal macrophages (MPM) and human peripheral blood monocytes (HBM) were infected by pathogenic Leptospira. The host cells were collected at 1h, 2h, and 4h after infection. The uninfected macrophage sample was designed as a negative control. Three biological replicates were designed for each sample type. There were totally 24 hybridizations (12 for MPM, and 12 for HBM) included in this submission.

first: Jinping

last: Zhao


title: Microarray analysis on brains of mice fed a normal diet and supplemented with either oil palm phenolics (OPP) or distilled water

status: Public on Aug 12 2012

city: Kajang

first: Soon-Sen

last: Leow

state: Selangor

address: No. 6, Persiaran Institusi, Bandar Baru Bangi

country: Malaysia

institute: Malaysian Palm Oil Board

summary: OPP (1500 ppm gallic acid equivalent (GAE)) was supplemented to BALB/c mice given a normal diet to observe for possible neuroprotective effects. Brains were harvested six weeks after the feeding regimen for gene expression studies. Results from the separate microarray data analysis carried out on the different organs show that OPP have neuroprotective properties in vivo.

overall_design: Total RNA obtained from brains of BALB/c mice given OPP (six weeks after the feeding regimen) were compared to controls given distilled water (three replicates in the treatment group versus three replicates in the control group)

first: Shamala

middle: D

last: Sekaran

first: Ravigadevi

last: Sambanthamurthi


title: Comparative analysis between wild type and hap5-null mutant in response to rapamycin treatment

status: Public on Mar 24 2013

city: Hsinchu

first: Po-Chen

last: Hsu

address: Room 404, Life Science Building I, No. 101, Section 2, Kuang-Fu Road

department: Institute of Molecular and Cellular Biology

institute: National Tsing Hua university

summary: To identify the potential Hap5-regulated genes, we treated wild type cells and hap5 mutant cells with rapamycin and their corespondent transcriptional profiles were compared. Total 608 significant genes were identified. Among them, 343 genes are upregulated in hap5 mutant in repsonse to rapamycin while 265 genes are downregulated in comparison with the wild type.

overall_design: Rapamycin induced gene expression in wild type and hap5-null mutant was measured after 4.5 h growth in YPD followed by 0.5 h of incubation in the presence of 0.2 ug/ml rapamycin.  Four independent experiments were performed for each strain.

first: Chung-Yu

last: Lan


title: Transcriptionfactor study during the zebrafish development

status: Public on Jul 31 2012

city: Eggenstein-Leopoldshafen

first: Jessica

last: Legradi

state: Baden-Wuerttemberg

address: Hermann-von-Helmholtz-Platz 1

department: Institute of Toxicology and Genomics ITG

institute: Karlsruhe Institute of Technology

summary: We used a self-designed microarray to study the gene-expression of transcription factors in 5 different developmental stages and two tissue-samples

overall_design: Three to 6 biological replicates were performed, labeled with two different colors

first: J

first: L

last: Yang

first: O

last: Armant

first: M

last: Maerz


title: Chemosensitization of DLBCL cells in vitro and in vivo by demethylating nucleoside analogues [gene expression]

status: Public on Jul 30 2012

city: New-York

first: thomas

last: clozel

state: NY

address: 1300 York Avenue

department: Hematology-Oncology

institute: Weill Cornell Medical College

summary: Silencing of genes that suppress the malignant phenotype by DNA methylation spurred an interest in the clinical use of epigenetic reprogramming agents. Single therapy is unlikely to be curative in the context of a heterogeneous disease such as Diffuse Large B cell Lymphomas (DLBCL). The combination of DNA demethylating drugs could increase the chance to respond to classical and new treatments. We found that DLBCL cell lines respond heterogeneously to DNA demethylating agents. In sensitive cell lines, 5-aza-2’-deoxycytidine induced a genomic signature similar to that of doxorubicin, the most important drug of the combinatorial chemotherapy regimen for DLBCL treatment. Accordingly, the combination of 5-aza-2’-deoxycytidine and doxorubicin proved to be synergistic in cell killing in vitro and in vivo for DLBCL cell lines individually responsive to these drugs. In doxorubicin resistant cell lines, long-term exposure to low-dose of 5-aza-2’-deoxycytidine induces DNA demethylation and subsequent doxorubicin sensitization in vitro and in vivo. This later effect correlates with SMAD1 demethylation. SMAD1 is epigenetically silenced in doxorubicin-resistant DLBCL cells and DLBCL patients with poor prognostic. In addition, we found that DNA demethylating agents can sensitize primary DLBCL cells to doxorubicin. Primary cells obtained from a DLBCL patient treated with 5-azacytidine shows SMAD1 demethylation and ex vivo sensitization to multiple drugs. Therefore, DNA demethylating drugs can reprogram otherwise resistant DLBCL cells to respond to chemotherapy agents without increasing the toxicity to normal tissues. Our data also indicate that DNA methylation and consequent suppression of SMAD1 expression represent a previously undescribed molecular mechanism of chemoresistance in DLBCL that can be further exploit for therapy.

overall_design: A microarray study using double-stranded cDNA from different DLBCL cell lines before any treatment. Two biological replicates by cell line. The gene expression will be used to find gene expression signatures between sensitive and resistant cell lines to chemotherapeutics.

first: Leandro

last: Cerchietti

first: Shao-Ning


title: MiR-374a Promotes Epithelial-Mesenchymal Transition (EMT) and Metastasis of Breast Cancer (miRNA dataset)

status: Public on Jul 14 2012

city: Guangzhou

first: Mengfeng

last: Li

state: Guangdong

address: 74# zhongshan 2 Rd.

department: Zhongshan school of medicine

institute: Sun Yat-sen University

summary: This experiment is designed to screen miRNAs that are deregulated during breast cancer metastasis. Comparatively analyzing miRNAs in parental MDA-MB-435 cells and cells obtained from its lung metastases, 23 miRNAs expressed differentially, among which 12 were elevated and 11 were down-regulated.

overall_design: Total RNA were extracted from parental MDA-MB-435 cells and cells obtained from its lung metastases after 30 days inoculation. Two biological replications for each treatment.

first: Junchao

last: Cai


title: Translational microarray analysis of liver samples treated with PredTox hepatotoxic compound FP014SC

status: Public on Jun 21 2012

city: Leicester

first: Kate

middle: Marie

last: Dudek

address: Hodgkin Building, Lancaster Road

department: Genomics Group

institute: MRC Toxicology Unit

summary: Liver samples were lysed (15 mM Tris-HCl (pH 8.0), 300 mM NaCl and 15 mM MgCl2, plus inhibitors (1 mg/ml heparin 100 µg/ml cycloheximide and 80 U RNAsin)), added  to a 10-50% sucrose gradient and ultracentrifuged at 182,000x g for 2 hours. The gradient was aliquotted into 16 1ml fractions and added to Tri reagent (Sigma). RNA was extracted as per the manufacturer's instructions and then sub-pooled into fractions corresponding to monosomes, light polysomes, medium polysomes and heavy polysomes, according to ribosomal density (more ribosomal occupancy = higher translational activity = heavier, and therefore, located in the more dense fractions). Microarray analysis was performed by hybridising control (vehicle treated) monosomes against test (high-dose treated) monosomes on one microarray, control light polysomes against test light polysomes on a second microarray, and so forth for each sub-pool of fractions. Following microarray analysis there were a maximum of four values for each mRNA, corresponding to the proportional representation of that mRNA within each sub-pool of fractions. By calculating the change in values, i.e. degree of slope, across the monosomal region, the light polysomal region, the medium polysomal region and the dense polysomal region it was possible to determine any translational change in activity. In addition, the overall transcriptional change could be analysed for each mRNA.

overall_design: Dual colour microarrays performed on n = 3 samples, implementing a dye swap design. Control samples were treated with vehicle only. The treated samples had 1120mg/kg of the compound (Tetraethyl[(3-hydroxy-2-pyridyl)amino]-methanediphosphonate) for 15 days.


title: Expression data of colon cancer by conventional microarray and by suppressive subtractive hybridization coupled to microarray

status: Public on Jun 27 2012

city: Cordoba

first: German

middle: Alexis

last: Gonzalez

state: Cordoba

address: Av. Armada Argentina 3555

department: School of Engineering

country: Argentina

institute: Catholic University of Cordoba

summary: Transcriptional profiling of colonic tumors compared to paired adjacent non-tumoral colonic tissue by two strategies. The first strategy is a conventional microarray assay, while the second strategy involves a previous Suppression Subtractive Hybridization (SSH) followed by a conventional microarray assay. We applied subtractive hybridization followed by microarray analysis to identify tumor-specific low-abundance transcripts. The goal was to determine differences in gene expression profile in tumor samples by both strategies.

overall_design: Two-condition experiment: tumoral vs. adjacent non-tumoral tissues. Biological replicates: 10 tumoral replicates, 10 non-tumoral replicates. The same replicates were used for both strategies (conventional, SSH). Human Universal Reference RNA (Clontech) was used as the reference.

first: Felipe

last: Benavente

first: Carolina

last: Bizama

first: Jaime

last: Espinoza

first: Edgardo

last: Salvatierra

first: Ana

last: Gutiérrez

first: Elmer

last: Fernández

first: Eduardo

last: Sagredo

first: Iván

last: Roa

first: Manuel

last: Gidekel

first: Osvaldo

last: Podhajcer


title: Array comparative genomic hybridization to identify genomic differences between male and female zebrafish.

status: Public on Apr 17 2012

city: Singapore

first: Woei Chang

last: Liew

address: 1 Research Link, NUS

country: Singapore

institute: Temasek Life Sciences Laboratory

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

overall_design: Refer to individual Series

first: Woei

first: Richard

last: Bartfai

first: Zijie

last: Lim

first: Rajini

last: Sreenivasan

first: Kellee

last: Siegfried

first: Laszlo

last: Orban


title: Gene expression data from advanced gastric and colon cancers: tumoral vs. matched non-tumoral tissues

first: German

last: Gonzalez


title: Expression data of gastric cancer by conventional microarray and by suppressive subtractive hybridization coupled to microarray

summary: Transcriptional profiling of gastric tumors compared to paired adjacent non-tumoral gastric tissue by two strategies. The first strategy is a conventional microarray assay, while the second strategy involves a previous Suppression Subtractive Hybridization (SSH) followed by a conventional microarray assay. We applied subtractive hybridization followed by microarray analysis to identify tumor-specific low-abundance transcripts. The goal was to determine differences in gene expression profile in tumor samples by both strategies.

overall_design: Two-condition experiment: tumoral vs. adjacent non-tumoral tissues. Biological replicates: 12 tumoral replicates, 12 non-tumoral replicates. The same replicates were used for both strategies (conventional, SSH). Human Universal Reference RNA (Clontech) was used as the reference.

first: Carolina

first: Felipe

last: Benavente


title: Early transcriptional programming links progression to hepatitis C virus-induced severe liver disease in transplant patients

status: Public on Feb 10 2012

city: Seattle

first: Michael

last: Katze

state: WA

address: Rosen Building 960 Republican St.

department: Microbiology

institute: University of Washington

summary: This study determined key transcriptional signatures associated with HCV recurrence and eventual development of severe liver disease in infected transplant patient liver biopsies over time following transplant. We identified molecular signatures associated with severe fibrosis and liver injury prior to histologic evidence of disease progression.

overall_design: 111 liver biopsy specimens collected longitudinally from 57 HCV-infected liver transplant patients following organ transplant. Several batches of pooled normal liver RNA samples (Utah normal pool; UNP) are included.

first: Angela

middle: L

last: Rasmussen

first: Nicolas

last: Tchitchek

first: Nathan

last: Susnow

first: Alexei

last: Krasnoselsky

first: Deborah

last: Diamond

first: Matthew

last: Yeh

first: Sean

last: Proll

first: Marcus

last: Korth

first: Kathie

last: Walters

first: Sharon

last: Lederer

first: Anne

last: Larson

first: Robert

last: Carithers

first: Arndt

last: Benecke

middle: G


title: Enterococcus faecalis OG1RF vs. Δrel mutants under control or mupirocin-treated conditions

status: Public on Dec 20 2011

city: Gainesville

first: Jessica

middle: K

last: Kajfasz

state: FL

address: 1395 Center Drive

department: Oral Biology

institute: University of Florida

summary: Transcriptional profiling to define the stringent response regulon and significance of basal (p)ppGpp levels in E. faecalis OG1RF.

overall_design: RNA was extracted from four replicate samples of each strain of interest and labeled with Cy3. For each replicate, labeled RNA was hybridized to slides along with Cy5-labeled reference RNA, extracted from E. faecalis OG1RF cells grown to mid-log.

first: Anthony

middle: O

last: Gaca

first: Jacqueline

last: Abranches

first: Jose

last: Lemos


title: ASH1_A549 vs. vector control_A549

status: Public on Oct 31 2011

city: Nagoya

first: Osada

last: Hirotaka

state: Aichi

address: 1-1 Kanokoden, Chikusa-ku

institute: Aichi Cancer Center Research Institute

summary: We performed genome-wide expression profiling of miRNAs in order to search for those significantly affected by ASH1 transduction in a lung cancer cell line.

overall_design: Microarray analysis was　conducted to examine miRNA expression profiles using a Human miRNA Microarray, pre-commercial version 6.0 (Agilent) with 470 miRNA probes, according to the manufacturer’s instructions. A549 cells were infected with an ASH1-expressing or empty lentivirus and harvested 4 days later.

first: Eri

last: Nishikawa

first: Hirotaka

last: Osada

first: Yasumasa

last: Okazaki

first: Chinatsu

last: Arima

first: Shuta

last: Tomida

first: Yoshio

last: Tatematsu

first: Ayumu

last: Taguchi

first: Yukako

last: Shimada

first: Kiyoshi

last: Yanagisawa

first: Yasushi

last: Yatabe

first: Shinya

last: Toyokuni

first: Yoshitaka

last: Sekido

first: Takashi

last: Takahashi


title: Behavioural fever and its underlying molecular mechanisms

status: Public on Oct 06 2011

city: Chile

first: Sebastian

last: Boltana

state: Bio BIO

address: Edmundo Larenas s/n, Barrio Universitario. Biotechnology Center

department: Oceanografia

country: Chile

institute: Universidad de Concepcion

summary: Fever implies a significant increase in corporal temperature that aids toward the resolution of infective processes such as viral disease. The majority of vertebrate species are not homeothermic therefore must rely upon the environment for temperature regulation. Here we show that in the zebrafish an artificial viral infection, induced by poly (I:C), induces a fever response regulated by the behavioral choice of temperature. We recorded 12 h of the diurnal cycle in zebrafish previously treated (first 12 hours dark cycle) with the pyrogen, poly (I:C) [10 μg⋅kg-1]. Fish, n=10, were held in a thermal gradient (36-180C) separated into 7 interconnected chambers. Presence or absence of individuals in each chamber was recorded each 15 minutes throughout the 12 hour period. After the experimental period we dissected whole brains for microarray analysis. In monitored zebrafish intraperitoneal treatment with poly (I:C) induced a febrile behaviour with significantly elevated temperature preference (T of about 32ºC) in stark contrast with the observed frequency for saline-injected fish (28ºC). Microarray analyses uncovered significant shifts in transcriptional activity that were highly directed in the poly (I:C)-treated fish housed in the thermal gradient. When compared to gene expression profiles from poly (I:C)-treated fish deprived of a thermal gradient we observed a less intense specific to the poly (I:C) challenge and interestingly observed a scattered generalised stress response. Our results highlight the influence of temperature preference in the development of the immune response in zebrafish.

overall_design: Fever induced gene expression in zebrafish brain was measured at 24 after intra peritoneal injection with 1mg*Kg-1 of Poly (I:C) in zebrafish. Four independent experiments were performed to explore the transcriptomic profile induced by animals injected by Poly (I:C) with/whithout thermal gradient, saline solution (PBS), or control using different animals for each experiment.

last: Boltaña

first: Simon

last: Mackenzie


title: Mating-type specific gene expression in the filamentous ascomycete Neurospora tetrasperma

status: Public on Jan 26 2011

city: Uppsala

first: Nicklas

last: Samils

address: Norbyägen 18D

department: Evolutionary Biology

country: Sweden

institute: Uppsala University

summary: Neurospora tetrasperma is a pseudohomothallic filamentous ascomycete with a large (~ 7 Mbp) region of suppressed recombination surrounding its mating-type (mat) locus. The suppressed recombination has lead to sequence divergence between the two mating-type chromosomes of wild-type heterokaryotic strains, while the remaining genome is largely homoallelic. In this study, we use microarray technology to manifest expression divergence linked to mating type in N. tetrasperma. N. tetrasperma and N. crassa, were grown on agar regimes inducing sexual growth (Synthetic Crossing medium) and vegetative growth (Vogel's Medium), respectively.

sample_organism:
Neurospora crassa
Neurospora tetrasperma

overall_design:
[SC]: Neurospora tetrasperma mat-A FGSC#1270; mat-a FGSC#1271; Mat-A FGSC#9033; mat-a FGSC#9034; N. crassa mat-A FGSC#2489 and mat-a FGSC 4200: Synthetic Crossing medium was used as a nutrient regime before sampling and processing

[Veg]: Neurospora tetrasperma mat-A FGSC#1270; mat-a FGSC#1271; Mat-A FGSC#9033; mat-a FGSC#9034; N. crassa mat-A FGSC#2489 and mat-a FGSC 4200: Vogel's Medium (Vegetative Medium) was used as a nutrient regime before sampling and processing

first: Magnus

last: Karlsson

first: Zheng

last: Wang

first: Takao

last: Kasuga

first: Eric

last: Bastiaans

first: Jeffrey

middle: P

last: Townsend

first: Hanna

last: Johannesson


title: Endothelial Response to Increased Shear Stress Frequency

status: Public on Dec 01 2011

city: Durham

first: Ji

last: Zhang

state: NC

address: Room 136 Hudson Hall

department: Department of Biomedical Engineering

institute: Duke University

summary:
Porcine endothelial cells were preconditioned by a basal level shear stress of 15 ± 15 dynes/cm2 at 1 Hz for 24 hours, and an acute increase in shear stress frequency (2 Hz) was then applied. The transcriptomics studies using microarray identified  genes that were sensitive to the elevated shear frequency.
keywords: adaptation, shear stress, frequency, microarray, gene expression

overall_design: Porcine endothelial cells were preconditioned by a basal level shear stress of 15 ± 15 dynes/cm2 at 1 Hz for 24 hours, and an acute increase in shear stress magnitude (2 Hz) was then applied. Gene expression at multiple time points was measured using microarray.

first: Morton

middle: H

last: Friedman


title: Chromosome conformation capture on chip (4C) at the Fab7 polycomb response element in the bithorax complex of Drosophila

status: Public on Dec 31 2010

city: Montpellier

first: Giacomo

last: Cavalli

email: itys.comet@bric.ku.dk, benjamin.leblanc@igh.cnrs.fr, frederic.bantignies@igh.cnrs.fr, giacomo.cavalli@igh.cnrs.fr

address: 141 rue de la Cardonille

country: France

institute: Institute of Human Genetics

summary: In Drosophila melanogaster, Hox genes are organized in an anterior and a posterior cluster, called Antennapedia complex and bithorax complex, located on the same chromosome arm and separated by 10 Mb of DNA. Both clusters are repressed by Polycomb group (PcG) proteins. Here, we show that genes of the two Hox complexes can interact within PcG bodies in the cell nucleus in tissues where they are corepressed. This colocalization increases during development and depends on PcG proteins. Hox gene contacts are part of a large gene interaction network that includes other PcG target genes. Moreover, they are conserved in the distantly related Drosophila virilis species, and mutations on one of the loci weaken silencing of genes in the other locus, resulting in the exacerbation of homeotic phenotypes in a sensitized genetic background. Thus, the three-dimensional organization of Polycomb target genes in the cell nucleus is an evolutionarily conserved phenomenon that stabilizes the maintenance of epigenetic gene silencing.

overall_design:
Chromosome Conformation Capture on Chip (4C) was used to establish chromosomal interactions involving the Fab7 polycomb response  element from the bithorax complex of Drosophila melanogaster. Larval brain and anterior discs from 3rd instar larvae were dissected to produce 2 replicate experiments with wild-type (WT-r1, WT-r2) and Fab-712 (F12-r1, F12-r2) fly lines.
The 3C was performed as previously described (Hagege et al., 2007; Miele and Dekker, 2009) with the main differences being the use of DpnII (New England Biolabs), a 4 bp cutter restriction enzyme, and a fixation in 3% para-formaldehyde for 30 min, maximizing sensitivity and resolution of contact detection.
The 4C DNA and DpnII digested genomic DNA (Control) samples were hybridized  using a tiling array mapping the whole chromosome 3R which includes the antenapedia and bithorax hox clusters.

first: Itys

last: Comet

first: Benjamin

last: Leblanc

first: Frederic

last: Bantignies


title: Interferon signaling related gene expression and clinical outcome of chronic hepatitis C

status: Public on Feb 11 2010

city: Yokohama

first: Hiroyuki

last: Oshima

address: 10-1 Daikoku-cho, Tsurumi-ku

institute: Mitsubishi Rayon Co.,LTD

summary: Chronic hepatitis C (CH) frequently developed liver cirrhosis and hepatocellular carcinoma without suitable treatment. However, peginterferon(IFN) and ribavirin therapy (combination therapy), which is present standard therapy, is imperfect effect because of several side effect. We attempt to catalog IFN related gene expression profile according to the response of combination therapy in to predict the outcome of drug-response before administration and establish the adequate treatment according to the individual clinical information.

overall_design:
Chip1 [GPL9960-related Samples]: We quantified expression profile of 200 IFN related genes by microarray in the liver biopsy specimen from 88 CH patients without history of anti-viral therapy, 10 fatty liver patients without HCV infection and 5 normal livers were also enrolled.

Chip2 [GPL9962-related Samples]: We quantified expression profile of 37 IFN related genes by microarray in the liver biopsy specimen from 88 CH patients without history of anti-viral therapy, 10 fatty liver patients without HCV infection and 5 normal livers were also enrolled.

first: Koji

last: Onomoto

first: Shiho

last: Nakahara

first: Hidenori

last: Toyoda

first: Masami

last: Tanaka

last: Kumada

first: Kunitada

last: Shimotohno

last: Fujita

first: Yoshiki

last: Murakami


title: MicroRNA profiling of advanced serous ovarian carcinoma

status: Public on Feb 15 2011

city: New York

first: Douglas

last: Levine

address: 1275 York Ave.

department: Surgery

institute: MSKCC

summary: Objectives: MicroRNAs (miRNAs) are a class of small non-coding RNAs that negatively regulate gene expression primarily through post-transcriptional modification. We tested the hypothesis that miRNA expression is associated with overall survival in advanced ovarian cancer.  Methods: Cases included newly diagnosed patients with stage III or IV serous ovarian cancer. RNA from a training set of 62 cases was hybridized to an miRNA microarray containing 470 mature human transcripts. Cox regression was performed to identify miRNAs associated with overall survival. External validation was performed using quantitative RT-PCR miRNA assays in an independent test set of 123 samples. MiRNA targets and associated biologic pathways were predicted in silico.   Results: Of all patients, 80% had high-grade, stage IIIC tumors and 64% underwent optimal cytoreduction. The median survival for the entire cohort was 49 ± 4 months. The training set identified 3 miRNAs associated with survival - miR-337, miR-410, and miR-645. An miRNA signature containing miR-410 and miR-645 was most strongly associated with overall survival in the training set (HR=2.96, 95% CI: 1.51-5.78). This miRNA survival signature (MiSS) was validated in the test set (HR=1.71, 95% CI: 1.05-2.78). The MiSS was independent of FIGO stage and surgical debulking.   Conclusions: The data suggest that an MiSS that contains miR-410 and miR-645 is negatively associated with overall survival in advanced serous ovarian cancer. This signature, when further validated, may be useful in individualizing care for the ovarian cancer patient. Pathway analyses identify biologically plausible mechanisms.

first: Karin

last: Shih


title: The Role of RelA of Streptococcus mutans in Global Control of Gene Expression

status: Public on Nov 14 2007

first: Robert

last: Burne

address: 1600 SW Archer Rd. BOX 100424

summary:
The production of (p)ppGpp by Streptococcus mutans UA159 is catalyzed by three gene products, RelA, RelP and RelQ.  Here, we investigate the role of the RelA (Rel) homologue of S. mutans in the stringent response and in global control of gene expression.  RelA of S. mutans was shown to synthesize pppGpp in vitro from GTP and ATP in the absence of added ribosomes, as well as in vivo in an E. coli relA-spoT mutant.  Mupirocin (MUP) was shown to induce high levels of (p)ppGpp production in S. mutans in a relA-dependent manner, with a concommitant reduction in GTP pools.
Keywords: (p)ppGpp, nutrient starvation, biofilm, virulence, stress

overall_design:
For microarray analysis, cells were grown in the chemically-defined medium FMC  to an optical density at 600 nm (OD600) of 0.3 and the cultures were divided into two aliquots.  To one aliquot, 500 ng ml-1 of MUP was added and the cells were incubated at 37oC for 20 minutes (MUP-treated cells), while the other aliquot was collected by centrifugation and immediately frozen (MUP-control cells).  Since MUP treatment resulted in rapid growth arrest, the final OD600 of experimental and control samples were essentially identical.

Four individual Cy3-labeled cDNA samples originating from four different cultures of UA159 and JLrelA strains grown under control (FMC) and experimental conditions (FMC containing MUP) were hybridized to the arrays along with Cy5-labeled reference cDNA, generating a total of 16 slides.  The ratios of the signals of the test RNAs with that of the uniform reference RNA were analyzed within and between slides.

first: Marcelle

last: Nascimento


title: Transcriptome comparison between CCR2+ and CCR2- classical monocytes in SIV-infected Macaca nemestrina

status: Public on Mar 01 2012

city: Baltimore

first: Lucio

last: Gama

state: MD - EUA

address: 733 N Broadway St - BRB 831

department: Molecular and Comparative Pathobiology

institute: Johns Hopkins School of Medicine

summary:
Monocytes have been categorized in three main subpopulations based on CD14 and CD16 surface expression. Classical monocytes are the most abundant subset in the blood. They express a CD14+CD16-CCR2+ phenotype, which confers on them the ability to migrate to inflammatory sites by quickly responding to CCL2 signaling. Here we identified and characterized the surge and expansion of a novel monocyte subset during SIV and HIV infection. They were undistinguishable from classical monocytes regarding CD14 and CD16 expression, but did not express surface CCR2. Transcriptome analysis of sorted cells confirmed that they represent a distinct subpopulation that expresses lower levels of inflammatory cytokines and activation markers than their CCR2+ counterparts. They exhibited impaired phagocytosis and deficient chemotaxis in response to CCL2 and CCL7, besides being refractory to SIV infection. We named these cells atypical CCR2- classical (ACC) monocytes, and believe they play an important role in AIDS pathogenesis, possibly reflecting an anti-inflammatory response against the extreme immune activation observed during SIV and HIV infection. Antiretroviral therapy caused this population to decline in both macaque and human subjects, suggesting that this atypical phenotype may be induced by viral replication.

Expression profiling by NanoString nCounter gene expression system.

overall_design: Classical monocytes (CD14++CD16-) from six SIV-infected macaques (day 14 post inoculation) were sorted in two groups according to CCR2 expression.

first: Janice

last: Clements

first: Esper

last: Kallas


title: Global Nucleosome Positioning in Arabidopsis using whole genome Tiling Microarray

status: Public on Nov 24 2010

city: Lucknow

first: Samir

last: Sawant

state: UP

address: Sikandrabad

department: Plant Molecular Biology Lab

country: India

institute: National Botanical Research Institute

summary: We identified genome-wide nucleosome occupancy positions in Arabidopsis thaliana by comparing hybridization intensity difference between salicylic acid (induced) and water treated (uninduced) conditions with reference to genomic DNA.

overall_design: Genome-wide study of nucleosome dynamics in Arabidopsis thaliana in response to Salicylic acid (3 replicates) vs. Control Water (3 replicates) condition with comparison to genomic hybridization (2 replicates) using Tiling  Microarray.

middle: V

first: Amol

last: Ranjan

first: Mala

last: Singh

first: Sumit

last: Bag

first: Deepti

last: Nigam

first: Shrikant

last: Mantri


title: The Role of CcpA in Gene Regulation in Streptococcus mutans

status: Public on May 01 2008

country: USA

summary:
CcpA is a global regulator of transcription in Gram-positive bacteria that controls gene expression in response to carbohydrate availability.  Using the fructan hydrolase (fruA) gene of S. mutans as a model, we demonstrate that CcpA does indeed play a major role in carbohydrate catabolite repression.  Using DNA microarrays, the expression of at least 170 genes differed in CcpA-deficient cells compared to the parental strains when cells are grown in the presence of glucose, but only 90 genes showed altered expression when cells were cultivated in the poorly-repressing substrate galactose.  Interestingly, 90 genes were differently expressed in wild-type S. mutans in response to cultivation in glucose versus galactose, but the expression of over 500 genes was altered when growth in glucose and galactose were compared in the CcpA-deficient strain.  The results support a major role for CcpA in regulation of gene expression, but reveal that a substantial CcpA-independent network exists in S. mutans to regulate gene expression in response to carbohydrate source.
Keywords: Catabolite repression, sugar transport, biofilm, virulence, stress response

overall_design:
A reference RNA that had been isolated from 2 liters of UA159 cells grown in BHI broth to OD600 = 0.5 was used in every experiment.  Our experimental conditions consisted of S. mutans UA159 and TW1 grown in TV supplemented with 0.5% glucose or 0.5% galactose and collected at the mid-exponential phase of growth (OD600 = 0.5).  All RNAs were purified as described above and used to generate cDNA according to the protocol provided by TIGR with the following minor modifications.  The amount of RNA in each reaction was increased to 10 ug and the molar ratio of dTTP:aa-dUTP was increased to 1:2.  SuperscriptIII Reverse transcriptase (Invitrogen, Gaithersburg, MD) was used to increase cDNA yields.  Purified cDNAs were coupled with indocarbocyanine (Cy3)-dUTP, while reference cDNA was coupled with indodicarbocyanine (Cy5)-dUTP (Amersham Biosciences, Piscataway, NJ).  Four individual Cy3-labeled cDNA samples originating from four different cultures of UA159 or TW1, grown in each experimental condition, were hybridized to the arrays along with Cy5-labeled reference cDNA, generating a total of 16 slides.  Hybridizations were carried out in a Maui 4-chamber hybridization system (BioMicro Systems, Salt Lake City, Utah) for 17 h at 42ºC.  The slides were then washed according to TIGR protocols and scanned using a GenePix Scanner (Axon Instruments Inc., Union City, CA).
After the slides were scanned, single-channel images were loaded into TIGR Spotfinder software and overlayed.  A spot grid was created according to TIGR specifications and manually adjusted to fit all spots within the grid, then the intensity values of each spot were measured.  Data was normalized using TIGR Microarray data analysis software (MIDAS), by using LOWESS and Iterative Log Mean Centering with default settings, followed by In-Slide Replicate Analysis.  Statistical analysis was carried out using BRB Array Tools with a cutoff P value of 0.001 for class prediction and class comparison.


title: Role of EIIABMan and EIIGlc in regulation of energy metabolism, biofilms, and competence in Streptococcus mutans

status: Public on Sep 01 2007

summary:
The phosphoenolpyruvate:sugar phosphotransferase system (PTS) is the major carbohydrate transport
system in oral streptococci. The mannose-PTS of Streptococcus mutans, which transports mannose and glucose,
is involved in carbon catabolite repression (CCR) and regulates the expression of known virulence genes. In
this study, we investigated the role of EIIGlc and EIIABMan in sugar metabolism, gene regulation, biofilm
formation, and competence. The results demonstrate that the inactivation of ptsG, encoding a putative EIIGlc,
did not lead to major changes in sugar metabolism or affect the phenotypes of interest. However, the loss of
EIIGlc was shown to have a significant impact on the proteome and to affect the expression of a known virulence
factor, fructan hydrolase (fruA). JAM1, a mutant strain lacking EIIABMan, had an impaired capacity to form
biofilms in the presence of glucose and displayed a decreased ability to be transformed with exogenous DNA.
Also, the lactose- and cellobiose-PTSs were positively and negatively regulated by EIIABMan, respectively.
Microarrays were used to investigate the profound phenotypic changes displayed by JAM1, revealing that
EIIABMan of S. mutans has a key regulatory role in energy metabolism, possibly by sensing the energy levels
of the cells or the carbohydrate availability and, in response, regulating the activity of transcription factors and
carbohydrate transporters.
Keywords: Biofilms growth & development, Energy Metabolism, Gene Expression Regulation, Bacterial

overall_design:
S. mutans UA159 microarrays were provided by The
Institute for Genomic Research (TIGR). The microarrays consisted of 1,948
70-mer oligonucleotides representing 1,960 open reading frames. The full 70-mer
complement is printed four times on the surface of each microarray slide. Additional
details regarding the arrays are available at TIGR's Streptococcus mutans UA159 Genome Page
A reference RNA that had been isolated from 2
liters of UA159 cells grown in BHI broth to an optical density at 600 nm of 0.5
was used in every experiment. The reference RNA was purified as above, aliquoted,
and stored at -80°C. Our experimental conditions consisted of S. mutans
UA159 and JAM1 grown in BHI broth and collected at the mid-exponential
phase of growth (optical density at 600 nm = 0.5).  Six individual Cy3-
labeled cDNA samples originating from six different cultures of UA159 or JAM1
were hybridized to the arrays along with Cy5-labeled reference cDNA, generating
a total of 12 slides.

first: Jacqueline

last: Abranches

first: Melissa

last: Candella

first: Zezhang

middle: T

last: Wen

first: Henry

last: Baker


title: Influence of BrpA on critical virulence attributes of Streptococcus mutans

status: Public on Aug 30 2008

summary: Streptococcus mutans, the primary etiological agent of human dental caries, has developed multiple mechanisms to colonize and form biofilms on the tooth surface. The brpA gene codes for a predicted surface-associated protein with apparent roles in biofilm formation, autolysis, and cell division. In this study, we used two models to further characterize the biofilm-forming characteristics of a BrpA-deficient mutant, strain TW14. Compared to those of the parent strain, UA159, TW14 formed long chains and sparse microcolonies on hydroxylapatite disks but failed to accumulate and form three-dimensional biofilms when grown on glucose as the carbohydrate source. The biofilm formation defect was also readily apparent by confocal laser scanning microscopy when flow cells were used to grow biofilms. When subjected to acid killing at pH 2.8 for 45 min, the survival rate of strain TW14 was more than 1 log lower than that of the wild-type strain. TW14 was at least 3 logs more susceptible to killing by 0.2% hydrogen peroxide than was UA159. The expression of more than 200 genes was found by microarray analysis to be altered in cells lacking BrpA (P < 0.01). These results suggest that the loss of BrpA can dramatically influence the transcriptome and significantly affects the regulation of acid and oxidative stress tolerance and biofilm formation in S. mutans, which are key virulence attributes of the organism.

overall_design:
RNA extraction. S. mutans strains were grown in 50 ml of BHI broth and harvested at an OD600 of [congruent with]0.5 by centrifugation at 3,800 × g at 4°C for 5 min. The pellets were quickly resuspended and treated with RNAprotect (QIAGEN, Inc., CA) by using the procedures recommended by the supplier. The treated cells were collected by centrifugation at 3,800 × g at 4°C for 10 min and stored at −80°C. RNA extractions were performed by using hot phenol as previously described (33). To remove all DNA, the purified RNAs were treated with DNaseI (Ambion, Inc., Austin, TX) and retrieved with the RNeasy purification kit (QIAGEN, Inc., CA). To prepare a reference RNA, S. mutans UA159 was grown in 3 liters of BHI until mid-exponential phase (OD600 [congruent with] 0.5) and total RNA was extracted as described above. The purified RNA was then aliquoted and stored at −80°C until use.

cDNA synthesis, Cy dye coupling and array hybridization. Array analysis was performed by using the whole-genome S. mutans microarrays that were obtained from PFGRC at TIGR. The S. mutans genome array consisted of 70-mer oligonucleotides representing 1,960 open reading frames from strain UA159. The full 70-mer complement was printed four times on the surface of the slides. Experiments, including cDNA synthesis, Cy dye coupling, hybridization, and washing, were performed by using protocols from the PFGRC with minor modifications. Briefly, cDNA synthesis was carried out in a total volume of 45 μl by mixing 10 μg total RNA with 3 μg random hexamers (Invitrogen Life Technologies, CA), 9 μl 5× first-strand buffer, 4.5 μl 0.1 M dithiothreitol, 2 μl 12.5 mM dNTP/aa-UTP with a 1.5- to-1 ratio of aa-dUTP to dTTP, and 3 μl Superscript III reverse transcriptase. This mixture was incubated at 42°C for 16 h. Following the completion of cDNA synthesis, RNA was hydrolyzed with NaOH and the aminoallyl-labeled cDNA samples were purified by using the QIAGEN QIAquick PCR purification kit, followed by drying in a speed vacuum. The samples were then resuspended in 5 μl 0.1 M NaCO3 and incubated with 5 μl of Cy dye (resuspended as recommended by the manufacturer) for 2 h in the dark. For all experiments, the reference RNA samples were coupled to Cy5 and both wild-type and the BrpA-deficient mutant RNA samples were coupled to Cy3. Uncoupled Cy dyes were removed by the QIAquick PCR purification kit, and the coupled samples were eluted in 100 μl elution buffer. Each Cy3-labeled experimental sample was mixed with a Cy5-labeled reference sample, and the mixtures were allowed to dry in a speed vacuum. To hybridize, the samples were resuspended in 60 μl hybridization buffer and heated at 90°C for 10 min. Following a quick centrifugation, the Cy dye-coupled probes were applied to microarray slides and incubated at 42°C in a water bath for 17 h. Slides were then washed and scanned by using an Axon GenePix 4000B scanner (Axon Instruments, Foster City, CA).

Array data normalization and statistical analysis. Data were collected from at least four separate array slides, which contained four copies of the genome, with RNA isolated from four independent experiments. The raw data were loaded into the TIGR Spotfinder program and further normalized with the TIGR microarray data analysis system using LOWESS iterative log mean centering parameters with the default settings, followed by in-slide replicate analysis. Spots that were missing or labeled as “bad” during the upstream processes in 50% of the slides were cut off in the output files. The ratios of channel A over channel B were then converted to log2 values and analyzed with BRB array tools (version 3.01, developed by Richard Simon and Amy Peng Lam, National Cancer Institute, Bethesda, MD). A pairwise Student's t test was used to analyze the mean log ratios of the BrpA-deficient strain and the wild type, and genes that were differentially expressed with the significance level of a P value of <0.001 were selected.


title: The CodY Regulon in Streptoccus mutans

status: Public on Oct 31 2008

summary:
In this report, codY mutant strains were constructed and used to demonstrate the relationship of (p)ppGpp synthesized by RelP and RelQ with the activation of CodY.  In addition, because CodY has not been studied in S. mutans, we used microarrays to demonstrate that this protein function as a global regulator of gene expression in S. mutans.
Keywords: stress response, gene knockout analysis

overall_design:
For microarray analysis, cells were grown in complete FMC to an optical density at 600 nm (OD600) of 0.3 collected by centrifugation, quick frozen in a dry-ice/ethanol bath, and stored at - 80∞C for RNA isolation.

RNA was isolated from exponential phase (OD600 0.3), as described previously.  All samples were digested with DNase I (Ambion, Austin, TX) and purified using the RNeasy mini kit column (Qiagen, Inc., Chatsworth, CA).  RNA concentrations were determined spectrophotometrically in triplicate and one µg of RNA was run in a formaldehyde gel to verify RNA quality.  Reverse transcription (RT) reaction was performed with 10 mg of RNA following the protocol provided by TIGR with minor modifications.  Transcriptome analysis was performed using the S. mutans UA159 microarrays provided by The Institute for Genomic Research (TIGR).  The microarrays consisted of 1,948 70-mer oligonucleotides representing 1,960 open reading frames printed four times on the surface of each microarray slide.  Additional details regarding the arrays are available at http://pfgrc.tigr.org/descriptions/S_mutans.shtml.  A reference RNA prepared from a large-scale culture of S. mutans UA159 cells that had been grown in BHI broth to an OD600 = 0.5 was used in every experiment.   cDNA labeling was performed according to TIGR protocols with minor modifications as described elsewhere.  Four cDNA samples originating from four independent cultures of UA159 and JLcodY strains were hybridized to the arrays along with the reference cDNA, generating a total of 16 slides.  Hybridizations were performed in a Maui hybridization chamber (BioMicro Systems, Salt Lake City, UT).  Additional details regarding array protocols are available at http://pfgrc.tigr.org/protocols/protocols.shtml.  Data from all individual experiments were analyzed using Spotfinder software and normalized using Midas according to TIGR specifications (http://www.tigr.org/software/).  Statistical analysis was carried out using BRB Arrays Tools (http://linus.nci.nih.gov/BRB-ArrayTools.html) with a P-value of 0.001.

first: Jose

last: Lemos


title: Heterochromatin integrity affects chromosome reorganization after centromere dysfunction

status: Public on Aug 22 2008

city: Kami

first: Kojiro

last: Ishii

state: Kochi

address: 185 Miyanokuchi, Tosayamada

institute: Kochi University of Technology

summary:
The centromere is essential for the inheritance of genetic information on eukaryotic chromosomes. Epigenetic regulation of centromere identity has been implicated in genome stability, karyotype evolution and speciation. However, little is known regarding the manner in which centromere dysfunction affects the chromosomal architectures. Here we show that in the fission yeast Schizosaccharomyces pombe, the conditional deletion of the centromere produces survivors that carry either neocentromere-acquired chromosome at the sub-telomeric region or acentric chromosome rescued by inter-telomere fusion with other chromosome. The ratio of neocentromere formation to telomere fusion is considerably decreased by inactivation of genes involved in RNA interference (RNAi)-dependent heterochromatin formation. By affecting the modes of chromosomal reorganization, the genomic distribution of heterochromatin may influence the fate of karyotype evolution.
Keywords: ChIP-chip

overall_design: Chromosomal distributions of Cnp1, Mis6-GFP, H3me2K9, Swi6, Mis12-GFP and Cnp3-GFP were examined by immunoprecipitating the chromatin using anti-Cnp1, anti-GFP, anti-H3me2K9 and anti-Swi6 antibodies from the fission yeast cells harboring functional centromere at either authentic or ectopic position in the chromosome I. Immunoprecipitated DNA and input DNA were amplified by LM-PCR and labeled with either Cy5 or Cy3 by random priming. Labeled DNA samples were mixed and hybridised onto commercially-distributed S. pombe oligo array from Agilent. Data processing was performed using Agilent scanner and softwares.

first: Yuki

last: Ogiyama

first: Yuji

last: Chikashige

first: Saeko

last: Soejima

first: Fumie

last: Masuda

first: Tatsuyuki

last: Kakuma

first: Yasushi

last: Hiraoka

first: Kohta

last: Takahashi


title: p53-repressed miRNAs are Involved with E2F in a Feed Forward Loop Promoting Proliferation

status: Public on Sep 23 2008

city: Oslo

first: Hilde

last: Johnsen

address: Montebello

department: Genetics

country: Norway

institute: Norwegian Radium Hospital

summary:
Analysis of miRNA expression in human breast cancer samples with Agilent's miRNA arrays.
These samples are part of a study where we have investigated the mammalian cell proliferation control network consisting of transcription regulators, E2F and p53, their targets, and a family of 14 microRNAs. We observed that indicative of their significance, expression of these microRNAs is down-regulated in senescent cells and in breast cancers harboring wild-type p53. These microRNAs are repressed by p53 in an E2F1-mediated manner.

Abstract of paper:
Normal cell growth is governed by a complicated biological system, featuring multiple levels of control, often deregulated in cancers. The role of microRNAs in the control of gene expression is now increasingly appreciated, yet their involvement in controlling cell proliferation is still not well understood. Here we investigated the mammalian cell proliferation control network consisting of transcription regulators, E2F and p53, their targets, and a family of 14 microRNAs. Indicative of their significance, expression of these microRNAs is down-regulated in senescent cells and in breast cancers harboring wild-type p53. These microRNAs are repressed by p53 in an E2F1-mediated manner. Furthermore, we show that these microRNAs silence anti-proliferative genes, which themselves are E2F1 targets. Thus, microRNAs and transcriptional regulators appear to cooperate in the framework of a multi-gene transcriptional and post-transcriptional feed-forward loop. Finally, we show that, similarly to p53 inactivation, overexpression of representative microRNAs promotes proliferation and delays senescence, manifesting the detrimental phenotypic consequence of perturbations in this circuit. Together these findings position microRNAs as novel key players in the mammalian cellular proliferation network.

Keywords: Breast Cancer, miRNA, p53.

overall_design: 18 Primary human breast cancer samples analyzed for their miRNA expression. From two to four replicates were performed for each sample. Quality check (QC) were performed with Feature Extraction 9.1.3.44 and arrays not passing QC were excluded

first: Ran

last: Brosh

first: Reut

last: Shalgi

first: Atar

last: Liran

first: Gilad

last: Landan

first: Katya

last: Korotayev

first: Giang

last: Nguyen

first: Espen

last: Enerly

first: Yosef

last: Buganim

first: Hilla

last: Solomon

first: Ido

last: Goldstein

first: Shalom

last: Madar

first: Naomi

last: Goldfinger

first: Anne-Lise

last: Børresen-Dale

first: Doron

last: Ginsberg

first: Curtis

last: Harris

first: Yitzhak

last: Pilpel

first: Moshe

last: Oren

first: Varda

last: Rotter