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Haynes BC, Maier EJ, Kramer MH, Wang PI, Brown H, Brent MR. Mapping functional transcription factor networks from gene expression data. Genome Res 2013; 23:1319-28. [PMID: 23636944 PMCID: PMC3730105 DOI: 10.1101/gr.150904.112] [Citation(s) in RCA: 34] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022]
Abstract
A critical step in understanding how a genome functions is determining which transcription factors (TFs) regulate each gene. Accordingly, extensive effort has been devoted to mapping TF networks. In Saccharomyces cerevisiae, protein–DNA interactions have been identified for most TFs by ChIP-chip, and expression profiling has been done on strains deleted for most TFs. These studies revealed that there is little overlap between the genes whose promoters are bound by a TF and those whose expression changes when the TF is deleted, leaving us without a definitive TF network for any eukaryote and without an efficient method for mapping functional TF networks. This paper describes NetProphet, a novel algorithm that improves the efficiency of network mapping from gene expression data. NetProphet exploits a fundamental observation about the nature of TF networks: The response to disrupting or overexpressing a TF is strongest on its direct targets and dissipates rapidly as it propagates through the network. Using S. cerevisiae data, we show that NetProphet can predict thousands of direct, functional regulatory interactions, using only gene expression data. The targets that NetProphet predicts for a TF are at least as likely to have sites matching the TF's binding specificity as the targets implicated by ChIP. Unlike most ChIP targets, the NetProphet targets also show evidence of functional regulation. This suggests a surprising conclusion: The best way to begin mapping direct, functional TF-promoter interactions may not be by measuring binding. We also show that NetProphet yields new insights into the functions of several yeast TFs, including a well-studied TF, Cbf1, and a completely unstudied TF, Eds1.
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Affiliation(s)
- Brian C Haynes
- Center for Genome Sciences and Systems Biology, Washington University, Saint Louis, Missouri 63108, USA
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2
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Drake JI, Gomez-Arroyo J, Dumur CI, Kraskauskas D, Natarajan R, Bogaard HJ, Fawcett P, Voelkel NF. Chronic carvedilol treatment partially reverses the right ventricular failure transcriptional profile in experimental pulmonary hypertension. Physiol Genomics 2013; 45:449-61. [PMID: 23632417 DOI: 10.1152/physiolgenomics.00166.2012] [Citation(s) in RCA: 50] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/12/2023] Open
Abstract
Right ventricular failure (RVF) is the most frequent cause of death in patients with pulmonary arterial hypertension (PAH); however, specific therapies targeted to treat RVF have not been developed. Chronic treatment with carvedilol has been shown to reduce established maladaptive right ventricle (RV) hypertrophy and to improve RV function in experimental PAH. However, the mechanisms by which carvedilol improves RVF are unknown. We have previously demonstrated by microarray analysis that RVF is characterized by a distinct gene expression profile when compared with functional, compensatory hypertrophy. We next sought to identify the effects of carvedilol on gene expression on a genome-wide basis. PAH and RVF were induced in male Sprague-Dawley rats by the combination of VEGF-receptor blockade and chronic hypoxia. After RVF was established, rats were treated with carvedilol or vehicle for 4 wk. RNA was isolated from RV tissue and hybridized for microarray analysis. An initial prediction analysis of carvedilol-treated RVs showed that the gene expression profile resembled the RVF prediction set. However, a more extensive analysis revealed a small group of genes differentially expressed after carvedilol treatment. Further analysis categorized these genes in pathways involved in cardiac hypertrophy, mitochondrial dysfunction, and protein ubiquitination. Genes encoding proteins in the cardiac hypertrophy and protein ubiquitination pathways were downregulated in the RV by carvedilol, while genes encoding proteins in the mitochondrial dysfunction pathway were upregulated by carvedilol. These gene expression changes may explain some of the mechanisms that underlie the functional improvement of the RV after carvedilol treatment.
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Affiliation(s)
- Jennifer I Drake
- Victoria Johnson Center for Lung Obstructive Disease Research, Virginia Commonwealth University, Richmond, Virginia, USA
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3
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THEME: a web tool for loop-design microarray data analysis. Comput Biol Med 2011; 42:228-34. [PMID: 22188791 DOI: 10.1016/j.compbiomed.2011.11.012] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/23/2010] [Revised: 10/06/2011] [Accepted: 11/26/2011] [Indexed: 11/22/2022]
Abstract
A number of recent studies have shown that loop-design is more efficient than reference control design. Data analysis for loop-design microarray experiments is commonly undertaken using linear models and statistical tests. These techniques require specialized knowledge in statistical programming. However, limited loop-design web-based tools are available. We have developed the THEME (Tsing Hua Engine of Microarray Experiment) that exploits all necessary data analysis tools for loop-design microarray studies. THEME allows users to construct linear models and to apply multiple user-defined statistical tests of hypotheses for detection of DEG (differentially expressed genes). Users can modify entries of design matrix for experimental design as well as that of contrast matrix for statistical tests of hypotheses. The output of multiple user-defined statistical tests of hypotheses, DEG lists, can be cross-validated. The web platform provides data assessment and visualization tools that significantly assist users when evaluating the performance of microarray experimental procedures. THEME is also a MIAME (Minimal Information About a Microarray Experiment) compliant system, which enables users to export formatted files for GEO (Gene Expression Omnibus) submission. THEME offers comprehensive web services to biologists for data analysis of loop-design microarray experiments. This web-based resource is especially useful for core facility service as well as collaboration projects when researchers are not at the same site. Data analysis procedures, starting from uploading raw data files to retrieving DEG lists, can be flexibly operated with natural workflows. These features make THEME a reliable and powerful on-line system for data analysis of loop-design microarrays. The THEME server is available at http://metadb.bmes.nthu.edu.tw/theme/.
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miR-10a overexpression is associated with NPM1 mutations and MDM4 downregulation in intermediate-risk acute myeloid leukemia. Exp Hematol 2011; 39:1030-1042.e7. [PMID: 21784052 DOI: 10.1016/j.exphem.2011.07.008] [Citation(s) in RCA: 34] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/02/2011] [Revised: 06/05/2011] [Accepted: 07/01/2011] [Indexed: 12/19/2022]
Abstract
OBJECTIVE The study investigated differential microRNA (miRNA) expression patterns in acute myeloid leukemia (AML) patients with intermediate-risk (IR) characteristics. After characterization and validation of miR-10a, which was specifically upregulated in nucleophosmin 1 (NPM1) mutant AML samples, functional consequences of miR-10a overexpression were further delineated in vitro. MATERIALS AND METHODS Microarray analysis of miRNAs in bone marrow samples from AML (IR) patients with NPM1 mutations and healthy donors was performed to detect differential expression patterns. After validation of miRNA expression specific for NPM1 mutation in AML patients by quantitative reverse transcription polymerase chain reaction, a functional target gene search was conducted using complementary DNA microarray data from samples transfected with miR-10a. Potential target gene validation was done using transient transfection of K562 cells followed by Western blotting and luciferase reporter assay. RESULTS In comparison with wild-type samples, NPM1 mutant AML samples were shown to markedly overexpress miR-10a. Subsequent in vitro miR-10a overexpression induced differential gene expression as determined by microarray analysis. Here the murine double minute 4 (MDM4) gene turned out as a candidate gene for miR-10a. Validation of MDM4 in leukemic cells revealed a robust negative relationship between miR-10a overexpression and MDM4 downregulation. Furthermore, we determined an inverse association between miR-10a and MDM4 expression in AML (IR) samples with respect to their NPM1 mutational status. CONCLUSIONS miR-10a expression is highly characteristic for AML (IR) patients with NPM1 mutations and may influence its biological properties in AML by interfering with the p53 machinery partly regulated by MDM4.
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Lewohl JM, Nunez YO, Dodd PR, Tiwari GR, Harris RA, Mayfield RD. Up-regulation of microRNAs in brain of human alcoholics. Alcohol Clin Exp Res 2011; 35:1928-37. [PMID: 21651580 DOI: 10.1111/j.1530-0277.2011.01544.x] [Citation(s) in RCA: 159] [Impact Index Per Article: 11.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022]
Abstract
BACKGROUND MicroRNAs (miRNAs) are small, noncoding oligonucleotides with an important role in posttranscriptional regulation of gene expression at the level of translation and mRNA degradation. Recent studies have revealed that miRNAs play important roles in a variety of biological processes, such as cell proliferation, neuronal differentiation, developmental timing, synapse function, and neurogenesis. A single miRNA can target hundreds of mRNA transcripts for either translation repression or degradation, but the function of many human miRNAs is not known. METHODS miRNA array analysis was performed on the prefrontal cortex of 27 individual human cases (14 alcoholics and 13 matched controls). Target genes for differentially expressed miRNAs were predicted using multiple target prediction algorithms and a consensus approach, and predicted targets were matched against differentially expressed mRNAs from the same samples. Over- and under-representation analysis was performed using hypergeometric probability and z-score tests. RESULTS Approximately 35 miRNAs were significantly up-regulated in the alcoholic group compared with controls. Target prediction showed a large degree of overlap with our published cDNA microarray data. Functional classification of the predicted target genes of the regulated miRNAs includes apoptosis, cell cycle, cell adhesion, nervous system development, and cell-cell signaling. CONCLUSIONS These data suggest that the reduced expression of genes in human alcoholic cases may be because of the up-regulated miRNAs. Cellular processes fundamental to neuronal plasticity appear to represent major targets of the suggested miRNA regulation.
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Affiliation(s)
- Joanne M Lewohl
- School of Medical Science & Griffith Health Institute, Griffith University, Southport, Australia
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6
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Mulligan MK, Rhodes JS, Crabbe JC, Mayfield RD, Harris RA, Ponomarev I. Molecular profiles of drinking alcohol to intoxication in C57BL/6J mice. Alcohol Clin Exp Res 2011; 35:659-70. [PMID: 21223303 DOI: 10.1111/j.1530-0277.2010.01384.x] [Citation(s) in RCA: 85] [Impact Index Per Article: 6.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/08/2023]
Abstract
BACKGROUND Alcohol addiction develops through a series of stages, and mechanistic studies are needed to understand the transition from initial drug use to sustained controlled alcohol consumption followed by abuse and physical dependence. The focus of this study was to examine the effects of voluntary alcohol consumption on brain gene expression profiles using a mouse model of binge drinking. The main goal was to identify alcohol-responsive genes and functional categories after a single episode of drinking to intoxication. METHODS We used a modification of a "Drinking In the Dark" (DID) procedure (Rhodes et al., 2005) that allows mice to experience physiologically relevant amounts of alcohol in a non-stressful environment and also allows for detection of alcohol-sensitive molecular changes in a dose-dependent manner. C57BL/6J male mice were exposed to either 20% ethanol solution or water (single bottle) starting 3 hours after lights off for 4 hours and brains were harvested immediately after the drinking session. cDNA microarrays were used to assess the effects of voluntary drinking on global gene expression in 6 brain regions. We employed three statistical approaches to analyze microarray data. RESULTS A commonly used approach that applies a strict statistical threshold identified the eight top statistically significant genes whose expression was significantly correlated with blood ethanol concentration (BEC) in one of the brain regions. We then used a systems network approach to examine brain region-specific transcriptomes and identify modules of co-expressed (correlated) genes. In each brain region, we identified alcohol-responsive modules, i.e., modules significantly enriched for genes whose expression was correlated with BEC. A functional over-representation analysis was then applied to examine the organizing principles of alcohol-responsive modules. Genes were clustered into modules according to their roles in different physiological processes, functional groups, and cell types, including blood circulation, signal transduction, cell-cell communication, and striatal neurons. Finally, a meta-analysis across all brain regions suggested a global role of increasing alcohol dose in coordination of brain blood circulation and reaction of astrocytes. CONCLUSIONS This study showed that acute drinking resulted in small but consistent changes in brain gene expression which occurred in a dose-dependent manner. We identified both general and region-specific changes, some of which represent adaptive changes in response to increasing alcohol dose, which may play a role in alcohol-related behaviours, such as tolerance and consumption. Our systems approach allowed us to estimate the functional values of individual genes in the context of their genetic networks and formulate new refined hypotheses. An integrative analysis including other alcohol studies suggested several top candidates for functional validation, including Mt2, Gstm1, Scn4b, Prkcz, and Park7.
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Affiliation(s)
- Megan K Mulligan
- University of Texas at Austin, Waggoner Center for Alcohol and Addiction Research, Austin, Texas, USA
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Reimand J, Vaquerizas JM, Todd AE, Vilo J, Luscombe NM. Comprehensive reanalysis of transcription factor knockout expression data in Saccharomyces cerevisiae reveals many new targets. Nucleic Acids Res 2010; 38:4768-77. [PMID: 20385592 PMCID: PMC2919724 DOI: 10.1093/nar/gkq232] [Citation(s) in RCA: 78] [Impact Index Per Article: 5.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/22/2023] Open
Abstract
Transcription factor (TF) perturbation experiments give valuable insights into gene regulation. Genome-scale evidence from microarray measurements may be used to identify regulatory interactions between TFs and targets. Recently, Hu and colleagues published a comprehensive study covering 269 TF knockout mutants for the yeast Saccharomyces cerevisiae. However, the information that can be extracted from this valuable dataset is limited by the method employed to process the microarray data. Here, we present a reanalysis of the original data using improved statistical techniques freely available from the BioConductor project. We identify over 100,000 differentially expressed genes-nine times the total reported by Hu et al. We validate the biological significance of these genes by assessing their functions, the occurrence of upstream TF-binding sites, and the prevalence of protein-protein interactions. The reanalysed dataset outperforms the original across all measures, indicating that we have uncovered a vastly expanded list of relevant targets. In summary, this work presents a high-quality reanalysis that maximizes the information contained in the Hu et al. compendium. The dataset is available from ArrayExpress (accession: E-MTAB-109) and it will be invaluable to any scientist interested in the yeast transcriptional regulatory system.
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Affiliation(s)
- Jüri Reimand
- EMBL-European Bioinformatics Institute, Wellcome Trust Genome Campus, Cambridge, UK.
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8
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Tagmount A, Wang M, Lindquist E, Tanaka Y, Teranishi KS, Sunagawa S, Wong M, Stillman JH. The porcelain crab transcriptome and PCAD, the porcelain crab microarray and sequence database. PLoS One 2010; 5:e9327. [PMID: 20174471 PMCID: PMC2824831 DOI: 10.1371/journal.pone.0009327] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/15/2009] [Accepted: 01/27/2010] [Indexed: 01/11/2023] Open
Abstract
Background With the emergence of a completed genome sequence of the freshwater crustacean Daphnia pulex, construction of genomic-scale sequence databases for additional crustacean sequences are important for comparative genomics and annotation. Porcelain crabs, genus Petrolisthes, have been powerful crustacean models for environmental and evolutionary physiology with respect to thermal adaptation and understanding responses of marine organisms to climate change. Here, we present a large-scale EST sequencing and cDNA microarray database project for the porcelain crab Petrolisthes cinctipes. Methodology/Principal Findings A set of ∼30K unique sequences (UniSeqs) representing ∼19K clusters were generated from ∼98K high quality ESTs from a set of tissue specific non-normalized and mixed-tissue normalized cDNA libraries from the porcelain crab Petrolisthes cinctipes. Homology for each UniSeq was assessed using BLAST, InterProScan, GO and KEGG database searches. Approximately 66% of the UniSeqs had homology in at least one of the databases. All EST and UniSeq sequences along with annotation results and coordinated cDNA microarray datasets have been made publicly accessible at the Porcelain Crab Array Database (PCAD), a feature-enriched version of the Stanford and Longhorn Array Databases. Conclusions/Significance The EST project presented here represents the third largest sequencing effort for any crustacean, and the largest effort for any crab species. Our assembly and clustering results suggest that our porcelain crab EST data set is equally diverse to the much larger EST set generated in the Daphnia pulex genome sequencing project, and thus will be an important resource to the Daphnia research community. Our homology results support the pancrustacea hypothesis and suggest that Malacostraca may be ancestral to Branchiopoda and Hexapoda. Our results also suggest that our cDNA microarrays cover as much of the transcriptome as can reasonably be captured in EST library sequencing approaches, and thus represent a rich resource for studies of environmental genomics.
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Affiliation(s)
- Abderrahmane Tagmount
- Romberg Tiburon Center and Department of Biology, San Francisco State University, Tiburon, California, United States of America
| | - Mei Wang
- Department of Energy Joint Genome Institute, Walnut Creek, California, United States of America
| | - Erika Lindquist
- Department of Energy Joint Genome Institute, Walnut Creek, California, United States of America
| | - Yoshihiro Tanaka
- Romberg Tiburon Center and Department of Biology, San Francisco State University, Tiburon, California, United States of America
| | - Kristen S. Teranishi
- Romberg Tiburon Center and Department of Biology, San Francisco State University, Tiburon, California, United States of America
| | - Shinichi Sunagawa
- School of Natural Sciences, University of California Merced, Merced, California, United States of America
| | - Mike Wong
- Center for Computing in the Life Sciences, San Francisco State University, San Francisco, California, United States of America
| | - Jonathon H. Stillman
- Romberg Tiburon Center and Department of Biology, San Francisco State University, Tiburon, California, United States of America
- Department of Integrative Biology, University of California, Berkeley, California, United States of America
- * E-mail:
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9
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Zhang Z, Townsend JP. The filamentous fungal gene expression database (FFGED). Fungal Genet Biol 2009; 47:199-204. [PMID: 20025988 DOI: 10.1016/j.fgb.2009.12.001] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/18/2009] [Revised: 11/18/2009] [Accepted: 12/09/2009] [Indexed: 01/25/2023]
Abstract
Filamentous fungal gene expression assays provide essential information for understanding systemic cellular regulation. To aid research on fungal gene expression, we constructed a novel, comprehensive, free database, the filamentous fungal gene expression database (FFGED), available at http://bioinfo.townsend.yale.edu. FFGED features user-friendly management of gene expression data, which are assorted into experimental metadata, experimental design, raw data, normalized details, and analysis results. Data may be submitted in the process of an experiment, and any user can submit multiple experiments, thus classifying the FFGED as an "active experiment" database. Most importantly, FFGED functions as a collective and collaborative platform, by connecting each experiment with similar related experiments made public by other users, maximizing data sharing among different users, and correlating diverse gene expression levels under multiple experimental designs within different experiments. A clear and efficient web interface is provided with enhancement by AJAX (Asynchronous JavaScript and XML) and through a collection of tools to effectively facilitate data submission, sharing, retrieval and visualization.
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Affiliation(s)
- Zhang Zhang
- Department of Ecology and Evolutionary Biology, Yale University, 165 Prospect Street, New Haven, CT 06520, USA.
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Xia XQ, McClelland M, Porwollik S, Song W, Cong X, Wang Y. WebArrayDB: cross-platform microarray data analysis and public data repository. ACTA ACUST UNITED AC 2009; 25:2425-9. [PMID: 19602526 DOI: 10.1093/bioinformatics/btp430] [Citation(s) in RCA: 36] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/13/2022]
Abstract
MOTIVATION Cross-platform microarray analysis is an increasingly important research tool, but researchers still lack open source tools for storing, integrating and analyzing large amounts of microarray data obtained from different array platforms. RESULTS An open source integrated microarray database and analysis suite, WebArrayDB (http://www.webarraydb.org), has been developed that features convenient uploading of data for storage in a MIAME (Minimal Information about a Microarray Experiment) compliant fashion, and allows data to be mined with a large variety of R-based tools, including data analysis across multiple platforms. Different methods for probe alignment, normalization and statistical analysis are included to account for systematic bias. Student's t-test, moderated t-tests, non-parametric tests and analysis of variance or covariance (ANOVA/ANCOVA) are among the choices of algorithms for differential analysis of data. Users also have the flexibility to define new factors and create new analysis models to fit complex experimental designs. All data can be queried or browsed through a web browser. The computations can be performed in parallel on symmetric multiprocessing (SMP) systems or Linux clusters. AVAILABILITY The software package is available for the use on a public web server (http://www.webarraydb.org) or can be downloaded. are available at Bioinformatics online.
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Affiliation(s)
- Xiao-Qin Xia
- Vaccine Research Institute San Diego, San Diego, CA 92121, USA.
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Structure and in vivo requirement of the yeast Spt6 SH2 domain. J Mol Biol 2009; 389:211-25. [PMID: 19371747 DOI: 10.1016/j.jmb.2009.04.016] [Citation(s) in RCA: 43] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/04/2008] [Revised: 04/03/2009] [Accepted: 04/07/2009] [Indexed: 11/24/2022]
Abstract
During transcription elongation through chromatin, the Ser2-phosphorylated C-terminal repeat domain of RNA polymerase II binds the C-terminal Src homology 2 (SH2) domain of the nucleosome re-assembly factor Spt6. This SH2 domain is unusual in its specificity to bind phosphoserine, rather than phosphotyrosine and because it is the only SH2 domain in the yeast genome. Here, we report the high-resolution crystal structure of the SH2 domain from Candida glabrata Spt6. The structure combines features from both structural subfamilies of SH2 domains, suggesting it resembles a common ancestor of all SH2 domains. Two conserved surface pockets deviate from those of canonical SH2 domains, and may explain the unusual phosphoserine specificity. Differential gene expression analysis reveals that the SH2 domain is required for normal expression of a subset of yeast genes, and is consistent with multiple functions of Spt6 in chromatin transcription.
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Reconstructing generalized logical networks of transcriptional regulation in mouse brain from temporal gene expression data. EURASIP JOURNAL ON BIOINFORMATICS & SYSTEMS BIOLOGY 2009:545176. [PMID: 19300527 PMCID: PMC3171431 DOI: 10.1155/2009/545176] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 06/01/2008] [Revised: 09/08/2008] [Accepted: 12/12/2008] [Indexed: 02/02/2023]
Abstract
Gene expression time course data can be used not only to detect differentially expressed genes but also to find temporal associations among genes. The problem of reconstructing generalized logical networks to account for temporal dependencies among genes and environmental stimuli from transcriptomic data is addressed. A network reconstruction algorithm was developed that uses statistical significance as a criterion for network selection to avoid false-positive interactions arising from pure chance. The multinomial hypothesis testing-based network reconstruction allows for explicit specification of the false-positive rate, unique from all extant network inference algorithms. The method is superior to dynamic Bayesian network modeling in a simulation study. Temporal gene expression data from the brains of alcohol-treated mice in an analysis of the molecular response to alcohol are used for modeling. Genes from major neuronal pathways are identified as putative components of the alcohol response mechanism. Nine of these genes have associations with alcohol reported in literature. Several other potentially relevant genes, compatible with independent results from literature mining, may play a role in the response to alcohol. Additional, previously unknown gene interactions were discovered that, subject to biological verification, may offer new clues in the search for the elusive molecular mechanisms of alcoholism.
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Killion PJ, Iyer VR. ArrayPlex: distributed, interactive and programmatic access to genome sequence, annotation, ontology, and analytical toolsets. Genome Biol 2008; 9:R159. [PMID: 19014503 PMCID: PMC2614491 DOI: 10.1186/gb-2008-9-11-r159] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/22/2008] [Revised: 09/22/2008] [Accepted: 11/12/2008] [Indexed: 11/12/2022] Open
Abstract
ArrayPlex is a software package that centrally provides a large number of flexible toolsets useful for functional genomics. ArrayPlex is a software package that centrally provides a large number of flexible toolsets useful for functional genomics, including microarray data storage, quality assessments, data visualization, gene annotation retrieval, statistical tests, genomic sequence retrieval and motif analysis. It uses a client-server architecture based on open source components, provides graphical, command-line, and programmatic access to all needed resources, and is extensible by virtue of a documented application programming interface. ArrayPlex is available at .
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Affiliation(s)
- Patrick J Killion
- Center for Systems and Synthetic Biology, Institute for Cellular and Molecular Biology, University of Texas at Austin, 1 University Station A4800, Austin, Texas 78712, USA.
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Wang P, Yu W, Hu Z, Jia L, Iyer VR, Sanders BG, Kline K. Involvement of JNK/p73/NOXA in vitamin E analog-induced apoptosis of human breast cancer cells. Mol Carcinog 2008; 47:436-45. [PMID: 18058804 DOI: 10.1002/mc.20400] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/08/2022]
Abstract
Microarray analyses of human MDA-MB-435 breast cancer cells treated with vitamin E analog 2,5,7,8-tetramethyl-2R-(4R,8R,12-trimethyltridecyl) chroman-6-yloxy acetic acid (alpha-TEA) showed over 400 genes to be modulated. Thirty-four genes deemed of interest based on potential involvement in anticancer activities of alpha-TEA fell into six categories: apoptosis related, signal transduction, cell cycle related, cell adhesion and motility, transcriptional regulators, and membrane traffic related. The gene (PMAIP1) for NOXA was studied further. NOXA mRNA and protein levels were elevated in a time and dose-dependent fashion following alpha-TEA treatment. Functional knockdowns using small interfering RNA (siRNA) showed NOXA to contribute to alpha-TEA-induced apoptosis. A correlation between alpha-TEA's ability to upregulate NOXA and induce apoptosis was seen among several human breast cancer cell lines. Efforts to identify upstream regulators of NOXA in alpha-TEA-induced apoptosis identified the necessity of both c-Jun N-terminal kinase (JNK) activation and p73 expression. Additionally, protein levels of full length p73 were decreased by JNK siRNA treatment, suggesting that the signal transduction module of JNK-p73-NOXA is involved in alpha-TEA induced apoptosis of human breast cancer cells. Taken together, these findings suggest a role for JNK activation in mediating full length p73 expression and add to our understanding of the mechanisms of anticancer actions of alpha-TEA, a potential chemotherapeutic agent.
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Affiliation(s)
- Pei Wang
- Section of Molecular Genetics and Microbiology, University of Texas at Austin, Austin, Texas 78712, USA
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Hughes B, Joshi I, Wareham J. Health 2.0 and Medicine 2.0: tensions and controversies in the field. J Med Internet Res 2008; 10:e23. [PMID: 18682374 PMCID: PMC2553249 DOI: 10.2196/jmir.1056] [Citation(s) in RCA: 143] [Impact Index Per Article: 8.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/29/2008] [Revised: 05/13/2008] [Accepted: 06/03/2008] [Indexed: 11/22/2022] Open
Abstract
BACKGROUND The term Web 2.0 became popular following the O'Reilly Media Web 2.0 conference in 2004; however, there are difficulties in its application to health and medicine. Principally, the definition published by O'Reilly is criticized for being too amorphous, where other authors claim that Web 2.0 does not really exist. Despite this skepticism, the online community using Web 2.0 tools for health continues to grow, and the term Medicine 2.0 has entered popular nomenclature. OBJECTIVE This paper aims to establish a clear definition for Medicine 2.0 and delineate literature that is specific to the field. In addition, we propose a framework for categorizing the existing Medicine 2.0 literature and identify key research themes, underdeveloped research areas, as well as the underlying tensions or controversies in Medicine 2.0's diverse interest groups. METHODS In the first phase, we employ a thematic analysis of online definitions, that is, the most important linked papers, websites, or blogs in the Medicine 2.0 community itself. In a second phase, this definition is then applied across a series of academic papers to review Medicine 2.0's core literature base, delineating it from a wider concept of eHealth. RESULTS The terms Medicine 2.0 and Health 2.0 were found to be very similar and subsume five major salient themes: (1) the participants involved (doctors, patients, etc); (2) its impact on both traditional and collaborative practices in medicine; (3) its ability to provide personalized health care; (4) its ability to promote ongoing medical education; and (5) its associated method- and tool-related issues, such as potential inaccuracy in enduser-generated content. In comparing definitions of Medicine 2.0 to eHealth, key distinctions are made by the collaborative nature of Medicine 2.0 and its emphasis on personalized health care. However, other elements such as health or medical education remain common for both categories. In addition, this emphasis on personalized health care is not a salient theme within the academic literature. Of 2405 papers originally identified as potentially relevant, we found 56 articles that were exclusively focused on Medicine 2.0 as opposed to wider eHealth discussions. Four major tensions or debates between stakeholders were found in this literature, including (1) the lack of clear Medicine 2.0 definitions, (2) tension due to the loss of control over information as perceived by doctors, (3) the safety issues of inaccurate information, and (4) ownership and privacy issues with the growing body of information created by Medicine 2.0. CONCLUSION This paper is distinguished from previous reviews in that earlier studies mainly introduced specific Medicine 2.0 tools. In addressing the field's definition via empirical online data, it establishes a literature base and delineates key topics for future research into Medicine 2.0, distinct to that of eHealth.
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Affiliation(s)
- Benjamin Hughes
- Department of Information Systems, Ramon Llull University, ESADE, 60-62 Av. Pedralbes, 08034 Barcelona, Spain.
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Mulligan MK, Ponomarev I, Boehm SL, Owen JA, Levin PS, Berman AE, Blednov YA, Crabbe JC, Williams RW, Miles MF, Bergeson SE. Alcohol trait and transcriptional genomic analysis of C57BL/6 substrains. GENES BRAIN AND BEHAVIOR 2008; 7:677-89. [DOI: 10.1111/j.1601-183x.2008.00405.x] [Citation(s) in RCA: 63] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
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Yao J, Chang C, Salmi ML, Hung YS, Loraine A, Roux SJ. Genome-scale cluster analysis of replicated microarrays using shrinkage correlation coefficient. BMC Bioinformatics 2008; 9:288. [PMID: 18564431 PMCID: PMC2459189 DOI: 10.1186/1471-2105-9-288] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/16/2007] [Accepted: 06/18/2008] [Indexed: 11/10/2022] Open
Abstract
Background Currently, clustering with some form of correlation coefficient as the gene similarity metric has become a popular method for profiling genomic data. The Pearson correlation coefficient and the standard deviation (SD)-weighted correlation coefficient are the two most widely-used correlations as the similarity metrics in clustering microarray data. However, these two correlations are not optimal for analyzing replicated microarray data generated by most laboratories. An effective correlation coefficient is needed to provide statistically sufficient analysis of replicated microarray data. Results In this study, we describe a novel correlation coefficient, shrinkage correlation coefficient (SCC), that fully exploits the similarity between the replicated microarray experimental samples. The methodology considers both the number of replicates and the variance within each experimental group in clustering expression data, and provides a robust statistical estimation of the error of replicated microarray data. The value of SCC is revealed by its comparison with two other correlation coefficients that are currently the most widely-used (Pearson correlation coefficient and SD-weighted correlation coefficient) using statistical measures on both synthetic expression data as well as real gene expression data from Saccharomyces cerevisiae. Two leading clustering methods, hierarchical and k-means clustering were applied for the comparison. The comparison indicated that using SCC achieves better clustering performance. Applying SCC-based hierarchical clustering to the replicated microarray data obtained from germinating spores of the fern Ceratopteris richardii, we discovered two clusters of genes with shared expression patterns during spore germination. Functional analysis suggested that some of the genetic mechanisms that control germination in such diverse plant lineages as mosses and angiosperms are also conserved among ferns. Conclusion This study shows that SCC is an alternative to the Pearson correlation coefficient and the SD-weighted correlation coefficient, and is particularly useful for clustering replicated microarray data. This computational approach should be generally useful for proteomic data or other high-throughput analysis methodology.
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Affiliation(s)
- Jianchao Yao
- Institute for Cellular and Molecular Biology and Department of Mathematics, University of Texas at Austin, Austin, Texas 78712, USA.
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18
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Killion PJ, Iyer VR. Microarray data visualization and analysis with the Longhorn Array Database (LAD). CURRENT PROTOCOLS IN BIOINFORMATICS 2008; Chapter 7:Unit 7.10. [PMID: 18428730 DOI: 10.1002/0471250953.bi0710s08] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/10/2022]
Abstract
The wide availability and use of DNA microarrays has brought the power of whole-genome functional characterization to a large variety of research environments. Microarrays, however, also introduce significant infrastructural and analytical concerns with respect to the long-term warehousing, annotation, and visualization of immense data sets. The Longhorn Array Database (LAD) is a MIAME-compliant microarray database that operates on PostgreSQL and Linux. It is a free and completely open-source software solution, and provides a systematic and proven environment in which vast experiment sets can be safely archived, securely accessed, biologically annotated, quantitatively analyzed, and visually explored. This unit provides the complete set of information needed to successfully deploy, configure, and use LAD for the purposes of two-color DNA microarray analysis and visualization.
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Wang Q, Szaniszlo PJ. WdStuAp, an APSES transcription factor, is a regulator of yeast-hyphal transitions in Wangiella (Exophiala) dermatitidis. EUKARYOTIC CELL 2007; 6:1595-605. [PMID: 17693595 PMCID: PMC2043362 DOI: 10.1128/ec.00037-07] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/20/2022]
Abstract
APSES transcription factors are well-known regulators of fungal cellular development and differentiation. To study the function of an APSES protein in the fungus Wangiella dermatitidis, a conidiogenous and polymorphic agent of human phaeohyphomycosis with yeast predominance, the APSES transcription factor gene WdSTUA was cloned, sequenced, disrupted, and overexpressed. Analysis showed that its derived protein was most similar to the APSES proteins of other conidiogenous molds and had its APSES DNA-binding domain located in the amino-terminal half. Deletion of WdSTUA in W. dermatitidis induced convoluted instead of normal smooth colony surface growth on the rich yeast maintenance agar medium yeast extract-peptone-dextrose agar (YPDA) at 37 degrees C. Additionally, deletion of WdSTUA repressed aerial hyphal growth, conidiation, and invasive hyphal growth on the nitrogen-poor, hypha-inducing agar medium potato dextrose agar (PDA) at 25 degrees C. Ectopic overexpression of WdSTUA repressed the convoluted colony surface growth on YPDA at 37 degrees C, and also strongly repressed hyphal growth on PDA at 25 degrees C and 37 degrees C. These new results provide additional insights into the diverse roles played by APSES factors in fungi. They also suggest that the transcription factor encoded by WdSTUA is both a positive and negative morphotype regulator in W. dermatitidis and possibly other of the numerous human pathogenic, conidiogenous fungi capable of yeast growth.
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Affiliation(s)
- Qin Wang
- Section of Molecular Genetics and Microbiology, School of Biological Science and Institute of Cellular and Molecular Biology, The University of Texas at Austin, Austin, TX 78712, USA
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20
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Boulette ML, Payne SM. Anaerobic regulation of Shigella flexneri virulence: ArcA regulates Fur and iron acquisition genes. J Bacteriol 2007; 189:6957-67. [PMID: 17660284 PMCID: PMC2045222 DOI: 10.1128/jb.00621-07] [Citation(s) in RCA: 47] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Invasion and plaque formation in epithelial monolayers are routinely used to assess the virulence of Shigella flexneri, a causative agent of dysentery. A modified plaque assay was developed to identify factors contributing to the virulence of S. flexneri under the anaerobic conditions present in the colon. This assay demonstrated the importance of the ferrous iron transport system Feo, as well as the global transcription factors Fur, ArcA, and Fnr, for Shigella plaque formation in anoxic environments. Transcriptional analyses of S. flexneri iron transport genes indicated that anaerobic conditions activated feoABC while repressing genes encoding two other iron transport systems, the ABC transporter Sit and the Iuc/Iut aerobactin siderophore synthesis and transport system. The anaerobic transcription factors ArcA and Fnr activated expression of feoABC, while ArcA repressed iucABCD iutA. Transcription of fur, encoding the iron-responsive transcriptional repressor of bacterial iron acquisition, was also repressed anaerobically in an ArcA-dependent manner.
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21
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Liu J, Lewohl JM, Harris RA, Dodd PR, Mayfield RD. Altered gene expression profiles in the frontal cortex of cirrhotic alcoholics. Alcohol Clin Exp Res 2007; 31:1460-6. [PMID: 17625000 DOI: 10.1111/j.1530-0277.2007.00444.x] [Citation(s) in RCA: 48] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/26/2022]
Abstract
BACKGROUND Cirrhosis is the result of chronic liver disease that causes scarring and dysfunction of the liver. The disease is a common concomitant condition resulting from sustained exposure to alcohol. Heavy alcohol use results in brain damage that is generally more severe in cirrhotic compared with noncirrhotic alcoholics. We examined, at the cellular level, gene expression in the frontal cortex of cirrhotic alcoholics. METHODS Gene expression profiles were compared between cirrhotic and noncirrhotic alcoholics using approximately 47,000 element cDNA microarrays. RESULTS Widespread differences in transcriptome patterns were observed in cirrhotic compared with noncirrhotic alcoholics and these differences in gene expression accurately distinguished cirrhotic from noncirrhotic alcoholics. Functionally related groups of genes were identified that are involved in cell adhesion, mitochondrial function, synaptic transmission, apoptosis, and cell proliferation. Both astrocytes and neuronal cells were affected at the transcriptional level. The regulated genes are involved in neurite growth, neuronal cell adhesion, synaptic vesicle release, and postsynaptic neurotransmission. CONCLUSIONS These changes in the transcriptome likely contribute to the more severe brain dysfunction in cirrhotic alcoholics.
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Affiliation(s)
- Jianwen Liu
- Waggoner Center for Alcohol and Addiction Research, University of Texas at Austin, Austin, Texas 78712, USA
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22
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Quantitative gene expression assessment identifies appropriate cell line models for individual cervical cancer pathways. BMC Genomics 2007; 8:117. [PMID: 17493265 PMCID: PMC1878486 DOI: 10.1186/1471-2164-8-117] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/06/2006] [Accepted: 05/10/2007] [Indexed: 12/17/2022] Open
Abstract
BACKGROUND Cell lines have been used to study cancer for decades, but truly quantitative assessment of their performance as models is often lacking. We used gene expression profiling to quantitatively assess the gene expression of nine cell line models of cervical cancer. RESULTS We find a wide variation in the extent to which different cell culture models mimic late-stage invasive cervical cancer biopsies. The lowest agreement was from monolayer HeLa cells, a common cervical cancer model; the highest agreement was from primary epithelial cells, C4-I, and C4-II cell lines. In addition, HeLa and SiHa cell lines cultured in an organotypic environment increased their correlation to cervical cancer significantly. We also find wide variation in agreement when we considered how well individual biological pathways model cervical cancer. Cell lines with an anti-correlation to cervical cancer were also identified and should be avoided. CONCLUSION Using gene expression profiling and quantitative analysis, we have characterized nine cell lines with respect to how well they serve as models of cervical cancer. Applying this method to individual pathways, we identified the appropriateness of particular cell lines for studying specific pathways in cervical cancer. This study will allow researchers to choose a cell line with the highest correlation to cervical cancer at a pathway level. This method is applicable to other cancers and could be used to identify the appropriate cell line and growth condition to employ when studying other cancers.
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23
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Hu Z, Killion PJ, Iyer VR. Genetic reconstruction of a functional transcriptional regulatory network. Nat Genet 2007; 39:683-7. [PMID: 17417638 DOI: 10.1038/ng2012] [Citation(s) in RCA: 307] [Impact Index Per Article: 17.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/26/2006] [Accepted: 03/01/2007] [Indexed: 11/09/2022]
Abstract
Although global analyses of transcription factor binding provide one view of potential transcriptional regulatory networks, regulation also occurs at levels distinct from transcription factor binding. Here, we use a genetic approach to identify targets of transcription factors in yeast and reconstruct a functional regulatory network. First, we profiled transcriptional responses in S. cerevisiae strains with individual deletions of 263 transcription factors. Then we used directed-weighted graph modeling and regulatory epistasis analysis to identify indirect regulatory relationships between these transcription factors, and from this we reconstructed a functional transcriptional regulatory network. The enrichment of promoter motifs and Gene Ontology annotations provide insight into the biological functions of the transcription factors.
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Affiliation(s)
- Zhanzhi Hu
- Center for Systems and Synthetic Biology, Institute for Cellular and Molecular Biology, Section of Molecular Genetics and Microbiology, University of Texas at Austin, 1 University Station A4800, Austin, Texas 78712, USA
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Tsoi LC, Zheng WJ. A method of microarray data storage using array data type. Comput Biol Chem 2007; 31:143-7. [PMID: 17392028 PMCID: PMC2709412 DOI: 10.1016/j.compbiolchem.2007.01.004] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/03/2006] [Accepted: 01/04/2007] [Indexed: 11/17/2022]
Abstract
A well-designed microarray database can provide valuable information on gene expression levels. However, designing an efficient microarray database with minimum space usage is not an easy task since designers need to integrate the microarray data with the information of genes, probe annotation, and the descriptions of each microarray experiment. Developing better methods to store microarray data can greatly improve the efficiency and usefulness of such data. A new schema is proposed to store microarray data by using array data type in an object-relational database management system--PostgreSQL. The implemented database can store all the microarray data from the same chip in an array data structure. The variable-length array data type in PostgreSQL can store microarray data from same chip. The implementation of our schema can help to increase the data retrieval and space efficiency.
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Affiliation(s)
- Lam C. Tsoi
- Bioinformatics Graduate Program, Hollings Cancer Center, Medical University of South Carolina, 135 Cannon Street, Suite 303, Charleston, SC 29425
| | - W. Jim Zheng
- Department of Biostatistics, Bioinformatics and Epidemiology, and Bioinformatics Core Facility, Hollings Cancer Center, Medical University of South Carolina, 135 Cannon Street, Suite 303, Charleston, SC 29425
- Corresponding author. Tel: +1 843 876-1123, Fax: +1 843 876-1126, Email addresses: Lam C. Tsoi – , W. Jim Zheng* –
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Abstract
Microarrays and related technologies have allowed investigators to ask biological questions in far greater detail than has previously been possible. Microarrays had a troubled beginning, but most of these problems resulted from the growing pains of this technology, which, like many new things, was initially more promise than delivery. Nevertheless, over the past few years, investigators have learned how to achieve optimal performance of technology, and now exciting discoveries are made using microarray-based research. Many of the advances have come from the realization that microarrays are not a magic tool but rather are like any other measurement device. Unless microarray experimentation is coupled with good experimental practices, it will not yield valid results or, worse yet, may lead to misleading results. In this chapter, we highlight some of the important steps that should be taken to successfully conduct a microarray study. These steps include a clearly stated biological question, experimental design, careful experimental conduct, complete statistical analysis, validation/verification of results, and dissemination of the data.
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Affiliation(s)
- Grier P Page
- Department of Biostatistics, University of Alabama at Birmingham, Hoover, AL, USA
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Hayes A, Castrillo JI, Oliver SG, Brass A, Zeef LAH. 9 Transcript Analysis: A Microarray Approach. METHODS IN MICROBIOLOGY 2007. [DOI: 10.1016/s0580-9517(06)36009-6] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 10/23/2022]
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Demeter J, Beauheim C, Gollub J, Hernandez-Boussard T, Jin H, Maier D, Matese JC, Nitzberg M, Wymore F, Zachariah ZK, Brown PO, Sherlock G, Ball CA. The Stanford Microarray Database: implementation of new analysis tools and open source release of software. Nucleic Acids Res 2006; 35:D766-70. [PMID: 17182626 PMCID: PMC1781111 DOI: 10.1093/nar/gkl1019] [Citation(s) in RCA: 130] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/15/2022] Open
Abstract
The Stanford Microarray Database (SMD; ) is a research tool and archive that allows hundreds of researchers worldwide to store, annotate, analyze and share data generated by microarray technology. SMD supports most major microarray platforms, and is MIAME-supportive and can export or import MAGE-ML. The primary mission of SMD is to be a research tool that supports researchers from the point of data generation to data publication and dissemination, but it also provides unrestricted access to analysis tools and public data from 300 publications. In addition to supporting ongoing research, SMD makes its source code fully and freely available to others under an Open Source license, enabling other groups to create a local installation of SMD. In this article, we describe several data analysis tools implemented in SMD and we discuss features of our software release.
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Affiliation(s)
- Janos Demeter
- Department of Biochemistry, Stanford University School of MedicineStanford, CA, USA
| | - Catherine Beauheim
- Department of Genetics, Stanford University School of MedicineStanford, CA, USA
| | | | | | - Heng Jin
- Department of Biochemistry, Stanford University School of MedicineStanford, CA, USA
| | - Donald Maier
- Department of Biochemistry, Stanford University School of MedicineStanford, CA, USA
| | - John C. Matese
- Lewis-Sigler Institute for Integrative Genomics, Princeton UniversityPrinceton, NJ, USA
| | - Michael Nitzberg
- Department of Biochemistry, Stanford University School of MedicineStanford, CA, USA
| | - Farrell Wymore
- Department of Biochemistry, Stanford University School of MedicineStanford, CA, USA
| | | | - Patrick O. Brown
- Department of Biochemistry, Stanford University School of MedicineStanford, CA, USA
- Howard Hughes Medical InstituteStanford, CA, USA
| | - Gavin Sherlock
- Department of Genetics, Stanford University School of MedicineStanford, CA, USA
| | - Catherine A. Ball
- Department of Biochemistry, Stanford University School of MedicineStanford, CA, USA
- To whom correspondence should be addressed. Tel: +1 650 724 3028; Fax: +1 650 724 3701;
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Liu H, Lin J, Roy K. Effect of 3D scaffold and dynamic culture condition on the global gene expression profile of mouse embryonic stem cells. Biomaterials 2006; 27:5978-89. [PMID: 16824594 DOI: 10.1016/j.biomaterials.2006.05.053] [Citation(s) in RCA: 100] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/27/2006] [Accepted: 05/19/2006] [Indexed: 10/24/2022]
Abstract
We have previously demonstrated that mouse embryonic stem (ES) cells differentiated on three-dimensional (3D), highly porous, tantalum-based scaffolds (Cytomatrixtrade mark) have significantly higher hematopoietic differentiation efficiency than those cultured under conventional two-dimensional (2D) tissue culture conditions. In addition, ES cell-seeded scaffolds cultured inside spinner bioreactors showed further enhancement in hematopoiesis compared to static conditions. In the present study, we evaluated how these various biomaterial-based culture conditions, e.g. 2D vs. 3D scaffolds and static vs. dynamic, influence the global gene expression profile of differentiated ES cells. We report that compared to 2D tissue culture plates, cells differentiated on porous, Cytomatrixtrade mark scaffolds possess significantly higher expression levels of extracellular matrix (ECM)-related genes, as well as genes that regulate cell growth, proliferation and differentiation. In addition, these differences in gene expression were more pronounced in 3D dynamic culture compared to 3D static culture. We report specific genes that are either uniquely expressed under each condition or are quantitatively regulated, i.e. over expressed or inhibited by a specific culture environment. We conclude that that biomaterial-based 3D cultures, especially under dynamic conditions, might favor efficient hematopoietic differentiation of ES cells by stimulating increased expression of specific ECM proteins, growth factors and cell adhesion related genes while significantly down-regulating genes that act to inhibit expression of these molecules.
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Affiliation(s)
- Hui Liu
- Department of Biomedical Engineering, The University of Texas at Austin, Austin, TX 78712, USA
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Marzolf B, Troisch P. SLIMarray: lightweight software for microarray facility management. SOURCE CODE FOR BIOLOGY AND MEDICINE 2006; 1:5. [PMID: 17147785 PMCID: PMC1636632 DOI: 10.1186/1751-0473-1-5] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 09/11/2006] [Accepted: 10/26/2006] [Indexed: 12/03/2022]
Abstract
Background Microarray core facilities are commonplace in biological research organizations, and need systems for accurately tracking various logistical aspects of their operation. Although these different needs could be handled separately, an integrated management system provides benefits in organization, automation and reduction in errors. Results We present SLIMarray (System for Lab Information Management of Microarrays), an open source, modular database web application capable of managing microarray inventories, sample processing and usage charges. The software allows modular configuration and is well suited for further development, providing users the flexibility to adapt it to their needs. SLIMarray Lite, a version of the software that is especially easy to install and run, is also available. Conclusion SLIMarray addresses the previously unmet need for free and open source software for managing the logistics of a microarray core facility.
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Affiliation(s)
- Bruz Marzolf
- Institute for Systems Biology, 1441 N. 34Street, Seattle, Washington, USA
| | - Pamela Troisch
- Institute for Systems Biology, 1441 N. 34Street, Seattle, Washington, USA
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Honoré P, Granjeaud S, Tagett R, Deraco S, Beaudoing E, Rougemont J, Debono S, Hingamp P. MicroArray Facility: a laboratory information management system with extended support for Nylon based technologies. BMC Genomics 2006; 7:240. [PMID: 16987406 PMCID: PMC1592093 DOI: 10.1186/1471-2164-7-240] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/29/2006] [Accepted: 09/20/2006] [Indexed: 01/08/2023] Open
Abstract
Background High throughput gene expression profiling (GEP) is becoming a routine technique in life science laboratories. With experimental designs that repeatedly span thousands of genes and hundreds of samples, relying on a dedicated database infrastructure is no longer an option. GEP technology is a fast moving target, with new approaches constantly broadening the field diversity. This technology heterogeneity, compounded by the informatics complexity of GEP databases, means that software developments have so far focused on mainstream techniques, leaving less typical yet established techniques such as Nylon microarrays at best partially supported. Results MAF (MicroArray Facility) is the laboratory database system we have developed for managing the design, production and hybridization of spotted microarrays. Although it can support the widely used glass microarrays and oligo-chips, MAF was designed with the specific idiosyncrasies of Nylon based microarrays in mind. Notably single channel radioactive probes, microarray stripping and reuse, vector control hybridizations and spike-in controls are all natively supported by the software suite. MicroArray Facility is MIAME supportive and dynamically provides feedback on missing annotations to help users estimate effective MIAME compliance. Genomic data such as clone identifiers and gene symbols are also directly annotated by MAF software using standard public resources. The MAGE-ML data format is implemented for full data export. Journalized database operations (audit tracking), data anonymization, material traceability and user/project level confidentiality policies are also managed by MAF. Conclusion MicroArray Facility is a complete data management system for microarray producers and end-users. Particular care has been devoted to adequately model Nylon based microarrays. The MAF system, developed and implemented in both private and academic environments, has proved a robust solution for shared facilities and industry service providers alike.
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Affiliation(s)
- Paul Honoré
- IPSOGEN SAS, Luminy Biotech Entreprises, 163 avenue de Luminy, Case 923, 13009 Marseille, France
| | - Samuel Granjeaud
- TAGC, INSERM ERM206, Parc Scientifique de Luminy, Case 928, 13288 Marseille Cedex 09, France
| | - Rebecca Tagett
- IPSOGEN SAS, Luminy Biotech Entreprises, 163 avenue de Luminy, Case 923, 13009 Marseille, France
| | - Stéphane Deraco
- IPSOGEN SAS, Luminy Biotech Entreprises, 163 avenue de Luminy, Case 923, 13009 Marseille, France
- Now at CNRS – DSI, Tour Gaïa, rue Pierre-Gilles de Gennes, BP 21902, 31319 LABEGE CEDEX, France
| | - Emmanuel Beaudoing
- TAGC, INSERM ERM206, Parc Scientifique de Luminy, Case 928, 13288 Marseille Cedex 09, France
- Now at Swiss Institute of Bioinformatics, CH-1015 Lausanne, Switzerland
| | - Jacques Rougemont
- TAGC, INSERM ERM206, Parc Scientifique de Luminy, Case 928, 13288 Marseille Cedex 09, France
- Now at Swiss Institute of Bioinformatics, CH-1015 Lausanne, Switzerland
| | - Stéphane Debono
- IPSOGEN SAS, Luminy Biotech Entreprises, 163 avenue de Luminy, Case 923, 13009 Marseille, France
| | - Pascal Hingamp
- TAGC, INSERM ERM206, Parc Scientifique de Luminy, Case 928, 13288 Marseille Cedex 09, France
- Now at IGS, CNRS UPR 2589, 163 Avenue de Luminy Case 934, 13288 Marseille Cedex 09, France
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Liu J, Lewohl JM, Harris RA, Iyer VR, Dodd PR, Randall PK, Mayfield RD. Patterns of gene expression in the frontal cortex discriminate alcoholic from nonalcoholic individuals. Neuropsychopharmacology 2006; 31:1574-82. [PMID: 16292326 DOI: 10.1038/sj.npp.1300947] [Citation(s) in RCA: 193] [Impact Index Per Article: 10.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
Alcohol dependence is characterized by tolerance, physical dependence, and craving. The neuroadaptations underlying these effects of chronic alcohol abuse are likely due to altered gene expression. Previous gene expression studies using human post-mortem brain demonstrated that several gene families were altered by alcohol abuse. However, most of these changes in gene expression were small. It is not clear if gene expression profiles have sufficient power to discriminate control from alcoholic individuals and how consistent gene expression changes are when a relatively large sample size is examined. In the present study, microarray analysis (approximately 47,000 elements) was performed on the superior frontal cortex of 27 individual human cases (14 well characterized alcoholics and 13 matched controls). A partial least squares statistical procedure was applied to identify genes with altered expression levels in alcoholics. We found that genes involved in myelination, ubiquitination, apoptosis, cell adhesion, neurogenesis, and neural disease showed altered expression levels. Importantly, genes involved in neurodegenerative diseases such as Alzheimer's disease were significantly altered suggesting a link between alcoholism and other neurodegenerative conditions. A total of 27 genes identified in this study were previously shown to be changed by alcohol abuse in previous studies of human post-mortem brain. These results revealed a consistent re-programming of gene expression in alcohol abusers that reliably discriminates alcoholic from non-alcoholic individuals.
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Affiliation(s)
- Jianwen Liu
- Waggoner Center for Alcohol and Addiction Research, University of Texas at Austin, Austin, TX 78712, USA
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Lu P, Rangan A, Chan SY, Appling DR, Hoffman DW, Marcotte EM. Global metabolic changes following loss of a feedback loop reveal dynamic steady states of the yeast metabolome. Metab Eng 2006; 9:8-20. [PMID: 17049899 DOI: 10.1016/j.ymben.2006.06.003] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/03/2005] [Revised: 05/27/2006] [Accepted: 06/20/2006] [Indexed: 11/16/2022]
Abstract
Metabolic enzymes control cellular metabolite concentrations dynamically in response to changing environmental and intracellular conditions. Such real-time feedback regulation suggests the global metabolome may sample distinct dynamic steady states, forming "basins of stability" in the energy landscape of possible metabolite concentrations and enzymatic activities. Using metabolite, protein and transcriptional profiling, we characterize three dynamic steady states of the yeast metabolome that form by perturbing synthesis of the universal methyl donor S-adenosylmethionine (AdoMet). Conversion between these states is driven by replacement of serine with glycine+formate in the media, loss of feedback inhibition control by the metabolic enzyme Met13, or both. The latter causes hyperaccumulation of methionine and AdoMet, and dramatic global compensatory changes in the metabolome, including differences in amino acid and sugar metabolism, and possibly in the global nitrogen balance, ultimately leading to a G1/S phase cell cycle delay. Global metabolic changes are not necessarily accompanied by global transcriptional changes, and metabolite-controlled post-transcriptional regulation of metabolic enzymes is clearly evident.
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Affiliation(s)
- Peng Lu
- Center for Systems and Synthetic Biology, University of Texas, 1 University Station, Austin, TX 78712-0159, USA
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Ponomarev I, Maiya R, Harnett MT, Schafer GL, Ryabinin AE, Blednov YA, Morikawa H, Boehm SL, Homanics GE, Berman AE, Berman A, Lodowski KH, Bergeson SE, Harris RA. Transcriptional signatures of cellular plasticity in mice lacking the alpha1 subunit of GABAA receptors. J Neurosci 2006; 26:5673-83. [PMID: 16723524 PMCID: PMC1894896 DOI: 10.1523/jneurosci.0860-06.2006] [Citation(s) in RCA: 51] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/26/2023] Open
Abstract
GABAA receptors mediate the majority of inhibitory neurotransmission in the CNS. Genetic deletion of the alpha1 subunit of GABAA receptors results in a loss of alpha1-mediated fast inhibitory currents and a marked reduction in density of GABAA receptors. A grossly normal phenotype of alpha1-deficient mice suggests the presence of neuronal adaptation to these drastic changes at the GABA synapse. We used cDNA microarrays to identify transcriptional fingerprints of cellular plasticity in response to altered GABAergic inhibition in the cerebral cortex and cerebellum of alpha1 mutants. In silico analysis of 982 mutation-regulated transcripts highlighted genes and functional groups involved in regulation of neuronal excitability and synaptic transmission, suggesting an adaptive response of the brain to an altered inhibitory tone. Public gene expression databases permitted identification of subsets of transcripts enriched in excitatory and inhibitory neurons as well as some glial cells, providing evidence for cellular plasticity in individual cell types. Additional analysis linked some transcriptional changes to cellular phenotypes observed in the knock-out mice and suggested several genes, such as the early growth response 1 (Egr1), small GTP binding protein Rac1 (Rac1), neurogranin (Nrgn), sodium channel beta4 subunit (Scn4b), and potassium voltage-gated Kv4.2 channel (Kcnd2) as cell type-specific markers of neuronal plasticity. Furthermore, transcriptional activation of genes enriched in Bergman glia suggests an active role of these astrocytes in synaptic plasticity. Overall, our results suggest that the loss of alpha1-mediated fast inhibition produces diverse transcriptional responses that act to regulate neuronal excitability of individual neurons and stabilize neuronal networks, which may account for the lack of severe abnormalities in alpha1 null mutants.
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Affiliation(s)
- Igor Ponomarev
- Waggoner Center for Alcohol and Addiction Research, University of Texas, Austin, Texas 78712, USA.
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Marzolf B, Deutsch EW, Moss P, Campbell D, Johnson MH, Galitski T. SBEAMS-Microarray: database software supporting genomic expression analyses for systems biology. BMC Bioinformatics 2006; 7:286. [PMID: 16756676 PMCID: PMC1524999 DOI: 10.1186/1471-2105-7-286] [Citation(s) in RCA: 45] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/01/2006] [Accepted: 06/06/2006] [Indexed: 11/10/2022] Open
Abstract
Background The biological information in genomic expression data can be understood, and computationally extracted, in the context of systems of interacting molecules. The automation of this information extraction requires high throughput management and analysis of genomic expression data, and integration of these data with other data types. Results SBEAMS-Microarray, a module of the open-source Systems Biology Experiment Analysis Management System (SBEAMS), enables MIAME-compliant storage, management, analysis, and integration of high-throughput genomic expression data. It is interoperable with the Cytoscape network integration, visualization, analysis, and modeling software platform. Conclusion SBEAMS-Microarray provides end-to-end support for genomic expression analyses for network-based systems biology research.
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Affiliation(s)
- Bruz Marzolf
- Institute for Systems Biology, 1441 N. 34Street, Seattle, Washington, USA
| | - Eric W Deutsch
- Institute for Systems Biology, 1441 N. 34Street, Seattle, Washington, USA
| | - Patrick Moss
- Institute for Systems Biology, 1441 N. 34Street, Seattle, Washington, USA
| | - David Campbell
- Institute for Systems Biology, 1441 N. 34Street, Seattle, Washington, USA
| | - Michael H Johnson
- Institute for Systems Biology, 1441 N. 34Street, Seattle, Washington, USA
| | - Timothy Galitski
- Institute for Systems Biology, 1441 N. 34Street, Seattle, Washington, USA
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Gu J, Iyer VR. PI3K signaling and miRNA expression during the response of quiescent human fibroblasts to distinct proliferative stimuli. Genome Biol 2006; 7:R42. [PMID: 16737555 PMCID: PMC1779520 DOI: 10.1186/gb-2006-7-5-r42] [Citation(s) in RCA: 48] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/28/2005] [Revised: 03/10/2006] [Accepted: 04/20/2006] [Indexed: 01/13/2023] Open
Abstract
Global transcriptional profiling of human fibroblasts from two different tissue sources reveals distinct as well as conserved responses to different growth stimuli. Background Serum treatment of quiescent human dermal fibroblasts induces proliferation, coupled with a complex physiological response that is indicative of their normal role in wound-healing. However, it is not known to what extent such complex transcriptional events are specific to a given cell type and signal, and how these global changes are coordinately regulated. We have profiled the global transcriptional program of human fibroblasts from two different tissue sources to distinct growth stimuli, and identified a striking conservation in their gene-expression signatures. Results We found that the wound-healing program of gene expression was not specific to the response of dermal fibroblasts to serum but was regulated more broadly. However, there were specific differences among different stimuli with regard to signaling pathways that mediate these transcriptional programs. Our data suggest that the PI3-kinase pathway is differentially involved in mediating the responses of cells to serum as compared with individual peptide growth factors. Expression profiling indicated that let7 and other miRNAs with similar expression profiles may be involved in regulating the transcriptional program in response to proliferative signals. Conclusion This study provides insights into how different stimuli use distinct as well as conserved signaling and regulatory mechanisms to mediate genome-wide transcriptional reprogramming during cell proliferation. Our results indicate that conservation of transcriptional programs and their regulation among different cell types may be much broader than previously appreciated.
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Affiliation(s)
- Jian Gu
- Section of Molecular Genetics and Microbiology, Institute for Cellular and Molecular Biology, Center for Systems and Synthetic Biology, University of Texas at Austin, 1 University Station A4800, Austin, TX 78712-0159, USA
| | - Vishwanath R Iyer
- Section of Molecular Genetics and Microbiology, Institute for Cellular and Molecular Biology, Center for Systems and Synthetic Biology, University of Texas at Austin, 1 University Station A4800, Austin, TX 78712-0159, USA
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Hermida L, Schaad O, Demougin P, Descombes P, Primig M. MIMAS: an innovative tool for network-based high density oligonucleotide microarray data management and annotation. BMC Bioinformatics 2006; 7:190. [PMID: 16597336 PMCID: PMC1459208 DOI: 10.1186/1471-2105-7-190] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/16/2005] [Accepted: 04/05/2006] [Indexed: 01/26/2023] Open
Abstract
Background The high-density oligonucleotide microarray (GeneChip) is an important tool for molecular biological research aiming at large-scale detection of small nucleotide polymorphisms in DNA and genome-wide analysis of mRNA concentrations. Local array data management solutions are instrumental for efficient processing of the results and for subsequent uploading of data and annotations to a global certified data repository at the EBI (ArrayExpress) or the NCBI (GeneOmnibus). Description To facilitate and accelerate annotation of high-throughput expression profiling experiments, the Microarray Information Management and Annotation System (MIMAS) was developed. The system is fully compliant with the Minimal Information About a Microarray Experiment (MIAME) convention. MIMAS provides life scientists with a highly flexible and focused GeneChip data storage and annotation platform essential for subsequent analysis and interpretation of experimental results with clustering and mining tools. The system software can be downloaded for academic use upon request. Conclusion MIMAS implements a novel concept for nation-wide GeneChip data management whereby a network of facilities is centered on one data node directly connected to the European certified public microarray data repository located at the EBI. The solution proposed may serve as a prototype approach to array data management between research institutes organized in a consortium.
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Affiliation(s)
- Leandro Hermida
- Biozentrum and Swiss Institute of Bioinformatics Klingelbergstrasse 50-70 CH-4056 Basel Switzerland
| | - Olivier Schaad
- Genomics Platform, NCCR Frontiers in Genetics, Geneva University Medical Center 1, Rue Michel-Servet CH-1211 Geneva Switzerland
| | - Philippe Demougin
- Biozentrum and Swiss Institute of Bioinformatics Klingelbergstrasse 50-70 CH-4056 Basel Switzerland
| | - Patrick Descombes
- Genomics Platform, NCCR Frontiers in Genetics, Geneva University Medical Center 1, Rue Michel-Servet CH-1211 Geneva Switzerland
| | - Michael Primig
- Biozentrum and Swiss Institute of Bioinformatics Klingelbergstrasse 50-70 CH-4056 Basel Switzerland
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Mey AR, Wyckoff EE, Kanukurthy V, Fisher CR, Payne SM. Iron and fur regulation in Vibrio cholerae and the role of fur in virulence. Infect Immun 2006; 73:8167-78. [PMID: 16299312 PMCID: PMC1307094 DOI: 10.1128/iai.73.12.8167-8178.2005] [Citation(s) in RCA: 149] [Impact Index Per Article: 7.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Regulation of iron uptake and utilization is critical for bacterial growth and for prevention of iron toxicity. In many bacterial species, this regulation depends on the iron-responsive master regulator Fur. In this study we report the effects of iron and Fur on gene expression in Vibrio cholerae. We show that Fur has both positive and negative regulatory functions, and we demonstrate Fur-independent regulation of gene expression by iron. Nearly all of the known iron acquisition genes were repressed by Fur under iron-replete conditions. In addition, genes for two newly identified iron transport systems, Feo and Fbp, were found to be negatively regulated by iron and Fur. Other genes identified in this study as being induced in low iron and in the fur mutant include those encoding superoxide dismutase (sodA), fumarate dehydratase (fumC), bacterioferritin (bfr), bacterioferritin-associated ferredoxin (bfd), and multiple genes of unknown function. Several genes encoding iron-containing proteins were repressed in low iron and in the fur mutant, possibly reflecting the need to reserve available iron for the most critical functions. Also repressed in the fur mutant, but independently of iron, were genes located in the V. cholerae pathogenicity island, encoding the toxin-coregulated pilus (TCP), and genes within the V. cholerae mega-integron. The fur mutant exhibited very weak autoagglutination, indicating a possible defect in expression or assembly of the TCP, a major virulence factor of V. cholerae. Consistent with this observation, the fur mutant competed poorly with its wild-type parental strain for colonization of the infant mouse gut.
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Affiliation(s)
- Alexandra R Mey
- The University of Texas, Section of Molecular Genetics and Microbiology, Austin, TX 78712-1095, USA
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Burgarella S, Cattaneo D, Pinciroli F, Masseroli M. MicroGen: a MIAME compliant web system for microarray experiment information and workflow management. BMC Bioinformatics 2005; 6 Suppl 4:S6. [PMID: 16351755 PMCID: PMC1866379 DOI: 10.1186/1471-2105-6-s4-s6] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/24/2022] Open
Abstract
Background Improvements of bio-nano-technologies and biomolecular techniques have led to increasing production of high-throughput experimental data. Spotted cDNA microarray is one of the most diffuse technologies, used in single research laboratories and in biotechnology service facilities. Although they are routinely performed, spotted microarray experiments are complex procedures entailing several experimental steps and actors with different technical skills and roles. During an experiment, involved actors, who can also be located in a distance, need to access and share specific experiment information according to their roles. Furthermore, complete information describing all experimental steps must be orderly collected to allow subsequent correct interpretation of experimental results. Results We developed MicroGen, a web system for managing information and workflow in the production pipeline of spotted microarray experiments. It is constituted of a core multi-database system able to store all data completely characterizing different spotted microarray experiments according to the Minimum Information About Microarray Experiments (MIAME) standard, and of an intuitive and user-friendly web interface able to support the collaborative work required among multidisciplinary actors and roles involved in spotted microarray experiment production. MicroGen supports six types of user roles: the researcher who designs and requests the experiment, the spotting operator, the hybridisation operator, the image processing operator, the system administrator, and the generic public user who can access the unrestricted part of the system to get information about MicroGen services. Conclusion MicroGen represents a MIAME compliant information system that enables managing workflow and supporting collaborative work in spotted microarray experiment production.
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Affiliation(s)
- Sarah Burgarella
- BioMedical Informatics Laboratory, Bioengineering Department, Politecnico di Milano, piazza Leonardo da Vinci 32, 20133 Milan, Italy
| | - Dario Cattaneo
- BioMedical Informatics Laboratory, Bioengineering Department, Politecnico di Milano, piazza Leonardo da Vinci 32, 20133 Milan, Italy
| | - Francesco Pinciroli
- BioMedical Informatics Laboratory, Bioengineering Department, Politecnico di Milano, piazza Leonardo da Vinci 32, 20133 Milan, Italy
| | - Marco Masseroli
- BioMedical Informatics Laboratory, Bioengineering Department, Politecnico di Milano, piazza Leonardo da Vinci 32, 20133 Milan, Italy
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Hancock D, Wilson M, Velarde G, Morrison N, Hayes A, Hulme H, Wood AJ, Nashar K, Kell DB, Brass A. maxdLoad2 and maxdBrowse: standards-compliant tools for microarray experimental annotation, data management and dissemination. BMC Bioinformatics 2005; 6:264. [PMID: 16269077 PMCID: PMC1298287 DOI: 10.1186/1471-2105-6-264] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/29/2005] [Accepted: 11/03/2005] [Indexed: 11/24/2022] Open
Abstract
Background maxdLoad2 is a relational database schema and Java® application for microarray experimental annotation and storage. It is compliant with all standards for microarray meta-data capture; including the specification of what data should be recorded, extensive use of standard ontologies and support for data exchange formats. The output from maxdLoad2 is of a form acceptable for submission to the ArrayExpress microarray repository at the European Bioinformatics Institute. maxdBrowse is a PHP web-application that makes contents of maxdLoad2 databases accessible via web-browser, the command-line and web-service environments. It thus acts as both a dissemination and data-mining tool. Results maxdLoad2 presents an easy-to-use interface to an underlying relational database and provides a full complement of facilities for browsing, searching and editing. There is a tree-based visualization of data connectivity and the ability to explore the links between any pair of data elements, irrespective of how many intermediate links lie between them. Its principle novel features are: • the flexibility of the meta-data that can be captured, • the tools provided for importing data from spreadsheets and other tabular representations, • the tools provided for the automatic creation of structured documents, • the ability to browse and access the data via web and web-services interfaces. Within maxdLoad2 it is very straightforward to customise the meta-data that is being captured or change the definitions of the meta-data. These meta-data definitions are stored within the database itself allowing client software to connect properly to a modified database without having to be specially configured. The meta-data definitions (configuration file) can also be centralized allowing changes made in response to revisions of standards or terminologies to be propagated to clients without user intervention. maxdBrowse is hosted on a web-server and presents multiple interfaces to the contents of maxd databases. maxdBrowse emulates many of the browse and search features available in the maxdLoad2 application via a web-browser. This allows users who are not familiar with maxdLoad2 to browse and export microarray data from the database for their own analysis. The same browse and search features are also available via command-line and SOAP server interfaces. This both enables scripting of data export for use embedded in data repositories and analysis environments, and allows access to the maxd databases via web-service architectures. Conclusion maxdLoad2 and maxdBrowse are portable and compatible with all common operating systems and major database servers. They provide a powerful, flexible package for annotation of microarray experiments and a convenient dissemination environment. They are available for download and open sourced under the Artistic License.
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Affiliation(s)
- David Hancock
- School of Computer Science, The University of Manchester, Kilburn Building, Oxford Road, Manchester, UK
| | - Michael Wilson
- School of Computer Science, The University of Manchester, Kilburn Building, Oxford Road, Manchester, UK
| | - Giles Velarde
- School of Chemistry, The University of Manchester, Faraday Building, PO Box 88, Sackville Street, Manchester, UK
| | - Norman Morrison
- School of Computer Science, The University of Manchester, Kilburn Building, Oxford Road, Manchester, UK
| | - Andrew Hayes
- Faculty of Life Sciences, The University of Manchester, Oxford Road, Manchester, UK
| | - Helen Hulme
- School of Computer Science, The University of Manchester, Kilburn Building, Oxford Road, Manchester, UK
| | - A Joseph Wood
- NERC Environmental Bioinformatics Centre, Oxford Centre for Ecology and Hydrology, Oxford, UK
| | - Karim Nashar
- School of Computer Science, The University of Manchester, Kilburn Building, Oxford Road, Manchester, UK
| | - Douglas B Kell
- School of Chemistry, The University of Manchester, Faraday Building, PO Box 88, Sackville Street, Manchester, UK
| | - Andy Brass
- School of Computer Science, The University of Manchester, Kilburn Building, Oxford Road, Manchester, UK
- Faculty of Life Sciences, The University of Manchester, Oxford Road, Manchester, UK
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Mey AR, Craig SA, Payne SM. Characterization of Vibrio cholerae RyhB: the RyhB regulon and role of ryhB in biofilm formation. Infect Immun 2005; 73:5706-19. [PMID: 16113288 PMCID: PMC1231101 DOI: 10.1128/iai.73.9.5706-5719.2005] [Citation(s) in RCA: 130] [Impact Index Per Article: 6.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Vibrio cholerae encodes a small RNA with homology to Escherichia coli RyhB. Like E. coli ryhB, V. cholerae ryhB is negatively regulated by iron and Fur and is required for repression of genes encoding the superoxide dismutase SodB and multiple tricarboxylic acid cycle enzymes. However, V. cholerae RyhB is considerably longer (>200 nucleotides) than the E. coli RNA (90 nucleotides), and it regulates the expression of a variety of genes that are not known to be regulated by RyhB in E. coli, including genes involved in motility, chemotaxis, and biofilm formation. A mutant with a deletion in ryhB had reduced chemotactic motility in low-iron medium and was unable to form wild-type biofilms. The defect in biofilm formation was suppressed by growing the mutant in the presence of excess iron or succinate. The wild-type strain showed reduced biofilm formation in iron-deficient medium, further supporting a role for iron in normal biofilm formation. The ryhB mutant was not defective for colonization in a mouse model and appeared to be at a slight advantage when competing with the wild-type parental strain. Other genes whose expression was influenced by RyhB included those encoding the outer membrane porins OmpT and OmpU, several iron transport systems, and proteins containing heme or iron-sulfur clusters. These data indicate that V. cholerae RyhB has diverse functions, ranging from iron homeostasis to the regulation of biofilm formation.
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Affiliation(s)
- Alexandra R Mey
- Institute for Cellular and Molecular Biology, The University of Texas, Austin, TX 78712-1095, USA
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Cahan P, Ahmad AM, Burke H, Fu S, Lai Y, Florea L, Dharker N, Kobrinski T, Kale P, McCaffrey TA. List of lists-annotated (LOLA): a database for annotation and comparison of published microarray gene lists. Gene 2005; 360:78-82. [PMID: 16140476 DOI: 10.1016/j.gene.2005.07.008] [Citation(s) in RCA: 43] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/11/2005] [Revised: 07/07/2005] [Accepted: 07/11/2005] [Indexed: 10/25/2022]
Abstract
Microarray profiling of RNA expression is a powerful tool that generates large lists of transcripts that are potentially relevant to a disease or treatment. However, because the lists of changed transcripts are embedded in figures and tables, they are typically inaccessible for search engines. Due to differences in gene nomenclatures, the lists are difficult to compare between studies. LOLA (Lists of Lists Annotated) is an internet-based database for comparing gene lists from microarray studies or other genomic-scale methods. It serves as a common platform to compare and reannotate heterogeneous gene lists from different microarray platforms or different genomic methodologies such as serial analysis of gene expression (SAGE) or proteomics. LOLA () provides researchers with a means to store, annotate, and compare gene lists produced from different studies or different analyses of the same study. It is especially useful in identifying potentially "high interest" genes which are reported as significant across multiple studies and species. Its application to the fields of stem cell, cancer, and aging research is demonstrated by comparing published papers.
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Affiliation(s)
- Patrick Cahan
- The George Washington University Medical Center, Department of Biochemistry and Molecular Biology, 2300 I Street NW. Ross Hall 541, Washington, D.C. 20037, United States
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Shingara J, Keiger K, Shelton J, Laosinchai-Wolf W, Powers P, Conrad R, Brown D, Labourier E. An optimized isolation and labeling platform for accurate microRNA expression profiling. RNA (NEW YORK, N.Y.) 2005; 11:1461-70. [PMID: 16043497 PMCID: PMC1370829 DOI: 10.1261/rna.2610405] [Citation(s) in RCA: 211] [Impact Index Per Article: 10.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/03/2023]
Abstract
MicroRNAs (miRNAs) are small, noncoding RNAs that regulate gene expression in both plants and animals. miRNA genes have been implicated in a variety of important biological processes, including development, differentiation, apoptosis, fat metabolism, viral infection, and cancer. Similar to protein-coding messenger RNAs, miRNA expression varies between tissues and developmental states. To acquire a better understanding of global miRNA expression in tissues and cells, we have developed isolation, labeling, and array procedures to measure the relative abundance of all of the known human mature miRNAs. The method relies on rapid isolation of RNA species smaller than ~40 nucleotides (nt), direct and homogenous enzymatic labeling of the mature miRNAs with amine modified ribonucleotides, and hybridization to antisense DNA oligonucleotide probes. A thorough performance study showed that this miRNA microarray system can detect subfemtomole amounts of individual miRNAs from <1 mug of total RNA, with 98% correlation between independent replicates. The system has been applied to compare the global miRNA expression profiles in 26 different normal human tissues. This comprehensive analysis identified miRNAs that are preferentially expressed in one or a few related tissues and revealed that human adult tissues have unique miRNA profiles. This implicates miRNAs as important components of tissue development and differentiation. Taken together, these results emphasize the immense potential of microarrays for sensitive and high-throughput analysis of miRNA expression in normal and disease states.
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Affiliation(s)
- Jaclyn Shingara
- Ambion, Inc., 2130 Woodward Street, Austin, TX 78744-1832, USA
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Abstract
Complete genomic sequences of several oral pathogens have been deciphered and multiple sources of independently annotated data are available for the same genomes. Different gene identification schemes and functional annotation methods used in these databases present a challenge for cross-referencing and the efficient use of the data. The Bioinformatics Resource for Oral Pathogens (BROP) aims to integrate bioinformatics data from multiple sources for easy comparison, analysis and data-mining through specially designed software interfaces. Currently, databases and tools provided by BROP include: (i) a graphical genome viewer (Genome Viewer) that allows side-by-side visual comparison of independently annotated datasets for the same genome; (ii) a pipeline of automatic data-mining algorithms to keep the genome annotation always up-to-date; (iii) comparative genomic tools such as Genome-wide ORF Alignment (GOAL); and (iv) the Oral Pathogen Microarray Database. BROP can also handle unfinished genomic sequences and provides secure yet flexible control over data access. The concept of providing an integrated source of genomic data, as well as the data-mining model used in BROP can be applied to other organisms. BROP can be publicly accessed at .
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Affiliation(s)
- Tsute Chen
- The Forsyth Institute, 140 Fenway, Boston, MA 02115, USA.
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Salmi ML, Bushart TJ, Stout SC, Roux SJ. Profile and analysis of gene expression changes during early development in germinating spores of Ceratopteris richardii. PLANT PHYSIOLOGY 2005; 138:1734-45. [PMID: 15965014 PMCID: PMC1176442 DOI: 10.1104/pp.105.062851] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/03/2023]
Abstract
Analysis of an expressed sequence tag library with more than 5,000 sequences from spores of the fern Ceratopteris richardii reveals that more than 3,900 of them represent distinct genes, and almost 70% of these have significant similarity to Arabidopsis (Arabidopsis thaliana) genes. Eight genes are common between three very different dormant plant systems, Ceratopteris spores, Arabidopsis seeds, and Arabidopsis pollen. We evaluated the pattern of mRNA abundance over the first 48 h of spore development using a microarray of cDNAs representing 3,207 distinct genes of C. richardii and determined the relative levels of RNA abundance for 3,143 of these genes using a Bayesian method of statistical analysis. More than 900 of them (29%) show a significant change between any of the five time points analyzed, and these have been annotated based on their sequence similarity with the Arabidopsis proteome. Novel data arising from these analyses identify genes likely to be critical for the germination and subsequent early development of diverse cells and tissues emerging from dormancy.
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Affiliation(s)
- Mari L Salmi
- Molecular Cell and Developmental Biology, University of Texas, Austin, Texas 78751, USA
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Abstract
This article represents the proceedings of a symposium at the 2004 International Society for Biomedical Research on Alcoholism in Mannheim, Germany, organized and co-chaired by Susan E. Bergeson and Wolfgang Sommer. The presentations and presenter were (1) Gene Expression in Brains of Alcohol-Preferring and Non-Preferring Rats, by Howard J. Edenberg (2) Candidate Treatment Targets for Alcoholism: Leads from Functional Genomics Approaches, by Wolfgang Sommer (3) Microarray Analysis of Acute and Chronic Alcohol Response in Brain, by Susan E. Bergeson (4) On the Integration of QTL and Gene Expression Analysis, by Robert J. Hitzemann (5) Microarray and Proteomic Analysis of the Human Alcoholic Brain, by Peter R. Dodd.
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Jenkins H, Johnson H, Kular B, Wang T, Hardy N. Toward supportive data collection tools for plant metabolomics. PLANT PHYSIOLOGY 2005; 138:67-77. [PMID: 15888680 PMCID: PMC1104162 DOI: 10.1104/pp.104.058875] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/22/2004] [Revised: 03/04/2005] [Accepted: 03/04/2005] [Indexed: 05/02/2023]
Abstract
Over recent years, a number of initiatives have proposed standard reporting guidelines for functional genomics experiments. Associated with these are data models that may be used as the basis of the design of software tools that store and transmit experiment data in standard formats. Central to the success of such data handling tools is their usability. Successful data handling tools are expected to yield benefits in time saving and in quality assurance. Here, we describe the collection of datasets that conform to the recently proposed data model for plant metabolomics known as ArMet (architecture for metabolomics) and illustrate a number of approaches to robust data collection that have been developed in collaboration between software engineers and biologists. These examples also serve to validate ArMet from the data collection perspective by demonstrating that a range of software tools, supporting data recording and data upload to central databases, can be built using the data model as the basis of their design.
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Affiliation(s)
- Helen Jenkins
- Department of Computer Science, University of Wales, Penglais, Aberystwyth, Ceredigion, Wales, SY23 3DB, United Kingdom
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Maurer M, Molidor R, Sturn A, Hartler J, Hackl H, Stocker G, Prokesch A, Scheideler M, Trajanoski Z. MARS: microarray analysis, retrieval, and storage system. BMC Bioinformatics 2005; 6:101. [PMID: 15836795 PMCID: PMC1090551 DOI: 10.1186/1471-2105-6-101] [Citation(s) in RCA: 45] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/03/2005] [Accepted: 04/18/2005] [Indexed: 01/27/2023] Open
Abstract
Background Microarray analysis has become a widely used technique for the study of gene-expression patterns on a genomic scale. As more and more laboratories are adopting microarray technology, there is a need for powerful and easy to use microarray databases facilitating array fabrication, labeling, hybridization, and data analysis. The wealth of data generated by this high throughput approach renders adequate database and analysis tools crucial for the pursuit of insights into the transcriptomic behavior of cells. Results MARS (Microarray Analysis and Retrieval System) provides a comprehensive MIAME supportive suite for storing, retrieving, and analyzing multi color microarray data. The system comprises a laboratory information management system (LIMS), a quality control management, as well as a sophisticated user management system. MARS is fully integrated into an analytical pipeline of microarray image analysis, normalization, gene expression clustering, and mapping of gene expression data onto biological pathways. The incorporation of ontologies and the use of MAGE-ML enables an export of studies stored in MARS to public repositories and other databases accepting these documents. Conclusion We have developed an integrated system tailored to serve the specific needs of microarray based research projects using a unique fusion of Web based and standalone applications connected to the latest J2EE application server technology. The presented system is freely available for academic and non-profit institutions. More information can be found at .
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Affiliation(s)
- Michael Maurer
- Institute for Genomics and Bioinformatics and Christian Doppler Laboratory for Genomics and Bioinformatics, Graz University of Technology, Petersgasse 14, 8010 Graz, Austria
| | - Robert Molidor
- Institute for Genomics and Bioinformatics and Christian Doppler Laboratory for Genomics and Bioinformatics, Graz University of Technology, Petersgasse 14, 8010 Graz, Austria
| | - Alexander Sturn
- Institute for Genomics and Bioinformatics and Christian Doppler Laboratory for Genomics and Bioinformatics, Graz University of Technology, Petersgasse 14, 8010 Graz, Austria
| | - Juergen Hartler
- Institute for Genomics and Bioinformatics and Christian Doppler Laboratory for Genomics and Bioinformatics, Graz University of Technology, Petersgasse 14, 8010 Graz, Austria
| | - Hubert Hackl
- Institute for Genomics and Bioinformatics and Christian Doppler Laboratory for Genomics and Bioinformatics, Graz University of Technology, Petersgasse 14, 8010 Graz, Austria
| | - Gernot Stocker
- Institute for Genomics and Bioinformatics and Christian Doppler Laboratory for Genomics and Bioinformatics, Graz University of Technology, Petersgasse 14, 8010 Graz, Austria
| | - Andreas Prokesch
- Institute for Genomics and Bioinformatics and Christian Doppler Laboratory for Genomics and Bioinformatics, Graz University of Technology, Petersgasse 14, 8010 Graz, Austria
| | - Marcel Scheideler
- Institute for Genomics and Bioinformatics and Christian Doppler Laboratory for Genomics and Bioinformatics, Graz University of Technology, Petersgasse 14, 8010 Graz, Austria
| | - Zlatko Trajanoski
- Institute for Genomics and Bioinformatics and Christian Doppler Laboratory for Genomics and Bioinformatics, Graz University of Technology, Petersgasse 14, 8010 Graz, Austria
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Tanino M, Debily MA, Tamura T, Hishiki T, Ogasawara O, Murakawa K, Kawamoto S, Itoh K, Watanabe S, de Souza SJ, Imbeaud S, Graudens E, Eveno E, Hilton P, Sudo Y, Kelso J, Ikeo K, Imanishi T, Gojobori T, Auffray C, Hide W, Okubo K. The Human Anatomic Gene Expression Library (H-ANGEL), the H-Inv integrative display of human gene expression across disparate technologies and platforms. Nucleic Acids Res 2005; 33:D567-72. [PMID: 15608263 PMCID: PMC540058 DOI: 10.1093/nar/gki104] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/26/2022] Open
Abstract
The Human Anatomic Gene Expression Library (H-ANGEL) is a resource for information concerning the anatomical distribution and expression of human gene transcripts. The tool contains protein expression data from multiple platforms that has been associated with both manually annotated full-length cDNAs from H-InvDB and RefSeq sequences. Of the H-Inv predicted genes, 18 897 have associated expression data generated by at least one platform. H-ANGEL utilizes categorized mRNA expression data from both publicly available and proprietary sources. It incorporates data generated by three types of methods from seven different platforms. The data are provided to the user in the form of a web-based viewer with numerous query options. H-ANGEL is updated with each new release of cDNA and genome sequence build. In future editions, we will incorporate the capability for expression data updates from existing and new platforms. H-ANGEL is accessible at http://www.jbirc.aist.go.jp/hinv/h-angel/.
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Affiliation(s)
- Motohiko Tanino
- Integrated Database Group, Japan Biological Information Research Center, Japan Biological Informatics Consortium, Time24 Building 10F, 2-45 Aomi, Koto-ku, Tokyo 135-0064, Japan.
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Ponomarev I, Schafer GL, Blednov YA, Williams RW, Iyer VR, Harris RA. Convergent analysis of cDNA and short oligomer microarrays, mouse null mutants and bioinformatics resources to study complex traits. GENES BRAIN AND BEHAVIOR 2005; 3:360-8. [PMID: 15544578 DOI: 10.1111/j.1601-183x.2004.00088.x] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/01/2022]
Abstract
Gene expression data sets have recently been exploited to study genetic factors that modulate complex traits. However, it has been challenging to establish a direct link between variation in patterns of gene expression and variation in higher order traits such as neuropharmacological responses and patterns of behavior. Here we illustrate an approach that combines gene expression data with new bioinformatics resources to discover genes that potentially modulate behavior. We have exploited three complementary genetic models to obtain convergent evidence that differential expression of a subset of genes and molecular pathways influences ethanol-induced conditioned taste aversion (CTA). As a first step, cDNA microarrays were used to compare gene expression profiles of two null mutant mouse lines with difference in ethanol-induced aversion. Mice lacking a functional copy of G protein-gated potassium channel subunit 2 (Girk2) show a decrease in the aversive effects of ethanol, whereas preproenkephalin (Penk) null mutant mice show the opposite response. We hypothesize that these behavioral differences are generated in part by alterations in expression downstream of the null alleles. We then exploited the WebQTL databases to examine the genetic covariance between mRNA expression levels and measurements of ethanol-induced CTA in BXD recombinant inbred (RI) strains. Finally, we identified a subset of genes and functional groups associated with ethanol-induced CTA in both null mutant lines and BXD RI strains. Collectively, these approaches highlight the phosphatidylinositol signaling pathway and identify several genes including protein kinase C beta isoform and preproenkephalin in regulation of ethanol- induced conditioned taste aversion. Our results point to the increasing potential of the convergent approach and biological databases to investigate genetic mechanisms of complex traits.
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Affiliation(s)
- I Ponomarev
- Waggoner Center for Alcohol and Addiction Research, The University of Texas at Austin, Austin, TX, USA.
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Kelley ML, Keiger KE, Lee CJ, Huibregtse JM. The global transcriptional effects of the human papillomavirus E6 protein in cervical carcinoma cell lines are mediated by the E6AP ubiquitin ligase. J Virol 2005; 79:3737-47. [PMID: 15731267 PMCID: PMC1075713 DOI: 10.1128/jvi.79.6.3737-3747.2005] [Citation(s) in RCA: 83] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/06/2004] [Accepted: 10/14/2004] [Indexed: 11/20/2022] Open
Abstract
The function of the human papillomavirus (HPV) E6 protein that is most clearly linked to carcinogenesis is the targeted degradation of p53, which is dependent on the E6AP ubiquitin ligase. Additional functions have been attributed to E6, including the stimulation of telomerase activity and the targeted degradation of other cellular proteins, but in most cases it is unclear whether these activities are also E6AP dependent. While E6 clearly influences the transcriptional program of HPV-positive cell lines through the inactivation of p53, it has been shown that at least a subset of its p53-independent functions are also reflected in the transcriptional program. For this study, we have determined the extent to which E6AP is involved in mediating the set of E6 functions that impact on the global transcriptional program of HPV-positive cell lines. The transcriptional profiles of approximately 31,000 genes were characterized for three cell lines (HeLa, Caski, and SiHa cells) after small interfering RNA (siRNA)-mediated silencing of E6 or E6AP. We found that E6 and E6AP siRNAs elicited nearly identical alterations in the transcriptional profile of each cell line. Some of the expression alterations were apparent secondary effects of p53 stabilization, while the basis of most other changes was not reconcilable with previously proposed E6 functions. While expression changes of the TERT gene (telomerase catalytic subunit) were not revealed by the array, telomerase repeat amplification protocol assays showed that both E6 and E6AP knockouts resulted in a suppression of telomerase activity. Together, these results suggest that E6AP mediates a broad spectrum of E6 functions, including virtually all functions that impact on the transcriptional program of HPV-positive cell lines.
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Affiliation(s)
- Melissa L Kelley
- Institute for Cellular and Molecular Biology, Section of Molecular Genetics and Microbiology, University of Texas at Austin, Austin, TX 78712, USA
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