Biomedical subjects

Philippe Demougin

Publications and source records attributed to Philippe Demougin.

7 recordsLinked to original sources

PubMed2006

MIMAS: an innovative tool for network-based high density oligonucleotide microarray data management and annotation.

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.

Database Management Systems↗

PubMed2006

Extraction of high-integrity RNA suitable for microarray gene expression analysis from long-term stored human thyroid tissues.

INTRODUCTION: Isolation of high-quality RNA from fresh-frozen thyroid tissues stored for more than a decade would open novel options for gene expression profiling. Herein, we describe successful extraction of high-integrity RNA from human thyroid tissues that were stored for more than a decade. METHODS: Seventy-nine samples (15 goitres, 20 follicular adenomas, 30 papillary carcinomas, 14 follicular carcinomas) that were shock-frozen in isopentane and stored for a median of 11 years (range 1-16 years) were processed using standard precipitation and column filtration techniques. RNA integrity was assessed by electrophoresis using the RNA integrity number (RIN) algorithm and by gene expression profiling determining the 3'/5' ratio of the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene and the percentage of transcripts detected on the Affymetrix U133 2.0 human genome GeneChip. RESULTS: The median RNA yield was 1.9 microg/mg tissue (papillary carcinoma 2.1 microg/mg, range 0.2-7.2 microg/mg; follicular carcinoma 2.4 microg/mg, range 0.2-3.2 microg/mg; goitre 1.4 microg/mg, range 0.1-5.4 microg/mg; follicular adenoma 1.6 microg/mg, range 0.1-6.2 microg/mg; p = 0.46) with an 8.6 (7.3-9.8) median RIN. The median GAPDH gene 3'/5' ratio was 1.43 (1.34-1.52) and the median percentage of present calls was 48.1% (42.7-52.0%). CONCLUSIONS: Age and entity independent RNA suitable for expression profiling can be extracted from long-term stored fresh-frozen human thyroid tissues.

Adenocarcinoma, Follicular↗

PubMed2005

Three-dimensional culture of melanoma cells profoundly affects gene expression profile: a high density oligonucleotide array study.

Growth in three-dimensional (3D) architectures has been suggested to play an important role in tumor expansion and in the resistance of cancers to treatment with drugs or cytokines or irradiation. To obtain an insight into underlying molecular mechanisms, we addressed gene expression profiles of NA8 melanoma cells cultured in bidimensional monolayers (2D) or in 3D multicellular tumor spheroids (MCTS). MCTS containing 10-30,000 cells were generated upon overnight culture in poly-Hydroxyethylmethacrylate (polyHEMA) coated plates. Kinetics of cell proliferation in MCTS was significantly slower than in monolayer cultures. Following long-term culture (>10 days), however, MCTS showed highly compact and organised cell growth in outer layers, with necrotic cores. Oligonucleotide microarray analysis of the expression of over 20,000 genes was performed on cells cultured in standard 2D, in the presence of collagen as model of extracellular matrix (ECM), or in MCTS. Gene expression profiles of cells cultured in 2D in the presence or absence of ECM were highly similar, with >/=threefold differences limited to five genes. In contrast, culture in MCTS resulted in the significant, >/=threefold, upregulation of the expression of >100 transcripts while 73 were >/=threefold downregulated. In particular, genes encoding CXCL1, 2, and 3 (GRO-alpha, -beta, and gamma), IL-8, CCL20 (MIP-3alpha), and Angiopoietin-like 4 were significantly upregulated, whereas basic FGF and CD49d encoding genes were significantly downregulated. Oligonucleotide chip data were validated at the gene and protein level by quantitative real-time PCR, ELISA, and cell surface staining assays. Taken together, our data indicate that structural modifications of the architecture of tumor cell cultures result in a significant upregulation of the expression of a number of genes previously shown to play a role in melanoma progression and metastatic process.

Cell Division↗

PubMed2005

Novel response to microtubule perturbation in meiosis.

During the mitotic cell cycle, microtubule depolymerization leads to a cell cycle arrest in metaphase, due to activation of the spindle checkpoint. Here, we show that under microtubule-destabilizing conditions, such as low temperature or the presence of the spindle-depolymerizing drug benomyl, meiotic budding yeast cells arrest in G(1) or G(2), instead of metaphase. Cells arrest in G(1) if microtubule perturbation occurs as they enter the meiotic cell cycle and in G(2) if cells are already undergoing premeiotic S phase. Concomitantly, cells down-regulate genes required for cell cycle progression, meiotic differentiation, and spore formation in a highly coordinated manner. Decreased expression of these genes is likely to be responsible for halting both cell cycle progression and meiotic development. Our results point towards the existence of a novel surveillance mechanism of microtubule integrity that may be particularly important during specialized cell cycles when coordination of cell cycle progression with a developmental program is necessary.

Benomyl↗

PubMed2004

Rank Difference Analysis of Microarrays (RDAM), a novel approach to statistical analysis of microarray expression profiling data.

BACKGROUND: A key step in the analysis of microarray expression profiling data is the identification of genes that display statistically significant changes in expression signals between two biological conditions. RESULTS: We describe a new method, Rank Difference Analysis of Microarrays (RDAM), which estimates the total number of truly varying genes and assigns a p-value to each signal variation. Information on a group of differentially expressed genes includes the sensitivity and the false discovery rate. We demonstrate the feasibility and efficiency of our approach by applying it to a large synthetic expression data set and to a biological data set obtained by comparing vegetatively-growing wild type and tor2-mutant yeast strains. In both cases we observed a significant improvement of the power of analysis when our method is compared to another popular nonparametric method. CONCLUSIONS: This study provided a valuable new statistical method to analyze microarray data. We conclude that the good quality of the results obtained by RDAM is mainly due to the quasi-perfect equalization of variation distribution, which is related to the standardization procedure used and to the measurement of variation by rank difference.

Gene Expression Profiling↗

PubMed2004

NPR1 kinase and RSP5-BUL1/2 ubiquitin ligase control GLN3-dependent transcription in Saccharomyces cerevisiae.

The GATA transcription factors GLN3 and GAT1 activate nitrogen-regulated genes in Saccharomyces cerevisiae. NPR1 is a protein kinase that controls post-Golgi sorting of amino acid permeases. In the presence of a good nitrogen source, TOR (target of rapamycin) maintains GLN3 and NPR1 phosphorylated and inactive by inhibiting the type 2A-related phosphatase SIT4. We identified NPR1 as a regulator of GLN3. Specifically, loss of NPR1 causes nuclear translocation and activation of GLN3, but not GAT1, in nitrogen-rich conditions. NPR1-mediated inhibition of GLN3 is independent of the phosphatase SIT4. We also demonstrate that the E3/E4 ubiquitin-protein ligase proteins RSP5 and BUL1/2 are required for GLN3 activation under poor nitrogen conditions. Thus, NPR1 and BUL1/2 antagonistically control GLN3-dependent transcription, suggesting a role for regulated ubiquitination in the control of nutrient-responsive transcription.

Adaptor Proteins, Signal Transducing↗

PubMed2004

Expression profiling of mammalian male meiosis and gametogenesis identifies novel candidate genes for roles in the regulation of fertility.

We report a comprehensive large-scale expression profiling analysis of mammalian male germ cells undergoing mitotic growth, meiosis, and gametogenesis by using high-density oligonucleotide microarrays and highly enriched cell populations. Among 11,955 rat loci investigated, 1268 were identified as differentially transcribed in germ cells at subsequent developmental stages compared with total testis, somatic Sertoli cells as well as brain and skeletal muscle controls. The loci were organized into four expression clusters that correspond to somatic, mitotic, meiotic, and postmeiotic cell types. This work provides information about expression patterns of approximately 200 genes known to be important during male germ cell development. Approximately 40 of those are included in a group of 121 transcripts for which we report germ cell expression and lack of transcription in three somatic control cell types. Moreover, we demonstrate the testicular expression and transcriptional induction in mitotic, meiotic, and/or postmeiotic germ cells of 293 as yet uncharacterized transcripts, some of which are likely to encode factors involved in spermatogenesis and fertility. This group also contains potential germ cell-specific targets for innovative contraceptives. A graphical display of the data is conveniently accessible through the GermOnline database at http://www.germonline.org.

Animals↗