29 results on '"Demay F"'
Search Results
2. Deciphering the molecular mechanisms sustaining the estrogenic activity of the two major dietary compounds zearalenone and apigenin in ER-positive breast cancer cell lines
- Author
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Lecomte, S., Demay, F., Pham, T.H., Moulis, S., Efstathiou, T., Chalmel, F., Pakdel, F., Institut de recherche en santé, environnement et travail (Irset), Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique )-Institut National de la Santé et de la Recherche Médicale (INSERM)-École des Hautes Études en Santé Publique [EHESP] (EHESP)-Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Université d'Angers (UA), École des Hautes Études en Santé Publique [EHESP] (EHESP), Nutrinov, Université d'Angers (UA)-Université de Rennes (UR)-École des Hautes Études en Santé Publique [EHESP] (EHESP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), and Troccaz, Olivier
- Subjects
[SDV.MHEP.EM] Life Sciences [q-bio]/Human health and pathology/Endocrinology and metabolism ,Endocrine disruption ,Breast Neoplasms ,Phytoestrogens ,lcsh:TX341-641 ,[SDV.CAN]Life Sciences [q-bio]/Cancer ,[SDV.TOX.TCA]Life Sciences [q-bio]/Toxicology/Toxicology and food chain ,[SDV.MHEP.EM]Life Sciences [q-bio]/Human health and pathology/Endocrinology and metabolism ,Article ,[SDV.AEN] Life Sciences [q-bio]/Food and Nutrition ,Breast cancer ,Receptors, Estrogen ,[SDV.TOX.TCA] Life Sciences [q-bio]/Toxicology/Toxicology and food chain ,[SDV.CAN] Life Sciences [q-bio]/Cancer ,Cell Line, Tumor ,Humans ,Zearalenone ,Estrogen receptor ,Female ,Estrogens, Non-Steroidal ,Gene expression ,Apigenin ,lcsh:Nutrition. Foods and food supply ,[SDV.AEN]Life Sciences [q-bio]/Food and Nutrition ,Dietary compounds - Abstract
The flavone apigenin and the mycotoxin zearalenone are two major compounds found in the human diet which bind estrogen receptors (ERs), and therefore influence ER activity. However, the underlying mechanisms are not well known. To unravel the molecular mechanisms that could explain the differential effect of zearalenone and apigenin on ER-positive breast cancer cell proliferation, gene-reporter assays, chromatin immunoprecipitation (ChIP) experiments, proliferation assays and transcriptomic analysis were performed. We found that zearalenone and apigenin transactivated ERs and promoted the expression of estradiol (E2)-responsive genes. However, zearalenone clearly enhanced cellular proliferation, while apigenin appeared to be antiestrogenic in the presence of E2 in both ER-positive breast cancer cell lines, MCF-7 and T47D. The transcriptomic analysis showed that both compounds regulate gene expression in the same way, but with differences in intensity. Two major sets of genes were identified, one set was linked to the cell cycle and the other set was linked to stress response and growth arrest. Our results show that the transcription dynamics in gene regulation induced by apigenin were somehow different with zearalenone and E2 and may explain the differential effect of these compounds on the phenotype of the breast cancer cell. Together, our results confirmed the potential health benefit effect of apigenin, while zearalenone appeared to be a true endocrine-disrupting compound.
- Published
- 2019
3. Effect of gonadotropin-releasing hormone on estrogen receptor messenger ribonucleic acid level in perifused pituitary cells
- Author
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Demay, F., Tiffoche, C., and Thieulant, M. L.
- Published
- 1996
- Full Text
- View/download PDF
4. Steroid-Independent Activation of ER by GnRH in Gonadotrope Pituitary Cells
- Author
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Demay, F, De Monti, M, Tiffoche, C, Vaillant, C, and Thieulant, M L
- Published
- 2001
5. Quantitative PCR with colorimetric detection
- Author
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Demay, F., primary, Darche, S., additional, Kourilsky, P., additional, and Delassus, S., additional
- Published
- 1996
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6. The Effect of Piracetam on Volunteers in a Low-Pressure Tank
- Author
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Demay, F and Bande, J
- Abstract
Twelve volunteers were subjected to a double-blind crossover test to evaluate their concentration in conditions of hypoxemia after treatment with piracetam or with a placebo.Piracetam was administered in doses of 4.8 g per day for 4 days and 7.2 g on the 5th day.The speed with which the test was carried out was not influenced to a statistically significant degree but the proportion of errors was. This effect of piracetam was more pronounced for the longer-lasting periods of hypoxemia.
- Published
- 1980
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7. Simple purification and characterization of soluble and homogenous ABC-F translation factors from Enterococcus faecium.
- Author
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Demay F, Hallier M, Georgeault S, Com E, Cattoir V, Goude R, and Gillet R
- Subjects
- ATP-Binding Cassette Transporters chemistry, Protein Biosynthesis, Anti-Bacterial Agents metabolism, Ribosomes metabolism, Escherichia coli genetics, Escherichia coli metabolism, Enterococcus faecium genetics, Enterococcus faecium metabolism
- Abstract
The family of ATP-binding cassette F proteins (ABC-F) is mainly made up of cytosolic proteins involved in regulating protein synthesis, and they are often part of a mechanism that confers resistance to ribosome-targeting antibiotics. The existing literature has emphasized the difficulty of purifying these recombinant proteins because of their very low solubility and stability. Here, we describe a rapid and efficient three-step purification procedure that allows for the production of untagged ABC-F proteins from Enterococcus faecium in the heterologous host Escherichia coli. After four purified ABC-F proteins were produced using this protocol, their biological activities were validated by in vitro experiment. In conclusion, our study provides an invaluable tool for obtaining large amounts of untagged and soluble ABC-F proteins that can then be used for in vitro experiments., (Copyright © 2023 Elsevier Inc. All rights reserved.)
- Published
- 2023
- Full Text
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8. Isolation of Leishmania Promastigote Flagella.
- Author
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Beneke T, Demay F, Wheeler RJ, and Gluenz E
- Subjects
- Centrifugation, Density Gradient, Cytoskeleton metabolism, Leishmania mexicana growth & development, Lipidomics, Mass Spectrometry, Microscopy, Electron, Protozoan Proteins analysis, Protozoan Proteins isolation & purification, Protozoan Proteins metabolism, Cell Fractionation methods, Flagella metabolism, Leishmania mexicana cytology, Life Cycle Stages
- Abstract
Eukaryotic flagella are conserved multifunctional organelles with roles in motility, intercellular interactions, and signal transduction. Leishmania possess a single flagellum at all stages of their life cycle. Flagella of promastigote forms in the fly are long and motile, with a canonical 9 + 2 microtubule axoneme and an extra-axonemal paraflagellar rod (PFR). This protocol describes a simple method for the isolation of Leishmania mexicana promastigote flagella, optimized to yield intact flagella that retain both the cytoskeletal elements (9 + 2 axoneme and PFR) and the surrounding membrane. The isolated flagella and deflagellated cell bodies are suitable for analysis by electron microscopy, protein mass spectrometry, and lipidomics.
- Published
- 2020
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9. Synthesis and evaluation of 1,3,4-oxadiazole derivatives for development as broad-spectrum antibiotics.
- Author
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Tresse C, Radigue R, Gomes Von Borowski R, Thepaut M, Hanh Le H, Demay F, Georgeault S, Dhalluin A, Trautwetter A, Ermel G, Blanco C, van de Weghe P, Jean M, Giard JC, and Gillet R
- Subjects
- Anti-Bacterial Agents chemical synthesis, Anti-Bacterial Agents toxicity, Cell Line, Tumor, Cell Survival drug effects, Drug Design, Drug Synergism, Gram-Negative Bacteria drug effects, Gram-Positive Bacteria drug effects, Humans, Microbial Sensitivity Tests, Oxadiazoles chemical synthesis, Oxadiazoles toxicity, Anti-Bacterial Agents pharmacology, Oxadiazoles pharmacology
- Abstract
The reality and intensity of antibiotic resistance in pathogenic bacteria calls for the rapid development of new antimicrobial drugs. In bacteria, trans-translation is the primary quality control mechanism for rescuing ribosomes arrested during translation. Because trans-translation is absent in eukaryotes but necessary to avoid ribosomal stalling and therefore essential for bacterial survival, it is a promising target either for novel antibiotics or for improving the activities of the protein synthesis inhibitors already in use. Oxadiazole derivatives display strong bactericidal activity against a large number of bacteria, but their effects on trans-translation were recently questioned. In this work, a series of new 1,3,4-oxadiazole derivatives and analogs were synthesized and assessed for their efficiency as antimicrobial agents against a wide range of gram-positive and gram-negative pathogenic strains. Despite the strong antimicrobial activity observed in these molecules, it turns out that they do not target trans-translation in vivo, but they definitely act on other cellular pathways., (Copyright © 2019 Elsevier Ltd. All rights reserved.)
- Published
- 2019
- Full Text
- View/download PDF
10. Genetic dissection of a Leishmania flagellar proteome demonstrates requirement for directional motility in sand fly infections.
- Author
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Beneke T, Demay F, Hookway E, Ashman N, Jeffery H, Smith J, Valli J, Becvar T, Myskova J, Lestinova T, Shafiq S, Sadlova J, Volf P, Wheeler RJ, and Gluenz E
- Subjects
- Animals, Flagella genetics, Leishmania genetics, Proteome genetics, Protozoan Proteins genetics, Flagella metabolism, Leishmania metabolism, Proteome metabolism, Protozoan Proteins metabolism, Psychodidae parasitology
- Abstract
The protozoan parasite Leishmania possesses a single flagellum, which is remodelled during the parasite's life cycle from a long motile flagellum in promastigote forms in the sand fly to a short immotile flagellum in amastigotes residing in mammalian phagocytes. This study examined the protein composition and in vivo function of the promastigote flagellum. Protein mass spectrometry and label free protein enrichment testing of isolated flagella and deflagellated cell bodies defined a flagellar proteome for L. mexicana promastigote forms (available via ProteomeXchange with identifier PXD011057). This information was used to generate a CRISPR-Cas9 knockout library of 100 mutants to screen for flagellar defects. This first large-scale knockout screen in a Leishmania sp. identified 56 mutants with altered swimming speed (52 reduced and 4 increased) and defined distinct mutant categories (faster swimmers, slower swimmers, slow uncoordinated swimmers and paralysed cells, including aflagellate promastigotes and cells with curled flagella and disruptions of the paraflagellar rod). Each mutant was tagged with a unique 17-nt barcode, providing a simple barcode sequencing (bar-seq) method for measuring the relative fitness of L. mexicana mutants in vivo. In mixed infections of the permissive sand fly vector Lutzomyia longipalpis, paralysed promastigotes and uncoordinated swimmers were severely diminished in the fly after defecation of the bloodmeal. Subsequent examination of flies infected with a single paralysed mutant lacking the central pair protein PF16 or an uncoordinated swimmer lacking the axonemal protein MBO2 showed that these promastigotes did not reach anterior regions of the fly alimentary tract. These data show that L. mexicana need directional motility for successful colonisation of sand flies., Competing Interests: The authors have declared that no competing interests exist.
- Published
- 2019
- Full Text
- View/download PDF
11. Deciphering the Molecular Mechanisms Sustaining the Estrogenic Activity of the Two Major Dietary Compounds Zearalenone and Apigenin in ER-Positive Breast Cancer Cell Lines.
- Author
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Lecomte S, Demay F, Pham TH, Moulis S, Efstathiou T, Chalmel F, and Pakdel F
- Subjects
- Cell Line, Tumor, Estrogens, Non-Steroidal toxicity, Female, Humans, Phytoestrogens, Receptors, Estrogen genetics, Apigenin toxicity, Breast Neoplasms metabolism, Receptors, Estrogen metabolism, Zearalenone toxicity
- Abstract
The flavone apigenin and the mycotoxin zearalenone are two major compounds found in the human diet which bind estrogen receptors (ERs), and therefore influence ER activity. However, the underlying mechanisms are not well known. To unravel the molecular mechanisms that could explain the differential effect of zearalenone and apigenin on ER-positive breast cancer cell proliferation, gene-reporter assays, chromatin immunoprecipitation (ChIP) experiments, proliferation assays and transcriptomic analysis were performed. We found that zearalenone and apigenin transactivated ERs and promoted the expression of estradiol (E2)-responsive genes. However, zearalenone clearly enhanced cellular proliferation, while apigenin appeared to be antiestrogenic in the presence of E2 in both ER-positive breast cancer cell lines, MCF-7 and T47D. The transcriptomic analysis showed that both compounds regulate gene expression in the same way, but with differences in intensity. Two major sets of genes were identified; one set was linked to the cell cycle and the other set was linked to stress response and growth arrest. Our results show that the transcription dynamics in gene regulation induced by apigenin were somehow different with zearalenone and E2 and may explain the differential effect of these compounds on the phenotype of the breast cancer cell. Together, our results confirmed the potential health benefit effect of apigenin, while zearalenone appeared to be a true endocrine-disrupting compound., Competing Interests: The authors declare they have no actual or potential competing financial interests.
- Published
- 2019
- Full Text
- View/download PDF
12. A Genetic Tool to Quantify trans-Translation Activity in Vivo.
- Author
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Macé K, Demay F, Guyomar C, Georgeault S, Giudice E, Goude R, Trautwetter A, Ermel G, Blanco C, and Gillet R
- Subjects
- Bacterial Proteins genetics, RNA, Bacterial genetics, RNA, Messenger genetics, RNA-Binding Proteins genetics, Ribosomes genetics, Bacteria metabolism, Bacterial Proteins metabolism, Protein Biosynthesis, RNA, Bacterial metabolism, RNA, Messenger metabolism, RNA-Binding Proteins metabolism, Ribosomes metabolism
- Abstract
In bacteria, trans-translation is the main quality control mechanism for rescuing ribosomes arrested during translation. This key process is universally conserved and plays a critical role in the viability and virulence of many pathogens. We developed a reliable in vivo double-fluorescence reporter system for the simultaneous quantification of both trans-translation and the associated proteolysis activities in bacteria. The assay was validated using mutant bacteria lacking tmRNA, SmpB, and the ClpP protease. Both antisense tmRNA-binding RNA and a peptide mimicking the SmpB C-terminal tail proved to be potent inhibitors of trans-translation in vivo. The double-fluorescent reporter was also tested with KKL-35, an oxadiazole derivative that is supposed to be a promising trans-translation inhibitor, and it surprisingly turns out that trans-translation is not the only target of KKL-35 in vivo., (Copyright © 2017 Elsevier Ltd. All rights reserved.)
- Published
- 2017
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13. Phytochemicals Targeting Estrogen Receptors: Beneficial Rather Than Adverse Effects?
- Author
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Lecomte S, Demay F, Ferrière F, and Pakdel F
- Subjects
- Animals, Epigenesis, Genetic genetics, Estrogen Antagonists pharmacology, Humans, Receptors, Estrogen antagonists & inhibitors, Signal Transduction drug effects, Signal Transduction genetics, Transcription, Genetic genetics, Phytochemicals pharmacology, Receptors, Estrogen metabolism
- Abstract
In mammals, the effects of estrogen are mainly mediated by two different estrogen receptors, ERα and ERβ. These proteins are members of the nuclear receptor family, characterized by distinct structural and functional domains, and participate in the regulation of different biological processes, including cell growth, survival and differentiation. The two estrogen receptor (ER) subtypes are generated from two distinct genes and have partially distinct expression patterns. Their activities are modulated differently by a range of natural and synthetic ligands. Some of these ligands show agonistic or antagonistic effects depending on ER subtype and are described as selective ER modulators (SERMs). Accordingly, a few phytochemicals, called phytoestrogens, which are synthesized from plants and vegetables, show low estrogenic activity or anti-estrogenic activity with potentially anti-proliferative effects that offer nutraceutical or pharmacological advantages. These compounds may be used as hormonal substitutes or as complements in breast cancer treatments. In this review, we discuss and summarize the in vitro and in vivo effects of certain phytoestrogens and their potential roles in the interaction with estrogen receptors., Competing Interests: The authors declare no conflict of interest.
- Published
- 2017
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14. Complementation of a manganese-dependent superoxide dismutase-deficient yeast strain with Pneumocystis carinii sod2 gene.
- Author
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Khalife S, Aliouat el M, Gantois N, Jakobczyk H, Demay F, Chabé M, Pottier M, Dabboussi F, Hamze M, Dei-Cas E, Standaert-Vitse A, and Aliouat-Denis CM
- Subjects
- Cloning, Molecular, Gene Expression, Saccharomyces cerevisiae chemistry, Genetic Complementation Test, Mitochondria enzymology, Pneumocystis carinii enzymology, Pneumocystis carinii genetics, Saccharomyces cerevisiae enzymology, Saccharomyces cerevisiae genetics, Superoxide Dismutase deficiency
- Abstract
Manganese-dependent superoxide dismutase (MnSOD) is one of the key enzymes involved in the cellular defense against oxidative stress. Previously, the Pneumocystis carinii sod2 gene (Pcsod2) was isolated and characterized. Based on protein sequence comparison, Pcsod2 was suggested to encode a putative MnSOD protein likely to be targeted into the mitochondrion. In this work, the Pcsod2 was cloned and expressed as a recombinant protein in EG110 Saccharomyces cerevisiae strain lacking the MnSOD-coding gene (Scsod2) in order to investigate the function and subcellular localization of P. carinii MnSOD (PcMnSOD). The Pcsod2 gene was amplified by PCR and cloned into the pYES2.1/V5-His-TOPO(®) expression vector. The recombinant construct was then transformed into EG110 strain. Once its expression had been induced, PcMnSOD was able to complement the growth defect of EG110 yeast cells that had been exposed to the redox-cycling compound menadione. N-term sequencing of the PcMnSOD protein allowed identifying the cleavage site of a mitochondrial targeting peptide. Immune-colocalization of PcMnSOD and yeast CoxIV further confirmed the mitochondrial localization of the PcMnSOD. Heterologous expression of PcMnSOD in yeast indicates that Pcsod2 encodes an active MnSOD, targeted to the yeast mitochondrion that allows the yeast cells to grow in the presence of reactive oxygen species (ROS)., (Copyright © 2014 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.)
- Published
- 2014
- Full Text
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15. The actin/MKL1 signalling pathway influences cell growth and gene expression through large-scale chromatin reorganization and histone post-translational modifications.
- Author
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Flouriot G, Huet G, Demay F, Pakdel F, Boujrad N, and Michel D
- Subjects
- Acetylation, Actins genetics, Cell Line, Tumor, Cell Proliferation, Chromatin chemistry, DNA-Binding Proteins genetics, Epithelial-Mesenchymal Transition genetics, Gene Expression, Histones genetics, Humans, Methylation, Oncogene Proteins, Fusion genetics, Signal Transduction, Trans-Activators, Actins metabolism, Chromatin Assembly and Disassembly, DNA-Binding Proteins metabolism, Histones metabolism, Oncogene Proteins, Fusion metabolism, Protein Processing, Post-Translational
- Abstract
In addition to soluble factors, mechanical constraints and extracellular matrix stiffness are important regulators of cell fate that are mediated by cytoskeletal modifications. The EMT (epithelial-mesenchymal transition) that occurs during normal development and malignant progression is a typical example of the phenotypic switch associated with profound actin remodelling and changes in gene expression. For instance, actin dynamics control motile cell functions in EMT, in part, through regulating the subcellular localization of the myocardin-related transcription factor MKL1 (megakaryoblastic leukaemia translocation 1), a co-activator of SRF (serum-responsive factor). In the present paper, we show that MKL1 participates also to the control of the cellular switch between growth and quiescence. Experimental disconnection between MKL1 and G-actin (globular actin), by using an MKL1 mutant or enhancing the F (filamentous)-/G-actin ratio, generates a widely open chromatin state and a global increase in biosynthetic activity, classically associated with cell growth. Conversely, G-actin accumulation favours nuclear condensation and cell quiescence. These large-scale chromatin changes rely upon extensive histone modifications, exemplified by that of H3K9 (H3 Lys9) shifting from trimethylation, a heterochromatin mark, to acetylation, a mark of euchromatin. The present study provides the first evidence for a global reversible hetero/euchromatinization phenomenon triggered by the actin/MKL1 signalling pathway.
- Published
- 2014
- Full Text
- View/download PDF
16. Activation of the MKL1/actin signaling pathway induces hormonal escape in estrogen-responsive breast cancer cell lines.
- Author
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Kerdivel G, Boudot A, Habauzit D, Percevault F, Demay F, Pakdel F, and Flouriot G
- Subjects
- Antineoplastic Agents, Hormonal pharmacology, Breast Neoplasms drug therapy, Drug Resistance, Neoplasm, Estradiol physiology, Estrogen Receptor alpha genetics, Estrogen Receptor alpha metabolism, Female, Humans, MCF-7 Cells, Neoplasms, Hormone-Dependent drug therapy, Neoplasms, Hormone-Dependent metabolism, Receptor, ErbB-2 genetics, Receptor, ErbB-2 metabolism, Receptors, Progesterone genetics, Receptors, Progesterone metabolism, Tamoxifen pharmacology, Trans-Activators, Transcription, Genetic, Actins metabolism, Breast Neoplasms metabolism, DNA-Binding Proteins metabolism, Oncogene Proteins, Fusion metabolism, Signal Transduction
- Abstract
Estrogen receptor alpha (ERα) is generally considered to be a good prognostic marker because almost 70% of ERα-positive tumors respond to anti-hormone therapies. Unfortunately, during cancer progression, mammary tumors can escape from estrogen control, resulting in resistance to treatment. In this study, we demonstrate that activation of the actin/megakaryoblastic leukemia 1 (MKL1) signaling pathway promotes the hormonal escape of estrogen-sensitive breast cancer cell lines. The actin/MKL1 signaling pathway is silenced in differentiated ERα-positive breast cancer MCF-7 and T47D cell lines and active in ERα-negative HMT-3522 T4-2 and MDA-MB-231 breast cancer cells, which have undergone epithelial-mesenchymal transition. We showed that MKL1 activation in MCF-7 cells, either by modulating actin dynamics or using MKL1 mutants, down-regulates ERα expression and abolishes E2-dependent cell growth. Interestingly, the constitutively active form of MKL1 represses PR and HER2 expression in these cells and increases the expression of HB-EGF, TGFβ, and amphiregulin growth factors in an E2-independent manner. The resulting expression profile (ER-, PR-, HER2-) typically corresponds to the triple-negative breast cancer expression profile., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)
- Published
- 2014
- Full Text
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17. Epigenetic memories: structural marks or active circuits?
- Author
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Nicol-Benoît F, Le-Goff P, Le-Dréan Y, Demay F, Pakdel F, Flouriot G, and Michel D
- Subjects
- Cybernetics, Biological Evolution, Chromatin Assembly and Disassembly physiology, Epigenesis, Genetic physiology, Gene Expression Regulation physiology, Gene Regulatory Networks physiology, Models, Genetic
- Abstract
A hallmark of living systems is the management and the storage of information through genetic and epigenetic mechanisms. Although the notion of epigenetics was originally given to any regulation beyond DNA sequence, it has often been restricted to chromatin modifications, supposed to behave as cis-markers, specifying the sets of genes to be expressed or repressed. This definition does not take into account the initial view of epigenetics, based on nonlinear interaction networks whose "attractors" can remain stable without need for any chromatin mark. In addition, most chromatin modifications are the steady state resultants of highly dynamic modification and de-modification activities and, as such, seem poorly appropriate to work as long-term memory keepers. Instead, the basic support of epigenetic memory could remain the attractors, to which chromatin modifications belong as do many other components. The influence of chromatin modifications in memory is highly questionable when envisioned as static structural marks, but can be recovered under the dynamic circuitry perspective, thanks to their self-templating properties. Beside their standard repressive or permissive functions, chromatin modifications can also influence transcription in multiple ways such as: (1) by randomizing or inversely stabilizing gene expression, (2) by mediating cooperativity between pioneer and secondary transcription factors, and (3) in the hysteresis and the ultrasensitivity of gene expression switches, allowing the cells to take unambiguous transcriptional decisions.
- Published
- 2012
- Full Text
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18. Epigenetic switch involved in activation of pioneer factor FOXA1-dependent enhancers.
- Author
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Sérandour AA, Avner S, Percevault F, Demay F, Bizot M, Lucchetti-Miganeh C, Barloy-Hubler F, Brown M, Lupien M, Métivier R, Salbert G, and Eeckhoute J
- Subjects
- Animals, Binding Sites genetics, Cell Differentiation genetics, Cell Line, Tumor, Chromatin metabolism, DNA Methylation genetics, DNA-Binding Proteins genetics, DNA-Binding Proteins metabolism, Histones metabolism, Humans, Mice, Models, Genetic, Neurons cytology, Neurons metabolism, Enhancer Elements, Genetic, Epigenomics, Hepatocyte Nuclear Factor 3-alpha genetics, Hepatocyte Nuclear Factor 3-alpha metabolism
- Abstract
Transcription factors (TFs) bind specifically to discrete regions of mammalian genomes called cis-regulatory elements. Among those are enhancers, which play key roles in regulation of gene expression during development and differentiation. Despite the recognized central regulatory role exerted by chromatin in control of TF functions, much remains to be learned regarding the chromatin structure of enhancers and how it is established. Here, we have analyzed on a genomic-scale enhancers that recruit FOXA1, a pioneer transcription factor that triggers transcriptional competency of these cis-regulatory sites. Importantly, we found that FOXA1 binds to genomic regions showing local DNA hypomethylation and that its cell-type-specific recruitment to chromatin is linked to differential DNA methylation levels of its binding sites. Using neural differentiation as a model, we showed that induction of FOXA1 expression and its subsequent recruitment to enhancers is associated with DNA demethylation. Concomitantly, histone H3 lysine 4 methylation is induced at these enhancers. These epigenetic changes may both stabilize FOXA1 binding and allow for subsequent recruitment of transcriptional regulatory effectors. Interestingly, when cloned into reporter constructs, FOXA1-dependent enhancers were able to recapitulate their cell type specificity. However, their activities were inhibited by DNA methylation. Hence, these enhancers are intrinsic cell-type-specific regulatory regions of which activities have to be potentiated by FOXA1 through induction of an epigenetic switch that includes notably DNA demethylation.
- Published
- 2011
- Full Text
- View/download PDF
19. Biological and biophysical properties of the histone deacetylase inhibitor suberoylanilide hydroxamic acid are affected by the presence of short alkyl groups on the phenyl ring.
- Author
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Oger F, Lecorgne A, Sala E, Nardese V, Demay F, Chevance S, Desravines DC, Aleksandrova N, Le Guével R, Lorenzi S, Beccari AR, Barath P, Hart DJ, Bondon A, Carettoni D, Simonneaux G, and Salbert G
- Subjects
- Blotting, Western, Caco-2 Cells, Cell Growth Processes drug effects, Enzyme-Linked Immunosorbent Assay, Hep G2 Cells, Histone Deacetylase Inhibitors chemical synthesis, Histone Deacetylases chemistry, Histone Deacetylases metabolism, Humans, Hydroxamic Acids chemical synthesis, Magnetic Resonance Spectroscopy, Mass Spectrometry, Microscopy, Fluorescence, Models, Molecular, Structure-Activity Relationship, Vorinostat, Histone Deacetylase Inhibitors chemistry, Histone Deacetylase Inhibitors pharmacology, Hydroxamic Acids chemistry, Hydroxamic Acids pharmacology
- Abstract
Inhibition of histone deacetylases (HDACs) leads to growth arrest, differentiation, or apoptosis of tumor cell lines, suggesting HDACs as promising targets for cancer therapy. At present, only one HDAC inhibitor (HDACi) is used in therapy: suberoylanilide hydroxamic acid (SAHA). Here, we describe the synthesis and biological evaluation of a new series of compounds derived from SAHA by substituting short alkyl chains at various positions of the phenyl ring. Such modifications induced variable effects ranging from partial loss of activity to increased potency. Through molecular modeling, we describe a possible interaction between HDAC7 proline 809, a residue that is strictly conserved within class 2 enzymes only, and the amide group of HDACi, while nuclear magnetic resonance experiments indicated that dimethyl m-substitution may stabilize the inhibitor in the active site. Our data provide novel information on the structure-activity relationship of HDACi and suggest new ways for developing second generation SAHA-like molecules.
- Published
- 2010
- Full Text
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20. Cyclical DNA methylation of a transcriptionally active promoter.
- Author
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Métivier R, Gallais R, Tiffoche C, Le Péron C, Jurkowska RZ, Carmouche RP, Ibberson D, Barath P, Demay F, Reid G, Benes V, Jeltsch A, Gannon F, and Salbert G
- Subjects
- Cell Line, Chromatin Immunoprecipitation, CpG Islands genetics, DNA (Cytosine-5-)-Methyltransferases antagonists & inhibitors, DNA (Cytosine-5-)-Methyltransferases metabolism, DNA Repair, Deamination, Estrogens pharmacology, Humans, Kinetics, Thymine DNA Glycosylase metabolism, Transcription, Genetic drug effects, Transcriptional Activation drug effects, Trefoil Factor-1, DNA Methylation drug effects, Gene Expression Regulation drug effects, Promoter Regions, Genetic genetics, Transcription, Genetic genetics, Transcriptional Activation genetics, Tumor Suppressor Proteins genetics
- Abstract
Processes that regulate gene transcription are directly under the influence of the genome organization. The epigenome contains additional information that is not brought by DNA sequence, and generates spatial and functional constraints that complement genetic instructions. DNA methylation on CpGs constitutes an epigenetic mark generally correlated with transcriptionally silent condensed chromatin. Replication of methylation patterns by DNA methyltransferases maintains genome stability through cell division. Here we present evidence of an unanticipated dynamic role for DNA methylation in gene regulation in human cells. Periodic, strand-specific methylation/demethylation occurs during transcriptional cycling of the pS2/TFF1 gene promoter on activation by oestrogens. DNA methyltransferases exhibit dual actions during these cycles, being involved in CpG methylation and active demethylation of 5mCpGs through deamination. Inhibition of this process precludes demethylation of the pS2 gene promoter and its subsequent transcriptional activation. Cyclical changes in the methylation status of promoter CpGs may thus represent a critical event in transcriptional achievement.
- Published
- 2008
- Full Text
- View/download PDF
21. Dynamics of estrogen receptor-mediated transcriptional activation of responsive genes in vivo: apprehending transcription in four dimensions.
- Author
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Métivier R, Huet G, Gallais R, Finot L, Petit F, Tiffoche C, Mérot Y, LePéron C, Reid G, Penot G, Demay F, Gannon F, Flouriot G, and Salbert G
- Subjects
- Gene Expression Regulation, Humans, Promoter Regions, Genetic genetics, Receptors, Estrogen physiology, Transcriptional Activation
- Published
- 2008
- Full Text
- View/download PDF
22. Deoxyribonucleic acid methyl transferases 3a and 3b associate with the nuclear orphan receptor COUP-TFI during gene activation.
- Author
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Gallais R, Demay F, Barath P, Finot L, Jurkowska R, Le Guével R, Gay F, Jeltsch A, Métivier R, and Salbert G
- Subjects
- Animals, COS Cells, Chlorocebus aethiops, DNA Methyltransferase 3A, Mice, Promoter Regions, Genetic, Transcriptional Activation, Vitronectin biosynthesis, Vitronectin genetics, DNA Methyltransferase 3B, COUP Transcription Factor I metabolism, DNA (Cytosine-5-)-Methyltransferases metabolism, Gene Expression Regulation physiology
- Abstract
Transcriptional activation of silent genes can require the erasure of epigenetic marks such as DNA methylation at CpGs (cytosine-guanine dinucleotide). Active demethylation events have been observed, and associated processes are repeatedly suspected to involve DNA glycosylases such as mCpG binding domain protein 4, thymine DNA glycosylase (TDG), Demeter, and repressor of silencing 1. A complete characterization of the molecular mechanisms occurring in metazoan is nonetheless awaited. Here, we report that activation of the endogenous vitronectin gene in P19 cells by the nuclear receptor chicken ovalbumin upstream promoter-transcription factor I (COUP-TFI) is observed in parallel with the recruitment of TDG and p68 RNA helicase, two components of a putative demethylation complex. Interestingly, when activated, the vitronectin gene was loaded with DNA methyltransferases 3a and 3b (Dnmt3a/b), and a strand-biased decrease in CpG methylation was detected. Dnmt3a was further found to associate with COUP-TFI and TDG in vivo, and cotransfection experiments demonstrated that Dnmt3a/b can enhance COUP-TFI-mediated activation of a methylated reporter gene. These results suggest that Dnmt3a/b could cooperate with the orphan receptor COUP-TFI to regulate transcription of the vitronectin gene.
- Published
- 2007
- Full Text
- View/download PDF
23. T-box factors: targeting to chromatin and interaction with the histone H3 N-terminal tail.
- Author
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Demay F, Bilican B, Rodriguez M, Carreira S, Pontecorvi M, Ling Y, and Goding CR
- Subjects
- Animals, COS Cells, Chlorocebus aethiops, DNA metabolism, HeLa Cells, Heterochromatin metabolism, Humans, Mitosis, Nucleosomes metabolism, Protein Binding, Protein Transport, Chromatin metabolism, Histones chemistry, Histones metabolism, T-Box Domain Proteins metabolism
- Abstract
T-box transcription factors play a crucial role in development where they are implicated in patterning and cell fate decisions. Tbx2 and Tbx3 have also been implicated in several cancers including melanoma, and can act as antisenescence factors through their ability to repress p19(ARF) and p21(CIP1) expression. Although several target genes for T-box factors have been identified, it is unknown whether this family of proteins can bind chromatin, a property that would facilitate the epigenetic reprogramming that occurs in both development and cancer progression. Here, we show that Tbx2 has the potential to recognize mitotic chromatin in a DNA-dependent fashion, can interact specifically with the histone H3 N-terminal tail, a property shared with Tbx4, Tbx5 and Tbx6, and can also recognize nucleosomal DNA, with binding to nucleosomes being antagonized by the presence of the histone tails. Strikingly, in vivo Tbx2 co-localization with pericentric heterochromatin appears to be regulated and ectopic expression of Tbx2 leads to severe mitotic defects. Taken together our results suggest that Tbx2, and most likely other members of the T-box family, are able to target chromatin and may indicate a role for the T-box factors in epigenetic reprogramming events.
- Published
- 2007
- Full Text
- View/download PDF
24. A neural-specific splicing event generates an active form of the Wiskott-Aldrich syndrome protein.
- Author
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Le Page Y, Demay F, and Salbert G
- Subjects
- Actins metabolism, Animals, COS Cells, COUP Transcription Factor I, Chlorocebus aethiops, DNA-Binding Proteins metabolism, Exons physiology, Genes, Reporter, Mutation, PC12 Cells, Protein Isoforms genetics, Protein Isoforms metabolism, Proteins metabolism, Rats, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins metabolism, Transcription Factors metabolism, Wiskott-Aldrich Syndrome Protein, Alternative Splicing physiology, Neurons metabolism, Proteins genetics
- Abstract
Actin polymerization is required for cellular events such as podosome, lamellipode or filopode formation in migrating cells, and members of the Wiskott-Aldrich syndrome protein (WASP) family have essential roles in regulating actin dynamics at the cell leading edge. However, WASP proteins need first to be activated in order to be able to target actin polymerization. Here, we show the occurrence of a neural-specific splicing event, which is favoured by the nuclear orphan receptor chicken ovalbumin upstream promoter-transcription factor I, and generates a truncated WASP protein deleted of exon 2-encoded amino acids. This deletion relocates the protein to the plasma membrane and induces the formation of actin-rich podosome-like structures that also contain paxillin and vinculin. Furthermore, expression of the truncated protein in PC12 cells, as well as in primary neurons, stimulates neuritogenesis. These data underscore the importance of the neural-specific splicing of WASP RNA during development.
- Published
- 2004
- Full Text
- View/download PDF
25. Leukemic transformation by the v-ErbA oncoprotein entails constitutive binding to and repression of an erythroid enhancer in vivo.
- Author
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Ciana P, Braliou GG, Demay FG, von Lindern M, Barettino D, Beug H, and Stunnenberg HG
- Subjects
- Alpharetrovirus, Animals, Base Sequence, Carbonic Anhydrases biosynthesis, Cell Differentiation, DNA Footprinting, Enhancer Elements, Genetic, Erythropoiesis, Gene Expression Regulation, Neoplastic, Histone Deacetylase Inhibitors, Hydroxamic Acids pharmacology, Introns, Molecular Sequence Data, Protein Binding, Receptors, Thyroid Hormone metabolism, Carbonic Anhydrases genetics, Cell Transformation, Neoplastic, Leukemia genetics, Oncogene Proteins v-erbA metabolism, Response Elements
- Abstract
v-ErbA, a mutated thyroid hormone receptor alpha (TRalpha), is thought to contribute to avian erythroblastosis virus (AEV)-induced leukemic transformation by constitutively repressing transcription of target genes. However, the binding of v-ErbA or any unliganded nuclear receptor to a chromatin-embedded response element as well as the role of the N-CoR-SMRT-HDAC co-repressor complex in mediating repression remain hypothetical. Here we identify a v-ErbA-response element, VRE, in an intronic DNase I hypersensitive site (HS2) of the chicken erythroid carbonic anhydrase II (CAII) gene. In vivo footprinting shows that v-ErbA is constitutively bound to this HS2-VRE in transformed, undifferentiated erythroblasts along with other transcription factors like GATA-1. Transfection assays show that the repressed HS2 region can be turned into a potent enhancer in v-ErbA-expressing cells by mutation of the VRE. Differentiation of transformed cells alleviates v-ErbA binding concomitant with activation of CAII transcription. Co-expression of a gag-TRalpha fusion protein in AEV-transformed cells and addition of ligand derepresses CAII transcription. Treatment of transformed cells with the histone deacetylase inhibitor, trichostatin A, derepresses the endogenous, chromatin-embedded CAII gene, while a transfected HS2-enhancer construct remains repressed. Taken together, our data suggest that v-ErbA prevents CAII activation by 'neutralizing' in cis the activity of erythroid transcription factors.
- Published
- 1998
- Full Text
- View/download PDF
26. In vitro induction of specific cytotoxic T lymphocytes using recombinant single-chain MHC class I/peptide complexes.
- Author
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Lone YC, Motta I, Mottez E, Guilloux Y, Lim A, Demay F, Levraud JP, Kourilsky P, and Abastado JP
- Subjects
- Animals, Cells, Cultured, Gene Rearrangement, T-Lymphocyte, HLA-A2 Antigen chemistry, Humans, Male, Mice, Mice, Inbred DBA, Rats, Recombinant Proteins immunology, HLA-A2 Antigen immunology, T-Lymphocytes, Cytotoxic immunology
- Abstract
We have previously described the production and purification of a murine single-chain, soluble recombinant major histocompatibility complex (MHC) class I molecule (SC-Kd). A similar strategy was devised to produce a recombinant HLA-A2.1 (SC-A2) molecule. The latter was composed of the first three domains of the HLA-A2.1 heavy chain connected to human beta 2-microglobulin through a spacer of 15 amino acids. Immunoaffinity-purified SC-A2 molecules-were correctly folded and biologically functional. They specifically bound HLA-A2-restricted peptides and induced a peptide-specific cytotoxic T lymphocyte (CTL) clone to proliferate and secrete interleukin-2. The ability of murine and human SC-MHC molecules to elicit primary CTLs in vitro was next investigated. When coated in high density onto beads, complexes of antigenic peptide and SC-Kd or SC-A2 molecules efficiently induced a specific primary CTL response in vitro. Furthermore, the structural features of these CTLs were characterized by T cell receptor-beta chain analysis, which revealed rearrangements very similar, if not identical, to those found in CTLs generated by in vivo immunization. Such single-chain, soluble recombinant MHC class I molecules should provide a useful tool in particular for peptide binding assays and for in vitro primary CTL induction to identify immunogenic peptides such as those derived from known tumor-associated antigens.
- Published
- 1998
- Full Text
- View/download PDF
27. Sex- and cell-specific expression of an estrogen receptor isoform in the pituitary gland.
- Author
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Demay F, Tiffoche C, and Thieulant ML
- Subjects
- Animals, Blotting, Northern, Female, Male, RNA, Messenger metabolism, Rats, Rats, Wistar, Sex Factors, Pituitary Gland metabolism, Receptors, Estrogen metabolism, Sex Distribution
- Abstract
The presence of multiple monomeric forms has been described for the estrogen receptor (ER) in the pituitary gland. We analyzed ER mRNA forms in male and female rat pituitary. A single 6.2-kb ER mRNA species was detected in the male rat pituitary, whereas the female rat pituitary exhibited two ER mRNA forms of 6.2 and 5.5 kb, respectively. The 6.2-kb mRNA was present throughout the different stages of the estrous cycle, while the 5.5-kb mRNA appeared to be restricted to proestrus, suggesting an acute regulation of ER transcription at this stage. The 5.5-kb ER mRNA could be rapidly induced either by 17 beta-estradiol replacement in ovariectomized adult female rats or by priming immature rats with pregnant-mare serum gonadotropin. Using enriched cell populations, an inverse and strong correlation was established between the presence of the 5.5-kb ER mRNA form and the number of gonadotropes. Conversely, the localization of the 5.5-kb mRNA form was demonstrated in lactotrope populations. In order to elucidate the structural modifications in the transiently expressed ER mRNA, a series of reverse-transcriptase polymerase chain reaction amplifications was carried out using several pairs of primers corresponding to the entire ER-coding region. The data showed that no alternative splicing was occurring in the ER-coding region involving a potential role of either 3'- or 5'-untranslated regions. Thus, ER presents a 17 beta-estradiol-dependent transcriptional mechanism triggered on proestrous day and specific to the female lactotropes.
- Published
- 1996
- Full Text
- View/download PDF
28. [Hypoglycemia and mesenchymal tumors].
- Author
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Verdy M, Cholette JP, Nadeau P, Fauteux JP, and Demay F
- Subjects
- Glucose Tolerance Test, Humans, Insulin analysis, Male, Mesothelioma metabolism, Middle Aged, Radioimmunoassay, Hypoglycemia etiology, Mesothelioma complications, Pleural Neoplasms complications, Retroperitoneal Neoplasms complications
- Published
- 1969
29. Ocular anomalies in de Lange syndrome.
- Author
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Milot J and Demay F
- Subjects
- Adolescent, Blepharoptosis complications, Child, De Lange Syndrome diagnosis, Eyebrows abnormalities, Eyelashes abnormalities, Eyelids abnormalities, Female, Humans, Infant, Male, Myopia complications, Nystagmus, Pathologic complications, Pigmentation Disorders complications, Sclera, Strabismus complications, De Lange Syndrome complications, Eye Abnormalities, Eye Diseases complications
- Published
- 1972
- Full Text
- View/download PDF
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