Descriptor: "MESH : Cell Line" - Searchworks@Jio Institute Digital Library Search Results

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85 results on '"MESH : Cell Line"'

1. Endophilin-A2 functions in membrane scission in clathrin-independent endocytosis

Author: Anne K. Kenworthy, Mijo Simunovic, Anne A. Schmidt, Valérie Chambon, Patricia Bassereau, Henri-François Renard, Christian Wunder, Cécile Sykes, Joël Lemière, Emmanuel Boucrot, Christophe Lamaze, Harvey T. McMahon, Maria Daniela Garcia-Castillo, Senthil Arumugam, Ludger Johannes, Bondidier, Martine, Chimie biologique des membranes et ciblage thérapeutique ( CBMCT - UMR 3666 / U1143 ), Université Paris Descartes - Paris 5 ( UPD5 ) -Institut Curie-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Physico-Chimie-Curie ( PCC ), Centre National de la Recherche Scientifique ( CNRS ) -INSTITUT CURIE-Université Pierre et Marie Curie - Paris 6 ( UPMC ), Department of Chemistry, University of Chicago, Université Paris Diderot - Paris 7 ( UPD7 ), Institute of Structural and Molecular Biology, Birkbeck College, Department of Molecular Physiology and Biophysics, Vanderbilt University School of Medicine. Nashville, Vanderbilt University School of Medicine. Nashville-Vanderbilt University School of Medicine. Nashville, Institut Jacques Monod ( IJM ), Université Paris Diderot - Paris 7 ( UPD7 ) -Centre National de la Recherche Scientifique ( CNRS ), Laboratory of Molecular Biology, Division of Cell Biology, Medical Research Council Laboratory of Molecular Biology, Cambridge CB2 0QH, UK, Agence Nationale pour la Recherche : (ANR-09-BLAN-283, ANR-10-LBX-0038, ANR-11 BSV2 014 03, ANR-12-BSV5-0014), Indo-French Centre for the Promotion of Advanced Science (project no. 3803), Marie Curie Actions — Networks for Initial Training (FP7-PEOPLE-2010-ITN), Marie Curie International Reintegration Grant (FP7-RG-277078), European Research Council advanced grant (project 340485), Royal Society (RG120481), Fondation ARC pour la Recherche sur le Cancer (DEQ20120323737), National Institutes of Health (RO1 GM106720), Ligue contre le Cancer, Comité de Paris (RS08/75-89), Fondation ARC pour la Recherche sur le Cancer, AXA Research Funds, Biological Sciences Research Council, Chateaubriand fellowship, France and Chicago Collaborating in the Sciences grant, Chimie biologique des membranes et ciblage thérapeutique (CBMCT - UMR 3666 / U1143), Université Paris Descartes - Paris 5 (UPD5)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Physico-Chimie-Curie (PCC), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut Curie [Paris]-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Université Paris Diderot - Paris 7 (UPD7), Vanderbilt University School of Medicine [Nashville], Institut Jacques Monod (IJM (UMR_7592)), Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS), Université Paris Descartes - Paris 5 (UPD5)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC), and Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de Chimie du CNRS (INC)
Subjects: MESH : Cell Line, MESH: Rats, MESH : Endocytosis, MESH : Cell Membrane, MESH: Shiga Toxin, Endocytic cycle, MESH : Actins, MESH : Dynamins, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, macromolecular substances, MESH: Acyltransferases, Biology, MESH: Actins, Endocytosis, environment and public health, Clathrin, Cell membrane, 03 medical and health sciences, 0302 clinical medicine, medicine, BAR domain, MESH: Animals, MESH: Clathrin, Endophilin-A2, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, MESH: Cholera Toxin, 030304 developmental biology, Dynamin, 0303 health sciences, MESH: Humans, MESH : Clathrin, Multidisciplinary, MESH : Rats, MESH : Cholera Toxin, [ SDV.BC ] Life Sciences [q-bio]/Cellular Biology, MESH : Humans, MESH: Cell Line, Cell biology, MESH: Dynamins, medicine.anatomical_structure, MESH : Shiga Toxin, MESH: Endocytosis, Amphiphysin, biology.protein, MESH : Animals, MESH : Acyltransferases, 030217 neurology & neurosurgery, MESH: Cell Membrane
Abstract: International audience; During endocytosis, energy is invested to narrow the necks of cargo-containing plasma membrane invaginations to radii at which the opposing segments spontaneously coalesce, thereby leading to the detachment by scission of endocytic uptake carriers. In the clathrin pathway, dynamin uses mechanical energy from GTP hydrolysis to this effect, assisted by the BIN/amphiphysin/Rvs (BAR) domain-containing protein endophilin. Clathrin-independent endocytic events are often less reliant on dynamin, and whether in these cases BAR domain proteins such as endophilin contribute to scission has remained unexplored. Here we show, in human and other mammalian cell lines, that endophilin-A2 (endoA2) specifically and functionally associates with very early uptake structures that are induced by the bacterial Shiga and cholera toxins, which are both clathrin-independent endocytic cargoes. In controlled in vitro systems, endoA2 reshapes membranes before scission. Furthermore, we demonstrate that endoA2, dynamin and actin contribute in parallel to the scission of Shiga-toxin-induced tubules. Our results establish a novel function of endoA2 in clathrin-independent endocytosis. They document that distinct scission factors operate in an additive manner, and predict that specificity within a given uptake process arises from defined combinations of universal modules. Our findings highlight a previously unnoticed link between membrane scaffolding by endoA2 and pulling-force-driven dynamic scission.
Published: 2014
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2. Flow Cytometric Analysis of the Expression Pattern of Peroxisomal Proteins, Abcd1, Abcd2, and Abcd3 in BV-2 Murine Microglial Cells

Author: Debbabi, Meryam, Nury, Thomas, Helali, Imen, Karym, El Mostafa, Geillon, Flore, Gondcaille, Catherine, Trompier, Doriane, Najid, Amina, Terreau, Sébastien, Bezine, Maryem, Zarrouk, Amira, Vejux, Anne, ANDREOLETTI, Pierre, Cherkaoui-Malki, Mustapha, Savary, Stéphane, Lizard, Gérard, Nutrition, Aliments Fonctionnels et Santé Vasculaire, Université de Monastir ( UM ), Laboratoire Bio-PeroxIL. Biochimie du peroxysome, inflammation et métabolisme lipidique [Dijon] ( BIO-PEROXIL ), Université de Bourgogne ( UB ) -Université Bourgogne Franche-Comté ( UBFC ), Laboratoire des Maladies Transmissibles et Substances Biologiquement Actives (LR99ES27), Université de Monastir, Faculté de pharmacie [Monastir] ( FPHM ), Laboratoire Bio-PeroxIL. Biochimie du peroxysome, inflammation et métabolisme lipidique [Dijon] (BIO-PEROXIL), Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Bourgogne Franche-Comté [COMUE] (UBFC), Faculté de Médecine de Monastir [Tunisie], Laboratoire des Maladies Transmissibles et Substances Biologiquement Actives [Monastir] (LR99ES27), Faculté de Pharmacie [Monastir] (FPHM)-Université de Monastir - University of Monastir (UM), Laboratoire des Venins et Biomolécules Thérapeutiques - Laboratory of Venoms and Therapeutic Biomolecules (LR11IPT08), Institut Pasteur de Tunis, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP), Université de Tunis El Manar (UTM), This work was supported by grants from the INSERM, the Université de Bourgogne, the European Leucodystrophies Association (ELA, and project number: 2010-03014), the Conseil Régional de Bourgogne, the Action Intégrée of the Comité Mixte Inter-universitaire Franco-Marocain (CMIFM, AIMA/14/310, EGIDE) from the PHC Volubilis/Toubkal program, Ministère des Affaires Etrangères, the Ministère de l’Enseignement et de la Recherche, and the Projet Sectoriel CNRST.
Subjects: MESH : Cell Line, MESH: Gene Expression, MESH : Cloning, Molecular, MESH : Flow Cytometry, MESH : Microscopy, Fluorescence, Gene Expression, MESH: Flow Cytometry, MESH: Microscopy, Fluorescence, MESH : Microglia, Cell Line, Mice, MESH : Mice, Peroxisomes, Animals, MESH: Animals, MESH: Cloning, Molecular, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH : ATP-Binding Cassette Transporters, Cloning, Molecular, MESH: Mice, [ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology, Flow cytometric analysis, Peroxisomal ABC transporters, Flow Cytometry, MESH: Peroxisomes, MESH: Cell Line, MESH: Microglia, MESH : Gene Expression, Microscopy, Fluorescence, MESH: ATP-Binding Cassette Transporters, ATP-Binding Cassette Transporters, Microglia, BV-2 microglial cells, MESH : Animals, MESH : Peroxisomes
Abstract: International audience; Microglial cells play important roles in neurodegenerative diseases including peroxisomal leukodystrophies. The BV-2 murine immortalized cells are widely used in the context of neurodegenerative researches. It is therefore important to establish the expression pattern of peroxisomal proteins by flow cytometry in these cells. So, the expression pattern of various peroxisomal transporters (Abcd1, Abcd2, Abcd3) contributing to peroxisomal β-oxidation was evaluated on BV-2 cells by flow cytometry and complementary methods (fluorescence microscopy, and RT-qPCR). By flow cytometry a strong expression of peroxisomal proteins (Abcd1, Abcd2, Abcd3) was observed. These data were in agreement with those obtained by fluorescence microscopy (presence of numerous fluorescent dots in the cytoplasm characteristic of a peroxisomal staining pattern) and RT-qPCR (high levels of Abcd1, Abcd2, and Abcd3 mRNAs). Thus, the peroxisomal proteins (Abcd1, Abcd2, Abcd3) are expressed in BV-2 cells, and can be analyzed by flow cytometry.
Published: 2017
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3. De novo mutations in SMCHD1 cause Bosma arhinia microphthalmia syndrome and abrogate nasal development

Author: Duangrurdee Wattanasirichaigoon, Frédérique Magdinier, Axel M. Hillmer, Mung Kei Kong, Sabine Sigaudy, Myriam Oufadem, Hicham Filali, Hülya Kayserili, Christopher T. Gordon, Denise Williams, Peter Nürnberg, Carine Bonnard, Stanislas Lyonnet, Camille Dion, Siham Chafai Elalaoui, Jeanne Amiel, Audrey S.M. Teo, Nobuhiko Okamoto, Bruno Reversade, Asif Javed, Dieter Meschede, Alex Magee, Abdelaziz Sefiani, Rachel E A Irving, Alexandra D. Gurzau, James M. Murphy, Nicola K. Ragge, Bernd Wollnik, Gökhan Tunçbilek, Koh-ichiro Yoshiura, Chalermpong Chatdokmaiprai, Tamara Beck, Hallvard Reigstad, Christine Bole-Feysot, Kelan Chen, Nicolas Lévy, Michael L. Cunningham, Nadine Rosin, Ruth McGowan, Holger Thiele, Janine Altmüller, Vinod Varghese, Nawfal Fejjal, Patrick Nitschké, Ilham Ratbi, Marnie E. Blewitt, Gökhan Yigit, Wolfgang Mühlbauer, Meriem Fikri, Shifeng Xue, S Faisal Ahmed, Imagine - Institut des maladies génétiques ( IMAGINE - U1163 ), Centre National de la Recherche Scientifique ( CNRS ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Université Paris Descartes - Paris 5 ( UPD5 ), Department of Human Genetics, Nagasaki University Graduate School of Biomedical Sciences, Department of Clinical Genetics, Leicester Royal Infirmary, Universität zu Köln, Génétique Médicale et Génomique Fonctionnelle ( GMGF ), Aix Marseille Université ( AMU ) -Assistance Publique - Hôpitaux de Marseille ( APHM ) - Hôpital de la Timone [CHU - APHM] ( TIMONE ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Département de génétique médicale [Hôpital de la Timone - APHM], Aix Marseille Université ( AMU ) -Assistance Publique - Hôpitaux de Marseille ( APHM ) - Hôpital de la Timone [CHU - APHM] ( TIMONE ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Plate Forme Paris Descartes de Bioinformatique ( BIP-D ), Université Paris Descartes - Paris 5 ( UPD5 ), Department of Medical Genetics, Istanbul University, Génétique et épigénétique des maladies métaboliques, neurosensorielles et du développement ( Inserm U781 ), Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), ACS - Amsterdam Cardiovascular Sciences, ARD - Amsterdam Reproduction and Development, Center for Reproductive Medicine, ACS - Heart failure & arrhythmias, ACS - Diabetes & metabolism, Imagine - Institut des maladies génétiques (IMAGINE - U1163), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Nagasaki University, University Hospitals Leicester-University Hospitals Leicester, Génétique Médicale et Génomique Fonctionnelle (GMGF), Aix Marseille Université (AMU)-Assistance Publique - Hôpitaux de Marseille (APHM)- Hôpital de la Timone [CHU - APHM] (TIMONE)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Aix Marseille Université (AMU)-Assistance Publique - Hôpitaux de Marseille (APHM)- Hôpital de la Timone [CHU - APHM] (TIMONE)-Institut National de la Santé et de la Recherche Médicale (INSERM), Plate Forme Paris Descartes de Bioinformatique (BIP-D), Université Paris Descartes - Paris 5 (UPD5), Génétique et épigénétique des maladies métaboliques, neurosensorielles et du développement (Inserm U781), Universität zu Köln = University of Cologne, and Roche, Stephane
Subjects: Male, 0301 basic medicine, MESH : Cell Line, Embryology, Chromosomal Proteins, Non-Histone, Xenopus, MESH : Child, Preschool, medicine.disease_cause, Epigenesis, Genetic, MESH: Choanal Atresia, MESH : Microphthalmos, Mice, Xenopus laevis, 0302 clinical medicine, MESH : Xenopus laevis, Microphthalmos, Facioscapulohumeral muscular dystrophy, Missense mutation, MESH: Animals, MESH : Female, MESH: Epigenesis, Genetic, MESH : Epigenesis, Genetic, Muscular dystrophy, [ SDV.GEN.GH ] Life Sciences [q-bio]/Genetics/Human genetics, nasal development, Genetics, Mutation, biology, Medical genetics, MESH: Genetic Predisposition to Disease, [SDV.BDD.EO] Life Sciences [q-bio]/Development Biology/Embryology and Organogenesis, MESH: Nose, food and beverages, Muscular Dystrophy, Facioscapulohumeral, 3. Good health, MESH : Muscular Dystrophy, Facioscapulohumeral, Genetic linkage study, Child, Preschool, MESH : Choanal Atresia, Female, Epigenetics, MESH : Nose, MESH : Chromosomal Proteins, Non-Histone, medicine.medical_specialty, MESH: Microphthalmos, MESH : Male, Mutation, Missense, MESH: Muscular Dystrophy, Facioscapulohumeral, MESH : Mice, Inbred C57BL, [SDV.GEN.GH] Life Sciences [q-bio]/Genetics/Human genetics, Nose, Choanal Atresia, Cell Line, 03 medical and health sciences, MESH: Mice, Inbred C57BL, MESH: Xenopus laevis, MESH: Chromosomal Proteins, Non-Histone, MESH : Mice, medicine, Bosma arhinia microphthalmia syndrome, Animals, Humans, Genetic Predisposition to Disease, Allele, MESH: Mice, MESH: Mutation, Missense, FSHD, MESH: Humans, SMCHD1, MESH: Child, Preschool, MESH : Humans, medicine.disease, biology.organism_classification, MESH: Male, MESH: Cell Line, Mice, Inbred C57BL, [SDV.BDD.EO]Life Sciences [q-bio]/Development Biology/Embryology and Organogenesis, 030104 developmental biology, [SDV.GEN.GH]Life Sciences [q-bio]/Genetics/Human genetics, [ SDV.BDD.EO ] Life Sciences [q-bio]/Development Biology/Embryology and Organogenesis, MESH : Genetic Predisposition to Disease, MESH : Animals, MESH: Female, 030217 neurology & neurosurgery, MESH : Mutation, Missense
Abstract: International audience; Bosma arhinia microphthalmia syndrome (BAMS) is an extremely rare and striking condition characterized by complete absence of the nose with or without ocular defects. We report here that missense mutations in the epigenetic regulator SMCHD1 mapping to the extended ATPase domain of the encoded protein cause BAMS in all 14 cases studied. All mutations were de novo where parental DNA was available. Biochemical tests and in vivo assays in Xenopus laevis embryos suggest that these mutations may behave as gain-of-function alleles. This finding is in contrast to the loss-of-function mutations in SMCHD1 that have been associated with facioscapulohumeral muscular dystrophy (FSHD) type 2. Our results establish SMCHD1 as a key player in nasal development and provide biochemical insight into its enzymatic function that may be exploited for development of therapeutics for FSHD.
Published: 2017
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4. Ochratoxin A and T-2 Toxin Induce Clonogenicity and Cell Migration in Human Colon Carcinoma and Fetal Lung Fibroblast Cell Lines

Author: Abassi, Haila, Ayed-Boussema, Imen, Shirley, Sarah, Abid, Salwa, Bacha, Hassen, Laboratoire de Recherche sur les Substances Biologiquement Compatibles [Monastir] ( LRSBC ), Faculté de médecine dentaire de Monastir [Tunisie] ( FMDM ), Lipides - Nutrition - Cancer (U866) ( LNC ), Université de Bourgogne ( UB ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Ecole Nationale Supérieure de Biologie Appliquée à la Nutrition et à l'Alimentation de Dijon ( ENSBANA ), Ministère Tunisien de l'Enseignement Supérieur et de la Recherche Scientifique et de la Technologie (Laboratoire de Recherche sur les Substances Biologiquement Compatibles (LRSBC), UMR 866 Lipides, Nutrition Cancer (Dijon), Laboratoire de Recherche sur les Substances Biologiquement Compatibles [Monastir] (LRSBC), Faculté de Médecine Dentaire de Monastir [Tunisie] (FMDM), Lipides - Nutrition - Cancer (U866) (LNC), and Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Ecole Nationale Supérieure de Biologie Appliquée à la Nutrition et à l'Alimentation de Dijon (ENSBANA)
Subjects: MESH : Cell Line, MESH: Cell Line, Tumor, Aflatoxin B1, MESH : Lung, [ SDV.TOX ] Life Sciences [q-bio]/Toxicology, MESH : Wound Healing, MESH: T-2 Toxin, MESH : Mycotoxins, MESH: Mycotoxins, Cell Migration, Cell Line, MESH : Aflatoxin B1, Cell Movement, Cell Line, Tumor, T2-Toxin, MESH : T-2 Toxin, MESH : Ochratoxins, Humans, MESH : Cell Movement, MESH: Lung, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Clonogenicity, MESH: Cell Movement, Lung, [ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH: Colonic Neoplasms, Wound Healing, MESH: Humans, MESH : Cell Line, Tumor, MESH: Aflatoxin B1, MESH : Humans, MESH: Ochratoxins, Ochratoxin A, Mycotoxins, Ochratoxins, MESH: Cell Line, MESH: Wound Healing, T-2 Toxin, MESH : Colonic Neoplasms, [SDV.TOX]Life Sciences [q-bio]/Toxicology, Colonic Neoplasms
Abstract: IF 3.02; International audience; T-2 toxin and Ochratoxin A (OTA) are toxic secondary metabolites produced by various fungi, and together they contaminate feedstuffs worldwide. T-2 toxin and OTA may exert carcinogenic action in rodent. Despite the various in vivo experiments, carcinogenicity of these two mycotoxins has not yet been proven for human. In this current study, we proposed to investigate, in Human colon carcinoma cells and fetal lung fibroblast-like cells transfected with MYC, the effect of T-2 toxin and OTA on cell clonogenicity and cell migration. Results of the present investigation showed that T2-toxin as well as OTA has an important clonogenic effect in all cell lines, suggesting that these mycotoxins could promote the transcription of c-myc gene. Furthermore, T-2 toxin and OTA enhanced the migration effect of HCT116 cells at very low concentrations, proposing that these mycotoxins may exhibit carcinogenesis-like properties in the studied cells.
Published: 2016
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5. Candida albicans is able to use M cells as a portal of entry across the intestinal barrier in vitro

Author: Albac, Sandrine, Schmitz, Antonin, Lopez-Alayon, Carolina, d'Enfert, Christophe, Sautour, Marc, Ducreux, Amandine, Labruère-Chazal, Catherine, Laue, Michael, Holland, Gudrun, Bonnin, Alain, Dalle, Frederic, Agroécologie [Dijon], Institut National de la Recherche Agronomique (INRA)-Université de Bourgogne (UB)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement, Biologie et Pathogénicité fongiques, Institut Pasteur [Paris]-Institut National de la Recherche Agronomique (INRA), Laboratoire de parasitologie mycologie (CHU de Dijon), Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon), Institut de Mathématiques de Bourgogne [Dijon] (IMB), Centre National de la Recherche Scientifique (CNRS)-Université de Franche-Comté (UFC), Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université de Bourgogne (UB), Robert Koch Institute [Berlin] (RKI), Agence Nationale de la Recherche. Grant Number: ANR-08-MIEN- 033-01 BIOASTER-Sanofi-Alliance pour les Sciences de la Vie et de la Santé (AVIESAN) Investissement d'Avenir. Grant Number: ANR-10-AIRT-03, ANR-08-MIEN-0033,KANJI,Colonisation et invasion de la muqueuse digestive par le champignon pathogène de l'homme Candida albicans(2008), ANR-10-AIRT-0005,NANOELEC,NANOELEC(2010), Institut National de la Recherche Agronomique (INRA)-Institut Pasteur [Paris], Université de Bourgogne (UB)-Centre National de la Recherche Scientifique (CNRS), ANR: ANR-10-AIRT-05,Programme Investissements d’Avenir, Institut National de la Recherche Agronomique ( INRA ) -Université de Bourgogne ( UB ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement, Institut National de la Recherche Agronomique ( INRA ) -Institut Pasteur [Paris], Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand ( CHU Dijon ), Institut de Mathématiques de Bourgogne [Dijon] ( IMB ), Université de Bourgogne ( UB ) -Centre National de la Recherche Scientifique ( CNRS ), Robert Koch Institute [Berlin] ( RKI ), ANR-08-MIEN-0033,KANJI,Colonisation et invasion de la muqueuse digestive par le champignon pathogène de l'homme Candida albicans ( 2008 ), ANR : ANR-10-AIRT-05,Programme Investissements d’Avenir, Biologie et Pathogénicité fongiques (BPF), Institut National de la Recherche Agronomique (INRA)-Institut Pasteur [Paris] (IP), and Université de Bourgogne (UB)-Université Bourgogne Franche-Comté [COMUE] (UBFC)-Centre National de la Recherche Scientifique (CNRS)
Subjects: MESH : Cell Line, MESH : Coculture Techniques, MESH : Microscopy, Fluorescence, [SDV]Life Sciences [q-bio], MESH: Microscopy, Fluorescence, MESH: Candidemia, Peyer's Patches, Mucosal immunity, Invasion, Candida albicans, MESH : Host-Pathogen Interactions, [ SDV.MP.MYC ] Life Sciences [q-bio]/Microbiology and Parasitology/Mycology, [SDV.MP.MYC]Life Sciences [q-bio]/Microbiology and Parasitology/Mycology, B-Lymphocytes, Endocytosis, MESH : Epithelial Cells, MESH : Peyer's Patches, MESH: Epithelial Cells, Host-Pathogen Interactions, MESH: Endocytosis, Adhesion, Transcytosis, Infection, MESH : Microscopy, Electron, Scanning, MESH : Candida albicans, MESH: Microscopy, Electron, Scanning, MESH : Endocytosis, MESH : B-Lymphocytes, Cell Line, MESH: Cell Adhesion, MESH: Coculture Techniques, MESH: B-Lymphocytes, Cell Adhesion, MESH : Candidemia, Humans, MESH: Humans, [ SDV ] Life Sciences [q-bio], MESH: Candida albicans, MESH : Humans, MESH: Host-Pathogen Interactions, Candidemia, Epithelial Cells, Coculture Techniques, MESH: Cell Line, Gastrointestinal Tract, MESH : Cell Adhesion, Microscopy, Fluorescence, Microscopy, Electron, Scanning, MESH: Peyer's Patches, MESH: Gastrointestinal Tract, MESH : Gastrointestinal Tract
Abstract: International audience; Candida albicans is the most frequent yeast responsible for systemic infections in humans. These infections mainly originate from the gastrointestinal tract where C. albicans can invade the gut epithelial barrier to gain access to the bloodstream. Along the gut, pathogens can use Microfold (M) cells as a portal of entry to cross the epithelial barrier. M cells are specialized cells mainly located in the follicule-associated epithelium of Peyer patches. In this study, we used scanning electron and fluorescence microscopy, adhesion and invasion assays and fungal mutants to investigate the interactions of C. albicans with M cells obtained in an established in vitro model whereby enterocyte-like Caco-2 cells co-cultured with the Raji B cell line undergo a phenotypic switch to morphologically and functionally resembling M cells. Our data demonstrate that C. albicans co-localizes with and invades preferentially M cells, providing evidence that the fungus can use M cells as a portal of entry into the intestinal barrier. In addition to active penetration, F-actin dependent endocytosis contributes to internalization of the fungus into M cells through a mechanism involving hypha-associated invasins including Ssa1 and Als3.
Published: 2016
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6. Patched dependence receptor triggers apoptosis through ubiquitination of caspase-9

Author: Patrick Mehlen, Chantal Thibert, Rémy Sadoul, Joanna Fombonne, Catherine Guix, Pierre-Antoine Bissey, Puisieux, Alain, Centre de Recherche en Cancérologie de Lyon (UNICANCER/CRCL), Centre Léon Bérard [Lyon]-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), INSERM U836, équipe 2, Neurodégénérescence et plasticité, Grenoble Institut des Neurosciences (GIN), Université Joseph Fourier - Grenoble 1 (UJF)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Joseph Fourier - Grenoble 1 (UJF)-Institut National de la Santé et de la Recherche Médicale (INSERM), This work was supported by institutional grants from the Ligue Contre le Cancer, INCA, ANR, and IP ApoSys., Institute for Advanced Biosciences / Institut pour l'Avancée des Biosciences (Grenoble) (IAB), Centre Hospitalier Universitaire [Grenoble] (CHU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Etablissement français du sang - Auvergne-Rhône-Alpes (EFS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre de Recherche en Cancérologie de Lyon ( CRCL ), Centre Léon Bérard [Lyon]-Université Claude Bernard Lyon 1 ( UCBL ), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Grenoble Institut des Neurosciences ( GIN ), and Université Joseph Fourier - Grenoble 1 ( UJF ) -CHU Grenoble-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Université Joseph Fourier - Grenoble 1 ( UJF ) -CHU Grenoble-Institut National de la Santé et de la Recherche Médicale ( INSERM )
Subjects: MESH : Cell Line, endocrine system diseases, Apoptosis, NEDD4, [SDV.BC.BC]Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC], Dependence receptor, MESH : Ubiquitination, 0302 clinical medicine, Ubiquitin, [SDV.BC.IC]Life Sciences [q-bio]/Cellular Biology/Cell Behavior [q-bio.CB], MESH: Caspase 3, MESH : Caspase 3, ComputingMilieux_MISCELLANEOUS, Caspase-9, 0303 health sciences, Multidisciplinary, biology, Caspase 3, MESH: Real-Time Polymerase Chain Reaction, Signal transducing adaptor protein, Biological Sciences, MESH : Mutagenesis, Site-Directed, Ubiquitin ligase, MESH: Mutagenesis, Site-Directed, 030220 oncology & carcinogenesis, Patched, MESH : Real-Time Polymerase Chain Reaction, [SDV.CAN]Life Sciences [q-bio]/Cancer, macromolecular substances, Real-Time Polymerase Chain Reaction, MESH: Two-Hybrid System Techniques, Cell Line, 03 medical and health sciences, Two-Hybrid System Techniques, signalisation, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, [ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology, 030304 developmental biology, posttranslational modification, MESH: Humans, MESH: Apoptosis, MESH : Humans, Ubiquitination, [SDV.BDD.MOR]Life Sciences [q-bio]/Development Biology/Morphogenesis, protease, MESH: Cell Line, [SDV.BDD.EO]Life Sciences [q-bio]/Development Biology/Embryology and Organogenesis, MESH : Two-Hybrid System Techniques, Mutagenesis, Site-Directed, biology.protein, Cancer research, MESH: Ubiquitination, MESH : Apoptosis
Abstract: International audience; Patched (Ptc), the main receptor for Sonic Hedgehog, is a tumor suppressor. Ptc has been shown to be a dependence receptor, and as such triggers apoptosis in the absence of its ligand. This apoptosis induction occurs through the recruitment by the Ptc intracellular domain of a caspase-activating complex, which includes the adaptor proteins DRAL and TUCAN, and the apical caspase-9. We show here that this caspase-activating complex also includes the E3 ubiquitin ligase NEDD4. We demonstrate that Ptc-mediated apoptosis and Ptc-induced caspase-9 activation require NEDD4. We show that Ptc, but not Bax, the prototypical inducer of the intrinsic cell-death pathway, triggers polyubiquitination of caspase-9. Moreover, a caspase-9 mutant that could not be ubiquitinated failed to mediate Ptc-induced apoptosis. Taken together, these data support the view that the Ptc dependence receptor specifically allows the activation of caspase-9 via its ubiquitination, which occurs via the recruitment by Ptc of NEDD4.
Published: 2012
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7. Characterization of Unique Signature Sequences in the Divergent Maternal Protein Bcl2l10

Author: Germain Gillet, Abdel Aouacheria, François Penin, Aurélie Cornut-Thibaut, Yannis Guillemin, Institut de biologie et chimie des protéines [Lyon] (IBCP), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Centre de Recherche en Cancérologie de Lyon (UNICANCER/CRCL), Centre Léon Bérard [Lyon]-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM), Laboratoire de Biologie Moléculaire de la Cellule (LBMC), École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Ganivet, Agnès, Institut de biologie et chimie des protéines [Lyon] ( IBCP ), Université Claude Bernard Lyon 1 ( UCBL ), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique ( CNRS ), Centre de Recherche en Cancérologie de Lyon ( CRCL ), Université de Lyon-Université de Lyon-Centre Léon Bérard [Lyon]-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Laboratoire de Biologie Moléculaire de la Cellule ( LBMC ), École normale supérieure - Lyon ( ENS Lyon ) -Université Claude Bernard Lyon 1 ( UCBL ), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), and École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL)
Subjects: MESH : Cell Line, MESH : Molecular Sequence Data, Amino Acid Motifs, MESH: Amino Acid Sequence, MESH: Base Sequence, MESH : Proto-Oncogene Proteins c-bcl-2, [ SDV.CAN ] Life Sciences [q-bio]/Cancer, MESH: INDEL Mutation, protein sequence, MESH: Amino Acid Motifs, MESH: Protein Structure, Tertiary, 0302 clinical medicine, INDEL Mutation, Databases, Genetic, genetic polymorphism, MESH : Female, MESH: Animals, MESH: Genetic Variation, [SDV.BDD]Life Sciences [q-bio]/Development Biology, Peptide sequence, MESH: Databases, Genetic, MESH: Evolution, Molecular, Sequence Deletion, Genetics, chemistry.chemical_classification, 0303 health sciences, MESH : Amino Acid Sequence, MESH : Sequence Alignment, apoptosis, Vertebrate, MESH: Sequence Deletion, Amino acid, Proto-Oncogene Proteins c-bcl-2, MESH: Calcium, 030220 oncology & carcinogenesis, Female, MESH : Protein Structure, Tertiary, MESH : Amino Acid Motifs, Molecular Sequence Data, MESH: Sequence Alignment, [SDV.CAN]Life Sciences [q-bio]/Cancer, Sequence alignment, Biology, Cell Line, MESH: Oocytes, Evolution, Molecular, 03 medical and health sciences, Bcl-2 family, MESH : Genetic Variation, biology.animal, evolution, [SDV.BDD] Life Sciences [q-bio]/Development Biology, Genetic variation, Homologous chromosome, Animals, Humans, MESH : HeLa Cells, MESH : Evolution, Molecular, Amino Acid Sequence, MESH : Calcium, MESH : Databases, Genetic, Indel, MESH : INDEL Mutation, Molecular Biology, Ecology, Evolution, Behavior and Systematics, 030304 developmental biology, MESH : Mutagenesis, Insertional, calcium, MESH: Humans, MESH: Molecular Sequence Data, Base Sequence, MESH : Sequence Deletion, MESH: Apoptosis, MESH : Humans, MESH : Oocytes, Genetic Variation, Protein Structure, Tertiary, MESH: Cell Line, Mutagenesis, Insertional, MESH: Mutagenesis, Insertional, MESH: Proto-Oncogene Proteins c-bcl-2, chemistry, MESH: HeLa Cells, Oocytes, MESH : Base Sequence, MESH : Animals, Sequence Alignment, MESH: Female, MESH : Apoptosis, HeLa Cells
Abstract: International audience; Insertions or deletions (indels) of amino acids residues have been recognized as an important source of genetic and structural divergence between paralogous Bcl-2 family members. However, these signature sequences have not so far been extensively investigated amongst orthologous Bcl-2 family proteins. Bcl2l10 is an antiapoptotic member of the Bcl-2 family that has evolved rapidly throughout the vertebrate lineage and which shows conserved abundant expression in eggs and oocytes. In this paper, we have unraveled two major sites of divergence between human Bcl2l10 and its vertebrate homologs. The first one provides length variation at the N-terminus (before the BH4 domain) and the second one is located between the predicted α5-α6 pore-forming helices, providing an unprecedented case in the superfamily of helix-bundled pore-forming proteins. These two particular indels were studied phylogenetically and through biochemical and cell biological techniques, including truncation and site-directed mutagenesis. While deletion of the N-terminal extension had no significant functional impact in HeLa cells, our results suggest that the human Bcl2l10 protein evolved a calcium-binding motif in its α5-α6 interhelical region by acquiring critical negatively charged residues. Considering the reliance of female eggs on calcium-dependent proteins and calcium-regulated processes and the exceptional longevity of oocytes in the primate lineage, we propose that this microstructural variation may be an adaptive feature associated with high maternal expression of this Bcl-2 family member.
Published: 2011
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8. Functional characterization of the AFF (AF4/FMR2) family of RNA-binding proteins: insights into the molecular pathology of FRAXE intellectual disability

Author: Mireille Melko, Samantha Zongaro, Jozef Gecz, Dominique Douguet, Barbara Bardoni, Céline Verheggen, Mounia Bensaid, Institut de pharmacologie moléculaire et cellulaire ( IPMC ), Université Nice Sophia Antipolis ( UNS ), Université Côte d'Azur ( UCA ) -Université Côte d'Azur ( UCA ) -Centre National de la Recherche Scientifique ( CNRS ), Institut de Génétique Moléculaire de Montpellier ( IGMM ), Centre National de la Recherche Scientifique ( CNRS ) -Université de Montpellier ( UM ), Department of Genetic Medicine Women's and Children's, University of Adelaide, Université Côte d'Azur ( UCA ), Institut de pharmacologie moléculaire et cellulaire (IPMC), Université Nice Sophia Antipolis (... - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Centre National de la Recherche Scientifique (CNRS), Institut de Génétique Moléculaire de Montpellier (IGMM), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), and COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)
Subjects: MESH : Cell Line, MESH : Molecular Sequence Data, MESH: Sequence Homology, Amino Acid, Gene Expression, RNA-binding protein, MESH: Amino Acid Sequence, MESH: Gene Order, Exon, 0302 clinical medicine, MESH : Protein Transport, Genes, Reporter, Transcription (biology), Gene Order, Genetics (clinical), Genetics, 0303 health sciences, MESH : Amino Acid Sequence, MESH : Sequence Alignment, RNA-Binding Proteins, General Medicine, MESH : Genes, Reporter, Phenotype, MESH : Sequence Homology, Amino Acid, DNA-Binding Proteins, Protein Transport, MESH : Fragile X Syndrome, 030220 oncology & carcinogenesis, RNA splicing, MESH : RNA Splicing, MESH : DNA-Binding Proteins, Intranuclear Space, MESH: Fragile X Syndrome, MESH: Protein Transport, MESH: Gene Expression, RNA Splicing, Molecular Sequence Data, MESH : RNA-Binding Proteins, MESH: Sequence Alignment, MESH: Intranuclear Space, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Biology, Cell Line, 03 medical and health sciences, MESH : Fibroblasts, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH : HeLa Cells, Amino Acid Sequence, [ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology, Molecular Biology, Gene, 030304 developmental biology, MESH: Molecular Sequence Data, MESH: Humans, MESH : Gene Order, Sequence Homology, Amino Acid, [ SDV.BC ] Life Sciences [q-bio]/Cellular Biology, MESH: Genes, Reporter, MESH : Humans, Alternative splicing, RNA, Fibroblasts, MESH: Cell Line, MESH : Gene Expression, MESH: RNA-Binding Proteins, MESH : Intranuclear Space, MESH: Fibroblasts, Fragile X Syndrome, MESH: HeLa Cells, MESH: RNA Splicing, Sequence Alignment, MESH: DNA-Binding Proteins, HeLa Cells
Abstract: International audience; The AFF (AF4/FMR2) family of genes includes four members: AFF1/AF4, AFF2/FMR2, AFF3/LAF4 and AFF4/AF5q31. AFF2/FMR2 is silenced in FRAXE intellectual disability, while the other three members have been reported to form fusion genes as a consequence of chromosome translocations with the myeloid/lymphoid or mixed lineage leukemia (MLL) gene in acute lymphoblastic leukemias (ALLs). All AFF proteins are localized in the nucleus and their role as transcriptional activators with a positive action on RNA elongation was primarily studied. We have recently shown that AFF2/FMR2 localizes to nuclear speckles, subnuclear structures considered as storage/modification sites of pre-mRNA splicing factors, and modulates alternative splicing via the interaction with the G-quadruplex RNA-forming structure. We show here that similarly to AFF2/FMR2, AFF3/LAF4 and AFF4/AF5q31 localize to nuclear speckles and are able to bind RNA, having a high apparent affinity for the G-quadruplex structure. Interestingly, AFF3/LAF4 and AFF4/AF5q31, like AFF2/FMR2, modulate, in vivo, the splicing efficiency of a mini-gene containing a G-quadruplex structure in one alternatively spliced exon. Furthermore, we observed that the overexpression of AFF2/3/4 interferes with the organization and/or biogenesis of nuclear speckles. These findings fit well with our observation that enlarged nuclear speckles are present in FRAXE fibroblasts. Furthermore, our findings suggest functional redundancy among the AFF family members in the regulation of splicing and transcription. It is possible that other members of the AFF family compensate for the loss of AFF2/FMR2 activity and as such explain the relatively mild to borderline phenotype observed in FRAXE patients.
Published: 2011
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9. Truncation of PITX2 differentially affects its activity on physiological targets

Author: Véronique Vieira, Maurice Menasche, James P. Herman, Thierry Brue, Alain Enjalbert, Marc Abitbol, Marie-Helene Quentien, Jean-Louis Dufier, Centre de recherche en neurobiologie - neurophysiologie de Marseille (CRN2M), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre de recherche thérapeutique en ophtalmologie, Université Paris Descartes - Paris 5 (UPD5)-EA2502, EA no 2502 du ministère de la Recherche, de l'Enseignement Supérieur et la Technologie, Université Paris Descartes - Paris 5 (UPD5)-CERTO, Service d'ophtalmologie [CHU Necker], Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-CHU Necker - Enfants Malades [AP-HP], Centre d'étude et de recherche thérapeutiques en ophtalmologie, Université Paris Descartes - Paris 5 (UPD5), Aix Marseille Université (AMU) - Institut National de la Santé et de la Recherche Médicale (INSERM) - Centre National de la Recherche Scientifique (CNRS), Université Paris Descartes - Paris 5 (UPD5) - EA2502, Université Paris Descartes - Paris 5 (UPD5) - CERTO, Assistance publique - Hôpitaux de Paris (AP-HP) - CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Necker - Enfants Malades [AP-HP], and Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)
Subjects: MESH : Cell Line, MESH : Transcription Factors, MESH : Transcription Factor Pit-1, Mutant, Electrophoretic Mobility Shift Assay, MESH : Blotting, Western, MESH : Promoter Regions, Genetic, medicine.disease_cause, MESH : Pituitary Gland, 0302 clinical medicine, Endocrinology, MESH : Prolactin, MESH: Pituitary Gland, MESH: Human Growth Hormone, MESH: Animals, MESH : Transcriptional Activation, MESH : Homeodomain Proteins, Eye Abnormalities, Promoter Regions, Genetic, 0303 health sciences, Mutation, MESH: Anterior Eye Segment, Human Growth Hormone, Eye Diseases, Hereditary, MESH: Transcription Factors, Transfection, MESH: Eye Abnormalities, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Cell biology, MESH : Electrophoretic Mobility Shift Assay, Pituitary Gland, MESH : Mutation, Transcription Factor Pit-1, hormones, hormone substitutes, and hormone antagonists, Transcriptional Activation, congenital, hereditary, and neonatal diseases and abnormalities, medicine.medical_specialty, MESH: Mutation, Blotting, Western, MESH: Prolactin, Biology, MESH : Anterior Eye Segment, Cell Line, [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], 03 medical and health sciences, stomatognathic system, Anterior Eye Segment, Internal medicine, MESH : Eye Abnormalities, MESH: Promoter Regions, Genetic, MESH: Homeodomain Proteins, medicine, MESH: Blotting, Western, Animals, Humans, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], MESH : Human Growth Hormone, Molecular Biology, Transcription factor, Gene, 030304 developmental biology, Homeodomain Proteins, Reporter gene, MESH: Humans, POU domain, MESH : Humans, Promoter, MESH: Cell Line, Prolactin, stomatognathic diseases, MESH: Electrophoretic Mobility Shift Assay, 030221 ophthalmology & optometry, MESH: Transcriptional Activation, MESH : Animals, sense organs, MESH: Transcription Factor Pit-1, Transcription Factors
Abstract: International audience; The bicoid-like transcription factor PITX2 has been previously described to interact with the pituitary-specific POU homeodomain factor POU1F1 (human ortholog of PIT-1) to achieve cell-specific expression of prolactin (PRL) and GH in pituitary somatolactotroph cells. In this work, we have investigated the functional properties of three PITX2 mutants reported in Axenfeld-Rieger syndrome patients relative to the regulation of these genes, using reporter genes under the control of human PRL (hPRL), hGH, or POU1F1 promoters transfected in nonpituitary and pituitary cell lines. Among the three mutations studied, Y167X and E101X introduce a premature stop codon, and F104L leads to an amino acid substitution. While PITX2(E101X) is not expressed in the cells following transfection, and PITX2(F104L) is functionally inactive, the PITX2(Y167X) mutant keeps its DNA-binding capacity and displays a markedly enhanced activation of the hPRL and POU1F1 promoters, but not of the hGH promoter. Y167X is the first mutation of PITX2 described to result in a differential effect on the activation of its different physiological targets, hPRL and POU1F1 on one hand and hGH on the other hand. The differential effect of the Y167X mutation might be linked to an interaction of PITX2 with different transcription factors or cofactors when bound to the hPRL and POU1F1 or the hGH promoters. These results might form the basis for the identification of the PITX2 protein complex necessary for the differential GH or PRL expression.
Published: 2010
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10. Meningococcal Type IV Pili Recruit the Polarity Complex to Cross the Brain Endothelium

Author: Xavier Nassif, Guillaume Duménil, Florence Miller, Mathieu Coureuil, Pierre-Olivier Couraud, Babette B. Weksler, Hervé Lécuyer, Christine Bernard, Ignacio A. Romero, Sandrine Bourdoulous, René-Marc Mège, Guillain Mikaty, INSERM U1002, Pathogénie des infections systémiques (UMR_S 570), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut Cochin (UMR_S567 / UMR 8104), Institut du Fer à Moulin, Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM), Weill Medical College of Cornell University [New York], Department of Biological Sciences, The Open University [Milton Keynes] (OU), Pathogénie des infections systémiques ( UMR_S 570 ), Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ) -Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Institut Cochin ( UMR_S567 / UMR 8104 ), Université Pierre et Marie Curie - Paris 6 ( UPMC ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), The Open University [Milton Keynes] ( OU ), and Coureuil, Mathieu
Subjects: MESH : Cell Line, Delta Catenin, Cell Cycle Proteins, blood brain barrier, Neisseria meningitidis, medicine.disease_cause, Cell junction, MESH : cdc42 GTP-Binding Protein, Bacterial Adhesion, MESH: Cadherins, MESH : Blood-Brain Barrier, MESH: Blood-Brain Barrier, MESH : Cell Cycle Proteins, 0302 clinical medicine, [ SDV.MP ] Life Sciences [q-bio]/Microbiology and Parasitology, Cell polarity, MESH: Endothelial Cells, MESH : Adaptor Proteins, Signal Transducing, cdc42 GTP-Binding Protein, MESH: Antigens, CD, Protein Kinase C, 0303 health sciences, Multidisciplinary, Cell adhesion molecule, MESH : Fimbriae, Bacterial, Brain, Cell Polarity, meningitis, Catenins, Cadherins, endothelial cells, 3. Good health, Cell biology, Endothelial stem cell, Intercellular Junctions, [SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology, medicine.anatomical_structure, Cdc42 GTP-Binding Protein, Blood-Brain Barrier, MESH : Intercellular Junctions, MESH: Cell Adhesion Molecules, MESH : Cadherins, MESH: Endothelium, Vascular, MESH: Membrane Proteins, MESH: Cell Polarity, MESH : Cell Polarity, MESH : Bacterial Adhesion, Biology, Blood–brain barrier, MESH: Phosphoproteins, Article, MESH: Neisseria meningitidis, MESH : Cell Adhesion Molecules, Cell Line, MESH: Fimbriae, Bacterial, MESH: Brain, 03 medical and health sciences, MESH: Cell Cycle Proteins, Antigens, CD, MESH : Antigens, CD, medicine, Humans, MESH : Protein Kinase C, MESH: Bacterial Adhesion, [SDV.MP] Life Sciences [q-bio]/Microbiology and Parasitology, MESH: Catenins, Adaptor Proteins, Signal Transducing, MESH: Adaptor Proteins, Signal Transducing, 030304 developmental biology, MESH: Humans, MESH: cdc42 GTP-Binding Protein, MESH : Endothelial Cells, MESH : Neisseria meningitidis, Cadherin, MESH : Humans, Membrane Proteins, Phosphoproteins, MESH: Protein Kinase C, MESH : Catenins, MESH: Cell Line, MESH : Brain, MESH : Endothelium, Vascular, MESH : Membrane Proteins, Fimbriae, Bacterial, Endothelium, Vascular, MESH : Phosphoproteins, Cell Adhesion Molecules, 030217 neurology & neurosurgery, MESH: Intercellular Junctions
Abstract: Breaking the Barrier Being able to deliver drugs into the brain to treat degenerative diseases such as Alzheimer's or Parkinson's requires the ability to traverse the blood-brain barrier (BBB). Understanding the formation of the very specific adherent junctions (AJ) and tight junctions present at the BBB cell junctions is a prerequisite to the design of such therapeutics. However, diminishing the expression of any one component involved in the formation of these intercellular junctions destroys them. Coureuil et al. (p. 83 , published online 11 June) exploited the specific recruitment of AJ proteins by Neisseria meningitidis to dissect this process. Adhesion of the bacteria to human brain endothelial cells recruited the polarity complex Par3/Par6/PKCζ required for the establishment of eukaryotic cell polarity and the formation of intercellular junctions. The bacterial recruitment of the polarity complex depleted junctional proteins at the cell-cell interface opening the intercellular junctions at the brainendothelial interface.
Published: 2009
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11. Interaction of the coiled‐coil domain with glycosaminoglycans protects angiopoietin‐like 4 from proteolysis and regulates its antiangiogenic activity

Author: Sylvie Ricard-Blum, Clément Faye, Corinne Ardidie-Robouant, Clémence Chomel, Laurent Muller, Stéphane Germain, Aurélie Cazes, A. Barret, Catherine Monnot, Elisa Gomez, Marine Bignon, Pathologie vasculaire et endocrinologie rénale - Chaire de médecine expérimentale (INSERM U36), Collège de France (CdF (institution))-Institut National de la Santé et de la Recherche Médicale (INSERM), Service d'anatomo-pathologie [CHU HEGP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Institut de biologie et chimie des protéines [Lyon] (IBCP), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Centre interdisciplinaire de recherche en biologie (CIRB), Labex MemoLife, École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Collège de France (CdF (institution))-Ecole Superieure de Physique et de Chimie Industrielles de la Ville de Paris (ESPCI Paris), Université Paris sciences et lettres (PSL)-École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM), This work was supported by the European Vascular Genomics Network (http://www.evgn.org) (contract LSHMCT-2003-503254), ANR-06-JCJC-0160,ANGPTL4,Role of ANGPTL4 during development and ischemic heart disease(2006), Pathologie vasculaire et endocrinologie rénale, Collège de France ( CdF ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Experimental Medicine Unit, Collège de France ( CdF ), Assistance publique - Hôpitaux de Paris (AP-HP)-Hôpital Européen Georges Pompidou [APHP] ( HEGP ), Institut de biologie et chimie des protéines [Lyon] ( IBCP ), Université Claude Bernard Lyon 1 ( UCBL ), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique ( CNRS ), Service d'hématologie A [CHU HEGP], and ANR-06-JCJC-0160,ANGPTL4,Role of ANGPTL4 during development and ischemic heart disease ( 2006 )
Subjects: MESH : Cell Line, MESH : Angiopoietins, Protein Conformation, Angiogenesis, MESH: Cricetinae, Biochemistry, Extracellular matrix, angiogenesis, MESH: Protein Structure, Tertiary, MESH: Protein Conformation, 0302 clinical medicine, MESH : Lipid Metabolism, Cricetinae, MESH: Animals, MESH : Protein Conformation, Glycosaminoglycans, MESH: Lipid Metabolism, 0303 health sciences, MESH : Neovascularization, Physiologic, medicine.diagnostic_test, Chemistry, MESH : Cricetinae, MESH : Extracellular Matrix, MESH : Protein Binding, MESH: Glycosaminoglycans, [ SDV.MHEP.CSC ] Life Sciences [q-bio]/Human health and pathology/Cardiology and cardiovascular system, 3. Good health, Cell biology, Endothelial stem cell, 030220 oncology & carcinogenesis, endothelial cell, MESH : Mutation, MESH : Protein Structure, Tertiary, MESH: Neovascularization, Physiologic, Protein Binding, Biotechnology, MESH: Mutation, extracellular matrix, Proteolysis, Neovascularization, Physiologic, Angiopoietin-like 4 Protein, MESH: Extracellular Matrix, Cell Line, Angiopoietin, 03 medical and health sciences, [SDV.MHEP.CSC]Life Sciences [q-bio]/Human health and pathology/Cardiology and cardiovascular system, MESH : Glycosaminoglycans, Genetics, medicine, Animals, Humans, MESH: Protein Binding, Molecular Biology, 030304 developmental biology, MESH: Humans, hypoxia, MESH : Humans, Lipid Metabolism, Protein Structure, Tertiary, MESH: Cell Line, Cell culture, MESH: Angiopoietins, Mutation, MESH : Animals, Proprotein Convertases, Angiopoietins
Abstract: International audience; Angiopoietin-like 4 (ANGPTL4) is involved in angiogenesis and lipid metabolism. It is secreted by liver and adipose tissues and cleaved to generate circulating coiled-coil domain (CCD) and fibrinogen-like domain (FLD) fragments. The full-length ANGPTL4 produced by hypoxic endothelial cells interacts with the extracellular matrix (ECM). The ECM-bound and soluble forms of ANGPTL4 have antiangiogenic properties. We carried out a structure-function analysis to investigate the regulation of ANGPTL4 bioactivity in endothelial cells. We found that the recombinant CCD binds to the ECM, whereas the FLD is released into the medium. The CCD, like the full-length ANGPTL4, binds to heparan and dermatan sulfates in surface plasmon resonance assays and inhibits endothelial cell adhesion, motility, and tubule-like formation. In endothelial cells, ANGPTL4 is processed in the secretion medium after release from the ECM. This processing is altered by the proprotein convertases inhibitor alpha1-PDX and abolished by the mutation of the (161)RRKR(164) cleavage site without modification of the ECM binding and release. These data suggest that the full-length form, which interacts with heparan sulfate proteoglycans via its CCD, is protected from proteolysis by proprotein convertases and constitutes the major active pool of ANGPTL4 in hypoxic endothelial cells.
Published: 2008
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12. Synthesis and Biological Evaluation of 1-Amino-2-Phosphonomethylcyclopropanecarboxylic Acids, New Group III Metabotropic Glutamate Receptor Agonists

Author: Cyril Goudet, Hugues-Olivier Bertrand, Isabelle Brabet, Michael J. Marino, Pauline Sibille, Florence Gaven, Sébastien Lopez, Jacques Neyton, Jean-Philippe Pin, Ornella Valenti, Francine Acher, Nadia Oueslati, Marianne Amalric, Laboratoire de Chimie et de Biochimie Pharmacologiques et Toxicologiques (LCBPT - UMR 8601), Université Paris Descartes - Paris 5 (UPD5)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Neurosciences Cognitives [Marseille] (LNC), Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU), Institut de Génomique Fonctionnelle (IGF), Université de Montpellier (UM)-Université Montpellier 1 (UM1)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Montpellier 2 - Sciences et Techniques (UM2)-Centre National de la Recherche Scientifique (CNRS), Department of Molecular Neurology, Merck Research Laboratories, Accelrys, Laboratoire de Neurobiologie (UMR 8544) (NEURO), École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), This work was supported by grants from the Fondation de France (Parkinson committee), RETINA-France, the french National Research Agency (ANR-05-NEUR-021-01), the french ministry of Education and Research and the Centre National de la Recherche Scientifique., ANR-05-NEUR-0021,MGluRs Park,Cible thérapeutique potentielle de nouveaux ligands des récepteurs métabotropiques du glutamate du groupe III dans le traitement de la maladie de Parkinson : de la conception à l'évaluation préclinique(2005), Laboratoire de Chimie et de Biochimie Pharmacologiques et Toxicologiques ( LCBPT - UMR 8601 ), Université Paris Descartes - Paris 5 ( UPD5 ) -Centre National de la Recherche Scientifique ( CNRS ), Laboratoire de Neurosciences Cognitives [Marseille] ( LNC ), Aix Marseille Université ( AMU ) -Centre National de la Recherche Scientifique ( CNRS ), Institut de Génomique Fonctionnelle ( IGF ), Centre National de la Recherche Scientifique ( CNRS ) -Université Montpellier 2 - Sciences et Techniques ( UM2 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Université Montpellier 1 ( UM1 ) -Université de Montpellier ( UM ), Laboratoire de Neurobiologie (UMR 8544) ( NEURO ), École normale supérieure - Paris ( ENS Paris ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), ANR PARK, ANR-05-NEUR-021-01,ANR PARK, ANR-05-NEUR-021-01, Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Université Montpellier 1 (UM1)-Université Montpellier 2 - Sciences et Techniques (UM2)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), École normale supérieure - Paris (ENS-PSL), Goudet, Cyril, and Neurosciences, neurologie et psychiatrie - Cible thérapeutique potentielle de nouveaux ligands des récepteurs métabotropiques du glutamate du groupe III dans le traitement de la maladie de Parkinson : de la conception à l'évaluation préclinique - - MGluRs Park2005 - ANR-05-NEUR-0021 - NEURO - VALID
Subjects: Male, Models, Molecular, MESH : Cell Line, Patch-Clamp Techniques, MESH: Amino Acids, MESH: Antiparkinson Agents, Molecular Conformation, MESH: Catalepsy, MESH: Rats, Sprague-Dawley, Pharmacology, Receptors, Metabotropic Glutamate, Synaptic Transmission, Basal Ganglia, Antiparkinson Agents, Rats, Sprague-Dawley, MESH: Basal Ganglia, MESH: Structure-Activity Relationship, 0302 clinical medicine, MESH : Structure-Activity Relationship, Drug Discovery, MESH: Animals, Amino Acids, MESH : Patch-Clamp Techniques, MESH : Haloperidol, Receptor, MESH : Synaptic Transmission, 0303 health sciences, MESH : Rats, Chemistry, Metabotropic glutamate receptor 4, MESH : Receptors, Metabotropic Glutamate, Metabotropic glutamate receptor 6, Stereoisomerism, MESH : Antiparkinson Agents, 3. Good health, HYDIA, MESH : Stereoisomerism, Biochemistry, Molecular Medicine, Metabotropic glutamate receptor 1, [SDV.NEU]Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC], MESH : Phosphonic Acids, MESH: Models, Molecular, Agonist, MESH: Rats, MESH : Models, Molecular, medicine.drug_class, MESH : Male, Organophosphonates, MESH: Phosphonic Acids, In Vitro Techniques, Catalepsy, MESH : Rats, Wistar, Receptors, N-Methyl-D-Aspartate, Cell Line, Injections, Structure-Activity Relationship, 03 medical and health sciences, MESH: Patch-Clamp Techniques, MESH: Synaptic Transmission, medicine, Animals, Humans, MESH: Injections, [SDV.NEU] Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC], Rats, Wistar, MESH: Receptors, Metabotropic Glutamate, MESH : Basal Ganglia, 030304 developmental biology, MESH: Receptors, N-Methyl-D-Aspartate, MESH: Molecular Conformation, MESH: Humans, MESH : Catalepsy, MESH : Humans, MESH: Rats, Wistar, medicine.disease, MESH : Injections, MESH: Stereoisomerism, MESH : Rats, Sprague-Dawley, MESH: Male, Rats, MESH: Cell Line, MESH: Haloperidol, MESH : Molecular Conformation, Metabotropic glutamate receptor, [ SDV.NEU ] Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC], Haloperidol, MESH : Amino Acids, MESH : Animals, MESH : Receptors, N-Methyl-D-Aspartate, 030217 neurology & neurosurgery
Abstract: International audience; Stereoisomers of 1-amino-2-phosphonomethylcyclopropanecarboxylic acid (APCPr), conformationally restricted analogues of L-AP4 (2-amino-4-phosphonobutyric acid), have been prepared and evaluated at recombinant group III metabotropic glutamate receptors. They activate these receptors over a broad range of potencies. The most potent isomer (1S,2R)-APCPr displays a similar pharmacological profile as that of L-AP4 (EC50 0.72, 1.95, >500, 0.34 microM at mGlu4, 6, 7, 8 receptors, respectively, and no effect at group I/II mGluRs). It was characterized on native receptors located in the basal ganglia (BG) where it induced a robust and reversible inhibition of synaptic transmission. It was tested in vivo in haloperidol-induced catalepsy, a model of Parkinsonian akinesia, by direct infusion in the globus pallidus of the BG. At a dose of 0.5 nmol/microL, catalepsy was significantly antagonized. This study reveals that (1S,2R)-APCPr is a potent group III mGluR agonist and confirms that these receptors may be considered as a therapeutic target in the Parkinson's disease.
Published: 2007
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13. Focal adhesion kinase is involved in rabies virus infection through its interaction with viral phosphoprotein P

Author: Jovan Nikolic, Hervé Bourhy, Cécile Lagaudrière-Gesbert, Danielle Blondel, Christoph Wirblich, Florence Larrous, Baptiste Fouquet, Rhabdovirus (RHABDO), Département Virologie (Dpt Viro), Institut de Biologie Intégrative de la Cellule (I2BC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Institut de Biologie Intégrative de la Cellule (I2BC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS), Centre National de Référence de la Rage-Dynamique des Lyssavirus et adaptation à l'hôte (CNR), Institut Pasteur [Paris], Centre Collaborateur de l'OMS pour la Rage - Dynamique des lyssavirus et adaptation à l'hôte (CC-OMS), Thomas Jefferson University, Philadelphia, This work was supported by a grant from Fondation pour la Recherche Médicale (FRM DEQ20120323711). Confocal microscopy was performed on the 'Plate-forme Imagerie et Biologie Cellulaire' of the CNRS campus, supported by the Institut Fédératif de Recherche 87 'La plante et son environnement' and the program ASTRE of the Conseil Général de l’Essonne, Rhabdovirus ( RHABDO ), Département Virologie ( Dpt Viro ), Institut de Biologie Intégrative de la Cellule ( I2BC ), Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Sud - Paris 11 ( UP11 ) -Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Sud - Paris 11 ( UP11 ) -Institut de Biologie Intégrative de la Cellule ( I2BC ), Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Sud - Paris 11 ( UP11 ) -Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Sud - Paris 11 ( UP11 ), Centre National de Référence de la Rage-Dynamique des Lyssavirus et adaptation à l'hôte ( CNR ), Centre Collaborateur de l'OMS pour la Rage - Dynamique des lyssavirus et adaptation à l'hôte ( CC-OMS ), and Institut Pasteur [Paris] (IP)
Subjects: MESH : Cell Line, DNA Mutational Analysis, MESH : Focal Adhesion Protein-Tyrosine Kinases, medicine.disease_cause, Virus Replication, Inclusion Bodies, Viral, MESH : Protein Interaction Mapping, Interferon, MESH : Rabies virus, Protein Interaction Mapping, MESH: Microscopy, Confocal, MESH: Animals, MESH: DNA Mutational Analysis, MESH : Host-Pathogen Interactions, Microscopy, Confocal, MESH: Focal Adhesion Protein-Tyrosine Kinases, MESH : Viral Structural Proteins, MESH : Protein Binding, 3. Good health, Cell biology, Virus-Cell Interactions, MESH : Mutagenesis, Site-Directed, MESH: Rabies virus, MESH : Virus Replication, MESH: Mutagenesis, Site-Directed, Host-Pathogen Interactions, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, MESH : Inclusion Bodies, Viral, medicine.drug, Protein Binding, Viral protein, Immunoprecipitation, Immunology, MESH : DNA Mutational Analysis, MESH: Viral Structural Proteins, Biology, MESH: Two-Hybrid System Techniques, Microbiology, [ SDV.MP.VIR ] Life Sciences [q-bio]/Microbiology and Parasitology/Virology, MESH: Phosphoproteins, Virus, Cell Line, MESH : Immunoprecipitation, Focal adhesion, Virology, Two-Hybrid System Techniques, medicine, Animals, Humans, MESH: Protein Binding, MESH : Microscopy, Confocal, Viral Structural Proteins, MESH: Humans, MESH: Immunoprecipitation, Rabies virus, MESH : Humans, MESH: Protein Interaction Mapping, MESH: Virus Replication, MESH: Host-Pathogen Interactions, Phosphoproteins, Molecular biology, MESH: Cell Line, Viral replication, MESH : Two-Hybrid System Techniques, Insect Science, Phosphoprotein, Focal Adhesion Protein-Tyrosine Kinases, Mutagenesis, Site-Directed, MESH: Inclusion Bodies, Viral, MESH : Animals, MESH : Phosphoproteins, Molecular Chaperones
Abstract: The rabies virus (RABV) phosphoprotein P is a multifunctional protein: it plays an essential role in viral transcription and replication, and in addition, RABV P has been identified as an interferon antagonist. Here, a yeast two-hybrid screen revealed that RABV P interacts with the focal adhesion kinase (FAK). The binding involved the 106-to-131 domain, corresponding to the dimerization domain of P and the C-terminal domain of FAK containing the proline-rich domains PRR2 and PRR3. The P-FAK interaction was confirmed in infected cells by coimmunoprecipitation and colocalization of FAK with P in Negri bodies. By alanine scanning, we identified a single mutation in the P protein that abolishes this interaction. The mutant virus containing a substitution of Ala for Arg in position 109 in P (P.R109A), which did not interact with FAK, is affected at a posttranscriptional step involving protein synthesis and viral RNA replication. Furthermore, FAK depletion inhibited viral protein expression in infected cells. This provides the first evidence of an interaction of RABV with FAK that positively regulates infection. IMPORTANCE Rabies virus exhibits a small genome that encodes a limited number of viral proteins. To maintain efficient virus replication, some of them are multifunctional, such as the phosphoprotein P. We and others have shown that P establishes complex networks of interactions with host cell components. These interactions have revealed much about the role of P and about host-pathogen interactions in infected cells. Here, we identified another cellular partner of P, the focal adhesion kinase (FAK). Our data shed light on the implication of FAK in RABV infection and provide evidence that P-FAK interaction has a proviral function.
Published: 2015
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14. A vlincRNA participates in senescence maintenance by relieving H2AZ-mediated repression at the INK4 locus

Author: Lazorthes, Sandra, Vallot, Céline, Briois, Sébastien, Aguirrebengoa, Marion, Thuret, Jean-Yves, St Laurent, Georges, Rougeulle, Claire, Kapranov, Philipp, Mann, Carl, Trouche, Didier, Nicolas, Estelle, Laboratoire de Biologie Cellulaire et Moléculaire du Contrôle de la Prolifération (LBCMCP), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre épigénétique et destin cellulaire (EDC (UMR_7216)), Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS), Institut de Biologie et de Technologies de Saclay (IBITECS), Université Paris-Saclay-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Sénescence et stabilité génomique (SEN), Département Biologie des Génomes (DBG), Institut de Biologie Intégrative de la Cellule (I2BC), Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Institut de Biologie Intégrative de la Cellule (I2BC), Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS), St Laurent Institute, St. Laurent Institute, Institute of Genomics, Huaqiao University School of Medicine, Xiamen, Academy of Biology and Biotechnology, Southern Federal University [Rostov-on-Don] (SFEDU), LBCMCP, Centre National de la Recherche Scientifique ( CNRS ), Centre épigénétique et destin cellulaire ( EDC ), Université Paris Diderot - Paris 7 ( UPD7 ) -Centre National de la Recherche Scientifique ( CNRS ), Institut de Biologie et de Technologies de Saclay ( IBITECS ), Université Paris-Saclay-Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ), Sénescence et stabilité génomique ( SEN ), Département Biologie des Génomes ( DBG ), Institut de Biologie Intégrative de la Cellule ( I2BC ), Université Paris-Sud - Paris 11 ( UP11 ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ) -Université Paris-Sud - Paris 11 ( UP11 ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ) -Institut de Biologie Intégrative de la Cellule ( I2BC ), Université Paris-Sud - Paris 11 ( UP11 ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ) -Université Paris-Sud - Paris 11 ( UP11 ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ), Woburn, Massachusetts, Southern Federal University - Rostov on Don, Université de Toulouse (UT)-Université de Toulouse (UT)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre de Biologie Intégrative (CBI), Université de Toulouse (UT)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), Centre épigénétique et destin cellulaire (EDC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay, Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Institut de Biologie Intégrative de la Cellule (I2BC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Université de Paris (UP), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre de Biologie Intégrative (CBI), and Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS)
Subjects: MESH : Cell Line, RNA, Untranslated, MESH : RNA, Untranslated, MESH: Humans, MESH : Chromatin, [ SDV ] Life Sciences [q-bio], MESH : Cell Aging, MESH : Heterochromatin, [SDV]Life Sciences [q-bio], MESH : Humans, Chromatin, Article, Cell Line, MESH: Cell Line, MESH: Chromatin, MESH: RNA, Untranslated, MESH: Heterochromatin, MESH: Cell Aging, Heterochromatin, Humans, Erratum, ComputingMilieux_MISCELLANEOUS, Cellular Senescence
Abstract: Non-coding RNAs (ncRNAs) play major roles in proper chromatin organization and function. Senescence, a strong anti-proliferative process and a major anticancer barrier, is associated with dramatic chromatin reorganization in heterochromatin foci. Here we analyze strand-specific transcriptome changes during oncogene-induced human senescence. Strikingly, while differentially expressed RNAs are mostly repressed during senescence, ncRNAs belonging to the recently described vlincRNA (very long intergenic ncRNA) class are mainly activated. We show that VAD, a novel antisense vlincRNA strongly induced during senescence, is required for the maintenance of senescence features. VAD modulates chromatin structure in cis and activates gene expression in trans at the INK4 locus, which encodes cell cycle inhibitors important for senescence-associated cell proliferation arrest. Importantly, VAD inhibits the incorporation of the repressive histone variant H2A.Z at INK4 gene promoters in senescent cells. Our data underline the importance of vlincRNAs as sensors of cellular environment changes and as mediators of the correct transcriptional response., Senescence is associated with chromatin reorganization in heterochromatin foci. Here the authors show that VAD, a very long intergenic non-coding RNA activated by senescence, inhibits the incorporation of the repressive histone variant H2A.Z to INK4 promoters in senescent cells.
Published: 2015
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15. In vitro release of vascular endothelial growth factor from gadolinium-doped biodegradable microspheres

Author: Michael F. Haller, Patrice Laquerriere, Anthony Z. Faranesh, Monet T. Nastley, Elliot R. McVeigh, Kam W. Leong, Cristina Perez de la Cruz, Laurent-Maquin, Dominique, Department of Biomedical Engineering [Baltimore] ( DBE ), Johns Hopkins University ( JHU ), Laboratory of Cardiac Energetics ( LCE ), National Heart-Lung, and Blood Institute-National Institutes of Health, Supramolecular Structure and Function Resource ( SSFR ), and Office of Research Services-National Institutes of Health
Subjects: MESH : Cell Line, Gadolinium DTPA, Vascular Endothelial Growth Factor A, MESH : Polyglycolic Acid, Polymers, Gadolinium, Contrast Media, law.invention, chemistry.chemical_compound, Polylactic Acid-Polyglycolic Acid Copolymer, law, [ SDV.IB ] Life Sciences [q-bio]/Bioengineering, chemistry.chemical_classification, Drug Carriers, MESH : Gadolinium DTPA, MESH : Cell Division, Infusion Pumps, Implantable, Polymer, Microspheres, Vascular endothelial growth factor, PLGA, Biodegradation, Environmental, MESH : Vascular Endothelial Growth Factor A, Drug delivery, MESH : Microspheres, Cell Division, MESH : Drug Carriers, MRI contrast agent, chemistry.chemical_element, Nanotechnology, In Vitro Techniques, Article, Cell Line, MESH : Biodegradation, Humans, Radiology, Nuclear Medicine and imaging, Lactic Acid, MESH : Polymers, [SDV.IB] Life Sciences [q-bio]/Bioengineering, MESH : Contrast Media, MESH : Cell Div, MESH : Humans, MESH : Infusion Pumps, Implantable, MESH : In Vitro, Biodegradable polymer, chemistry, MESH : Lactic Acid, Electron microscope, Polyglycolic Acid, Biomedical engineering
Abstract: A drug delivery vehicle was constructed that could be visualized noninvasively with MRI. The biodegradable polymer poly(DL-lactic-co-glycolic acid) (PLGA) was used to fabricate microspheres containing vascular endothelial growth factor (VEGF) and the MRI contrast agent gadolinium diethylenetriamine pentaacetic acid (Gd-DTPA). The microspheres were characterized in terms of size, drug and contrast agent encapsulation, and degradation rate. The PLGA microspheres had a mean diameter of 48 +/- 18 microm. The gadolinium loading was 17 +/- 3 microg/mg polymer and the VEGF loading was 163 +/- 22 ng/mg polymer. Electron microscopy revealed that the Gd was dispersed throughout the microspheres and it was confirmed that the Gd loading was sufficient to visualize the microspheres under MRI. VEGF and Gd-DTPA were released from the microspheres in vitro over a period of approximately 6 weeks in three phases: a burst, followed by a slow steady-state, then a rapid steady-state. Biodegradable Gd-doped microspheres can be effectively used to deliver drugs in a sustained manner, while being monitored noninvasively with MRI.
Published: 2004
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16. Selective mRNA degradation by antisense oligonucleotide-2,5A chimeras: Involvement of RNase H and RNase L

Author: Anna Ubysz, Ian Robbins, Bernard Lebleu, Sylvie Vichier-Guerre, Robert W. Sobol, Guillaume Mitta, Bernard Rayner, Institut de Génétique Moléculaire de Montpellier (IGMM), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), LCBO-Laboratoire de Chimie Bio-organique, Université Montpellier 2 - Sciences et Techniques (UM2), Institut de Génétique Moléculaire de Montpellier ( IGMM ), Université de Montpellier ( UM ) -Centre National de la Recherche Scientifique ( CNRS ), Université Montpellier 2 - Sciences et Techniques ( UM2 ), and Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)
Subjects: MESH : Cell Line, MESH : Protein Biosynthesis, MESH : RNA, Messenger, MESH: Ribonuclease, Pancreatic, MESH : Ribonuclease, Pancreatic, MESH : Models, Biological, MESH : Ribonuclease H, Calf Thymus, Biochemistry, MESH: Nucleic Acid Hybridization, Mice, MESH: Oligonucleotides, Antisense, MESH : Tumor Cells, Cultured, Gene expression, Tumor Cells, Cultured, MESH: Animals, MESH: Ribonuclease H, Calf Thymus, 0303 health sciences, MESH: Spodoptera, biology, 030302 biochemistry & molecular biology, Nucleic Acid Hybridization, Translation (biology), General Medicine, MESH : Nucleic Acid Hybridization, 3. Good health, MESH: Protein Biosynthesis, Antisense oligonucleotides, [SDV.OT]Life Sciences [q-bio]/Other [q-bio.OT], RNase P, Ribonuclease H, Spodoptera, MESH : Spodoptera, Models, Biological, Cell Line, 03 medical and health sciences, MESH : Mice, Animals, Humans, MESH: Tumor Cells, Cultured, RNA, Messenger, [ SDV.OT ] Life Sciences [q-bio]/Other [q-bio.OT], RNase H, MESH: Mice, MESH: RNA, Messenger, 030304 developmental biology, MESH: Humans, Oligonucleotide, MESH : Humans, MESH : Oligonucleotides, Antisense, MESH: Models, Biological, RNA, Ribonuclease, Pancreatic, Oligonucleotides, Antisense, MESH: Cell Line, RNase MRP, Protein Biosynthesis, biology.protein, MESH : Animals
Abstract: Antisense oligonucleotides (ON) allow the specific control of gene expression and phosphorothioate derivatives are currently being evaluated for possible clinical applications. Numerous second generation ON analogues with improved pharmacological properties have been described. Most of them, however, do not recruit RNase H, which is known to increase ON potency by eliciting the specific degradation of the target RNA. Silverman, Torrence and colleagues, have conjugated 2.5 A to natural antisense ON and demonstrated the preferential cleavage of a target RNA in cell-free and intact cell experiments. We have established for the first time that RNase H-incompetent ON, viz, α-anomeric ON analogues, can be converted into sequence-specific nucleases upon conjugation to 2,5 A. The use of α-ON and β-ON-2,5 A chimeras has allowed us to delineate the part played by RNase H and RNase L in target RNA degradation and translation arrest. Finally, the present studies have revealed limitations which are encountered in the choice of a suitable target for such ON-2,5 A chimeras.
Published: 1998
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17. Single-cell analysis of thymocyte differentiation: identification of transcription factor interactions and a major stochastic component in αβ-lineage commitment

Author: Sophie Ezine, Benedita Rocha, Serge M. Candéias, Amine Boudil, Agnès Legrand, Lamia Skhiri, Laetitia Gautreau-Rolland, Marie Bedora-Faure, Valérie Pasqualetto, Différenciation et physiologie des lymphocytes T (U1020), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Paris Descartes - Paris 5 (UPD5), Laboratoire de Chimie et Biologie des Métaux (LCBM - UMR 5249), Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Laboratoire de Chimie et Biologie des Métaux ( LCBM - UMR 5249 ), Université Joseph Fourier - Grenoble 1 ( UJF ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Centre National de la Recherche Scientifique ( CNRS ) -Université Grenoble Alpes ( UGA ), Université Paris Descartes - Paris 5 ( UPD5 ), Différenciation et physiologie des lymphocytes T ( U1020 ), Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), and Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG)
Subjects: MESH : Cell Line, MESH : Transcription Factors, Cellular differentiation, lcsh:Medicine, MESH : Thymocytes, Mice, 0302 clinical medicine, Single-cell analysis, Gene expression, [ SDV.IMM ] Life Sciences [q-bio]/Immunology, MESH: Animals, lcsh:Science, 0303 health sciences, Multidisciplinary, Thymocytes, GATA3, Cell Differentiation, MESH: Transcription Factors, medicine.anatomical_structure, MESH : Single-Cell Analysis, MESH: Stochastic Processes, [SDV.IMM]Life Sciences [q-bio]/Immunology, Single-Cell Analysis, MESH : Cell Differentiation, Research Article, MESH: Cell Differentiation, MESH: Thymocytes, T cell, MESH : Cell Lineage, MESH : Stochastic Processes, Biology, Cell Line, 03 medical and health sciences, MESH: Gene Expression Profiling, MESH : Mice, medicine, Animals, Cell Lineage, Transcription factor, MESH: Mice, 030304 developmental biology, Stochastic Processes, MESH : Gene Expression Profiling, Gene Expression Profiling, lcsh:R, MESH: Cell Lineage, Molecular biology, MESH: Cell Line, Gene expression profiling, Cell culture, lcsh:Q, MESH : Animals, 030215 immunology, Transcription Factors, MESH: Single-Cell Analysis
Abstract: International audience; T cell commitment and $\alpha\beta$/$\gamma\delta$ lineage specification in the thymus involves interactions between many different genes. Characterization of these interactions thus requires a multiparameter analysis of individual thymocytes. We developed two efficient single-cell methods: (i) the quantitative evaluation of the co-expression levels of nine different genes, with a plating efficiency of 99-100% and a detection limit of 2 mRNA molecules/cell; and (ii) single-cell differentiation cultures, in the presence of OP9 cells transfected with the thymus Notch1 ligand DeltaL4. We show that during T cell commitment, Gata3 has a fundamental, dose-dependent role in maintaining Notch1 expression, with thymocytes becoming T-cell-committed when they co-express Notch1, Gata3 and Bc11b. Of the transcription factor expression patterns studied here, only that of Bcl11b was suggestive of a role in Pu1 down-regulation. Individual thymocytes became $\alpha\beta$$\gamma\delta$-lineage-committed at very different stages (from the TN2a stage onwards). However, 20% of TN3 cells are not $\alpha\beta$/$\gamma\delta$ lineage committed and TN4 cells comprise two main subpopulations with different degrees of maturity. The existence of a correlation between differentiation potential and expression of the pre-TCR showed that 83% of $\alpha\beta$-committed cells do not express the pre-TCR and revealed a major stochastic component in $\alpha\beta$-lineage specification.
Published: 2013
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18. Cell-penetrating anti-GFAP VHH and corresponding fluorescent fusion protein VHH-GFP spontaneously cross the blood-brain barrier and specifically recognize astrocytes: application to brain imaging

Author: Jean-Pierre Bourgeois, Anne-Marie Le Sourd, Fabienne Glacial, Pierre Lafaye, Susanna Celli, Pierre-Olivier Couraud, Babette B. Weksler, Ignacio A. Romero, Tengfei Li, Salah Mecheri, François Rougeon, Production de Protéines Recombinantes et d'Anticorps (Plate-Forme), Institut Pasteur [Paris] (IP), Gènes, Synapses et Cognition (CNRS - UMR3571 ), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Cochin (IC UM3 (UMR 8104 / U1016)), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Biologie des Interactions Hôte-Parasite, Division of Hematology and Oncology, Weill Medical College of Cornell University [New York], Department of Life, Health and Chemical Sciences, The Open University [Milton Keynes] (OU), Génétique et Biochimie du Développement, Institut Pasteur [Paris], Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre National de la Recherche Scientifique (CNRS)-Institut Pasteur [Paris], Gènes, Synapses et Cognition, Institut Pasteur [Paris]-Université Paris Diderot - Paris 7 ( UPD7 ) -Centre National de la Recherche Scientifique ( CNRS ), Institut National de la Santé et de la Recherche Médicale ( INSERM ), Institut Cochin ( UM3 (UMR 8104 / U1016) ), Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Institut Pasteur [Paris]-Centre National de la Recherche Scientifique ( CNRS ), and The Open University [Milton Keynes] ( OU )
Subjects: MESH : Cell Line, [SDV.IB.IMA]Life Sciences [q-bio]/Bioengineering/Imaging, Plasmodium berghei, MESH : Immunohistochemistry, [SDV.NEU.NB]Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC]/Neurobiology, Fluorescent Antibody Technique, Biochemistry, law.invention, Green fluorescent protein, MESH : Blood-Brain Barrier, MESH: Blood-Brain Barrier, Mice, 0302 clinical medicine, law, MESH: Glial Fibrillary Acidic Protein, MESH : Female, MESH: Animals, MESH: Fluorescent Antibody Technique, [ SDV.IB.IMA ] Life Sciences [q-bio]/Bioengineering/Imaging, 0303 health sciences, biology, Glial fibrillary acidic protein, intrabodies, MESH: Enzyme-Linked Immunosorbent Assay, Brain, MESH : Enzyme-Linked Immunosorbent Assay, MESH : Glial Fibrillary Acidic Protein, Immunohistochemistry, nanobodies, MESH : Mutagenesis, Site-Directed, 3. Good health, MESH: Mutagenesis, Site-Directed, medicine.anatomical_structure, Blood-Brain Barrier, [ SDV.NEU.NB ] Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC]/Neurobiology, MESH : Single-Domain Antibodies, Recombinant DNA, Female, Antibody, Biotechnology, MESH: Cell Line, Tumor, MESH : Recombinant Fusion Proteins, Recombinant Fusion Proteins, [ SDV.IMM.IMM ] Life Sciences [q-bio]/Immunology/Immunotherapy, fluobodies, Enzyme-Linked Immunosorbent Assay, MESH : Mice, Inbred C57BL, Astrocytoma, Blood–brain barrier, MESH: Single-Domain Antibodies, camelids, Cell Line, 03 medical and health sciences, MESH: Brain, Antigen, MESH: Mice, Inbred C57BL, Cell Line, Tumor, MESH : Mice, Glial Fibrillary Acidic Protein, Genetics, medicine, MESH: Recombinant Fusion Proteins, MESH: Plasmodium berghei, Animals, Humans, Molecular Biology, MESH: Mice, MESH : Plasmodium berghei, 030304 developmental biology, MESH: Humans, MESH : Cell Line, Tumor, MESH : Humans, MESH : Astrocytes, MESH : Astrocytoma, MESH: Immunohistochemistry, [SDV.IMM.IMM]Life Sciences [q-bio]/Immunology/Immunotherapy, Single-Domain Antibodies, central nervous system, Fusion protein, Molecular biology, MESH: Cell Line, MESH: Astrocytes, Mice, Inbred C57BL, MESH : Fluorescent Antibody Technique, MESH: Astrocytoma, MESH : Brain, Astrocytes, biology.protein, Mutagenesis, Site-Directed, Paratope, MESH : Animals, BBB, MESH: Female, 030217 neurology & neurosurgery
Abstract: International audience; Antibodies normally do not cross the blood-brain barrier (BBB) and cannot bind an intracellular cerebral antigen. We demonstrate here for the first time that a new class of antibodies can cross the BBB without treatment. Camelids produce native homodimeric heavy-chain antibodies, the paratope being composed of a single-variable domain called VHH. Here, we used recombinant VHH directed against human glial fibrillary acidic protein (GFAP), a specific marker of astrocytes. Only basic VHHs (e.g., pI=9.4) were able to cross the BBB in vitro (7.8 vs. 0% for VHH with pI=7.7). By intracarotid and intravenous injections into live mice, we showed that these basic VHHs are able to cross the BBB in vivo, diffuse into the brain tissue, penetrate into astrocytes, and specifically label GFAP. To analyze their ability to be used as a specific transporter, we then expressed a recombinant fusion protein VHH-green fluorescent protein (GFP). These "fluobodies" specifically labeled GFAP on murine brain sections, and a basic variant (pI=9.3) of the fusion protein VHH-GFP was able to cross the BBB and to label astrocytes in vivo. The potential of VHHs as diagnostic or therapeutic agents in the central nervous system now deserves attention.
Published: 2012
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19. Live-Cell Imaging Reveals Multiple Interactions between Epstein-Barr Virus Nuclear Antigen 1 and Cellular Chromatin during Interphase and Mitosis

Author: Maïté Coppey-Moisan, Tristan Piolot, Vincent Maréchal, Gabriel Dos Reis, Christophe Klein, Nathalie Jourdan, Marc Tramier, Frédérique Quignon, Aude Jobart-Malfait, Adaptation Biologique et Vieillissement = Biological Adaptation and Ageing (B2A), Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS)-Institut de Biologie Paris Seine (IBPS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Institut Jacques Monod (IJM (UMR_7592)), Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS), Centre de Recherche des Cordeliers (CRC (UMR_S 872)), Université Pierre et Marie Curie - Paris 6 (UPMC)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut de Génétique et Développement de Rennes (IGDR), Université de Rennes (UR)-Centre National de la Recherche Scientifique (CNRS)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), Centre de Recherche des Cordeliers ( CRC (UMR_S 872) ), Université Pierre et Marie Curie - Paris 6 ( UPMC ) -Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Institut Jacques Monod ( IJM ), Université Paris Diderot - Paris 7 ( UPD7 ) -Centre National de la Recherche Scientifique ( CNRS ), Institut de Génétique et Développement de Rennes ( IGDR ), Université de Rennes 1 ( UR1 ), Université de Rennes ( UNIV-RENNES ) -Université de Rennes ( UNIV-RENNES ) -Centre National de la Recherche Scientifique ( CNRS ) -Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), UPMC, INSERM, Université Paris Descartes - Paris 5 (UPD5)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Centre National de la Recherche Scientifique (CNRS)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), and De Villemeur, Hervé
Subjects: MESH : Cell Line, viruses, [SDV]Life Sciences [q-bio], Gene Expression, MESH : Hepatocytes, MESH: Hepatocytes, Chromatin bridge, 0302 clinical medicine, MESH : Protein Transport, hemic and lymphatic diseases, MESH : Protein Stability, 0303 health sciences, MESH : Chromatin, MESH : Epstein-Barr Virus Nuclear Antigens, MESH : Interphase, Protein Stability, RNA-Binding Proteins, MESH : Protein Binding, Cell cycle, MESH : Mitosis, Chromatin, Virus-Cell Interactions, Protein Transport, 030220 oncology & carcinogenesis, Premature chromosome condensation, Interphase, MESH: Cell Nucleolus, MESH : Carrier Proteins, MESH: Epstein-Barr Virus Nuclear Antigens, Cell Nucleolus, Plasmids, Protein Binding, MESH: Protein Transport, MESH: Gene Expression, Immunology, Mitosis, MESH: Carrier Proteins, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Biology, Microbiology, Cell Line, MESH: Chromatin, MESH: Interphase, 03 medical and health sciences, G2 phase, Prophase, MESH: Plasmids, MESH: Protein Stability, Virology, MESH: HMGB2 Protein, HMGB2 Protein, Humans, MESH: Protein Binding, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, 030304 developmental biology, MESH: Humans, [ SDV.BC ] Life Sciences [q-bio]/Cellular Biology, MESH : Humans, MESH: Mitosis, MESH : HMGB2 Protein, Molecular biology, MESH: Cell Line, MESH : Gene Expression, Epstein-Barr Virus Nuclear Antigens, MESH : Plasmids, Insect Science, MESH : Cell Nucleolus, Hepatocytes, Carrier Proteins
Abstract: Epstein-Barr virus (EBV) establishes a life-long latent infection in humans. In proliferating latently infected cells, EBV genomes persist as multiple episomes that undergo one DNA replication event per cell cycle and remain attached to the mitotic chromosomes. EBV nuclear antigen 1 (EBNA-1) binding to the episome and cellular genome is essential to ensure proper episome replication and segregation. However, the nature and regulation of EBNA-1 interaction with chromatin has not been clearly elucidated. This activity has been suggested to involve EBNA-1 binding to DNA, duplex RNA, and/or proteins. EBNA-1 binding protein 2 (EBP2), a nucleolar protein, has been proposed to act as a docking protein for EBNA-1 on mitotic chromosomes. However, there is no direct evidence thus far for EBP2 being associated with EBNA-1 during mitosis. By combining video microscopy and Förster resonance energy transfer (FRET) microscopy, we demonstrate here for the first time that EBNA-1 and EBP2 interact in the nucleoplasm, as well as in the nucleoli during interphase. However, in strong contrast to the current proposed model, we were unable to observe any interaction between EBNA-1 and EBP2 on mitotic chromosomes. We also performed a yeast double-hybrid screening, followed by a FRET analysis, that led us to identify HMGB2 (high-mobility group box 2), a well-known chromatin component, as a new partner for EBNA-1 on chromatin during interphase and mitosis. Although the depletion of HMGB2 partly altered EBNA-1 association with chromatin in HeLa cells during interphase and mitosis, it did not significantly impact the maintenance of EBV episomes in Raji cells.
Published: 2012
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20. CXCR3 antagonism of SDF-1(5-67) restores trabecular function and prevents retinal neurodegeneration in a rat model of ocular hypertension

Author: William Rostène, Julie Frugier, Françoise Baleux, David Godefroy, Isabelle Célérier, Serge Picaud, Françoise Brignole-Baudouin, Julie Degardin, Jeffrey K. Harrison, Christophe Baudouin, José Sahel, Alexandre Denoyer, Institut de la Vision, Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre de recherche des Cordeliers - Equipe 20 'ingénierie des connaissances en santé', Centre de Recherche des Cordeliers (CRC (UMR_S 872)), Université Paris Descartes - Paris 5 (UPD5)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Université Paris Descartes - Paris 5 (UPD5)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), University of Florida [Gainesville] (UF), Université Paris Descartes - Paris 5 (UPD5), Université Paris Descartes - Faculté de Pharmacie de Paris (UPD5 Pharmacie), Chimie des biomolécules, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Centre Hospitalier National d'Ophtalmologie des Quinze-Vingts (CHNO), Centre National de la Recherche Scientifique ( CNRS ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Université Pierre et Marie Curie - Paris 6 ( UPMC ), CIC Quinze-Vingts, Assistance publique - Hôpitaux de Paris (AP-HP)-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre Hospitalier National d'Ophtalmologie des Quinze-Vingts, Centre de Recherche des Cordeliers ( CRC (UMR_S 872) ), Université Pierre et Marie Curie - Paris 6 ( UPMC ) -Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ) -Université Pierre et Marie Curie - Paris 6 ( UPMC ) -Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Department of Pharmacology & Therapeutics, College of Medicine, University of Florida, Laboratory of Toxicology, Faculty of Pharmaceutical and Biological Sciences, Université Paris Descartes - Paris 5 ( UPD5 ), Chimie des Biomolécules, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique ( CNRS ), This work was supported by INSERM grants for AD, DG, JD, WR and CB, by a UPMC grant for IC and JF, and by an NIH grant for JKH (AI058256)., Université Pierre et Marie Curie - Paris 6 ( UPMC ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Assistance publique - Hôpitaux de Paris (AP-HP) ( APHP ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre Hospitalier National d'Ophtalmologie des Quinze-Vingts, Université Pierre et Marie Curie - Paris 6 (UPMC)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Université Pierre et Marie Curie - Paris 6 (UPMC)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Faculté de Pharmacie de Paris - Université Paris Descartes (UPD5 Pharmacie), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), and Godefroy, David
Subjects: Retinal degeneration, Male, Visual System, Ocular hypertension, Apoptosis, MESH : Cytoprotection, MESH : Receptors, CXCR4, MESH : Receptors, CXCR3, 0302 clinical medicine, MESH: Animals, lcsh:Science, MESH: Stress, Physiological, MESH : Glaucoma, 0303 health sciences, Neurodegeneration, Anatomy, MESH : Chemokine CXCL12, Sensory Systems, 3. Good health, [SDV] Life Sciences [q-bio], MESH : Retinal Degeneration, Medicine, MESH: Chemokine CXCL12, MESH: Glaucoma, medicine.medical_specialty, MESH: Enzyme Activation, Receptors, CXCR4, Receptors, CXCR3, MESH: Retinal Degeneration, Ocular Anatomy, Caspase 3, MESH: Receptors, CXCR4, MESH: Receptors, CXCR3, 03 medical and health sciences, Humans, Rats, Long-Evans, MESH: Vision, Ocular, Biology, Intraocular Pressure, MESH: Humans, [ SDV ] Life Sciences [q-bio], lcsh:R, MESH : Humans, Glaucoma, medicine.disease, MESH : Disease Models, Animal, eye diseases, Chemokine CXCL12, MESH: Cell Line, MESH: Ocular Hypertension, Enzyme Activation, MESH: Cytoprotection, Ophthalmology, Endocrinology, chemistry, 030221 ophthalmology & optometry, Rat, lcsh:Q, Ocular Hypertension, Trabecular meshwork, MESH: Disease Models, Animal, MESH : Apoptosis, Neuroscience, MESH : Cell Line, Intraocular pressure, Anatomy and Physiology, genetic structures, [SDV]Life Sciences [q-bio], MESH : Vision, Ocular, lcsh:Medicine, chemistry.chemical_compound, MESH: Caspase 3, Molecular Cell Biology, MESH : Caspase 3, MESH : Rats, Long-Evans, MESH : Intraocular Pressure, Multidisciplinary, Cell Death, MESH : Rats, Retinal Degeneration, Animal Models, MESH : Trabecular Meshwork, medicine.anatomical_structure, Research Article, MESH: Rats, MESH : Male, Cell Line, Model Organisms, MESH: Intraocular Pressure, MESH: Rats, Long-Evans, Stress, Physiological, Trabecular Meshwork, Ocular System, Internal medicine, medicine, MESH : Ocular Hypertension, Animals, MESH : Stress, Physiological, Vision, Ocular, 030304 developmental biology, MESH: Apoptosis, [ SDV.BC.BC ] Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC], Retinal, MESH: Male, Rats, MESH: Trabecular Meshwork, Disease Models, Animal, Cytoprotection, MESH : Animals, sense organs, MESH : Enzyme Activation
Abstract: International audience; Glaucoma, the most common cause of irreversible blindness, is a neuropathy commonly initiated by pathological ocular hypertension due to unknown mechanisms of trabecular meshwork degeneration. Current antiglaucoma therapy does not target the causal trabecular pathology, which may explain why treatment failure is often observed. Here we show that the chemokine CXCL12, its truncated form SDF-1(5-67), and the receptors CXCR4 and CXCR3 are expressed in human glaucomatous trabecular tissue and a human trabecular cell line. SDF-1(5-67) is produced under the control of matrix metallo-proteinases, TNF-α, and TGF-β2, factors known to be involved in glaucoma. CXCL12 protects in vitro trabecular cells from apoptotic death via CXCR4 whereas SDF-1(5-67) induces apoptosis through CXCR3 and caspase activation. Ocular administration of SDF-1(5-67) in the rat increases intraocular pressure. In contrast, administration of a selective CXCR3 antagonist in a rat model of ocular hypertension decreases intraocular pressure, prevents retinal neurodegeneration, and preserves visual function. The protective effect of CXCR3 antagonism is related to restoration of the trabecular function. These data demonstrate that proteolytic cleavage of CXCL12 is involved in trabecular pathophysiology, and that local administration of a selective CXCR3 antagonist may be a beneficial therapeutic strategy for treating ocular hypertension and subsequent retinal degeneration.
Published: 2012
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21. Induction of GADD34 is necessary for dsRNA-dependent interferon-β production and participates in the control of Chikungunya virus infection

Author: Evelina Gatti, Till Wenger, Nuno Cláudio, Enrico K. Schmidt, Alexandre Dalet, Delphine Judith, Thérèse Couderc, Philippe Pierre, Voahirana Camosseto, Giovanna Clavarino, Marc Lecuit, Centre d'Immunologie de Marseille - Luminy (CIML), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Laboratoire Adaptation et pathogénie des micro-organismes [Grenoble] (LAPM), Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS), Biologie des Infections - Biology of Infection, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre d'infectiologie Necker-Pasteur [CHU Necker], Institut Pasteur [Paris] (IP)-CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Microorganismes et Barrières de l'Hôte (Equipe avenir), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU), Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF), Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Pasteur [Paris]-CHU Necker - Enfants Malades [AP-HP], Centre d'Immunologie de Marseille - Luminy ( CIML ), Aix Marseille Université ( AMU ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Laboratoire Adaptation et pathogénie des micro-organismes [Grenoble] ( LAPM ), Université Joseph Fourier - Grenoble 1 ( UJF ) -Centre National de la Recherche Scientifique ( CNRS ), Biologie des Infections, Institut Pasteur [Paris]-Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Assistance publique - Hôpitaux de Paris (AP-HP)-CHU Necker - Enfants Malades [AP-HP], Equipe avenir Microorganismes et Barrières de l'Hôte, and Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale ( INSERM )
Subjects: MESH : Cell Line, MESH: Interferon Type I, MESH: 3T3 Cells, viruses, MESH: Thapsigargin, MESH: Unfolded Protein Response, Mice, eIF-2 Kinase, [ SDV.MP ] Life Sciences [q-bio]/Microbiology and Parasitology, Protein Phosphatase 1, MESH : eIF-2 Kinase, Protein biosynthesis, MESH: Animals, Phosphorylation, Biology (General), MESH: Alphavirus Infections, 0303 health sciences, Protein translation, MESH: Protein Phosphatase 1, Messenger RNA, 030302 biochemistry & molecular biology, MESH : Thapsigargin, 3T3 Cells, 3. Good health, MESH : Activating Transcription Factor 4, [SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology, Interferon Type I, MESH : Unfolded Protein Response, Thapsigargin, Cytokines, Chikungunya infection, Chikungunya virus, MESH : Poly I-C, Research Article, medicine.drug, MESH : 3T3 Cells, MESH : Interleukin-6, QH301-705.5, MESH : Protein Phosphatase 1, Immunology, Immunoblotting, Biology, MESH : Chikungunya virus, Microbiology, MESH: Poly I-C, Cell Line, MESH: eIF-2 Kinase, 03 medical and health sciences, MESH : Interferon-beta, MESH : Interferon Type I, MESH: RNA, Double-Stranded, Virology, MESH : Mice, MESH : Fibroblasts, Genetics, medicine, MESH: Activating Transcription Factor 4, Animals, Protein kinase A, Molecular Biology, MESH: Mice, MESH : Alphavirus Infections, RNA, Double-Stranded, 030304 developmental biology, EIF-2 kinase, MESH: Interferon-beta, Alphavirus Infections, Interleukin-6, MESH : RNA, Double-Stranded, Protein phosphatase 1, MESH: Chikungunya virus, Interferon-beta, Fibroblasts, RC581-607, Activating Transcription Factor 4, Molecular biology, Protein kinase R, MESH: Interleukin-6, MESH: Cell Line, Poly I-C, MESH: Fibroblasts, Unfolded Protein Response, Unfolded protein response, biology.protein, Chikungunya Fever, Parasitology, MESH : Animals, Immunologic diseases. Allergy, Protein synthesis, Interferon type I
Abstract: Nucleic acid sensing by cells is a key feature of antiviral responses, which generally result in type-I Interferon production and tissue protection. However, detection of double-stranded RNAs in virus-infected cells promotes two concomitant and apparently conflicting events. The dsRNA-dependent protein kinase (PKR) phosphorylates translation initiation factor 2-alpha (eIF2α) and inhibits protein synthesis, whereas cytosolic DExD/H box RNA helicases induce expression of type I-IFN and other cytokines. We demonstrate that the phosphatase-1 cofactor, growth arrest and DNA damage-inducible protein 34 (GADD34/Ppp1r15a), an important component of the unfolded protein response (UPR), is absolutely required for type I-IFN and IL-6 production by mouse embryonic fibroblasts (MEFs) in response to dsRNA. GADD34 expression in MEFs is dependent on PKR activation, linking cytosolic microbial sensing with the ATF4 branch of the UPR. The importance of this link for anti-viral immunity is underlined by the extreme susceptibility of GADD34-deficient fibroblasts and neonate mice to Chikungunya virus infection., Author Summary Nucleic acids detection by multiple molecular sensors results in type-I interferon production, which protects cells and tissues from viral infections. At the intracellular level, the detection of double-stranded RNA by one of these sensors, the dsRNA-dependent protein kinase also leads to the profound inhibition of protein synthesis. We describe here that the inducible phosphatase 1 co-factor Ppp1r15a/GADD34, a well known player in the endoplasmic reticulum unfolded protein response (UPR), is activated during double-stranded RNA detection and is absolutely necessary to allow cytokine production in cells exposed to poly I:C or Chikungunya virus. Our data shows that the cellular response to nucleic acids can reveal unanticipated connections between innate immunity and fundamental stress pathways, such as the ATF4 branch of the UPR.
Published: 2012
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22. c-mip down-regulates NF-κB activity and promotes apoptosis in podocytes

Author: Vincent Audard, Marina Candelier, Laure-Hélène Noël, André Pawlak, Georges Guellaen, Nabila Hamdaoui, Dominique Desvaux, Shao-yu Zhang, Philippe Lang, Djillali Sahali, Qingfeng Fan, Virginie Ory, Guellaen, Georges, Institut Mondor de Recherche Biomédicale (IMRB), Institut National de la Santé et de la Recherche Médicale (INSERM)-IFR10-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12), Service d'histologie, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Henri Mondor-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12), Institut Cochin (IC UM3 (UMR 8104 / U1016)), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Service d'anatomie pathologique [CHU Necker], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Service de néphrologie et transplantation, Institut francilien de recheche en nephrologie et transplantation, IFRNT, This work was supported in part by an Avenir Program from INSERM, a grant from the French Kidney Foundation, Association pour l'Utilisation du Rein Artificiel (AURA), Assistance Publique des Hôpitaux de Paris (AP-HP, Programme hospitalier de recherche clinique) and a grant from the Fondation pour la Recherche Médicale (FRM)., Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12) - Institut National de la Santé et de la Recherche Médicale (INSERM) - IFR10, Assistance publique - Hôpitaux de Paris (AP-HP) - Hôpital Henri Mondor - Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12), Institut Cochin (UM3 (UMR 8104 / U1016)), Université Paris Descartes - Paris 5 (UPD5) - Institut National de la Santé et de la Recherche Médicale (INSERM) - Centre National de la Recherche Scientifique (CNRS), Assistance publique - Hôpitaux de Paris (AP-HP) - CHU Necker - Enfants Malades [AP-HP], and Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12)-IFR10
Subjects: Male, MESH : Cell Line, Nephrotic Syndrome, MESH: NF-kappa B, Apoptosis, Idiopathic Nephrotic Syndrome, MESH: Down-Regulation, Podocyte, MESH : Down-Regulation, Mice, chemistry.chemical_compound, 0302 clinical medicine, MESH: Transcription Factor RelA, MESH: Caspase 3, MESH: Up-Regulation, MESH: Microscopy, Confocal, MESH : Caspase 3, MESH: Animals, MESH: In Situ Nick-End Labeling, MESH : Up-Regulation, 0303 health sciences, Microscopy, Confocal, Caspase 3, Podocytes, NF-kappa B, MESH : Adult, MESH : Podocytes, Pathophysiology, MESH : Mice, Transgenic, Up-Regulation, medicine.anatomical_structure, 030220 oncology & carcinogenesis, MESH : NF-kappa B, MESH : In Situ Nick-End Labeling, MESH : Carrier Proteins, Adult, MESH: Mice, Transgenic, MESH : Nephrotic Syndrome, MESH : Male, Down-Regulation, Mice, Transgenic, MESH: Carrier Proteins, Biology, Cell Line, Pathology and Forensic Medicine, 03 medical and health sciences, In vivo, MESH : Mice, MESH: Podocytes, In Situ Nick-End Labeling, medicine, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, Animals, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH : Microscopy, Confocal, MESH: Mice, Adaptor Proteins, Signal Transducing, 030304 developmental biology, MESH: Humans, MESH: Apoptosis, MESH : Transcription Factor RelA, MESH : Humans, Transcription Factor RelA, NF-κB, MESH: Adult, MESH: Male, MESH: Cell Line, Antiapoptotic protein, chemistry, Cancer research, MESH : Animals, MESH: Nephrotic Syndrome, Carrier Proteins, Spatial relationship, MESH : Apoptosis
Abstract: International audience; The mechanisms of podocyte disorders in cases of idiopathic nephrotic syndrome (INS) are complex and remain incompletely elucidated. The abnormal regulation of NF-κB may play a key role in the pathophysiology of these podocyte diseases, but at present, NF-κB has not been thoroughly investigated. In this study, we report that induction of c-mip in podocytes of patients with INS is associated with a down-regulation of RelA, a potent antiapoptotic factor that belongs to the NF-κB family. Overexpression of c-mip in differentiated podocytes promotes apoptosis by inducing caspase-3 activity and up-regulating the proapoptotic protein Bax, whereas the overall levels of the antiapoptotic protein Bcl-2 was concomitantly decreased. The associated overexpression of RelA prevented the proapoptotic effects of c-mip. In addition, the targeted induction of c-mip in podocytes in vivo inhibited the expression of the RelA protein and increased the Bax/Bcl-2 ratio. The expression of both c-mip and active caspase-3 increased in focal and segmental glomerulosclerosis biopsies, and both proteins displayed a close spatial relationship. These results suggest that alterations in NF-κB activity might result from the up-regulation of c-mip and are likely to contribute to podocyte disorders in cases of INS.
Published: 2012
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23. Class-specific effector functions of therapeutic antibodies

Author: Pascal, Virginie, Laffleur, Brice, Cogné, Michel, Contrôle de la Réponse Immune B et des Lymphoproliférations ( CRIBL ), Université de Limoges ( UNILIM ) -Génomique, Environnement, Immunité, Santé, Thérapeutique ( GEIST FR CNRS 3503 ) -Centre National de la Recherche Scientifique ( CNRS ), Université de Limoges ( UNILIM ), Contrôle de la Réponse Immune B et des Lymphoproliférations (CRIBL), Université de Limoges (UNILIM)-Génomique, Environnement, Immunité, Santé, Thérapeutique (GEIST FR CNRS 3503)-Centre National de la Recherche Scientifique (CNRS), and Université de Limoges (UNILIM)
Subjects: MESH : Cell Line, MESH: Immunoglobulins, Cell Survival, Immunoglobulins, Apoptosis, Antibodies, MESH : Antibodies, Cell Line, MESH: Antibodies, Monoclonal, MESH : Immunoglobulins, Mice, MESH : Mice, Animals, Humans, [ SDV.IMM ] Life Sciences [q-bio]/Immunology, MESH: Animals, MESH: Mice, MESH : Cell Survival, MESH: Humans, MESH: Antibodies, MESH: Apoptosis, MESH : Humans, Antibodies, Monoclonal, MESH: Cell Line, MESH: Cell Survival, MESH : Antibodies, Monoclonal, [SDV.IMM]Life Sciences [q-bio]/Immunology, MESH : Animals, MESH : Apoptosis
Abstract: International audience; Physiology usually combines polyclonal antibodies of multiple classes in a single humoral response. Beyond their common ability to bind antigens, these various classes of human immunoglobulins carry specific functions which can each serve specific goals. In many cases, the function of a monoclonal therapeutic antibody may thus be modulated according to the class of its constant domains. Depending on the immunoglobulin class, different functional assays will be used in order to evaluate the functional activity of a monoclonal antibody.
Published: 2012
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24. Variable beta-glucans production by different states of Eurotium amstelodami explains differences in inflammatory responses in airway cells

Author: Stéphane Bretagne, Sandrine Roussel, Anne-Pauline Bellanger, Bénédicte Rognon, Laurence Millon, Françoise Botterel, Gabriel Reboux, Laboratoire Chrono-environnement ( LCE ), Université Bourgogne Franche-Comté ( UBFC ) -Centre National de la Recherche Scientifique ( CNRS ) -Université de Franche-Comté ( UFC ), Laboratoire de Parasitologie-Mycologie, Assistance publique - Hôpitaux de Paris (AP-HP)-Hôpital Henri Mondor-Université Paris-Est Créteil Val-de-Marne - Paris 12 ( UPEC UP12 ), Service de parasitologie [Mondor], Laboratoire Chrono-environnement - CNRS - UBFC (UMR 6249) (LCE), Université de Franche-Comté (UFC), Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université Bourgogne Franche-Comté [COMUE] (UBFC)-Centre National de la Recherche Scientifique (CNRS), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Henri Mondor-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12), Centre National de la Recherche Scientifique (CNRS)-Université de Franche-Comté (UFC), and Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université Bourgogne Franche-Comté [COMUE] (UBFC)
Subjects: MESH: Inflammation, MESH : Cell Line, Chemokine, MESH : Spores, Fungal, beta-Glucans, MESH : beta-Glucans, MESH : Cytokines, MESH: Eurotium, Conidium, Immunology and Allergy, MESH : Fungal Proteins, [SDV.MP.MYC]Life Sciences [q-bio]/Microbiology and Parasitology/Mycology, [ SDV.MP.MYC ] Life Sciences [q-bio]/Microbiology and Parasitology/Mycology, MESH : Eurotium, 0303 health sciences, Fungal protein, MESH: Cytokines, biology, MESH: beta-Glucans, General Medicine, Spores, Fungal, MESH: Chemokines, MESH : Epithelial Cells, MESH : Air Pollution, Indoor, MESH: Epithelial Cells, Air Pollution, Indoor, Cytokines, Proteoglycans, MESH: Fungal Proteins, medicine.symptom, Chemokines, Microbiology (medical), Hypha, Hyphae, Inflammation, Fungus, Pathology and Forensic Medicine, Microbiology, Cell Line, Fungal Proteins, 03 medical and health sciences, MESH: Hyphae, MESH: Spores, Fungal, Eurotium, medicine, Humans, MESH : Chemokines, Interleukin 8, 030304 developmental biology, A549 cell, MESH: Humans, MESH : Inflammation, 030306 microbiology, fungi, MESH : Humans, Epithelial Cells, biology.organism_classification, MESH: Cell Line, Immunology, biology.protein, MESH : Hyphae, MESH: Air Pollution, Indoor
Abstract: International audience; Eurotium amstelodami, a mold frequently identified in housing and farm air samples, is a suspected cause of respiratory diseases such as allergic alveolitis, atopic asthma, and organic dust toxic syndrome. This fungus is present in the air in three different states (ascospores, conidia, and hyphae). The aim of this study was to test in vitro the differential inflammatory response of airway cells exposed to 1,3 betaglucanase-treated protein extract (BGPE), from E. amstelodami ascospores, conidia, and hyphae. Confluent cells from the A549 cell line were inoculated with calibrated BGPE issued from the three fungal forms. The levels of eight cytokines and chemokines involved in inflammatory responses were measured after 8 h of exposure. Beta-d-glucan (BDG) was quantified in total fungal extract as well as in the BGPE from the three fungal states. Hyphal BGPE were the only ones to induce a marked inflammatory response and they contain higher quantities of BDG. The present study adds to the growing body of evidence that beta-glucan from fungal hyphae play a crucial role in respiratory diseases.
Published: 2011
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25. Improved antiangiogenic and antitumour activity of the combination of the natural flavonoid fisetin and cyclophosphamide in Lewis lung carcinoma-bearing mice

Author: Daniel Scherman, Guy G. Chabot, Johanne Seguin, Yasmine S. Touil, Unité de pharmacologie chimique et génétique et d'imagerie (UPCGI - UMR 8151 / UMR_S 1022 ), Université Paris Descartes - Paris 5 (UPD5)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Ecole Nationale Supérieure de Chimie de Paris - Chimie ParisTech-PSL (ENSCP), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut de Chimie du CNRS (INC), Ecole Nationale Supérieure de Chimie de Paris - Chimie ParisTech-PSL (ENSCP), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Chabot, Guy, Ecole Nationale Supérieure de Chimie de Paris- Chimie ParisTech-PSL (ENSCP)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), and Ecole Nationale Supérieure de Chimie de Paris- Chimie ParisTech-PSL (ENSCP) - Université Paris Descartes - Paris 5 (UPD5) - Institut National de la Santé et de la Recherche Médicale (INSERM) - Centre National de la Recherche Scientifique (CNRS)
Subjects: MESH: Tumor Burden, Flavonols, MESH : Antineoplastic Combined Chemotherapy Protocols, MESH: Cell Cycle, Pharmacology, Toxicology, MESH : Flavonoids, Mice, 0302 clinical medicine, Cell Movement, Antineoplastic Combined Chemotherapy Protocols, MESH : Cell Proliferation, MESH : Cell Movement, Pharmacology (medical), MESH: Animals, MESH: Endothelial Cells, MESH: Cell Movement, ComputingMilieux_MISCELLANEOUS, MESH : Cell Survival, 0303 health sciences, MESH : Tumor Burden, Neovascularization, Pathologic, Cell Cycle, Tubulin Modulators, endothelial cells, Tumor Burden, 3. Good health, Endothelial stem cell, MESH: Antineoplastic Combined Chemotherapy Protocols, Oncology, MESH: Cell Survival, 030220 oncology & carcinogenesis, Growth inhibition, fisetin, MESH: Antineoplastic Agents, Phytogenic, Article, 03 medical and health sciences, [SDV.CAN] Life Sciences [q-bio]/Cancer, In vivo, MESH : Carcinoma, Lewis Lung, MESH : Cell Cycle, MESH : Angiogenesis Inhibitors, Humans, Antineoplastic Agents, Alkylating, EA⋅hy 926, MESH: Humans, MESH : Endothelial Cells, MESH : Humans, MESH : Neovascularization, Pathologic, MESH: Cyclophosphamide, MESH: Cell Line, Mice, Inbred C57BL, chemistry, Lewis lung carcinoma, cyclophosphamide, MESH: Female, MESH: Flavonoids, MESH: NIH 3T3 Cells, MESH : Cell Line, Cancer Research, Angiogenesis, MESH : NIH 3T3 Cells, Angiogenesis Inhibitors, MESH: Antineoplastic Agents, Alkylating, MESH : Antineoplastic Agents, Phytogenic, Carcinoma, Lewis Lung, chemistry.chemical_compound, angiogenesis, MESH : Female, Cytotoxicity, MESH : Cyclophosphamide, MESH: Angiogenesis Inhibitors, MESH: Carcinoma, Lewis Lung, cytotoxicity, Female, EA*hy 926 endothelial cells, Cell Survival, MESH : Antineoplastic Agents, Alkylating, [SDV.CAN]Life Sciences [q-bio]/Cancer, MESH : Mice, Inbred C57BL, Biology, Cell Line, MESH: Mice, Inbred C57BL, MESH: Cell Proliferation, MESH : Mice, MESH: Tubulin Modulators, Animals, flavonoid, MESH: Mice, Cell Proliferation, 030304 developmental biology, Flavonoids, Cell growth, MESH : Tubulin Modulators, Antineoplastic Agents, Phytogenic, NIH 3T3 Cells, MESH : Animals, antitumour activity, MESH: Neovascularization, Pathologic, Fisetin
Abstract: International audience; PURPOSE: The natural flavonoid fisetin was recently identified as a lead compound that stabilizes endothelial cell microtubules. In this study, we investigated the antiproliferative and antiangiogenic properties of fisetin in vitro and in vivo. METHODS: Fisetin cytotoxicity was evaluated using Lewis lung carcinoma cells (LLC), endothelial cells and NIH 3T3 cells. Endothelial cell (EC) migration and capillary-like structure formation were evaluated using EAhy 926 cells. In vivo tumour growth inhibition studies were performed using LLC-bearing mice treated with fisetin and/or cyclophosphamide (CPA). RESULTS: The fisetin IC(50) was 59 μM for LLC and 77 μM for EC cells, compared to 210 μM for normal NIH 3T3 cells (24 h). Fisetin inhibited EC migration and capillary-like structure formation at non-cytotoxic concentrations (22-44 μM). In mice, fisetin inhibited angiogenesis assessed using the Matrigel plug assay. In LLC-bearing mice, fisetin produced a 67% tumour growth inhibition (223 mg/kg, intraperitoneal), similar to the 66% produced by low-dose CPA (30 mg/kg, subcutaneous). When fisetin and CPA were combined, however, a marked improvement in antitumour activity was observed (92% tumour growth inhibition), with low systemic toxicity. Tumour histology showed decreased microvessel density with either fisetin or CPA alone, and a dramatic decrease after the fisetin/CPA combination. CONCLUSIONS: We have shown that fisetin not only displays in vitro and in vivo antiangiogenic properties, but also can markedly improve the in vivo antitumour effect of CPA. We propose that this drug combination associating a non-toxic dietary flavonoid with a cytotoxic agent could advantageously be used in the treatment of solid tumours.
Published: 2011
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26. Importance of viral genomic composition in modulating glycoprotein content on the surface of influenza virus particles

Author: Jean-Paul Rolland, Christine Moriscot, Emmanuel Giudice, Bruno Lina, Vincent Moules, Olivier Terrier, Guy Schoehn, Emilie Frobert, Thomas Julien, M. Bouscambert-Duchamp, Yi Pu Lin, Béatrice Riteau, Olivier Ferraris, Alan J. Hay, Daniel Thomas, Alexandra Erny, Matthieu Yver, Manuel Rosa-Calatrava, Virologie et pathologie humaine (VirPath), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon, Inserm European Associated Laboratory, University of Dundee, Unit for Virus Host-Cell Interactions [Grenoble] (UVHCI), Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF), Institut de biologie structurale (IBS - UMR 5075 ), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Interactions cellulaires et moléculaires (ICM), Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Centre National de la Recherche Scientifique (CNRS), World Health Organization Collaborating Centre Medical Research Council/National Institute of Medical Research, CNRS, Virologie et pathologie humaine ( VirPath ), Université Claude Bernard Lyon 1 ( UCBL ), Unit of Virus Host Cell Interactions ( UVHCI ), Université Joseph Fourier - Grenoble 1 ( UJF ) -Centre National de la Recherche Scientifique ( CNRS ), Institut de biologie structurale ( IBS - UMR 5075 ), Université Joseph Fourier - Grenoble 1 ( UJF ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Centre National de la Recherche Scientifique ( CNRS ) -Université Grenoble Alpes ( UGA ), Interactions cellulaires et moléculaires ( ICM ), Université de Rennes 1 ( UR1 ), Université de Rennes ( UNIV-RENNES ) -Université de Rennes ( UNIV-RENNES ) -IFR140-Centre National de la Recherche Scientifique ( CNRS ), Université Joseph Fourier - Grenoble 1 (UJF)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), and Université de Rennes (UR)-Centre National de la Recherche Scientifique (CNRS)
Subjects: MESH : Cell Line, viruses, Hemagglutinin Glycoproteins, Influenza Virus, law.invention, MESH: Dogs, Influenza A Virus, H1N1 Subtype, MESH : Dogs, law, Virus Components, MESH: Animals, MESH : Neuraminidase, MESH : Viral Proteins, chemistry.chemical_classification, 0303 health sciences, biology, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], 030302 biochemistry & molecular biology, Spike density, MESH: Hemagglutinin Glycoproteins, Influenza Virus, MESH: Neuraminidase, Neuraminidase (NA), Orthomyxoviridae, Cryo-electron microscopy (cryo-EM), MESH : Influenza A Virus, H1N1 Subtype, MESH : RNA Replicase, Recombinant DNA, MESH: Virion, MESH: Cryoelectron Microscopy, MESH: RNA Replicase, MESH : Influenza A Virus, H3N2 Subtype, Neuraminidase, MESH: Influenza A Virus, H3N2 Subtype, Virus, Cell Line, MESH: Influenza A Virus, H1N1 Subtype, Viral Proteins, 03 medical and health sciences, Dogs, Virology, MESH : Virion, Animals, Humans, Polymerase Gene, Haemagglutinin (HA), 030304 developmental biology, MESH: Humans, Host (biology), Influenza A Virus, H3N2 Subtype, Cryoelectron Microscopy, MESH : Humans, Virion, MESH : Cryoelectron Microscopy, RNA-Dependent RNA Polymerase, biology.organism_classification, MESH: Viral Proteins, Influenza, Viral glycoprotein, MESH: Cell Line, MESH : Hemagglutinin Glycoproteins, Influenza Virus, chemistry, MESH : Animals, Glycoprotein, [ SDV.BBM.BS ] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM]
Abstract: International audience; Despite progress in our knowledge of the internal organisation of influenza virus particles, little is known about the determinants of their morphology and, more particularly, of the actual abundance of structural proteins at the virion level. To address these issues, we used cryo-EM to focus on viral (and host) factors that might account for observed differences in virion morphology and characteristics such as size, shape and glycoprotein (GP) spike density. Twelve recombinant viruses were characterised in terms of their morphology, neuraminidase activity and virus growth. The genomic composition was shown to be important in determining the GP spike density. In particular, polymerase gene segments and especially PB1/PB2 were shown to have a prominent influence in addition to that for HA in determining GP spike density, a feature consistent with a functional link between these virus components important for virus fitness.
Published: 2011
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27. Coenzyme Q as an antiadipogenic factor

Author: Luc Van Gaal, Anne Galinier, Louis Casteilla, Luc Pénicaud, Bart Staels, Sandy Bour, Maria-Carmen Carmona, Sylvie Caspar-Bauguil, Institut des Maladies Métaboliques et Cardiovasculaires (I2MC), Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Université de Toulouse (UT)-Institut National de la Santé et de la Recherche Médicale (INSERM), Neurobiologie, plasticité tissulaire et métabolisme énergétique (NPTME), Université de Toulouse (UT)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS), Antwerp University Hospital [Edegem] (UZA), Récepteurs nucléaires, maladies cardiovasculaires et diabète - U 1011 (RNMCD), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Centre des Sciences du Goût et de l'Alimentation [Dijon] (CSGA), Institut National de la Recherche Agronomique (INRA)-Université de Bourgogne (UB)-Centre National de la Recherche Scientifique (CNRS), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre National de la Recherche Scientifique (CNRS)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Institut National de la Recherche Agronomique (INRA)-Université de Bourgogne (UB), Antwerp University Hospital, Récepteurs nucléaires, maladies cardiovasculaires et diabète (EGID), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille, Droit et Santé-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Institut National de la Recherche Agronomique (INRA)-Université de Bourgogne (UB)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Centre National de la Recherche Scientifique (CNRS), Institut des Maladies Métaboliques et Cardiovasculaires ( I2MC ), Université Paul Sabatier - Toulouse 3 ( UPS ) -Hôpital de Rangueil-Institut National de la Santé et de la Recherche Médicale ( INSERM ), Neurobiologie, plasticité tissulaire et métabolisme énergétique ( NPTME ), Université Paul Sabatier - Toulouse 3 ( UPS ) -Centre National de la Recherche Scientifique ( CNRS ), Antwerp University Hospital [Edegem], Récepteurs nucléaires, maladies cardiovasculaires et diabète ( EGID ), Université de Lille, Droit et Santé-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Institut Pasteur de Lille, Réseau International des Instituts Pasteur ( RIIP ) -Réseau International des Instituts Pasteur ( RIIP ) -Centre Hospitalier Régional Universitaire [Lille] ( CHRU Lille ), Centre des Sciences du Goût et de l'Alimentation [Dijon] ( CSGA ), and Institut National de la Recherche Agronomique ( INRA ) -Université de Bourgogne ( UB ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Centre National de la Recherche Scientifique ( CNRS )
Subjects: Male, Antioxidant, muscle, medicine.medical_treatment, Cellular differentiation, coq(10) deficiency, MESH : Adipocytes, White adipose tissue, Biochemistry, Antioxidants, Mice, 0302 clinical medicine, q(10) deficiency, Adipocyte, MESH: Animals, Enzyme Inhibitors, MESH: Mice, Obese, MESH : Body Weight, General Environmental Science, 0303 health sciences, Adipogenesis, MESH : Adipogenesis, food and beverages, MESH : Mice, Obese, MESH : Ubiquinone, MESH : alpha-Tocopherol, MESH : Antioxidants, MESH : Cell Differentiation, MESH: Adipose Tissue, medicine.medical_specialty, MESH: Alkyl and Aryl Transferases, MESH : Adipose Tissue, MESH : Quinones, 03 medical and health sciences, Humans, Molecular Biology, MESH: Adipocytes, Alkyl and Aryl Transferases, MESH: Humans, [ SDV ] Life Sciences [q-bio], MESH : Humans, MESH: Antioxidants, MESH : Organ Size, MESH: Body Weight, MESH: Cell Line, Mice, Inbred C57BL, Endocrinology, chemistry, Human medicine, Reactive Oxygen Species, MESH: Female, MESH : Dietary Fats, 030217 neurology & neurosurgery, MESH : Cell Line, obesity, Physiology, Ubiquinone, MESH: Organ Size, [SDV]Life Sciences [q-bio], Clinical Biochemistry, alpha-Tocopherol, MESH : Reactive Oxygen Species, Adipose tissue, Mice, Obese, rosiglitazone, chemistry.chemical_compound, Adipocytes, oxidative stress, MESH: Obesity, MESH : Female, MESH: Ubiquinone, Quinones, MESH: Reactive Oxygen Species, Cell Differentiation, Organ Size, adipose tissue, MESH: alpha-Tocopherol, MESH: Enzyme Inhibitors, MESH: Dietary Fats, MESH : Obesity, Female, MESH: Cell Differentiation, uncoupling protein 2, MESH : Male, MESH : Mice, Inbred C57BL, mitochondrial encephalomyopathy, Biology, MESH: Quinones, Cell Line, MESH: Mice, Inbred C57BL, Internal medicine, MESH : Mice, expression, medicine, Animals, MESH: Mice, MESH : Alkyl and Aryl Transferases, 030304 developmental biology, MESH : Enzyme Inhibitors, Body Weight, Cell Biology, Dietary Fats, MESH: Male, Cell culture, Coenzyme Q – cytochrome c reductase, General Earth and Planetary Sciences, MESH : Animals, MESH: Adipogenesis
Abstract: WOS: 000285876900007; International audience; Coenzyme Q (CoQ) is not only the single antioxidant synthesized in humans but also an obligatory element of mitochondrial functions. We have previously reported CoQ deficiency in white adipose tissue of ob/ob mice. We sought to determine (i) whether this deficit exists in all species and its relevance in human obesity and (ii) to what extent CoQ could be involved in adipocyte differentiation. Here we identified in rodents as well as in humans a specific very strong nonlinear negative correlation between CoQ content in subcutaneous adipose tissue and obesity indexes. This striking correlation reveals a threshold value similar in both species. This relative deficit in CoQ content in adipose tissue rapidly took place during the time course of high-fat-diet-induced obesity in mice. Adipocyte differentiation was assessed in vitro using the preadipocyte 3T3-F442A cell line. When CoQ synthesis was inhibited by a pharmacological approach using chlorobenzoic acid, this strongly triggered adipose differentiation. In contrast, adipogenesis was strongly inhibited when a long-term increase in CoQ content was obtained by overexpressing human 4-hydroxy benzoate acid polyprenyltransferase gene. Altogether, these data suggest that a strict level of CoQ remains essential for adipocyte differentiation, and its impairment is associated with obesity.
Published: 2011
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28. Herpes simplex virus type 1-derived recombinant and amplicon vectors

Author: Fraefel , Cornel, Marconi , Peggy, Epstein , Alberto L, Centre de génétique et de physiologie moléculaire et cellulaire ( CGPhiMC ), Université Claude Bernard Lyon 1 ( UCBL ), and Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique ( CNRS )
Subjects: MESH : Cell Line, MESH : Chromosomes, Artificial, Bacterial, MESH : Cell Culture Techniques, MESH : Escherichia coli, MESH : Cloning, Molecular, viruses, MESH : Humans, MESH : Recombination, Genetic, MESH : Helper Viruses, [ SDV.BBM.BM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, MESH : Herpesvirus 1, Human, MESH : Genetic Vectors, MESH : DNA, Recombinant, MESH : Plasmids, MESH : Titrimetry, MESH : Virion, MESH : Electroporation, MESH : Animals, MESH : Transfection
Abstract: International audience; Herpes simplex virus type 1 (HSV-1) is a human pathogen whose lifestyle is based on a long-term dual interaction with the infected host, being able to establish both lytic and latent infections. The virus genome is a 153 kbp double-stranded DNA molecule encoding more than 80 genes. The interest of HSV-1 as gene transfer vector stems from its ability to infect many different cell types, both quiescent and proliferating cells, the very high packaging capacity of the virus capsid, the outstanding neurotropic adaptations that this virus has evolved, and the fact that it never integrates into the cellular chromosomes, thus avoiding the risk of insertional mutagenesis. Two types of vectors can be derived from HSV-1, recombinant vectors and amplicon vectors, and different methodologies have been developed to prepare large stocks of each type of vector. This chapter summarizes (1) the two approaches most commonly used to prepare recombinant vectors through homologous recombination, either in eukaryotic cells or in bacteria, and (2) the two methodologies currently used to generate helper-free amplicon vectors, either using a bacterial artificial chromosome (BAC)-based approach or a Cre/loxP site-specific recombination strategy.
Published: 2011
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29. Interactions with M cells and macrophages as key steps in the pathogenesis of enterohemorrhagic Escherichia coli infections

Author: Benoit Chassaing, Nathalie Pradel, Pierre Sauvanet, Lucie Etienne-Mesmin, Jérémy Denizot, Stéphanie Blanquet-Diot, Arlette Darfeuille-Michaud, Valérie Livrelli, Laboratoire de Microbiologie et Biotechnologie des Environnements Chauds, Université de la Méditerranée - Aix-Marseille 2-Université de Provence - Aix-Marseille 1, Evolution des bactéries pathogènes et susceptibilité génétique de l'hôte (JE2526), Institut National de la Recherche Agronomique (INRA)-Université d'Auvergne - Clermont-Ferrand I (UdA), Microbiologie Environnement Digestif Santé - Clermont Auvergne (MEDIS), INRA Clermont-Ferrand-Theix-Université Clermont Auvergne (UCA), Microbiologie Environnement Digestif Santé (MEDIS), INRA Clermont-Ferrand-Theix-Université Clermont Auvergne [2017-2020] (UCA [2017-2020]), Ministere de l'Enseignement Superieur et de la Recherche [JE2526], and Institut National de Recherche Agronomique [USC-2018]
Subjects: Male, MESH: Escherichia coli O157: genetics,metabolism,physiology, Apoptosis, Pathogenesis, Shiga Toxins, Mice, [SDV.MHEP.MI]Life Sciences [q-bio]/Human health and pathology/Infectious diseases, Toxins, MESH: Animals, Intestinal Mucosa, MESH : Ileum: microbiology,pathology, MESH: Microbial Viability, 0303 health sciences, MESH : Cell Survival, Microscopy, Confocal, 3. Good health, POUVOIR PATHOGENE, MESH: Cell Survival, Medical Microbiology, MESH: Microscopy, Electron, Transmission, Medicine, MESH: Bacterial Translocation, Science, LONG POLAR FIMBRIAE, PEYERS-PATCHES, CROHNS-DISEASE, SALMONELLA-TYPHIMURIUM, TISSUE TROPISM, O157-H7, EXPRESSION, STRAINS, TRANSLOCATION, COLONIZATION, MESH: Vero Cells, Microbiology, MESH : Green Fluorescent Proteins: genetics,metabolism, 03 medical and health sciences, Microscopy, Electron, Transmission, Ileum, In vivo, MESH: Ileum: microbiology,pathology, Humans, MESH : Microscopy, Confocal, Biology, Microbial Pathogens, Escherichia coli, MESH: Intestinal Mucosa: microbiology,pathology, Microbial Viability, MESH: Humans, 030306 microbiology, Macrophages, MESH: Host-Pathogen Interactions, MESH : Humans, MESH : Caco-2 Cells, MESH: Cercopithecus aethiops, [SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, In vitro, MESH: Cell Line, MESH : Escherichia coli O157: genetics,metabolism,physiology, Bacterial Translocation, BACTERIOSE, Vero cell, MESH : Bacterial Translocation, Bacterial pathogens, MESH : Cercopithecus aethiops, MESH: Shiga Toxins: metabolism, MESH : Apoptosis, MESH : Vero Cells, MESH : Cell Line, Time Factors, MESH : Models, Biological, [ SDV.MP.BAC ] Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, medicine.disease_cause, MESH : Macrophages: microbiology,pathology,ultrastructure, Peyer's Patches, Intestinal mucosa, Gastrointestinal tract, hemic and lymphatic diseases, INFECTION, Chlorocebus aethiops, Gram Negative, MESH: Microscopy, Confocal, Enterohaemorrhagic Escherichia coli, MESH : Host-Pathogen Interactions, MESH: Peyer's Patches: microbiology,pathology, Microfold cell, Escherichia Coli, Multidisciplinary, Host-Pathogen Interaction, MESH : Microscopy, Electron, Transmission, Host-Pathogen Interactions, MESH: Caco-2 Cells, Research Article, MESH : Time Factors, Cell Survival, MESH: Macrophages: microbiology,pathology,ultrastructure, MESH : Male, Green Fluorescent Proteins, Escherichia coli O157, Escherichia coli infections, Models, Biological, Cell Line, MESH : Peyer's Patches: microbiology,pathology, MESH : Mice, medicine, Animals, MESH: Green Fluorescent Proteins: genetics,metabolism, MESH : Shiga Toxins: metabolism, Vero Cells, MESH: Mice, IMMUNITE, 030304 developmental biology, Hemolytic-uremic syndrome, MESH: Apoptosis, MESH : Intestinal Mucosa: microbiology,pathology, MESH: Time Factors, MESH: Models, Biological, MESH : Microbial Viability, DIARRHEE, MODELISATION, MESH: Male, Emerging Infectious Diseases, Cell culture, MESH : Animals, Caco-2 Cells, Ex vivo
Abstract: Enterohemorrhagic Escherichia coli (EHEC) are food-borne pathogens that can cause serious infections ranging from diarrhea to hemorrhagic colitis (HC) and hemolytic-uremic syndrome (HUS). Translocation of Shiga-toxins (Stx) from the gut lumen to underlying tissues is a decisive step in the development of the infection, but the mechanisms involved remain unclear. Many bacterial pathogens target the follicle-associated epithelium, which overlies Peyer's patches (PPs), cross the intestinal barrier through M cells and are captured by mucosal macrophages. Here, translocation across M cells, as well as survival and proliferation of EHEC strains within THP-1 macrophages were investigated using EHEC O157:H7 reference strains, isogenic mutants, and 15 EHEC strains isolated from HC/HUS patients. We showed for the first time that E. coli O157:H7 strains are able to interact in vivo with murine PPs, to translocate ex vivo through murine ileal mucosa with PPs and across an in vitro human M cell model. EHEC strains are also able to survive and to produce Stx in macrophages, which induce cell apoptosis and Stx release. In conclusion, our results suggest that the uptake of EHEC by M cells and underlying macrophages in the PP may be a critical step in Stx translocation and release in vivo. A new model for EHEC infection in humans is proposed that could help in a fuller understanding of EHEC-associated diseases.
Published: 2011
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30. Cellular transcriptional profiling in human lung epithelial cells infected by different subtypes of influenza A viruses reveals an overall down-regulation of the host p53 pathway

Author: Catherine Nguyen, Laurence Josset, Jean-Christophe Bourdon, Bruno Lina, Jean-Jacques Diaz, Olivier Ferraris, Virginie Marcel, Olivier Terrier, Julien Textoris, Manuel Rosa-Calatrava, Gaëlle Cartet, Division of Medical Sciences, University of Dundee-Centre for Oncology and Molecular Medicine, Virologie et pathologie humaine ( VirPath ), Université Claude Bernard Lyon 1 ( UCBL ), Université de Lyon-Université de Lyon, Laboratoire de Virologie Centre de Biologie et de Pathologie Est, Hospices Civils de Lyon ( HCL ), Technologies avancées pour le génôme et la clinique ( TAGC ), Aix Marseille Université ( AMU ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Centre de génétique et de physiologie moléculaire et cellulaire ( CGPhiMC ), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique ( CNRS ), This work was supported by University Claude Bernard of Lyon, the Civil hospitals of Lyon, the AVIESAN alliance and by a grant of the 'Programme de Recherche A (H1N1)' co-ordinated by the Institut de Microbiologie et Maladies Infectieuses (INSERM)., Virologie et pathologie humaine (VirPath), Université Claude Bernard Lyon 1 (UCBL), Hospices Civils de Lyon (HCL), Technologies avancées pour le génôme et la clinique (TAGC), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre de génétique et de physiologie moléculaire et cellulaire (CGPhiMC), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), and BMC, Ed.
Subjects: MESH: Signal Transduction, MESH : Cell Line, MESH : Blotting, Western, medicine.disease_cause, MESH: Down-Regulation, Transcriptome, MESH : Down-Regulation, [SDV.MHEP.MI]Life Sciences [q-bio]/Human health and pathology/Infectious diseases, MESH: Reverse Transcriptase Polymerase Chain Reaction, Gene expression, Influenza A virus, MESH : Influenza, Human, MESH: Tumor Suppressor Protein p53, 0303 health sciences, Reverse Transcriptase Polymerase Chain Reaction, MESH: Influenza, Human, MESH : Reverse Transcriptase Polymerase Chain Reaction, 3. Good health, MESH : Epithelial Cells, MESH : Influenza A virus, [ SDV.MHEP.MI ] Life Sciences [q-bio]/Human health and pathology/Infectious diseases, Infectious Diseases, MESH: Epithelial Cells, [SDV.MHEP.MI] Life Sciences [q-bio]/Human health and pathology/Infectious diseases, Signal Transduction, Blotting, Western, MESH: Influenza A virus, Down-Regulation, Biology, Cell Line, lcsh:Infectious and parasitic diseases, 03 medical and health sciences, MESH: Gene Expression Profiling, Virology, Influenza, Human, medicine, Humans, MESH: Blotting, Western, lcsh:RC109-216, Gene, 030304 developmental biology, MESH : Signal Transduction, MESH: Humans, 030306 microbiology, Gene Expression Profiling, Research, MESH : Gene Expression Profiling, MESH : Humans, Epithelial Cells, Influenza A virus subtype H5N1, MESH: Cell Line, Gene expression profiling, MESH : Tumor Suppressor Protein p53, Viral replication, Cell culture, Tumor Suppressor Protein p53
Abstract: BackgroundInfluenza viruses can modulate and hijack several cellular signalling pathways to efficiently support their replication. We recently investigated and compared the cellular gene expression profiles of human lung A549 cells infected by five different subtypes of human and avian influenza viruses (Jossetet al.Plos One 2010). Using these transcriptomic data, we have focused our analysis on the modulation of the p53 pathway in response to influenza infection.ResultsOur results were supported by both RT-qPCR and western blot analyses and reveal multiple alterations of the p53 pathway during infection. A down-regulation of mRNA expression was observed for the main regulators of p53 protein stability during infection by the complete set of viruses tested, and a significant decrease in p53 mRNA expression was also observed in H5N1 infected cells. In addition, several p53 target genes were also down-regulated by these influenza viruses and the expression of their product reduced.ConclusionsOur data reveal that influenza viruses cause an overall down-regulation of the host p53 pathway and highlight this pathway and p53 protein itself as important viral targets in the altering of apoptotic processes and in cell-cycle regulation.
Published: 2011
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31. Herpes simplex virus type 1-derived recombinant and amplicon vectors

Author: Fraefel , Cornel, Marconi , Peggy, Epstein , Alberto L, Centre de génétique et de physiologie moléculaire et cellulaire ( CGPhiMC ), Université Claude Bernard Lyon 1 ( UCBL ), and Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique ( CNRS )
Subjects: MESH : Cell Line, MESH : Chromosomes, Artificial, Bacterial, MESH : Cell Culture Techniques, MESH : Escherichia coli, MESH : Cloning, Molecular, viruses, MESH : Humans, MESH : Recombination, Genetic, MESH : Helper Viruses, [ SDV.BBM.BM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, MESH : Herpesvirus 1, Human, MESH : Genetic Vectors, MESH : DNA, Recombinant, MESH : Plasmids, MESH : Titrimetry, MESH : Virion, MESH : Electroporation, MESH : Animals, MESH : Transfection
Abstract: International audience; Herpes simplex virus type 1 (HSV-1) is a human pathogen whose lifestyle is based on a long-term dual interaction with the infected host, being able to establish both lytic and latent infections. The virus genome is a 153 kbp double-stranded DNA molecule encoding more than 80 genes. The interest of HSV-1 as gene transfer vector stems from its ability to infect many different cell types, both quiescent and proliferating cells, the very high packaging capacity of the virus capsid, the outstanding neurotropic adaptations that this virus has evolved, and the fact that it never integrates into the cellular chromosomes, thus avoiding the risk of insertional mutagenesis. Two types of vectors can be derived from HSV-1, recombinant vectors and amplicon vectors, and different methodologies have been developed to prepare large stocks of each type of vector. This chapter summarizes (1) the two approaches most commonly used to prepare recombinant vectors through homologous recombination, either in eukaryotic cells or in bacteria, and (2) the two methodologies currently used to generate helper-free amplicon vectors, either using a bacterial artificial chromosome (BAC)-based approach or a Cre/loxP site-specific recombination strategy.
Published: 2011
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32. A non-covalent peptide-based strategy for siRNA delivery

Author: Crombez , Laurence, Divita , Gilles, Centre de recherches de biochimie macromoléculaire ( CRBM ), Université Montpellier 1 ( UM1 ) -Université Montpellier 2 - Sciences et Techniques ( UM2 ) -IFR122-Centre National de la Recherche Scientifique ( CNRS ), Centre de recherche en Biologie Cellulaire (CRBM), and Université Montpellier 2 - Sciences et Techniques (UM2)-Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)-Université Montpellier 1 (UM1)
Subjects: MESH : Cell Line, MESH : Molecular Sequence Data, MESH : RNA, Messenger, MESH : Drug Carriers, Blotting, Western, Molecular Sequence Data, Cell-Penetrating Peptides, MESH: Amino Acid Sequence, MESH : Blotting, Western, MESH: Base Sequence, Transfection, MESH : RNA, Small Interfering, Cell Line, MESH: Cell Adhesion, MESH : Cell-Penetrating Peptides, MESH: RNA, Small Interfering, MESH : Gene Silencing, Cell Adhesion, Animals, Humans, MESH: Blotting, Western, MESH: Animals, MESH: Gene Silencing, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Amino Acid Sequence, Gene Silencing, RNA, Messenger, RNA, Small Interfering, [ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH: RNA, Messenger, MESH: Cell-Penetrating Peptides, Drug Carriers, MESH: Humans, MESH: Molecular Sequence Data, Base Sequence, MESH: Transfection, MESH : Amino Acid Sequence, MESH : Humans, MESH: Cell Line, MESH: Drug Carriers, MESH : Cell Adhesion, MESH : Base Sequence, MESH : Animals, MESH : Transfection
Abstract: International audience; The development of short-interfering RNA (siRNA) has provided great hope for therapeutic targeting of specific genes responsible for pathological disorders. However, the poor cellular uptake of siRNA together with the low permeability of the cell membrane to negatively charged molecules, remain major obstacles to clinical development. So far there is no universal method for siRNA delivery as they all present several limitations. Several non-viral strategies have been proposed to improve the delivery of synthetic siRNAs in both cultured cells and in vivo. Cell-penetrating peptides (CPPs) or protein transduction domains (PTD) constitute very promising tools for non-invasive cellular import of siRNA and non-covalent CPP/PTD-based strategies have been successfully applied for ex vivo and in vivo delivery of therapeutic siRNA molecules. We recently described a new peptide-based system, CADY, for efficient delivery of siRNA in both primary and suspension cell lines. CADY is a secondary amphiphatic peptide able to form stable non-covalent complexes with siRNA and to improve their cellular uptake independently of the endosomal pathway. This chapter describes easy to handle protocols for the use of the CADY-nanoparticle technology for the delivery of siRNA into both adherent and suspension cell lines. It will also highlight different critical points in the peptide/siRNA complex preparation and transfection protocols, in order to obtain siRNA-associated interfering response at low nanomolar concentration.
Published: 2011
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33. A non-covalent peptide-based strategy for ex vivo and in vivo oligonucleotide delivery

Author: Crombez , Laurence, Morris , May C, Heitz , Frederic, Divita , Gilles, Centre de recherches de biochimie macromoléculaire ( CRBM ), Université Montpellier 1 ( UM1 ) -Université Montpellier 2 - Sciences et Techniques ( UM2 ) -IFR122-Centre National de la Recherche Scientifique ( CNRS ), Centre de recherche en Biologie Cellulaire (CRBM), and Université Montpellier 2 - Sciences et Techniques (UM2)-Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)-Université Montpellier 1 (UM1)
Subjects: Male, Peptide Nucleic Acids, MESH : Cell Line, MESH : Molecular Sequence Data, MESH : RNA, Messenger, Oligonucleotides, Cell-Penetrating Peptides, MESH: Amino Acid Sequence, MESH : RNA, Small Interfering, Polyethylene Glycols, Mice, Drug Delivery Systems, MESH: Oligonucleotides, MESH : Cell-Penetrating Peptides, MESH: RNA, Small Interfering, MESH: Animals, MESH : Female, RNA, Small Interfering, MESH: Cell-Penetrating Peptides, MESH: Peptides, MESH : Peptides, MESH : Amino Acid Sequence, MESH : Peptide Nucleic Acids, MESH : Polyethylene Glycols, RNA-Binding Proteins, MESH : Mice, Nude, MESH : Protein Binding, MESH: Oligopeptides, MESH : Drug Delivery Systems, Female, MESH : Transfection, Oligopeptides, MESH : Prostatic Neoplasms, Protein Binding, MESH : Oligopeptides, MESH: Cyclin B1, MESH : Male, Molecular Sequence Data, MESH: Drug Delivery Systems, MESH : RNA-Binding Proteins, Mice, Nude, MESH : Oligonucleotides, Transfection, Cell Line, MESH: Cell Adhesion, MESH: Peptide Nucleic Acids, MESH : Mice, Cell Adhesion, MESH: Mice, Nude, Animals, Humans, MESH: Protein Binding, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Amino Acid Sequence, RNA, Messenger, Cyclin B1, MESH: Mice, [ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH: RNA, Messenger, MESH: Humans, MESH: Molecular Sequence Data, MESH: Transfection, MESH : Humans, Prostatic Neoplasms, MESH : Disease Models, Animal, MESH: Male, MESH: Cell Line, MESH : Cyclin B1, Disease Models, Animal, MESH : Cell Adhesion, MESH: RNA-Binding Proteins, MESH: Polyethylene Glycols, MESH : Nanoparticles, MESH: Prostatic Neoplasms, Nanoparticles, MESH : Animals, MESH: Disease Models, Animal, Peptides, MESH: Female, MESH: Nanoparticles
Abstract: International audience; The dramatic acceleration in identification of new nucleic acid-based therapeutic molecules such as short interfering RNA (siRNA) and peptide-nucleic acid (PNA) analogues has provided new perspectives for therapeutic targeting of specific genes responsible for pathological disorders. However, the poor cellular uptake of nucleic acids together with the low permeability of the cell membrane to negatively charged molecules remain major obstacles to their clinical development. Several non-viral strategies have been proposed to improve the delivery of synthetic short oligonucleotides both in cultured cells and in vivo. Cell-penetrating peptides constitute very promising tools for non-invasive cellular import of oligonucleotides and analogs. We recently described a non-covalent strategy based on short amphiphatic peptides (MPG8/PEP3) that have been successfully applied ex vivo and in vivo for the delivery of therapeutic siRNA and PNA molecules. PEP3 and MPG8 form stable nanoparticles with PNA analogues and siRNA, respectively, and promote their efficient cellular uptake, independently of the endosomal pathway, into a wide variety of cell lines, including primary and suspension lines, without any associated cytotoxicity. This chapter describes easy-to-handle protocols for the use of MPG-8 or PEP-3-nanoparticle technologies for PNA and siRNA delivery into adherent and suspension cell lines as well as in vivo into cancer mouse models.
Published: 2011
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34. The NIP7 protein is required for accurate pre-rRNA processing in human cells

Author: Cédric Hesling, Patricia P. Coltri, Beatriz A. Castilho, Ruth Rimokh, Luis Gustavo Morello, Nilson Ivo Tonin Zanchin, Centro Nacional de Pesquisa em Energia e materiais, Centra Nacional de Pesquisa, Centre de Recherche en Cancérologie de Lyon (UNICANCER/CRCL), Centre Léon Bérard [Lyon]-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM), Department of Microbiology, Immunology and Parasitology, Universidade Federal de São Paulo, Equipe 10, Oncogénèse et progression tumorale, Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre Léon Bérard [Lyon]-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre de Recherche en Cancérologie de Lyon (UNICANCER/CRCL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centro nacional de Perquisa em Energia e Materiai, centro nacional de Perquisa, Centre de Recherche en Cancérologie de Lyon ( CRCL ), Université Claude Bernard Lyon 1 ( UCBL ), Université de Lyon-Université de Lyon-Centre Léon Bérard [Lyon]-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), universidade federal de sao paulo, Universidade Federal de Sao Paulo, Centre Léon Bérard [Lyon]-Université Claude Bernard Lyon 1 ( UCBL ), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre Léon Bérard [Lyon]-Université Claude Bernard Lyon 1 ( UCBL ), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre de Recherche en Cancérologie de Lyon ( CRCL ), and Université de Lyon-Université de Lyon-Centre Léon Bérard [Lyon]-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS )
Subjects: Poly U, MESH : Cell Line, Ribosome, [ SDV.CAN ] Life Sciences [q-bio]/Cancer, 0302 clinical medicine, RNA Precursors, MESH : RNA, MESH : Cell Nucleus Structures, RNA Processing, Post-Transcriptional, MESH : RNA Precursors, MESH : RNA Processing, Post-Transcriptional, 0303 health sciences, NIP7, MESH : RNA, Ribosomal, Nuclear Proteins, Cell biology, MESH: Poly U, Gene Knockdown Techniques, 030220 oncology & carcinogenesis, MESH: HEK293 Cells, Eukaryotic Ribosome, MESH : Poly A-U, MESH: RNA Processing, Post-Transcriptional, MESH: RNA Precursors, [SDV.CAN]Life Sciences [q-bio]/Cancer, Biology, Cell Line, 03 medical and health sciences, MESH: RNA, Genetics, Humans, Eukaryotic Small Ribosomal Subunit, MESH: Cell Nucleus Structures, MESH: Poly A-U, RRNA processing, 030304 developmental biology, MESH: Humans, Eukaryotic Large Ribosomal Subunit, MESH : Humans, MESH : Nuclear Proteins, Ribosomal RNA, SBDS, Molecular biology, MESH: Gene Knockdown Techniques, Cell Nucleus Structures, MESH : HEK293 Cells, MESH: Cell Line, HEK293 Cells, MESH : Poly U, RNA, Ribosomal, Polyribosomes, MESH : Polyribosomes, MESH: RNA, Ribosomal, Poly A-U, RNA, MESH : Gene Knockdown Techniques, MESH: Nuclear Proteins, MESH: Polyribosomes
Abstract: International audience; Eukaryotic ribosome biogenesis requires the function of a large number of trans-acting factors which interact transiently with the nascent pre-rRNA and dissociate as the ribosomal subunits proceed to maturation and export to the cytoplasm. Loss-of-function mutations in human trans-acting factors or ribosome components may lead to genetic syndromes. In a previous study, we have shown association between the SBDS (Shwachman-Bodian-Diamond syndrome) and NIP7 proteins and that downregulation of SBDS in HEK293 affects gene expression at the transcriptional and translational levels. In this study, we show that downregulation of NIP7 affects pre-rRNA processing, causing an imbalance of the 40S/60S subunit ratio. We also identified defects at the pre-rRNA processing level with a decrease of the 34S pre-rRNA concentration and an increase of the 26S and 21S pre-rRNA concentrations, indicating that processing at site 2 is particularly slower in NIP7-depleted cells and showing that NIP7 is required for maturation of the 18S rRNA. The NIP7 protein is restricted to the nuclear compartment and co-sediments with complexes with molecular masses in the range of 40S-80S, suggesting an association to nucleolar pre-ribosomal particles. Downregulation of NIP7 affects cell proliferation, consistently with an important role for NIP7 in rRNA biosynthesis in human cells.
Published: 2011
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35. Herpes simplex virus type 1-derived recombinant and amplicon vectors

Author: Fraefel , Cornel, Marconi , Peggy, Epstein , Alberto L, Centre de génétique et de physiologie moléculaire et cellulaire ( CGPhiMC ), Université Claude Bernard Lyon 1 ( UCBL ), and Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique ( CNRS )
Subjects: MESH : Cell Line, MESH : Chromosomes, Artificial, Bacterial, MESH : Cell Culture Techniques, MESH : Escherichia coli, MESH : Cloning, Molecular, viruses, MESH : Humans, MESH : Recombination, Genetic, MESH : Helper Viruses, [ SDV.BBM.BM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, MESH : Herpesvirus 1, Human, MESH : Genetic Vectors, MESH : DNA, Recombinant, MESH : Plasmids, MESH : Titrimetry, MESH : Virion, MESH : Electroporation, MESH : Animals, MESH : Transfection
Abstract: International audience; Herpes simplex virus type 1 (HSV-1) is a human pathogen whose lifestyle is based on a long-term dual interaction with the infected host, being able to establish both lytic and latent infections. The virus genome is a 153 kbp double-stranded DNA molecule encoding more than 80 genes. The interest of HSV-1 as gene transfer vector stems from its ability to infect many different cell types, both quiescent and proliferating cells, the very high packaging capacity of the virus capsid, the outstanding neurotropic adaptations that this virus has evolved, and the fact that it never integrates into the cellular chromosomes, thus avoiding the risk of insertional mutagenesis. Two types of vectors can be derived from HSV-1, recombinant vectors and amplicon vectors, and different methodologies have been developed to prepare large stocks of each type of vector. This chapter summarizes (1) the two approaches most commonly used to prepare recombinant vectors through homologous recombination, either in eukaryotic cells or in bacteria, and (2) the two methodologies currently used to generate helper-free amplicon vectors, either using a bacterial artificial chromosome (BAC)-based approach or a Cre/loxP site-specific recombination strategy.
Published: 2011
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36. Herpes simplex virus type 1-derived recombinant and amplicon vectors

Author: Fraefel , Cornel, Marconi , Peggy, Epstein , Alberto L, Centre de génétique et de physiologie moléculaire et cellulaire ( CGPhiMC ), Université Claude Bernard Lyon 1 ( UCBL ), and Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique ( CNRS )
Subjects: MESH : Cell Line, MESH : Chromosomes, Artificial, Bacterial, MESH : Cell Culture Techniques, MESH : Escherichia coli, MESH : Cloning, Molecular, viruses, MESH : Humans, MESH : Recombination, Genetic, MESH : Helper Viruses, [ SDV.BBM.BM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, MESH : Herpesvirus 1, Human, MESH : Genetic Vectors, MESH : DNA, Recombinant, MESH : Plasmids, MESH : Titrimetry, MESH : Virion, MESH : Electroporation, MESH : Animals, MESH : Transfection
Abstract: International audience; Herpes simplex virus type 1 (HSV-1) is a human pathogen whose lifestyle is based on a long-term dual interaction with the infected host, being able to establish both lytic and latent infections. The virus genome is a 153 kbp double-stranded DNA molecule encoding more than 80 genes. The interest of HSV-1 as gene transfer vector stems from its ability to infect many different cell types, both quiescent and proliferating cells, the very high packaging capacity of the virus capsid, the outstanding neurotropic adaptations that this virus has evolved, and the fact that it never integrates into the cellular chromosomes, thus avoiding the risk of insertional mutagenesis. Two types of vectors can be derived from HSV-1, recombinant vectors and amplicon vectors, and different methodologies have been developed to prepare large stocks of each type of vector. This chapter summarizes (1) the two approaches most commonly used to prepare recombinant vectors through homologous recombination, either in eukaryotic cells or in bacteria, and (2) the two methodologies currently used to generate helper-free amplicon vectors, either using a bacterial artificial chromosome (BAC)-based approach or a Cre/loxP site-specific recombination strategy.
Published: 2011
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37. Purification of ribosomes from human cell lines

Author: Stéphane Pourpe, Sabine Hacot, Stephane Belin, Gabriel Therizols, Jean-Jacques Diaz, Lionel Daudignon, Manuel Rosa-Calatrava, Hichem C. Mertani, Centre de génétique et de physiologie moléculaire et cellulaire ( CGPhiMC ), Centre National de la Recherche Scientifique ( CNRS ) -Université Claude Bernard Lyon 1 ( UCBL ), Université de Lyon-Université de Lyon, Institut de biologie et chimie des protéines [Lyon] ( IBCP ), Université Claude Bernard Lyon 1 ( UCBL ), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique ( CNRS ), Virologie et Pathologie Humaine ( VirPath ), École normale supérieure - Lyon ( ENS Lyon ) -Université Claude Bernard Lyon 1 ( UCBL ), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Centre de génétique et de physiologie moléculaire et cellulaire (CGPhiMC), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Institut de biologie et chimie des protéines [Lyon] (IBCP), Virologie et Pathologie Humaine (VirPath), École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), and Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)
Subjects: MESH : Cell Line, MESH : Protein Biosynthesis, Ribosome biogenesis, Biology, Cell Fractionation, Ribosome, Cell Line, Ribosomal protein, Humans, Eukaryotic Small Ribosomal Subunit, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Ribosome profiling, [ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH : Ribosomes, MESH: Humans, MESH : Humans, Translation (biology), General Medicine, Ribosomal RNA, Cell biology, MESH: Cell Line, Protein Biosynthesis, MESH: Protein Biosynthesis, MESH : Cell Fractionation, MESH: Cell Fractionation, Eukaryotic Ribosome, Ribosomes, MESH: Ribosomes
Abstract: International audience; Highly conserved during evolution, the ribosome is the central effector of protein synthesis. In mammalian cells, the ribosome is a macromolecular complex composed of four different ribosomal RNAs (rRNA) and about 80 ribosomal proteins. Requiring more than 200 factors, ribosome biogenesis is a highly complex process that takes place mainly within the nucleoli of eukaryotic cells. Crystallographic data suggest that the ribosome is a ribozyme, in which the rRNA catalyses the peptide bond formation and ensures quality control of the translation. Ribosomal proteins are involved in this molecular mechanism; nonetheless, their role is still not fully characterized. Recent studies suggest that ribosomes themselves and/or the mechanisms underlying their synthesis, processing, and assembly play a key role in the establishment and progression of several human pathologies. The protocol described here is simple, efficient, and robust, and allows one to purify high-quality ribosomes from human cultured cell lines. Ribosomes purified with this protocol are adequate for most of the subsequent analyses of their RNA and protein content.
Published: 2010
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38. Effects of oxysterols on cell viability, inflammatory cytokines, VEGF, and reactive oxygen species production on human retinal cells: cytoprotective effects and prevention of VEGF secretion by resveratrol

Author: Franck Ménétrier, Am Bron, Catherine Creuzot-Garcher, T. Khalfaoui, Jeannine Lherminier, L. Malvitte, Dominique Delmas, Kévin Ragot, Soeli Charbonnier, Norbert Latruffe, Mauhamad Baarine, Brice Dugas, Gérard Lizard, Service d'Ophtalmologie (CHU de Dijon), Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand ( CHU Dijon ), Lipides - Nutrition - Cancer (U866) ( LNC ), Université de Bourgogne ( UB ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Ecole Nationale Supérieure de Biologie Appliquée à la Nutrition et à l'Alimentation de Dijon ( ENSBANA ), FLAveur, VIsion et Comportement du consommateur ( FLAVIC ), Etablissement National d'Enseignement Supérieur Agronomique de Dijon ( ENESAD ) -Institut National de la Recherche Agronomique ( INRA ) -Université de Bourgogne ( UB ), Agroécologie [Dijon], Institut National de la Recherche Agronomique ( INRA ) -Université de Bourgogne ( UB ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement, This work was supported by grants from the INSERM, and the University Hospital of Dijon (CHU de Dijon)., Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon), Lipides - Nutrition - Cancer (U866) (LNC), Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Ecole Nationale Supérieure de Biologie Appliquée à la Nutrition et à l'Alimentation de Dijon (ENSBANA), FLAveur, VIsion et Comportement du consommateur (FLAVIC), Institut National de la Recherche Agronomique (INRA)-Université de Bourgogne (UB)-Etablissement National d'Enseignement Supérieur Agronomique de Dijon (ENESAD), Institut National de la Recherche Agronomique (INRA)-Université de Bourgogne (UB)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement, Etablissement National d'Enseignement Supérieur Agronomique de Dijon (ENESAD)-Institut National de la Recherche Agronomique (INRA)-Université de Bourgogne (UB), and Latruffe, Norbert
Subjects: MESH: Cell Death, MESH: Inflammation, Vascular Endothelial Growth Factor A, MESH : Cell Line, inflammatory cytokines, medicine.medical_treatment, MESH : Cytokines, Medicine (miscellaneous), MESH : Reactive Oxygen Species, Wine, Resveratrol, Pharmacology, resveratrol, MESH : Cytoprotection, Antioxidants, chemistry.chemical_compound, 0302 clinical medicine, MESH: Cholesterol, MESH: Stilbenes, Stilbenes, polycyclic compounds, [ SHS.INFO ] Humanities and Social Sciences/Library and information sciences, MESH : Cholesterol, Phospholipids, chemistry.chemical_classification, reactive oxygen species, 0303 health sciences, MESH: Cytokines, MESH : Cell Survival, Nutrition and Dietetics, Cell Death, MESH: Retina, food and beverages, MESH: Reactive Oxygen Species, phospholipidosis, Cytoprotection, VEGF, 3. Good health, Cytokine, Cholesterol, MESH: Cell Survival, MESH : Vascular Endothelial Growth Factor A, Cytokines, oxysterols, MESH : Antioxidants, lipids (amino acids, peptides, and proteins), medicine.symptom, Programmed cell death, Cell Survival, [SHS.INFO]Humanities and Social Sciences/Library and information sciences, MESH : Wine, Inflammation, Biology, [SHS.INFO] Humanities and Social Sciences/Library and information sciences, Retina, MESH : Phospholipids, Proinflammatory cytokine, Cell Line, MESH : Stilbenes, 03 medical and health sciences, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, medicine, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Viability assay, [ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology, 030304 developmental biology, MESH: Phospholipids, Reactive oxygen species, MESH: Humans, MESH : Inflammation, MESH: Vascular Endothelial Growth Factor A, MESH: Antioxidants, MESH : Humans, MESH : Retina, MESH: Wine, MESH: Cell Line, MESH: Cytoprotection, chemistry, MESH : Cell Death, Immunology, 030221 ophthalmology & optometry, caspase independent cell death, ARPE-19 cells
Abstract: Oxysterols are assumed to play important roles in age-related macular degeneration, a major cause of blindness. So we characterized the cytotoxic, oxidative, inflammatory, and angiogenic activities of oxysterols (7β-hydroxycholesterol (7β-OH), 7-ketocholesterol (7KC), 25-hydroxycholesterol (25-OH)) in human retinal ARPE-19 cells, and evaluated the protective effects of resveratrol (Rsv: 1 μM), a polyphenol from red wine.ARPE-19 cells were treated with 7β-OH, 7KC, or 25-OH (5-40 μg/mL; 24-48 h) without or with Rsv. Cell viability was determined using trypan blue and the MTT assay. Cell death was characterized by electron microscopy and in situ detection of activated caspases with fluorochrome-labeled inhibitors of caspases. Reactive oxygen species (ROS) production was measured with hydroethidine. ELISA methods and a cytometric bead assay were used to quantify cytokines involved in inflammation (IL-8, IL-1β, IL-6, IL-10, IL-12p70, TNF-α, MCP-1) and VEGF.7β-OH and 7KC triggered a caspase-independent cell death process associated with the presence of multilamellar cytoplasmic structures evocating phospholipidosis, increased ROS production, and IL-8 secretion. 7β-OH enhanced VEGF secretion. No cytotoxic effects were identified with 25-OH, which highly stimulated ROS production, MCP-1, and VEGF secretion. With oxysterols, no IL-10, TNF-α, and IL-12p70 secretion were detected. 25-OH induced IL-8 secretion through the MEK/ERK½ signaling pathway, and Rsv showed cytoprotective activities and inhibited VEGF secretion.7β-OH, 7KC, and 25-OH have cytotoxic, oxidative, inflammatory, and/or angiogenic activities on ARPE-19 cells. As Rsv has some protective effects against oxysterol-induced cell death and VEGF secretion it could be valuable in ARMD treatment.
Published: 2010
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39. Dlx homeobox gene family expression in osteoclasts

Author: Dominique Heymann, Frédéric Lézot, Alba Bolaños, Beatriz Castaneda, D. Hotton, Ariane Berdal, Bethan Thomas, Agamemnon E. Grigoriadis, G.F. Carles, Claudine Blin-Wakkach, Paul T. Sharpe, Centre de Recherche des Cordeliers ( CRC (UMR_S 872) ), Université Pierre et Marie Curie - Paris 6 ( UPMC ) -Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Génétique, physiopathologie et ingénierie du tissu osseux ( GéPITOS ), Université Nice Sophia Antipolis ( UNS ), Université Côte d'Azur ( UCA ) -Université Côte d'Azur ( UCA ) -Centre National de la Recherche Scientifique ( CNRS ), Physiopathologie de la Résorption Osseuse et Thérapie des Tumeurs Osseuses Primitives, Université de Nantes ( UN ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Centre de Recherche des Cordeliers (CRC (UMR_S 872)), Université Pierre et Marie Curie - Paris 6 (UPMC)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Génétique, physiopathologie et ingénierie du tissu osseux (GéPITOS), Université Nice Sophia Antipolis (... - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Centre National de la Recherche Scientifique (CNRS)-Université Côte d'Azur (UCA), Physiopathologie des Adaptations Nutritionnelles (PhAN), Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Université Nice Sophia Antipolis (1965 - 2019) (UNS), Institut National de la Recherche Agronomique (INRA)-Université de Nantes (UN), Université Côte d'Azur (UCA)-Université Côte d'Azur (UCA)-Centre National de la Recherche Scientifique (CNRS), and Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)
Subjects: Male, MESH : Cell Line, MESH : Transcription Factors, Physiology, Cellular differentiation, Clinical Biochemistry, Osteoclasts, Mandible, Mice, 0302 clinical medicine, MESH : Acid Phosphatase, Osteogenesis, MESH: Osteoclasts, MESH: Reverse Transcriptase Polymerase Chain Reaction, MESH: Gene Expression Regulation, Developmental, MESH : Osteogenesis, MESH: Animals, MESH : Homeodomain Proteins, MESH : Matrix Metalloproteinase 9, MESH: Osteogenesis, Tartrate-resistant acid phosphatase, Regulation of gene expression, MESH : Isoenzymes, 0303 health sciences, Reverse Transcriptase Polymerase Chain Reaction, DLX2, MESH : Reverse Transcriptase Polymerase Chain Reaction, Gene Expression Regulation, Developmental, Cell Differentiation, MESH: Transcription Factors, DLX6, MESH : beta-Galactosidase, DLX5, Isoenzymes, MESH: beta-Galactosidase, Matrix Metalloproteinase 9, Multigene Family, 030220 oncology & carcinogenesis, MESH: Isoenzymes, MESH : Cell Differentiation, MESH: Cell Differentiation, musculoskeletal diseases, MESH : Mandible, MESH : Male, Acid Phosphatase, MESH : Multigene Family, MESH: Mandible, Biology, Cell Line, MESH: Acid Phosphatase, MESH: Gene Expression Profiling, 03 medical and health sciences, MESH: Homeodomain Proteins, MESH : Mice, Animals, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH: Mice, [ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology, 030304 developmental biology, Homeodomain Proteins, Tartrate-Resistant Acid Phosphatase, Gene Expression Profiling, MESH : Gene Expression Profiling, MESH: Matrix Metalloproteinase 9, Cell Biology, beta-Galactosidase, Molecular biology, MESH: Male, MESH: Cell Line, Gene expression profiling, MESH : Osteoclasts, MESH: Multigene Family, Homeobox, MESH : Animals, MESH : Gene Expression Regulation, Developmental, Transcription Factors
Abstract: International audience; Skeletal growth and homeostasis require the finely orchestrated secretion of mineralized tissue matrices by highly specialized cells, balanced with their degradation by osteoclasts. Time- and site-specific expression of Dlx and Msx homeobox genes in the cells secreting these matrices have been identified as important elements in the regulation of skeletal morphology. Such specific expression patterns have also been reported in osteoclasts for Msx genes. The aim of the present study was to establish the expression patterns of Dlx genes in osteoclasts and identify their function in regulating skeletal morphology. The expression patterns of all Dlx genes were examined during the whole osteoclastogenesis using different in vitro models. The results revealed that Dlx1 and Dlx2 are the only Dlx family members with a possible function in osteoclastogenesis as well as in mature osteoclasts. Dlx5 and Dlx6 were detected in the cultures but appear to be markers of monocytes and their derivatives. In vivo, Dlx2 expression in osteoclasts was examined using a Dlx2/LacZ transgenic mouse. Dlx2 is expressed in a subpopulation of osteoclasts in association with tooth, brain, nerve, and bone marrow volumetric growths. Altogether the present data suggest a role for Dlx2 in regulation of skeletal morphogenesis via functions within osteoclasts.
Published: 2010
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40. Glycosaminoglycan mimetics trigger IP3-dependent intracellular calcium release in myoblasts

Author: Isabelle Guillet-Deniau, Dulce Papy-Garcia, Isabelle Martelly, José Courty, Guy Raymond, Dominique Singabraya, Aurélie Vandebrouck, Christian Cognard, Bruno Constantin, Croissance cellulaire, réparation et régénération tissulaires (CRRET), Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12)-Centre National de la Recherche Scientifique (CNRS), Institut de physiologie et biologie cellulaires (IPBC), Université de Poitiers-Centre National de la Recherche Scientifique (CNRS), Institut Cochin (UMR_S567 / UMR 8104), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5), Institut de Physiologie et Biologie Cellulaires (IPBC), Institut Cochin (IC UM3 (UMR 8104 / U1016)), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Laboratoire CRRET, EAC/CNRS-7149, Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12), Laboratoire de recherche sur la croissance cellulaire, la réparation et la régénération tissulaires ( CRRET ), Université Paris-Est Créteil Val-de-Marne - Paris 12 ( UPEC UP12 ) -Centre National de la Recherche Scientifique ( CNRS ), Institut de physiologie et biologie cellulaires ( IPBC ), Université de Poitiers-Centre National de la Recherche Scientifique ( CNRS ), Institut Cochin ( UMR_S567 / UMR 8104 ), and Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS )
Subjects: MESH : Calcium, Dietary, MESH : Cell Line, MESH: Signal Transduction, Cytoplasm, MESH: Heparin, [SDV]Life Sciences [q-bio], Cellular homeostasis, [SDV.BC.BC]Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC], Muscle Development, Calcium in biology, MESH: Ryanodine Receptor Calcium Release Channel, Myoblasts, Mice, 0302 clinical medicine, MESH : Receptor, Serotonin, 5-HT2A, [SDV.BC.IC]Life Sciences [q-bio]/Cellular Biology/Cell Behavior [q-bio.CB], Receptor, Serotonin, 5-HT2A, MESH: Animals, MESH : Ryanodine Receptor Calcium Release Channel, Glycosaminoglycans, 0303 health sciences, MESH : Rats, Ryanodine receptor, Myogenesis, MESH: Regeneration, MESH: Satellite Cells, Skeletal Muscle, Cell Differentiation, MESH: Glycosaminoglycans, Cell biology, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Biochemistry, MESH: Calcium, MESH: Muscle Development, MESH : Cell Differentiation, MESH: Calcium, Dietary, Intracellular, Signal Transduction, MESH: Cell Differentiation, MESH : Cytoplasm, Satellite Cells, Skeletal Muscle, MESH: Rats, Inositol Phosphates, MESH : Muscle Development, MESH : Inositol Phosphates, chemistry.chemical_element, MESH : Regeneration, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Biology, Calcium, MESH : Rats, Wistar, MESH : Myoblasts, Cell Line, 03 medical and health sciences, MESH : Satellite Cells, Skeletal Muscle, MESH : Mice, MESH : Glycosaminoglycans, [SDV.MHEP.PHY]Life Sciences [q-bio]/Human health and pathology/Tissues and Organs [q-bio.TO], Animals, Regeneration, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH: Myoblasts, Rats, Wistar, MESH : Calcium, Molecular Biology, MESH: Mice, 030304 developmental biology, MESH : Signal Transduction, Calcium metabolism, Heparin, MESH: Cytoplasm, MESH: Receptor, Serotonin, 5-HT2A, MESH: Clone Cells, [ SDV.BC.BC ] Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC], MESH : Clone Cells, Ryanodine Receptor Calcium Release Channel, MESH: Rats, Wistar, Inositol trisphosphate receptor, MESH: Inositol Phosphates, Clone Cells, Rats, MESH: Cell Line, Calcium, Dietary, MESH : Heparin, chemistry, MESH : Animals, 030217 neurology & neurosurgery
Abstract: International audience; Glycosaminoglycans (GAG) are sulfated polysaccharides that play an important role in regulating cell functions. GAG mimetics called RGTAs (for ReGeneraTing Agents) have been shown to stimulate tissue repair. In particular they accelerate myogenesis, in part via their heparin-mimetic property towards growth factors. RGTAs also increase activity of calcium-dependent intracellular protease suggesting an effect on calcium cellular homeostasis. This effect was presently investigated on myoblasts in vitro using one member of the RGTA family molecule named OTR4120. We have shown that OTR4120 or heparin induced transient increases of intracellular calcium concentration ([Ca(2+)]i) in pre-fusing myoblasts from both mouse SolD7 cell line and rat skeletal muscle satellite cells grown in primary culture by mobilising sarcoplasmic reticulum store. This [Ca(2+)]i was not mediated by ryanodine receptors but instead resulted from stimulation of the Inositol-3 phosphate-phospholipase C activation pathway. OTR4120-induced calcium transient was not mediated through an ATP, nor a tyrosine kinase, nor an acetylcholine receptor but principally through serotonin 5-HT2A receptor. This original finding shows that the GAG mimetic can induce calcium signal through serotonin receptors and the IP3 pathway may be relevant to its ability to favour myoblast differentiation. It supports a novel and unexpected function of GAGs in the regulation of calcium homeostasis.
Published: 2010
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41. The human COP9 signalosome protects ubiquitin-conjugating enzyme 3 (UBC3/Cdc34) from beta-transducin repeat-containing protein (betaTrCP)-mediated degradation

Author: Emmanuel Sechet, Elisabetta Bianchi, Lars Rogge, Maria Elena Fernandez-Sanchez, Florence Margottin-Goguet, Immunorégulation, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Institut Cochin (IC UM3 (UMR 8104 / U1016)), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Pasteur [Paris] - Centre National de la Recherche Scientifique (CNRS), Institut Cochin (UM3 (UMR 8104 / U1016)), Université Paris Descartes - Paris 5 (UPD5) - Institut National de la Santé et de la Recherche Médicale (INSERM) - Centre National de la Recherche Scientifique (CNRS), and Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS)
Subjects: MESH : Cell Line, MESH : Hela Cells, Oligonucleotides, Ubiquitin-conjugating enzyme, Biochemistry, NEDD8, MESH: Down-Regulation, Substrate Specificity, MESH : Down-Regulation, 0302 clinical medicine, Ubiquitin, MESH: Transducin, MESH: Oligonucleotides, MESH: Reverse Transcriptase Polymerase Chain Reaction, MESH : RNA, MESH : Ubiquitin, MESH: Lentivirus, chemistry.chemical_classification, 0303 health sciences, biology, MESH : Substrate Specificity, Reverse Transcriptase Polymerase Chain Reaction, MESH : Reverse Transcriptase Polymerase Chain Reaction, Ubiquitin-Protein Ligase Complexes, Ubiquitin ligase, MESH : Lentivirus, Protein Synthesis and Degradation, 030220 oncology & carcinogenesis, MESH : Peptide Hydrolases, [SDV.IMM]Life Sciences [q-bio]/Immunology, Transducin, Plasmids, MESH: Ubiquitin, [SDV.IMM] Life Sciences [q-bio]/Immunology, MESH: Peptide Hydrolases, MESH : Transducin, MESH : Multiprotein Complexes, Down-Regulation, MESH : Oligonucleotides, Anaphase-Promoting Complex-Cyclosome, Cell Line, 03 medical and health sciences, Downregulation and upregulation, MESH: Plasmids, MESH: RNA, Humans, COP9 signalosome, Molecular Biology, 030304 developmental biology, MESH: Humans, COP9 Signalosome Complex, MESH : Humans, fungi, Lentivirus, MESH: Ubiquitin-Protein Ligase Complexes, Cell Biology, MESH: Multiprotein Complexes, MESH: Cell Line, MESH: Hela Cells, Enzyme, chemistry, MESH : Plasmids, Multiprotein Complexes, Ubiquitin-Conjugating Enzymes, biology.protein, RNA, MESH: Substrate Specificity, MESH : Ubiquitin-Protein Ligase Complexes, HeLa Cells, Peptide Hydrolases
Abstract: International audience; The COP9 signalosome (CSN) is an essential multisubunit complex that regulates the activity of cullin-RING ubiquitin ligases by removing the ubiquitin-like peptide NEDD8 from cullins. Here, we demonstrate that the CSN can affect other components of the ubiquitination cascade. Down-regulation of human CSN4 or CSN5 induced proteasome-mediated degradation of the ubiquitin-conjugating enzyme UBC3/Cdc34. UBC3 was targeted for ubiquitination by the cullin-RING ubiquitin ligase SCF(betaTrCP). This interaction required the acidic C-terminal extension of UBC3, which is absent in ubiquitin-conjugating enzymes of the UBCH5 family. Conversely, the UBC3 acidic domain was sufficient to impart sensitivity to SCF(betaTrCP)-mediated ubiquitination to UBCH5 enzymes. Our work indicates that the CSN is necessary to ensure the stability of selected ubiquitin-conjugating enzymes and uncovers a novel pathway of regulation of ubiquitination processes.
Published: 2010
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42. Expression of the HLA-C2-specific activating killer-cell Ig-like receptor KIR2DS1 on NK and T cells

Author: Coralie Frassati, Laurent Gauthier, Aude Magerus-Chatinet, Nicolas Anfossi, Eric Vivier, Pascale Andre, Catherine Farnarier, Nicolas Schleinitz, Céline Cognet, Frédéric Rieux-Laucat, Centre d'Immunologie de Marseille - Luminy ( CIML ), Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Aix Marseille Université ( AMU ) -Centre National de la Recherche Scientifique ( CNRS ), Hôpital de la Conception [CHU - APHM] ( LA CONCEPTION ), Innate Pharma, Marseille, Innate Pharma, Etablissement Français du Sang - Alpes-Méditerranée ( EFS - Alpes-Méditerranée ), Etablissement Français du Sang, Developpement Normal et Pathologique du Système Immunitaire, Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Centre d'Immunologie de Marseille - Luminy (CIML), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Hôpital de la Conception [CHU - APHM] (LA CONCEPTION), Etablissement Français du Sang - Alpes-Méditerranée (EFS - Alpes-Méditerranée), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU), Hôpital de la Conception [CHU - APHM] (LA CONCEPTION ), Service de Chirurgie, Assistance Publique - Hôpitaux de Marseille ( APHM ) -Hospices Civiles de Marseille-Hôpital de la Conception [CHU - APHM] ( LA CONCEPTION ), UMR 6578 : Adaptabilité Biologique et Culturelle ( UAABC ), Université de la Méditerranée - Aix-Marseille 2-Centre National de la Recherche Scientifique ( CNRS ), Laboratoire d'Informatique, Signal et Image, Electronique et Télécommunication ( LISITE ), and institut supérieur d'éléctronique de paris
Subjects: MESH : Cell Line, T-Lymphocytes, MESH : DNA, MESH: Flow Cytometry, MESH : Genotype, NK cells, Lymphocyte Activation, MESH: Genotype, Interleukin 21, Mice, 0302 clinical medicine, Receptors, KIR, Immunology and Allergy, [ SDV.IMM ] Life Sciences [q-bio]/Immunology, MESH: Animals, 0303 health sciences, MESH : HLA-C Antigens, MESH: DNA, 2DL1, Natural killer T cell, Flow Cytometry, Killer Cells, Natural, MESH: Leukocytes, Mononuclear, medicine.anatomical_structure, Interleukin 12, [SDV.IMM]Life Sciences [q-bio]/Immunology, MESH : Killer Cells, Natural, KIRs, MESH: Killer Cells, Natural, Genotype, MESH : Flow Cytometry, T cell, Immunology, chemical and pharmacologic phenomena, HLA-C Antigens, Biology, Natural killer cell, Cell Line, 03 medical and health sciences, KIR2DL1, MESH: Receptors, KIR, MESH : Mice, medicine, Animals, Humans, MESH: Lymphocyte Activation, MESH: Mice, MESH : Lymphocyte Activation, 030304 developmental biology, MESH : T-Lymphocytes, Lymphokine-activated killer cell, MESH: Humans, MESH : Humans, MESH : Receptors, KIR, KIR+T cells, DNA, MESH: Cell Line, MESH: HLA-C Antigens, 2DS1, MESH : Leukocytes, Mononuclear, MESH: T-Lymphocytes, [ SHS.ANTHRO-BIO ] Humanities and Social Sciences/Biological anthropology, Leukocytes, Mononuclear, MESH : Animals, CD8, 030215 immunology
Abstract: International audience; Killer Ig-like receptors (KIRs) are MHC class I-specific receptors expressed by Natural Killer (NK) and T cell subsets. KIRs either inhibit (KIR-L) or activate (KIR-S) lymphocyte functions. Inhibitory KIR2DL1 and activating KIR2DS1 share ligand specificity for the HLA-C2 group, consistent with their almost identical extracytoplasmic domain. This homology hampered the distinction between KIR2DL1 and KIR2DS1. We report here the characterization of the KIR2DS1(+) subsets among primary human NK and T cells. Regardless of the host HLA-C genotype, around 10% of circulating NK cells expressed KIR2DS1 in absence of KIR2DL1. In HLA-C2 individuals, KIR2DS1 was not able to induce NK cell education (i.e., the acquisition of NK cell competence) nor to interfere with KIR2DL1-induced NK cell education. KIR2DS1 was also present on rare oligoclonal TCRalphabeta(+)CD8alpha(+) and TCRalphabeta(+)CD4(-)CD8(-) subsets. As KIR2DS1 has been associated with autoimmunity and hematopoietic stem cell transplantation, these results pave the way to dissect the function of KIR2DS1 in these clinical conditions.
Published: 2010
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43. A network of hydrogen bonds on the surface of TLR2 controls ligand positioning and cell signaling

Author: Andrey V. Kajava, Thierry Vasselon, Centre de recherches de biochimie macromoléculaire ( CRBM ), Université Montpellier 1 ( UM1 ) -Université Montpellier 2 - Sciences et Techniques ( UM2 ) -IFR122-Centre National de la Recherche Scientifique ( CNRS ), Institut de Génétique Moléculaire de Montpellier ( IGMM ), Université de Montpellier ( UM ) -Centre National de la Recherche Scientifique ( CNRS ), Centre de recherche en Biologie Cellulaire (CRBM), Université Montpellier 2 - Sciences et Techniques (UM2)-Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)-Université Montpellier 1 (UM1), Institut de Génétique Moléculaire de Montpellier (IGMM), and Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)
Subjects: MESH: Signal Transduction, MESH : Cell Line, Protein Conformation, MESH : Toll-Like Receptor 2, Plasma protein binding, Ligands, Biochemistry, MESH : Protein Multimerization, Mixed Function Oxygenases, CSK Tyrosine-Protein Kinase, MESH: Protein Conformation, 0302 clinical medicine, Protein structure, MESH: Ligands, Ternary complex, MESH : Mixed Function Oxygenases, MESH : Protein Conformation, 0303 health sciences, MESH : Ligands, MESH: Protein Multimerization, Pattern recognition receptor, MESH: Toll-Like Receptor 2, MESH : Protein Binding, MESH: Mixed Function Oxygenases, Protein-Tyrosine Kinases, Lipids, src-Family Kinases, Ectodomain, MESH : Proto-Oncogene Proteins, MESH : Amidine-Lyases, Signal transduction, Protein Binding, Signal Transduction, Cell signaling, Stereochemistry, MESH : Multiprotein Complexes, Biology, MESH: Protein-Tyrosine Kinases, Cell Line, 03 medical and health sciences, Proto-Oncogene Proteins, MESH : Protein-Tyrosine Kinases, MESH: Protein Binding, Humans, MESH : Hydrogen Bonding, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH: Hydrogen Bonding, Molecular Biology, [ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology, 030304 developmental biology, MESH : Signal Transduction, MESH: Humans, Ligand, MESH : Humans, Hydrogen Bonding, Cell Biology, MESH: Multiprotein Complexes, Toll-Like Receptor 2, MESH: Cell Line, MESH: Proto-Oncogene Proteins, MESH: Amidine-Lyases, Multiprotein Complexes, Amidine-Lyases, Protein Multimerization, 030215 immunology
Abstract: International audience; TLR2 is a pattern recognition receptor that functions in association with TLR1 or TLR6 to mediate innate immune responses to a variety of conserved microbial products. In the present study, the ectodomain of TLR2 was extensively mutated, and the mutants were assessed for their ability to bind and to mediate cellular responses to triacylated lipopeptide Pam(3)CSK(4). This analysis provides evidence that the recently published crystal structure of the TLR2-TLR1-Pam(3)CSK(4) complex represents a functional signal-inducing complex. Furthermore, we report that extended H-bond networks on the surface of TLR2 are critical for signaling in response to Pam(3)CSK(4) and to other di- and tri-acylated TLR2-TLR6 and TLR2-TLR1 ligands. Based on this finding, we suggest a dynamic model for TLR2-mediated recognition of these ligands in which TLR2 fluctuates between a conformation that is more suitable for binding of the fatty acyl moieties of the ligands and a conformation that favors, via a specific orientation of the ligand head group, formation of a signal-inducing ternary complex.
Published: 2010
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44. Production and characterisation of a neutralising chimeric antibody against botulinum neurotoxin A

Author: Prigent, Julie, Mazuet, Christelle, Boquet, Didier, Lamourette, Patricia, Volland, Hervé, Popoff, Michel, Créminon, Christophe, Simon, Stéphanie, Moreno, Edgardo, Institut de Biologie et de Technologies de Saclay (IBITECS), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay, Bactéries anaérobies et Toxines, Institut Pasteur [Paris] (IP), Laboratoire d'Ingénierie des Anticorps pour la Santé (LIAS), Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Universidad Nacional, Heredia (UNA), Escuela de Medicina Veterinaria, This study was supported by a grant from the PhD programme of the CEA (Commissariat à l'Energie Atomique et aux Energies Renouvelables) (Julie Prigent) and funded by the IMASSA (Institut de Médecine Aérospatiale du Service de Santé des Armées) and NRBC (Nuclear Radiological Biological and Chemical Risks) programmes., Institut Pasteur [Paris], Institut de Biologie et de Technologies de Saclay ( IBITECS ), Université Paris-Saclay-Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ), Laboratoire d'Ingénierie des Anticorps pour la Santé ( LIAS ), Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ), and Universidad Nacional, Heredia ( UNA )
Subjects: MESH : Cell Line, [ SDV.TOX ] Life Sciences [q-bio]/Toxicology, lcsh:Medicine, MESH : Blotting, Western, medicine.disease_cause, Antibody production, Biochemistry, Neutralization, MESH: Antibodies, Neutralizing, Infectious Diseases/Bacterial Infections, Botulinum toxin, [ SDV.IMM ] Life Sciences [q-bio]/Immunology, Toxins, Botulism, MESH: Animals, Botulinum Toxins, Type A, Enzyme-linked immunoassays, lcsh:Science, 0303 health sciences, Multidisciplinary, MESH: Spodoptera, MESH : Antibodies, Neutralizing, 3. Good health, [SDV.TOX]Life Sciences [q-bio]/Toxicology, [SDV.IMM]Life Sciences [q-bio]/Immunology, Antibody, Research Article, MESH : Botulinum Toxins, Type A, medicine.drug_class, Immunology, Blotting, Western, Biology, Spodoptera, Monoclonal antibody, MESH : Spodoptera, Antibodies, Cell Line, 03 medical and health sciences, Affinity chromatography, medicine, Animals, MESH: Blotting, Western, 030304 developmental biology, Antiserum, 030306 microbiology, Toxin, MESH: Botulinum Toxins, Type A, lcsh:R, medicine.disease, Virology, Antibodies, Neutralizing, MESH: Cell Line, biology.protein, Clostridium botulinum, lcsh:Q, MESH : Animals, Antitoxins, Cloning
Abstract: International audience; Botulinum neurotoxins, produced by Clostridium botulinum bacteria, are the causative agent of botulism. This disease only affects a few hundred people each year, thus ranking it among the orphan diseases. However, botulinum toxin type A (BoNT/A) is the most potent toxin known to man. Due to their potency and ease of production, these toxins were classified by the Centers for Disease Control and Prevention (CDC) as Category A biothreat agents. For several biothreat agents, like BoNT/A, passive immunotherapy remains the only possible effective treatment allowing in vivo neutralization, despite possible major side effects. Recently, several mouse monoclonal antibodies directed against a recombinant fragment of BoNT/A were produced in our laboratory and most efficiently neutralised the neurotoxin. In the present work, the most powerful one, TA12, was selected for chimerisation. The variable regions of this antibody were thus cloned and fused with the constant counterparts of human IgG1 (kappa light and gamma 1 heavy chains). Chimeric antibody production was evaluated in mammalian myeloma cells (SP2/0-Ag14) and insect cells (Sf9). After purifying the recombinant antibody by affinity chromatography, the biochemical properties of chimeric and mouse antibody were compared. Both have the same very low affinity constant (close to 10 pM) and the chimeric antibody exhibited a similar capacity to its parent counterpart in neutralising the toxin in vivo. Its strong affinity and high neutralising potency make this chimeric antibody interesting for immunotherapy treatment in humans in cases of poisoning, particularly as there is a probable limitation of the immunological side effects observed with classical polyclonal antisera from heterologous species.
Published: 2010
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45. Human cord blood-derived hematopoietic and neural-like stem/progenitor cells are attracted by the neurotransmitter GABA

Author: Frédéric Deschaseaux, Pierre Tiberghien, Norbert Balon, Vincent Zangiacomi, Stéphane Maddens, Claudine Versaux-Botteri, Centre de Recherche des Cordeliers ( CRC (UMR_S 872) ), Université Pierre et Marie Curie - Paris 6 ( UPMC ) -Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Interactions hôte-greffon-tumeur, ingénierie cellulaire et génique - UFC (UMR INSERM 1098) ( HOTE GREFFON ), Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Etablissement français du sang [Bourgogne-France-Comté] ( EFS [Bourgogne-France-Comté] ) -Université de Franche-Comté ( UFC ), Centre de Recherche des Cordeliers (CRC (UMR_S 872)), Université Paris Descartes - Paris 5 (UPD5)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Interactions hôte-greffon-tumeur, ingénierie cellulaire et génique - UFC (UMR INSERM 1098) (RIGHT), Institut National de la Santé et de la Recherche Médicale (INSERM)-Etablissement français du sang [Bourgogne-Franche-Comté] (EFS [Bourgogne-Franche-Comté])-Université de Franche-Comté (UFC), Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université Bourgogne Franche-Comté [COMUE] (UBFC), Université Pierre et Marie Curie - Paris 6 (UPMC)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Etablissement français du sang [Bourgogne-Franche-Comté] (EFS BFC)-Université de Franche-Comté (UFC), and Saas, Philippe
Subjects: MESH: Chemotaxis, MESH : Cell Line, Baclofen, MESH: Neurons, Fluorescent Antibody Technique, Gene Expression, MESH: Flow Cytometry, MESH: gamma-Aminobutyric Acid, MESH: GABA Antagonists, MESH: Hematopoietic Stem Cells, GABA Antagonists, 0302 clinical medicine, MESH: Reverse Transcriptase Polymerase Chain Reaction, [ SDV.IMM ] Life Sciences [q-bio]/Immunology, AC133 Antigen, MESH: Receptors, GABA-B, MESH: Antigens, CD, MESH: Fluorescent Antibody Technique, MESH: Receptors, GABA-A, Cells, Cultured, gamma-Aminobutyric Acid, Neurons, MESH : GABA Antagonists, 0303 health sciences, Reverse Transcriptase Polymerase Chain Reaction, MESH: Peptides, MESH : Peptides, Chemotaxis, Stem Cells, MESH : Reverse Transcriptase Polymerase Chain Reaction, MESH : Bicuculline, Cell Differentiation, Hematology, Fetal Blood, Flow Cytometry, MESH : gamma-Aminobutyric Acid, 3. Good health, Cell biology, Endothelial stem cell, MESH: Baclofen, Haematopoiesis, Cord blood, MESH : Stem Cells, [SDV.IMM]Life Sciences [q-bio]/Immunology, MESH : Cell Differentiation, MESH : Colony-Forming Units Assay, Adult stem cell, MESH: Cells, Cultured, MESH: Cell Differentiation, MESH: Gene Expression, [SDV.IMM] Life Sciences [q-bio]/Immunology, MESH : Flow Cytometry, MESH: Glycoproteins, MESH: Stem Cells, Biology, Bicuculline, MESH : Neurons, Cell Line, Colony-Forming Units Assay, 03 medical and health sciences, MESH: Bicuculline, Antigens, CD, MESH : Cells, Cultured, MESH : Antigens, CD, MESH : Baclofen, Humans, MESH: Colony-Forming Units Assay, MESH: Fetal Blood, GABA-A Receptor Antagonists, Progenitor cell, Glycoproteins, 030304 developmental biology, Interleukin 3, MESH : Fetal Blood, MESH: Humans, MESH : Hematopoietic Stem Cells, MESH : Humans, MESH : Receptors, GABA-A, MESH : Receptors, GABA-B, Cell Biology, Hematopoietic Stem Cells, Receptors, GABA-A, MESH : Glycoproteins, MESH: Cell Line, MESH : Gene Expression, MESH : Fluorescent Antibody Technique, Receptors, GABA-B, nervous system, MESH : Chemotaxis, Cell culture, Immunology, Peptides, GABA-B Receptor Antagonists, 030217 neurology & neurosurgery, Developmental Biology, Homing (hematopoietic)
Abstract: International audience; Migration of stem/progenitor cells is a crucial event for homing toward tissue where cells need to be renewed. The neurotransmitter gamma-aminobutyric acid (GABA) has been shown to have a crucial role in migration of neuronal stem/progenitor cells. Since human umbilical cord blood (HUCB) contains stem/progenitor cells able to generate either neuronal or hematopoietic cells, we evaluated the effect of GABA on this type of cells. While whole fraction of mononuclear cells expressed GABA(A) and GABA(B) receptor subunits (GABA-R), only GABA(B)R subunits were found to be expressed on immature CD133+ cells. Functional experiments revealed that both cell fractions of HUCB were attracted by a gradient of GABA concentration and furthermore were blocked by specific antagonists of GABA(A)R and GABA(B)R bicuculline and saclofen, respectively. Moreover, through GABA(B)R activation the migrating fraction was highly enriched by both hematopoietic progenitors and cells able to generate neuron- like cells in culture. Therefore, GABA is a potent chemoattractant of HUCB stem/progenitor cells specifically through GABA(B)R activation.
Published: 2009
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46. Membrane-associated Hsp72 from tumor-derived exosomes mediates STAT3-dependent immunosuppressive function of mouse and human myeloid-derived suppressor cells

Author: Alain Rouleau, Jean-Paul Remy-Martin, David Lanneau, Benoit Simon, Lionel Apetoh, Christophe Borg, Gabriele Multhoff, Grégoire Mignot, Cédric Rébé, Bruno Chauffert, François Ghiringhelli, François Martin, Fanny Chalmin, Eric Solary, Sylvain Ladoire, Mélanie Bruchard, Carmen Garrido, Arlette Hamman, Bernhard Ryffel, Wilfrid Boireau, Julie Vincent, Aurélie de Thonel, Laurence Zitvogel, Lipides - Nutrition - Cancer (U866) ( LNC ), Université de Bourgogne ( UB ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Ecole Nationale Supérieure de Biologie Appliquée à la Nutrition et à l'Alimentation de Dijon ( ENSBANA ), Centre Régional de Lutte contre le cancer - Centre Georges-François Leclerc ( CRLCC - CGFL ), Interactions hôte-greffon-tumeur, ingénierie cellulaire et génique - UFC ( HOTE GREFFON ), Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Etablissement français du sang [Bourgogne-France-Comté] ( EFS [Bourgogne-France-Comté] ) -Université de Franche-Comté ( UFC ), Franche-Comté Électronique Mécanique, Thermique et Optique - Sciences et Technologies ( FEMTO-ST ), Université de Technologie de Belfort-Montbeliard ( UTBM ) -Ecole Nationale Supérieure de Mécanique et des Microtechniques ( ENSMM ) -Centre National de la Recherche Scientifique ( CNRS ) -Université de Franche-Comté ( UFC ), Department of Radiation Oncology [Munich], Ludwig-Maximilians-Universität München, Role des Cellules Dendritiques Dans la Regulation des Effecteurs de l'Immunite Antitumorale, Université Paris-Sud - Paris 11 ( UP11 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Immunologie et embryologie moléculaires ( IEM ), Université d'Orléans ( UO ) -Centre National de la Recherche Scientifique ( CNRS ), Center for Neurologic Diseases, Harvard Medical School [Boston] ( HMS ), This work was supported by special grants from the Fondation de France, the Association pour la Recherche sur le Cancer, the Institut National de la Santé et de la Recherche Médicale, and the Ligue Nationale Contre le Cancer (Region Bourgogne). F. Chalmin was supported by the Conseil Régional Bourgogne/INSERM, J. Vincent by the Institut National contre le Cancer. G. Mignot was funded by the Association pour la Recherche sur le Cancer. C. Garrido and E. Solary lead teams supported by the 'Ligue Nationale Contre le Cancer.' L. Apetoh is supported by EMBO., Lipides - Nutrition - Cancer (U866) (LNC), Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Ecole Nationale Supérieure de Biologie Appliquée à la Nutrition et à l'Alimentation de Dijon (ENSBANA), Centre Régional de Lutte contre le cancer Georges-François Leclerc [Dijon] (UNICANCER/CRLCC-CGFL), UNICANCER, Interactions hôte-greffon-tumeur, ingénierie cellulaire et génique - UFC (UMR INSERM 1098) (RIGHT), Institut National de la Santé et de la Recherche Médicale (INSERM)-Etablissement français du sang [Bourgogne-Franche-Comté] (EFS [Bourgogne-Franche-Comté])-Université de Franche-Comté (UFC), Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université Bourgogne Franche-Comté [COMUE] (UBFC), Franche-Comté Électronique Mécanique, Thermique et Optique - Sciences et Technologies (UMR 6174) (FEMTO-ST), Université de Technologie de Belfort-Montbeliard (UTBM)-Ecole Nationale Supérieure de Mécanique et des Microtechniques (ENSMM)-Université de Franche-Comté (UFC), Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université Bourgogne Franche-Comté [COMUE] (UBFC)-Centre National de la Recherche Scientifique (CNRS), Ludwig-Maximilians-Universität München (LMU), Université Paris-Sud - Paris 11 (UP11)-Institut National de la Santé et de la Recherche Médicale (INSERM), Immunologie et Embryologie Moléculaires (IEM), Université d'Orléans (UO)-Centre National de la Recherche Scientifique (CNRS), Harvard Medical School [Boston] (HMS), Centre épigénétique et destin cellulaire (EDC (UMR_7216)), Centre National de la Recherche Scientifique (CNRS)-Université de Paris (UP), Mignot, Grégoire, and Université de Technologie de Belfort-Montbeliard (UTBM)-Ecole Nationale Supérieure de Mécanique et des Microtechniques (ENSMM)-Centre National de la Recherche Scientifique (CNRS)-Université de Franche-Comté (UFC)
Subjects: MAPK/ERK pathway, MESH : Cell Line, [SDV]Life Sciences [q-bio], T-Lymphocytes, HSP72 Heat-Shock Proteins, Exosomes, T-Lymphocytes, Regulatory, MESH: Amiloride, Amiloride, MESH : T-Lymphocytes, Regulatory, Mice, 0302 clinical medicine, Neoplasms, [ SDV.IMM ] Life Sciences [q-bio]/Immunology, MESH: Animals, MESH: Neoplasms, ComputingMilieux_MISCELLANEOUS, MESH : Cyclophosphamide, 0303 health sciences, education.field_of_study, MESH: Immunosuppression, MESH : Mice, Nude, General Medicine, 3. Good health, Cell biology, medicine.anatomical_structure, 030220 oncology & carcinogenesis, [SDV.IMM]Life Sciences [q-bio]/Immunology, MESH : HSP72 Heat-Shock Proteins, Research Article, MESH: Cell Line, Tumor, [SDV.IMM] Life Sciences [q-bio]/Immunology, T cell, Population, Mice, Nude, Biology, Exosome, MESH: HSP72 Heat-Shock Proteins, Cell Line, 03 medical and health sciences, MESH : Amiloride, Cell Line, Tumor, MESH : Mice, medicine, MESH: Mice, Nude, Animals, Humans, MESH: Exosomes, education, Autocrine signalling, MESH: Mice, Cyclophosphamide, 030304 developmental biology, MESH : T-Lymphocytes, Immunosuppression Therapy, MESH: Humans, MESH : Cell Line, Tumor, MESH: T-Lymphocytes, Regulatory, MESH : Humans, MESH: Cyclophosphamide, MESH : Exosomes, MESH : Neoplasms, Microvesicles, MESH: Cell Line, TLR2, MESH: T-Lymphocytes, MESH : Immunosuppression, Myeloid-derived Suppressor Cell, MESH : Animals
Abstract: International audience; Myeloid-derived suppressor cells (MDSCs) have been identified in humans and mice as a population of immature myeloid cells with the ability to suppress T cell activation. They accumulate in tumor-bearing mice and humans and have been shown to contribute to cancer development. Here, we have isolated tumor-derived exosomes (TDEs) from mouse cell lines and shown that an interaction between TDE-associated Hsp72 and MDSCs determines the suppressive activity of the MDSCs via activation of Stat3. In addition, tumor-derived soluble factors triggered MDSC expansion via activation of Erk. TDE-associated Hsp72 triggered Stat3 activation in MDSCs in a TLR2/MyD88-dependent manner through autocrine production of IL-6. Importantly, decreasing exosome production using dimethyl amiloride enhanced the in vivo antitumor efficacy of the chemotherapeutic drug cyclophosphamide in 3 different mouse tumor models. We also demonstrated that this mechanism is relevant in cancer patients, as TDEs from a human tumor cell line activated human MDSCs and triggered their suppressive function in an Hsp72/TLR2-dependent manner. Further, MDSCs from cancer patients treated with amiloride, a drug used to treat high blood pressure that also inhibits exosome formation, exhibited reduced suppressor functions. Collectively, our findings show in both mice and humans that Hsp72 expressed at the surface of TDEs restrains tumor immune surveillance by promoting MDSC suppressive functions.
Published: 2009
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47. Peptides that mimic the amino-terminal end of the rabies virus phosphoprotein have antiviral activity

Author: Christophe Prehaud, Noël Tordo, Stéphanie Méhouas, Corinne Jallet, Rob W.H. Ruigrok, Guillaume Castel, Eléonore Real, Yves Jacob, M. Chteoui, Grégory Caignard, Stratégies Antivirales, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique ( CNRS ), Génomique Virale et Vaccination, Neuro-Immunologie Virale, Immunologie, génétique et traitement des maladies métaboliques et du diabète ( UMR_S 561 ), Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Université Paris Descartes - Paris 5 ( UPD5 ), Laboratory of Neurogenetics and Behavior, The Rockefeller University, Génétique, Papillomavirus et Cancer Humain, Institut Pasteur [Paris], Unit for Virus Host-Cell Interactions [Grenoble] ( UVHCI ), Université Joseph Fourier - Grenoble 1 ( UJF ) -European Molecular Biology Laboratory [Grenoble] ( EMBL ) -Centre National de la Recherche Scientifique ( CNRS ), Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Immunologie, génétique et traitement des maladies métaboliques et du diabète (UMR_S 561), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Rockefeller University [New York], Unit of Virus Host Cell Interactions (UVHCI), Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), Institut Pasteur [Paris] (IP), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Descartes - Paris 5 (UPD5), Unit for Virus Host-Cell Interactions [Grenoble] (UVHCI), and Université Joseph Fourier - Grenoble 1 (UJF)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Centre National de la Recherche Scientifique (CNRS)
Subjects: MESH : Cell Line, viruses, MESH: Neurons, MESH: Cricetinae, Virus Replication, medicine.disease_cause, [ SDV.MP ] Life Sciences [q-bio]/Microbiology and Parasitology, Cricetinae, MESH : Rabies virus, MESH: Animals, Mononegavirales, Neurons, 0303 health sciences, biology, MESH: Peptides, MESH : Peptides, 030302 biochemistry & molecular biology, MESH : Viral Structural Proteins, MESH : Cricetinae, MESH: Rabies virus, 3. Good health, MESH : Antiviral Agents, MESH : Virus Replication, [SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology, Vesicular stomatitis virus, MESH: Antiviral Agents, MESH: Cell Line, Tumor, Immunology, MESH: Viral Structural Proteins, MESH: Two-Hybrid System Techniques, MESH: Phosphoproteins, Antiviral Agents, Microbiology, Virus, MESH : Neurons, Cell Line, MESH : Immunoprecipitation, 03 medical and health sciences, Cell Line, Tumor, Two-Hybrid System Techniques, Virology, Vaccines and Antiviral Agents, medicine, Animals, Humans, Immunoprecipitation, Lyssavirus, 030304 developmental biology, Viral Structural Proteins, MESH: Humans, MESH: Immunoprecipitation, MESH : Cell Line, Tumor, MESH: Virus Replication, Rabies virus, MESH : Humans, Rhabdoviridae, Phosphoproteins, biology.organism_classification, Molecular biology, MESH: Cell Line, Nucleoprotein, Viral replication, MESH : Two-Hybrid System Techniques, Insect Science, MESH : Animals, MESH : Phosphoproteins, Peptides, Molecular Chaperones
Abstract: We wanted to develop a therapeutic approach against rabies disease by targeting the lyssavirus transcription/replication complex. Because this complex (nucleoprotein N-RNA template processed by the L polymerase and its cofactor, the phosphoprotein P) is similar to that of other negative-strand RNA viruses, we aimed to design broad-spectrum antiviral drugs that could be used as a complement to postexposure vaccination and immunotherapy. Recent progress in understanding the structure/function of the rabies virus P, N, and L proteins predicts that the amino-terminal end of P is an excellent target for destabilizing the replication complex because it interacts with both L (for positioning onto the N-RNA template) and N (for keeping N soluble, as needed for viral RNA encapsidation). Thus, peptides mimicking various lengths of the amino-terminal end of P have been evaluated, as follows: (i) for binding properties to the N-P-L partners by the two-hybrid method; (ii) for their capacity to inhibit the transcription/replication of a rabies virus minigenome encoding luciferase in BHK-21-T7 cells; and (iii) for their capacity to inhibit rabies virus infection of BHK-21-T7 cells and of two derivatives of the neuronal SK-N-SH cell line. Peptides P60 and P57 (the first 60 and first 57 NH 2 residues of P, respectively) exhibited a rapid, strong, and long-lasting inhibitory potential on luciferase expression (>95% from 24 h to 55 h). P42 was less efficient in its inhibition level (75% for 18 to 30 h) and duration (40% after 48 h). The most promising peptides were synthesized in tandem with the Tat sequence, allowing cell penetration. Their inhibitory effects were observed on BHK-21-T7 cells infected with rabies virus and Lagos bat virus but not with vesicular stomatitis virus. In neuronal cells, a significant inhibition of both nucleocapsid inclusions and rabies virus release was observed.
Published: 2009
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48. Drug development in oncology assisted by noninvasive optical imaging

Author: Didier Boturyn, Philippe Rizo, Stéphane Roux, Anne-Charlotte Faure, Jérôme Boutet, Sandrine Dufort, Lucie Sancey, Anne Koenig, Véronique Josserand, Pascal Dumy, Michelle Keramidas, Claire Rome, Stéphanie Foillard, Jean-Luc Coll, Christian Righini, Olivier Tillement, Institut d'oncologie/développement Albert Bonniot de Grenoble (INSERM U823), Institut National de la Santé et de la Recherche Médicale (INSERM)-EFS-CHU Grenoble-Université Joseph Fourier - Grenoble 1 (UJF), UF Cancérologie Biologique et Biothérapie, Université Joseph Fourier - Grenoble 1 (UJF)-CHU Grenoble, INSERM U823, équipe 5 (cibles diagnostiques ou thérapeutiques et vectorisation de drogues dans le cancer du poumon), Institut National de la Santé et de la Recherche Médicale (INSERM)-EFS-CHU Grenoble-Université Joseph Fourier - Grenoble 1 (UJF)-Institut National de la Santé et de la Recherche Médicale (INSERM)-EFS-CHU Grenoble-Université Joseph Fourier - Grenoble 1 (UJF), Laboratoire de Physico-Chimie des Matériaux Luminescents (LPCML), Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon, Département de Chimie Moléculaire (DCM), Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Joseph Fourier - Grenoble 1 (UJF), Commissariat à l'énergie atomique et aux énergies alternatives - Laboratoire d'Electronique et de Technologie de l'Information (CEA-LETI), Direction de Recherche Technologique (CEA) (DRT (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), INSERM, INCA, ARC, ANR, EMIL and N2L NoE of the 6th FWP, ANR-06-BLAN-0071,PEPVEC4THER,Peptide based nanostructure for the delivery of therapeutics: from mechanism to application(2006), Institut d'oncologie/développement Albert Bonniot de Grenoble ( INSERM U823 ), Université Joseph Fourier - Grenoble 1 ( UJF ) -CHU Grenoble-EFS-Institut National de la Santé et de la Recherche Médicale ( INSERM ), Université Joseph Fourier - Grenoble 1 ( UJF ) -CHU Grenoble, Université Joseph Fourier - Grenoble 1 ( UJF ) -CHU Grenoble-EFS-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Université Joseph Fourier - Grenoble 1 ( UJF ) -CHU Grenoble-EFS-Institut National de la Santé et de la Recherche Médicale ( INSERM ), Laboratoire de Physico-Chimie des Matériaux Luminescents ( LPCML ), Université Claude Bernard Lyon 1 ( UCBL ), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique ( CNRS ), Département de Chimie Moléculaire ( DCM ), Université Joseph Fourier - Grenoble 1 ( UJF ) -Centre National de la Recherche Scientifique ( CNRS ) -Université Grenoble Alpes ( UGA ), Laboratoire d'Electronique et des Technologies de l'Information ( CEA-LETI ), Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Grenoble Alpes [Saint Martin d'Hères], ANR-06-BLAN-0071,PEPVEC4THER,Peptide based nanostructure for the delivery of therapeutics: from mechanism to application ( 2006 ), Hurbin, Amandine, Programme 'blanc' - Peptide based nanostructure for the delivery of therapeutics: from mechanism to application - - PEPVEC4THER2006 - ANR-06-BLAN-0071 - BLANC - VALID, Université Joseph Fourier - Grenoble 1 (UJF)-CHU Grenoble-EFS-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Joseph Fourier - Grenoble 1 (UJF)-CHU Grenoble-EFS-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Joseph Fourier - Grenoble 1 (UJF)-CHU Grenoble-EFS-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), and Université Joseph Fourier - Grenoble 1 (UJF)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)
Subjects: MESH : Cell Line, Oncology, MESH : Microscopy, Fluorescence, MESH : Diagnostic Imaging, Pharmaceutical Science, MESH: Microscopy, Fluorescence, MESH : Radiation Oncology, [ SDV.CAN ] Life Sciences [q-bio]/Cancer, Metastasis, Mice, Drug Delivery Systems, 0302 clinical medicine, Neoplasms, Drug Discovery, MESH : Female, MESH: Animals, MESH: Neoplasms, MESH : Drug Discovery, 0303 health sciences, Optical Devices, MESH : Mice, Nude, Primary tumor, MESH: Radiation Oncology, 3. Good health, MESH : Optical Devices, MESH : Antineoplastic Agents, Drug development, 030220 oncology & carcinogenesis, Drug delivery, MESH : Drug Delivery Systems, Female, Diagnostic Imaging, medicine.medical_specialty, MESH: Cell Line, Tumor, MESH: Drug Delivery Systems, Cancer therapy, Mice, Nude, Antineoplastic Agents, Context (language use), [SDV.CAN]Life Sciences [q-bio]/Cancer, Cell Line, 03 medical and health sciences, optical imaging, Optical imaging, [SDV.CAN] Life Sciences [q-bio]/Cancer, MESH: Drug Discovery, Cell Line, Tumor, Internal medicine, MESH : Mice, medicine, MESH: Mice, Nude, Animals, Humans, MESH: Mice, 030304 developmental biology, MESH: Optical Devices, MESH: Humans, MESH : Cell Line, Tumor, business.industry, tumor targeting, MESH: Diagnostic Imaging, MESH : Humans, Cancer, medicine.disease, MESH : Neoplasms, MESH: Cell Line, Microscopy, Fluorescence, Radiation Oncology, MESH: Antineoplastic Agents, MESH : Animals, business, MESH: Female
Abstract: International audience; Early and accurate detection of tumors, like the development of targeted treatments, is a major field of research in oncology. The generation of specific vectors, capable of transporting a drug or a contrast agent to the primary tumor site as well as to the remote (micro-) metastasis would be an asset for early diagnosis and cancer therapy. Our goal was to develop new treatments based on the use of tumor-targeted delivery of large biomolecules (DNA, siRNA, peptides, or nanoparticles), able to induce apoptosis while dodging the specific mechanisms developed by tumor cells to resist this programmed cell death. Nonetheless, the insufficient effectiveness of the vectorization systems is still a crucial issue. In this context, we generated new targeting vectors for drug and biomolecules delivery and developed several optical imaging systems for the follow-up and evaluation of these vectorization systems in live mice. Based on our recent work, we present a brief overview of how noninvasive optical imaging in small animals can accelerate the development of targeted therapeutics in oncology.
Published: 2009
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49. Anoikis resistance of mesothelioma cells

Author: Jocelyne Fleury-Feith, Annie Renier, Daubriac J, Saint-Albin A, Josephine Galipon, Laurence Kheuang, Françoise Galateau-Sallé, Marco Giovannini, Marie-Claude Jaurand, Genomique Fonctionnelle des Tumeurs Solides, Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Université Paris Descartes - Paris 5 ( UPD5 ) -IFR105-Université Paris Diderot - Paris 7 ( UPD7 ), Laboratoire d'histologie et de biologie tumorale, Université Pierre et Marie Curie - Paris 6 ( UPMC ) -Assistance publique - Hôpitaux de Paris (AP-HP)-CHU Tenon [APHP], Institut Mondor de recherche biomédicale ( IMRB ), Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Université Paris-Est Créteil Val-de-Marne - Paris 12 ( UPEC UP12 ), Cancers et Populations : Facteurs de Risque, Depistage, Pratiques Diagnostiques et Therapeutiques, Université de Caen Normandie ( UNICAEN ), Normandie Université ( NU ) -Normandie Université ( NU ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Ligue Nationale Contre le Cancer (comité de l'Oise, France), R07103HH Agence Française de Sécurité Sanitaire de l'Environnement et du Travail RD-2004-015. JD is a fellow of the French Ministry of Research and the Fondation pour la Recherche Médicale, 05 9 31/ANR 05 SEST029-01,05 9 31/ANR 05 SEST029-01, Génomique Fonctionnelle des Tumeurs Solides (U1162), Université Paris 13 (UP13)-Université Paris Diderot - Paris 7 (UPD7)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), CHU Tenon [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Institut Mondor de recherche biomédicale (IMRB), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12), Université de Caen Normandie (UNICAEN), Normandie Université (NU)-Normandie Université (NU)-Institut National de la Santé et de la Recherche Médicale (INSERM), ANR-05-SEST-0029,Modélisation génétique du mésothéliome pour la mise en évidence de marqueurs d'exposition et de marqueurs d'effets de l'exposition a des fibres minérales dans le mésothéliome et dans le cancer broncho-pulmonaire dû aux fibres d'amiante(2005), and Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)
Subjects: MESH: Signal Transduction, MESH : Cell Line, MAPK/ERK pathway, Mesothelioma, MESH : Mesothelioma, MESH : Pleural Neoplasms, MESH: Anoikis, MESH : RNA, Small Interfering, [ SDV.CAN ] Life Sciences [q-bio]/Cancer, 0302 clinical medicine, MESH: RNA, Small Interfering, Medicine, Anoikis, RNA, Small Interfering, Extracellular Signal-Regulated MAP Kinases, MESH: Extracellular Signal-Regulated MAP Kinases, MESH : Anisomycin, 0303 health sciences, Bcl-2-Like Protein 11, MESH : JNK Mitogen-Activated Protein Kinases, MESH: G0 Phase, SAPK/JNK, Cell cycle, MESH : Extracellular Signal-Regulated MAP Kinases, Cell aggregation, Cell biology, MESH : Proto-Oncogene Proteins c-akt, medicine.anatomical_structure, 030220 oncology & carcinogenesis, MESH : Proto-Oncogene Proteins, MESH: Membrane Proteins, MESH : Apoptosis Regulatory Proteins, Anisomycin, Signal Transduction, Pleural Neoplasms, MESH: Anisomycin, [SDV.CAN]Life Sciences [q-bio]/Cancer, Resting Phase, Cell Cycle, MESH: Pleural Neoplasms, Cell Line, 03 medical and health sciences, MESH : G0 Phase, Proto-Oncogene Proteins, G0, Humans, Bim, Molecular Biology, Protein kinase B, PI3K/AKT/mTOR pathway, 030304 developmental biology, MESH : Signal Transduction, MESH: Humans, MESH: Mesothelioma, MESH: Proto-Oncogene Proteins c-akt, MESH: Apoptosis Regulatory Proteins, business.industry, MESH : Humans, JNK Mitogen-Activated Protein Kinases, Membrane Proteins, MESH: JNK Mitogen-Activated Protein Kinases, Cell Biology, Pleural cavity, MESH: Cell Line, MESH: Proto-Oncogene Proteins, cell junction, MESH : Anoikis, MESH : Membrane Proteins, Cell culture, business, Apoptosis Regulatory Proteins, Proto-Oncogene Proteins c-akt
Abstract: International audience; Pleural fluid accumulation is a frequent clinical observation in diffuse malignant pleural mesothelioma (MPM). The cytological analysis of pleural fluid often reveals the presence of free spheroid aggregates of malignant cells, giving rise to the question of the ability of non-adherent tumor cells to resist the loss of anchorage-induced apoptosis (termed as anoikis), and to develop new tumor foci in the pleural cavity. Here, we show that MPM cells cultured under non-adherent conditions form well-organized aggregates composed of viable cells, which progressively enter in G(0). Although the PI3K/Akt, ERK and SAPK/JNK signaling pathways are activated in adherent MPM cells, loss of anchorage results in the inactivation of these pathways. By comparison, we show that the non-tumoral mesothelial cells MeT-5A enter anoikis in an SAPK/JNK-, Bim- and caspase-9-dependent pathway. The survival of MPM cells can be reversed by activating SAPK/JNK with anisomycin, according to a Bim-dependent mitochondrial pathway. Finally, our findings show that impairment of cell aggregation activates SAPK/JNK and Bim and induces anoikis. Our results underline the importance of intercellular contacts in the anoikis resistance of MPM cells.
Published: 2009
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50. Aminopeptidase A contributes to the N-terminal truncation of amyloid beta-peptide

Author: Marie-Claude Fournie-Zaluski, Frédéric Checler, Bernard P. Roques, Jean Sevalle, Audrey Amoyel, Philippe Robert, Institut de pharmacologie moléculaire et cellulaire (IPMC), Université Nice Sophia Antipolis (... - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Centre National de la Recherche Scientifique (CNRS), Centre Mémoire de Ressources et de Recherche [Nice] (CMRR Nice), Université Côte d'Azur (UCA)-Centre Hospitalier Universitaire de Nice (CHU Nice)-Université Nice Sophia Antipolis (... - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA), Pharmaleads, Université Paris Descartes - Paris 5 (UPD5), Institut de pharmacologie moléculaire et cellulaire ( IPMC ), Université Nice Sophia Antipolis ( UNS ), Université Côte d'Azur ( UCA ) -Université Côte d'Azur ( UCA ) -Centre National de la Recherche Scientifique ( CNRS ), Centre Mémoire de Ressources et de Recherche [Nice] ( CMRR Nice ), Université Côte d'Azur ( UCA ) -Université Côte d'Azur ( UCA ) -CHU Nice, Université Paris Descartes - Paris 5 ( UPD5 ), Université Côte d'Azur (UCA)-Université Côte d'Azur (UCA)-Centre National de la Recherche Scientifique (CNRS), and Université Côte d'Azur (UCA)-Université Côte d'Azur (UCA)-Centre Hospitalier Universitaire de Nice (CHU Nice)
Subjects: MESH : Cell Line, MESH : Glutamyl Aminopeptidase, cell free amyloid-β production, Peptide, Biochemistry, Aminopeptidase, MESH: Amyloid beta-Protein, MESH: Mice, Knockout, Mice, chemistry.chemical_compound, MESH: Protein Structure, Tertiary, 0302 clinical medicine, inhibitors, Amyloid precursor protein, MESH: Animals, MESH: Peptide Fragments, Mice, Knockout, chemistry.chemical_classification, MESH: Glutamyl Aminopeptidase, 0303 health sciences, biology, MESH : Peptides, MESH: Peptides, MESH : Amyloid beta-Protein, cell death, [SDV.NEU]Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC], MESH : Protein Structure, Tertiary, Intracellular, amyloid-β degradation, Proteases, Amyloid beta, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Glutamyl Aminopeptidase, Cell Line, 03 medical and health sciences, Cellular and Molecular Neuroscience, Amastatin, MESH : Mice, mental disorders, Animals, Humans, APLP2, MESH: Mice, 030304 developmental biology, Amyloid beta-Peptides, MESH: Humans, [ SDV.BC ] Life Sciences [q-bio]/Cellular Biology, MESH : Humans, MESH : Peptide Fragments, Peptide Fragments, N-terminally truncated amyloid-β fragments, Protein Structure, Tertiary, MESH: Cell Line, chemistry, [ SDV.NEU ] Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC], biology.protein, MESH : Mice, Knockout, aminopeptidase A, MESH : Animals, Peptides, 030217 neurology & neurosurgery
Abstract: International audience; Several lines of data previously indicated that N-terminally truncated forms of amyloid-beta (Abeta) peptides are likely the earliest and more abundant species immunohistochemically detectable in Alzheimer's disease-affected brains. It is noteworthy that the free N-terminal residue of full-length Abeta (fl-Abeta) is an aspartyl residue, suggesting that Abeta could be susceptible to exopeptidasic attack by aminopeptidase A (APA)-like proteases. In this context, we have examined whether APA could target Abeta peptides in both cell-free and cellular models. We first show that the general aminopeptidase inhibitor amastatin as well as two distinct aminopeptidase A inhibitors EC33 and pl302 both significantly increase the recovery of genuine fl-Abeta peptides generated by cells over-expressing Swedish-mutated beta amyloid precursor protein (APP) while the aminopeptidase N blocker pl250 did not modify fl-Abeta recovery. In agreement with this observation, we establish that over-expressed APA drastically reduces, in a calcium dependent manner, fl-Abeta but not APP IntraCellular Domain in a cell-free model of Abeta production. In agreement with the above data, we show that recombinant APA degrades fl-Abeta in a pl302-sensitive manner. Interestingly, we also show that EC33 and pl302 lower staurosporine-stimulated activation of caspase-3 in wild-type fibroblasts but not in betaAPP/beta-amyloid precursor protein-like protein 2 (APLP2) double knockout fibroblasts, suggesting that protecting endogenous fl-Abeta physiological production triggers neuroprotective phenotype. By contrast, EC33 does not modify staurosporine-induced caspase-3 activation in wild-type and Swedish-mutated betaAPP-HEK293 expressing cells that display exacerbated production of Abeta. Overall, our data establish that APA contributes to the N-terminal truncation of Abeta and suggest that this cleavage is likely abrogating a protective function associated with physiological but not supraphysiological levels of genuine fl-Abeta peptides.
Published: 2009
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