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15 results on '"MESH : Cell Adhesion"'

1. Interplay of RhoA and mechanical forces in collective cell migration driven by leader cells

Author

Sylvie Coscoy, François Amblard, A. Buguin, Myriam Reffay, Benoit Ladoux, Jacques Camonis, Pascal Silberzan, Olivier Cochet-Escartin, Maria Carla Parrini, Physico-Chimie-Curie (PCC), Centre National de la Recherche Scientifique (CNRS)-Institut Curie-Université Pierre et Marie Curie - Paris 6 (UPMC), CNRS UMR 7057 - Laboratoire Matières et Systèmes Complexes (MSC) (MSC), Centre National de la Recherche Scientifique (CNRS), Unité de génétique et biologie des cancers (U830), Université Paris Descartes - Paris 5 (UPD5)-Institut Curie-Institut National de la Santé et de la Recherche Médicale (INSERM), Matière et Systèmes Complexes (MSC (UMR_7057)), Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS), Institut Jacques Monod (IJM (UMR_7592)), Mechanobiology Institute [Singapore] (MBI), National University of Singapore (NUS), C'nano Ile de France, Association Christelle Bouillot, Association pour la Recherche sur le Cancer, Labex CelTisPhyBio, Institut Curie Programme Incitatif et Coopératif 'Physique de la Cellule', Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut Curie [Paris]-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Matière et Systèmes Complexes (MSC), 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 Curie [Paris]-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de Chimie du CNRS (INC), Centre National de la Recherche Scientifique (CNRS)-Université Paris Diderot - Paris 7 (UPD7), Physico-Chimie-Curie ( PCC ), Centre National de la Recherche Scientifique ( CNRS ) -INSTITUT CURIE-Université Pierre et Marie Curie - Paris 6 ( UPMC ), CNRS UMR 7057 - Laboratoire Matières et Systèmes Complexes (MSC) ( MSC ), Centre National de la Recherche Scientifique ( CNRS ), Unité de génétique et biologie des cancers ( U830 ), Université Paris Descartes - Paris 5 ( UPD5 ) -Institut Curie-Institut National de la Santé et de la Recherche Médicale ( INSERM ), Matière et Systèmes Complexes ( MSC ), Université Paris Diderot - Paris 7 ( UPD7 ) -Centre National de la Recherche Scientifique ( CNRS ), Institut Jacques Monod ( IJM ), Mechanobiology Institute [Singapore] ( MBI ), and National University of Singapore ( NUS )

Subjects
rac1 GTP-Binding Protein, RHOA, MESH: Fluorescence Resonance Energy Transfer, GTPase, Madin Darby Canine Kidney Cells, MESH: Dogs, 0302 clinical medicine, MESH : Dogs, MESH : Protein Transport, Cell Movement, Fluorescence Resonance Energy Transfer, MESH : Cell Movement, MESH: Animals, Pseudopodia, MESH: Cell Movement, Physics, 0303 health sciences, Tractive force, biology, Biomechanical Phenomena, Cell biology, Actin Cytoskeleton, Protein Transport, MESH: rhoA GTP-Binding Protein, MESH: Protein Transport, MESH: Biomechanical Phenomena, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, MESH: Cell Adhesion, Mechanical traction, MESH : rac1 GTP-Binding Protein, 03 medical and health sciences, Dogs, Cell Adhesion, Animals, Cell adhesion, MESH : Biomechanical Phenomena, MESH : rhoA GTP-Binding Protein, 030304 developmental biology, MESH : Madin Darby Canine Kidney Cells, [ SDV.BC ] Life Sciences [q-bio]/Cellular Biology, MESH: rac1 GTP-Binding Protein, Collective cell migration, MESH: Madin Darby Canine Kidney Cells, Cell Biology, MESH : Cell Adhesion, Multicellular organism, MESH : Fluorescence Resonance Energy Transfer, MESH : Actin Cytoskeleton, MESH: Pseudopodia, MESH : Pseudopodia, biology.protein, MESH : Animals, MESH: Actin Cytoskeleton, rhoA GTP-Binding Protein, 030217 neurology & neurosurgery
Abstract

International audience; The leading front of a collectively migrating epithelium often destabilizes into multicellular migration fingers where a cell initially similar to the others becomes a leader cell while its neighbours do not alter. The determinants of these leader cells include mechanical and biochemical cues, often under the control of small GTPases. However, an accurate dynamic cartography of both mechanical and biochemical activities remains to be established. Here, by mapping the mechanical traction forces exerted on the surface by MDCK migration fingers, we show that these structures are mechanical global entities with the leader cells exerting a large traction force. Moreover, the spatial distribution of RhoA differential activity at the basal plane strikingly mirrors this force cartography. We propose that RhoA controls the development of these fingers through mechanical cues: the leader cell drags the structure and the peripheral pluricellular acto-myosin cable prevents the initiation of new leader cells.

Published
2014
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2. Sex differences in the GSK3β-mediated survival of adherent leukemic progenitors

Author

Jessica Bertrand, Bernard Payrastre, N Gallay, Nathalie Vergnolle, Véronique Maguer-Satta, Mathieu Despeaux, Cécile Demur, Ezzeddine Bourogaa, S Joly, P Bonnevialle, F Louache, Claire Racaud-Sultan, ANDRA, Laboratoire d'Hématologie [Purpan], Université Paul Sabatier - Toulouse 3 ( UPS ) -CHU Toulouse [Toulouse]-Hôpital Purpan [Toulouse], Equipe 11, 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 ) -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 ), Centre de Physiopathologie Toulouse Purpan, Université Paul Sabatier - Toulouse 3 ( UPS ) -IFR30-IFR150-Institut National de la Santé et de la Recherche Médicale ( INSERM ), Departement /u563 : Oncogenèse, Signalisation et Innovation thérapeutique, Université Paul Sabatier - Toulouse 3 ( UPS ) -Institut Claudius Regaud-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-CHU Toulouse [Toulouse]-Hôpital Purpan [Toulouse], CHU Toulouse [Toulouse], 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)-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), Centre de Physiopathologie Toulouse Purpan (CPTP), 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), 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)-Université Toulouse III - Paul Sabatier (UT3), and 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)-Institut Claudius Regaud

Subjects
Male, MESH : Aged, Antigens, CD34, MESH : Blotting, Western, [ SDV.CAN ] Life Sciences [q-bio]/Cancer, Glycogen Synthase Kinase 3, 0302 clinical medicine, MESH : Indoles, MESH : Tumor Cells, Cultured, GSK-3, Tumor Cells, Cultured, Protein Phosphatase 2, MESH : GTP-Binding Proteins, Cells, Cultured, MESH: Glycogen Synthase Kinase 3, MESH: Etoposide, Etoposide, MESH: Indoles, MESH : Maleimides, Aged, 80 and over, MESH : Cell Survival, 0303 health sciences, education.field_of_study, MESH: Middle Aged, MESH: Protein Phosphatase 2, Neoplasm Proteins, MESH: Cell Survival, MESH: Young Adult, 030220 oncology & carcinogenesis, Neoplastic Stem Cells, RNA Interference, MESH: Cells, Cultured, MESH : Protein Phosphatase 2, MESH : Glycogen Synthase Kinase 3, MESH: Antineoplastic Agents, Phytogenic, Blotting, Western, MESH : Young Adult, Receptors, Cell Surface, MESH: Cell Adhesion, 03 medical and health sciences, MESH : Neoplasm Proteins, MESH: Sex Factors, Genetics, MESH: Blotting, Western, Humans, MESH : Middle Aged, MESH: Tumor Cells, Cultured, Progenitor cell, MESH : Aged, 80 and over, Clonogenic assay, education, Molecular Biology, Aged, MESH: Interleukin-3 Receptor alpha Subunit, MESH: Humans, Glycogen Synthase Kinase 3 beta, MESH : Hematopoietic Stem Cells, MESH : Humans, MESH : Leukemia, MESH: Adult, MESH: Antigens, CD34, Protein phosphatase 2, Hematopoietic Stem Cells, MESH: Neoplastic Stem Cells, Immunology, MESH: Female, MESH: Neoplasm Proteins, Cancer Research, Indoles, MESH : Receptors, Cell Surface, MESH : Interleukin-3 Receptor alpha Subunit, MESH : Antineoplastic Agents, Phytogenic, MESH: Hematopoietic Stem Cells, Maleimides, MESH: Aged, 80 and over, MESH : Female, MESH: Maleimides, MESH: Receptors, Cell Surface, MESH: Aged, Leukemia, Myeloid leukemia, MESH : Adult, Middle Aged, medicine.anatomical_structure, MESH : Antigens, CD34, Female, Stem cell, Adult, MESH: GTP-Binding Proteins, Cell Survival, MESH : Male, MESH : Sex Factors, MESH: RNA Interference, Population, Interleukin-3 Receptor alpha Subunit, [SDV.CAN]Life Sciences [q-bio]/Cancer, MESH : Etoposide, Biology, Receptors for Activated C Kinase, Young Adult, MESH : Neoplastic Stem Cells, Sex Factors, GTP-Binding Proteins, MESH: Leukemia, MESH : Cells, Cultured, Cell Adhesion, medicine, 030304 developmental biology, Antineoplastic Agents, Phytogenic, MESH: Male, MESH : Cell Adhesion, Cancer research, Bone marrow, MESH : RNA Interference
Abstract

International audience; Therapeutic resistance of acute myeloid leukemia stem cells, enriched in the CD34(+)38(-)123(+) progenitor population, is supported by extrinsic factors such as the bone marrow niche. Here, we report that when adherent onto fibronectin or osteoblast components, CD34(+)38(-)123(+) progenitors survive through an integrin-dependent activation of glycogen synthase kinase 3β (GSK3β) by serine 9-dephosphorylation. Strikingly, GSK3β-mediated survival was restricted to leukemic progenitors from female patients. GSK3β inhibition restored sensitivity to etoposide, and impaired the clonogenic capacities of adherent leukemic progenitors from female patients. In leukemic progenitors from female but not male patients, the scaffolding protein RACK1, activated downstream of α(5)β(1)-integrin engagement, was specifically upregulated and controlled GSK3β activation through the phosphatase protein phosphatase 2A (PP2A). In a mirrored manner, survival of adherent progenitors (CD34(+)38(-)) from male but not female healthy donors was partially dependent on this pathway. We conclude that the GSK3β-dependent survival pathway might be sex-specific in normal immature population and flip-flopped upon leukemogenesis. Taken together, our results strengthen GSK3β as a promising target for leukemic stem cell therapy and reveal gender differences as a new parameter in anti-leukemia therapy.

Published
2011
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3. 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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4. A protocol to quantify mammary early common progenitors from long-term mammosphere culture

Author

Véronique Maguer-Satta, Emmanuel Delay, Elodie Bachelard-Cascales, Marion Chapellier, 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)-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), 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 ) -Université Claude Bernard Lyon 1 ( UCBL ), 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 )

Subjects
MESH: Neoplasm Proteins, Time Factors, Cellular differentiation, Cell Culture Techniques, MESH : Aged, MESH : Receptors, Cell Surface, MESH : Blotting, Western, MESH : Interleukin-3 Receptor alpha Subunit, MESH : Antineoplastic Agents, Phytogenic, [ SDV.CAN ] Life Sciences [q-bio]/Cancer, MESH: Hematopoietic Stem Cells, Mice, 0302 clinical medicine, MESH: Aged, 80 and over, MESH : Indoles, MESH : Tumor Cells, Cultured, MESH : Female, MESH : GTP-Binding Proteins, MESH: Glycogen Synthase Kinase 3, MESH: Maleimides, MESH: Receptors, Cell Surface, MESH: Etoposide, MESH : Maleimides, MESH: Aged, MESH: Indoles, MESH : Cell Survival, 0303 health sciences, MESH: Middle Aged, Stem Cells, MESH: Protein Phosphatase 2, Cell Differentiation, General Medicine, MESH : Adult, Cell biology, MESH: Cell Survival, MESH: Young Adult, 030220 oncology & carcinogenesis, MESH : Antigens, CD34, Stem cell, MESH: Cells, Cultured, Adult, MESH: GTP-Binding Proteins, MESH : Protein Phosphatase 2, MESH : Glycogen Synthase Kinase 3, MESH : Male, MESH : Sex Factors, MESH: RNA Interference, MESH: Antineoplastic Agents, Phytogenic, MESH : Young Adult, [SDV.CAN]Life Sciences [q-bio]/Cancer, MESH : Etoposide, Biology, MESH: Cell Adhesion, Colony-Forming Units Assay, 03 medical and health sciences, MESH : Neoplasm Proteins, MESH : Neoplastic Stem Cells, MESH: Sex Factors, Spheroids, Cellular, MESH : Cells, Cultured, MESH: Leukemia, Animals, Humans, MESH: Blotting, Western, MESH : Middle Aged, MESH: Tumor Cells, Cultured, Progenitor cell, Mammary Glands, Human, MESH : Aged, 80 and over, 030304 developmental biology, Progenitor, MESH: Interleukin-3 Receptor alpha Subunit, MESH: Humans, MESH : Hematopoietic Stem Cells, Stem cell population, MESH : Humans, MESH : Leukemia, MESH: Adult, Cell Biology, MESH: Antigens, CD34, MESH: Neoplastic Stem Cells, MESH: Male, MESH : Cell Adhesion, Cell culture, Immunology, NIH 3T3 Cells, MESH : RNA Interference, MESH: Female, Developmental Biology
Abstract

International audience; Here we describe a protocol established in our laboratory to quantify early common progenitors/stem cells grown as spheres in non-adherent culture conditions. This protocol is based on the combination of two functional tests: the mammosphere assay to maintain and enrich early mammary progenitors/stem cells, and the epithelial colony-forming cells (E-CFC) assay to identify and quantify further differentiated epithelial progenitors. Primary spheres mainly contain progenitors and rare stem/early common progenitor cells while secondary and tertiary spheres contain progenitor cells derived from the early common progenitor/stem cell population maintained through passages and partially differentiated. Spheres are enzymatically and mechanically dissociated; the derived cells are subsequently plated on irradiated NIH-3T3 fibroblasts for further processing, as in the E-CFC assay. The principle of this assay is to quantify the number of epithelial colonies generated by cells present in the different sequential spheres. This assay has therefore been named ECP-DC, standing for Early Common Progenitor-Derived Colonies assay.

Published
2012
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5. Myrip couples the capture of secretory granules by the actin-rich cell cortex and their attachment to the plasma membrane

Author

Tim Zeiske, François Darchen, Patricia Meireles, Sébastien Huet, Nathanael Larochette, Isabelle Fanget, Ouardane Jouannot, Claire Desnos, 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 ), Laboratoire de dynamique membranaire et maladies neurologiques (UMR 8192), Université Paris Descartes - Paris 5 (UPD5)-Centre National de la Recherche Scientifique (CNRS), 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 ), 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 ), Laboratoire de dynamique membranaire et maladies neurologiques ( UMR 8192 ), Centre National de la Recherche Scientifique ( CNRS ) -Université Paris Descartes - Paris 5 ( UPD5 ), and De Villemeur, Hervé

Subjects
MESH : Vesicular Transport Proteins, MESH: Vesicular Transport Proteins, MESH: Secretory Vesicles, Vesicular Transport Proteins, MESH : Actins, 0302 clinical medicine, Myosin, MESH : Secretory Vesicles, 0303 health sciences, MESH: Exocytosis, General Neuroscience, MESH : Protein Binding, Articles, Cell biology, Biochemistry, Protein Binding, MESH: Cell Line, Tumor, MESH : Cell Membrane, Exocyst, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Biology, MESH: Actins, MESH : Exocytosis, Exocytosis, MESH: Cell Adhesion, 03 medical and health sciences, stomatognathic system, MESH : Enterochromaffin Cells, Microtubule, Cell Line, Tumor, Cell cortex, Cell Adhesion, Enterochromaffin Cells, Molecular motor, Humans, MESH: Protein Binding, MESH: Enterochromaffin Cells, RAB27, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, Actin, 030304 developmental biology, MESH: Humans, MESH : Cell Line, Tumor, [ SDV.BC ] Life Sciences [q-bio]/Cellular Biology, Secretory Vesicles, MESH : Humans, Cell Membrane, Actins, MESH : Cell Adhesion, 030217 neurology & neurosurgery, MESH: Cell Membrane
Abstract

International audience; Exocytosis of secretory granules (SGs) requires their delivery to the actin-rich cell cortex followed by their attachment to the plasma membrane (PM). How these reactions are executed and coordinated is still unclear. Myrip, which is also known as Slac-2c, binds to the SG-associated GTPase Rab27 and is thought to promote the delivery of SGs to the PM by recruiting the molecular motor myosin Va. Myrip also interacts with actin and the exocyst complex, suggesting that it may exert multiple roles in the secretory process. By combining total internal reflection fluorescence microscopy, single-particle tracking, a photoconversion-based assay, and mathematical modeling, we show that, in human enterochromaffin cells, Myrip (1) inhibits a class of SG motion characterized by fast and directed movement, suggesting that it facilitates the dissociation of SGs from microtubules; (2) enhances their motion toward the PM and the probability of SG attachment to the PM; and (3) increases the characteristic time of immobilization at the PM, indicating that it is a component of the molecular machinery that tether SGs to the PM. Remarkably, while the first two effects of Myrip depend on its ability to recruit myosin Va on SGs, the third is myosin Va independent but relies on the C-terminal domain of Myrip. We conclude that Myrip couples the retention of SGs in the cell cortex, their transport to the PM, and their attachment to the PM, and thus promotes secretion. These three steps of the secretory process are thus intimately coordinated.

Published
2012
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6. 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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7. 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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8. What makes cells move: requirements and obstacles for spontaneous cell motility

Author

Urszula Hibner, Fabien Binamé, Géraldine Pawlak, Pierre Roux, Institut de Génétique Moléculaire de Montpellier ( IGMM ), Université de Montpellier ( UM ) -Centre National de la Recherche Scientifique ( CNRS ), 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), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), 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), Laboratoire de Géophysique Interne et Tectonophysique (LGIT), Observatoire des Sciences de l'Univers de Grenoble (OSUG), Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP)-Institut national de recherche en sciences et technologies pour l'environnement et l'agriculture (IRSTEA)-Université Joseph Fourier - Grenoble 1 (UJF)-Institut national des sciences de l'Univers (INSU - CNRS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP)-Institut national de recherche en sciences et technologies pour l'environnement et l'agriculture (IRSTEA)-Université Joseph Fourier - Grenoble 1 (UJF)-Institut national des sciences de l'Univers (INSU - CNRS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Laboratoire Central des Ponts et Chaussées (LCPC)-Institut des Sciences de la Terre (ISTerre), Université Grenoble Alpes (UGA)-Centre National de la Recherche Scientifique (CNRS)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-PRES Université de Grenoble-Institut de recherche pour le développement [IRD] : UR219-Institut national des sciences de l'Univers (INSU - CNRS)-Institut Français des Sciences et Technologies des Transports, de l'Aménagement et des Réseaux (IFSTTAR)-Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS)-PRES Université de Grenoble-Institut de recherche pour le développement [IRD] : UR219-Institut Français des Sciences et Technologies des Transports, de l'Aménagement et des Réseaux (IFSTTAR), Association pour la recherche sur le cancer, La Ligue contre le cancer, Agence Nationale de Recherche sur le Sida et les hépatites virales et Institut National du Cancer, Centre National de la Recherche Scientifique ( CNRS ) -Université de Montpellier ( UM ), Laboratoire de Géophysique Interne et Tectonophysique ( LGIT ), Observatoire des Sciences de l'Univers de Grenoble ( OSUG ), Université Joseph Fourier - Grenoble 1 ( UJF ) -Institut national des sciences de l'Univers ( INSU - CNRS ) -Centre National de la Recherche Scientifique ( CNRS ) -Université Grenoble Alpes ( UGA ) -Université Joseph Fourier - Grenoble 1 ( UJF ) -Institut national des sciences de l'Univers ( INSU - CNRS ) -Centre National de la Recherche Scientifique ( CNRS ) -Université Grenoble Alpes ( UGA ) -Laboratoire Central des Ponts et Chaussées ( LCPC ) -Institut des Sciences de la Terre ( ISTerre ), Université Grenoble Alpes ( UGA ) -Centre National de la Recherche Scientifique ( CNRS ) -Université Savoie Mont Blanc ( USMB [Université de Savoie] [Université de Chambéry] ) -PRES Université de Grenoble-Institut de recherche pour le développement [IRD] : UR219-Institut national des sciences de l'Univers ( INSU - CNRS ) -Institut Français des Sciences et Technologies des Transports, de l'Aménagement et des Réseaux ( IFSTTAR ) -Université Joseph Fourier - Grenoble 1 ( UJF ) -Centre National de la Recherche Scientifique ( CNRS ) -Université Savoie Mont Blanc ( USMB [Université de Savoie] [Université de Chambéry] ) -PRES Université de Grenoble-Institut de recherche pour le développement [IRD] : UR219-Institut Français des Sciences et Technologies des Transports, de l'Aménagement et des Réseaux ( IFSTTAR ), 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: Signal Transduction, MESH : Hepatocytes, MESH : Models, Biological, Mechanotransduction, Cellular, MESH : Mechanotransduction, Cellular, MESH: Hepatocytes, 0302 clinical medicine, Cell Movement, MESH : Cell Movement, MESH: Animals, Mechanotransduction, MESH: Cell Movement, 0303 health sciences, Amoeboid movement, Systems Biology, Cell Polarity, Cell migration, Molecular machine, Cell biology, 030220 oncology & carcinogenesis, MESH: Systems Biology, MESH : Imaging, Three-Dimensional, MESH: Cell Polarity, MESH : Cell Polarity, Biotechnology, Signal Transduction, Systems biology, Context (language use), MESH: Imaging, Three-Dimensional, Biology, In Vitro Techniques, Models, Biological, MESH: Cell Adhesion, 03 medical and health sciences, Imaging, Three-Dimensional, Cell Adhesion, Animals, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, [SDU.STU.GM]Sciences of the Universe [physics]/Earth Sciences/Geomorphology, Molecular Biology, [ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology, 030304 developmental biology, MESH : Systems Biology, MESH : Signal Transduction, MESH: Humans, [ SDU.STU.GM ] Sciences of the Universe [physics]/Earth Sciences/Geomorphology, MESH: Mechanotransduction, Cellular, Mesenchymal stem cell, MESH : Humans, MESH: Models, Biological, Multicellular organism, MESH : Cell Adhesion, Hepatocytes, MESH : Animals
Abstract

International audience; Movement of individual cells and of cellular cohorts, chains or sheets requires physical forces that are established through interactions of cells with their environment. In vivo, migration occurs extensively during embryonic development and in adults during wound healing and tumorigenesis. In order to identify the molecular events involved in cell movement, in vitro systems have been developed. These have contributed to the definition of a number of molecular pathways put into play in the course of migratory behaviours, such as mesenchymal and amoeboid movement. More recently, our knowledge of migratory modes has been enriched by analyses of cells exploring and moving through three-dimensional (3D) matrices. While the cells' morphologies differ in 2D and 3D environments, the basic mechanisms that put a cellular body into motion are remarkably similar. Thus, in both 2D and 3D, the polarity of the migrating cell is initially defined by a specific subcellular localization of signalling molecules and components of molecular machines required for motion. While the polarization can be initiated either in response to extracellular signalling or be a chance occurrence, it is reinforced and sustained by positive feedback loops of signalling molecules. Second, adhesion to a substratum is necessary to generate forces that will propel the cell engaged in either mesenchymal or ameboid migration. For collective cell movement, intercellular coordination constitutes an additional requirement: a cell cohort remains stationary if individual cells pull in opposite directions. Finally, the availability of space to move into is a general requirement to set cells into motion. Lack of free space is probably the main obstacle for migration of most healthy cells in an adult multicellular organism. Thus, the requirements for cell movement are both intrinsic to the cell, involving coordinated signalling and interactions with molecular machines, and extrinsic, imposed by the physicochemical nature of the environment. In particular, the geometry and stiffness of the support act on a range of signalling pathways that induce specific cell migratory responses. These issues are discussed in the present review in the context of published work and our own data on collective migration of hepatocyte cohorts.

Published
2010
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9. Foxp3+ T cells induce perforin-dependent dendritic cell death in tumor-draining lymph nodes

Author

Tim Sparwasser, Alexandre Boissonnas, Luc Fetler, Alix Scholer-Dahirel, Bernard Malissen, Adrien Kissenpfennig, Luigia Pace, Fabien Valet, Virginie Simon-Blancal, Sebastian Amigorena, Immunité et cancer (U932), Université Paris Descartes - Paris 5 (UPD5)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Cancer et génome: Bioinformatique, biostatistiques et épidémiologie d'un système complexe, Institut Curie [Paris]-MINES ParisTech - École nationale supérieure des mines de 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 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), Hannover Medical School [Hannover] (MHH), Physico-Chimie-Curie (PCC), Centre National de la Recherche Scientifique (CNRS)-Institut Curie [Paris]-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de Chimie du CNRS (INC), Université Paris Descartes - Paris 5 (UPD5)-Institut Curie-Institut National de la Santé et de la Recherche Médicale (INSERM), Cancer et génôme: Bioinformatique, biostatistiques et épidémiologie d'un système complexe, MINES ParisTech - École nationale supérieure des mines de Paris-Institut Curie-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre National de la Recherche Scientifique (CNRS)-Institut Curie-Université Pierre et Marie Curie - Paris 6 (UPMC), Mines Paris - PSL (École nationale supérieure des mines de Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut Curie [Paris]-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Immunité et cancer ( U932 ), Université Paris Descartes - Paris 5 ( UPD5 ) -Institut Curie-Institut National de la Santé et de la Recherche Médicale ( INSERM ), MINES ParisTech - École nationale supérieure des mines de Paris-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -INSTITUT CURIE, 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 ), Hannover Medical School [Hannover] ( MHH ), Physico-Chimie-Curie ( PCC ), and Centre National de la Recherche Scientifique ( CNRS ) -INSTITUT CURIE-Université Pierre et Marie Curie - Paris 6 ( UPMC )

Subjects
MESH: Cell Death, MESH : Fibrosarcoma, Fibrosarcoma, Priming (immunology), MESH: Lymph Nodes, MESH: T-Lymphocyte Subsets, Lymphocyte Activation, T-Lymphocytes, Regulatory, MESH: Mice, Knockout, MESH : T-Lymphocytes, Regulatory, Interleukin 21, Mice, 0302 clinical medicine, Cell Movement, T-Lymphocyte Subsets, MESH : Cell Movement, [ SDV.IMM ] Life Sciences [q-bio]/Immunology, Immunology and Allergy, Cytotoxic T cell, MESH : Female, MESH: Animals, MOLIMMUNO, MESH: Cell Movement, MESH : Lymph Nodes, Mice, Knockout, 0303 health sciences, Cell Death, MESH: Dendritic Cells, MESH: Fibrosarcoma, Forkhead Transcription Factors, hemic and immune systems, MESH : Mice, Transgenic, 3. Good health, MESH : Forkhead Transcription Factors, Infectious Diseases, medicine.anatomical_structure, [SDV.IMM]Life Sciences [q-bio]/Immunology, Female, MESH: Cell Line, Tumor, MESH: Mice, Transgenic, T cell, Immunology, Mice, Transgenic, chemical and pharmacologic phenomena, MESH : Mice, Inbred C57BL, Biology, TREG CELLS, MESH: Cell Adhesion, 03 medical and health sciences, MESH: Mice, Inbred C57BL, MESH: Forkhead Transcription Factors, Cell Line, Tumor, MESH : Mice, medicine, Lymph node stromal cell, Cell Adhesion, MESH : Neoplasm Transplantation, Animals, MESH : Perforin, Antigen-presenting cell, MESH: Lymphocyte Activation, MESH: Mice, MESH : Lymphocyte Activation, 030304 developmental biology, MESH : Cell Line, Tumor, Perforin, MESH: T-Lymphocytes, Regulatory, Dendritic cell, Dendritic Cells, MESH : T-Lymphocyte Subsets, MESH: Perforin, Mice, Inbred C57BL, MESH : Cell Adhesion, CELLIMMUNO, MESH : Cell Death, MESH : Dendritic Cells, Cancer research, MESH : Mice, Knockout, MESH : Animals, Lymph Nodes, MESH: Female, CD8, Neoplasm Transplantation, MESH: Neoplasm Transplantation, 030215 immunology
Abstract

International audience; Regulatory T (Treg) cells limit the onset of effective antitumor immunity, through yet-ill-defined mechanisms. We showed the rejection of established ovalbumin (OVA)-expressing MCA101 tumors required both the adoptive transfer of OVA-specific CD8(+) T cell receptor transgenic T cells (OTI) and the neutralization of Foxp3(+) T cells. In tumor-draining lymph nodes, Foxp3(+) T cell neutralization induced a marked arrest in the migration of OTI T cells, increased numbers of dendritic cells (DCs), and enhanced OTI T cell priming. Using an in vitro cytotoxic assay and two-photon live microscopy after adoptive transfer of DCs, we demonstrated that Foxp3(+) T cells induced the death of DCs in tumor-draining lymph nodes, but not in the absence of tumor. DC death correlated with Foxp3(+) T cell-DC contacts, and it was tumor-antigen and perforin dependent. We conclude that Foxp3(+) T cell-dependent DC death in tumor-draining lymph nodes limits the onset of CD8(+) T cell responses.

Published
2010
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10. Autophosphorylation-independent and -dependent functions of focal adhesion kinase during development

Author

Corsi, Jean-Marc, Houbron, Christophe, Billuart, Pierre, Brunet, Isabelle, Bouvrée, Karine, Eichmann, Anne, Girault, Jean-Antoine, Enslen, Hervé, Institut du Fer à Moulin, Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Cochin (UMR_S567 / UMR 8104), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Angiogénèse embryonnaire et pathologique, 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)-Labex MemoLife, Université Paris sciences et lettres (PSL)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Pierre et Marie Curie - Paris 6 (UPMC), ARC 3746, Université Pierre et Marie Curie - Paris 6 ( UPMC ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Institut Cochin ( UMR_S567 / UMR 8104 ), 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 ) -Collège de France ( CdF ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Université Pierre et Marie Curie - Paris 6 (UPMC)-Centre interdisciplinaire de recherche en biologie (CIRB), École normale supérieure - Paris (ENS-PSL), Université Paris sciences et lettres (PSL)-École normale supérieure - Paris (ENS-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)-Labex MemoLife, and Université Paris sciences et lettres (PSL)-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)

Subjects
MESH: Mutation, MESH: Mice, Transgenic, MESH : Male, proliferation, MESH : Focal Adhesion Protein-Tyrosine Kinases, MESH : Mice, Inbred C57BL, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, MESH: Cell Adhesion, MESH : Phosphorylation, vascularization, MESH: Mice, Inbred C57BL, MESH : Mice, MESH : Embryo, Mammalian, MESH: Animals, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, [ SDV.BDD ] Life Sciences [q-bio]/Development Biology, [ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology, development, MESH: Mice, [SDV.BDD]Life Sciences [q-bio]/Development Biology, MESH: Focal Adhesion Protein-Tyrosine Kinases, MESH: Phosphorylation, [ SDV.BC ] Life Sciences [q-bio]/Cellular Biology, MESH: Biological Markers, MESH: Embryo, Mammalian, tyrosine kinase, MESH: Integrins, MESH : Mice, Transgenic, MESH: Male, MESH : Biological Markers, MESH : Cell Adhesion, Adhesion, MESH : Animals, MESH : Mutation, MESH : Integrins
Abstract

International audience; Focal adhesion kinase (FAK) regulates numerous cellular functions and is critical for processes ranging from embryo development to cancer progression. Although autophosphorylation on Tyr-397 appears required for FAK functions in vitro, its role in vivo has not been established. We addressed this question using a mutant mouse (fakDelta) deleted of exon 15, which encodes Tyr-397. The resulting mutant protein FAKDelta is an active kinase expressed at normal levels. Our results demonstrate that the requirement for FAK autophosphorylation varies during development. FAK(Delta/Delta) embryos developed normally up to embryonic day (E) 12.5, contrasting with the lethality at E8.5 of FAK-null embryos. Thus, autophosphorylation on Tyr-397 is not required for FAK to achieve its functions until late mid-gestation. However, FAK(Delta/Delta) embryos displayed hemorrhages, edema, delayed artery formation, vascular remodeling defects, multiple organ abnormalities, and overall developmental retardation at E13.5-14.5, and died thereafter demonstrating that FAK autophosphorylation is also necessary for normal development. Fibroblasts derived from mutant embryos had a normal stellate morphology and expression of focal adhesion proteins, Src family members, p53, and Pyk2. In contrast, in FAK(Delta/Delta) fibroblasts and endothelial cells, spreading and lamellipodia formation were altered with an increased size and number of focal adhesions, enriched in FAKDelta. FAK mutation also decreased fibroblast proliferation. These results show that the physiological functions of FAK in vivo are achieved through both autophosphorylation-independent and autophosphorylation-dependent mechanisms.

Published
2009
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11. Erk5 controls Slug expression and keratinocyte activation during wound healing

Author

Pierre Savagner, Valérie Arnoux, Annie L'Helgoualc'h, Mayssaa Nassour, Robert A. Hipskind, Genotypes et Phenotypes Tumoraux, Université Montpellier 1 (UM1)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut de Génétique Moléculaire de Montpellier (IGMM), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Ligue Regionale contre le Cancer (Languedoc-Roussillon), Groupement des Entreprises Francaises dans la Lutte contre le Cancer, the Fondation de France and the Association pour la Recherche sur le Cancer (VA and RAH)., Université Montpellier 1 ( UM1 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Institut de Génétique Moléculaire de Montpellier ( IGMM ), Université de Montpellier ( UM ) -Centre National de la Recherche Scientifique ( CNRS ), Le Ster, Yves, and Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)

Subjects
[SDV.MHEP.PHY] Life Sciences [q-bio]/Human health and pathology/Tissues and Organs [q-bio.TO], MESH : Wound Healing, MESH : Promoter Regions, Genetic, MESH : RNA, Small Interfering, migration, [ SDV.CAN ] Life Sciences [q-bio]/Cancer, Mice, MESH : Phosphorylation, MESH : Cytoskeleton, 0302 clinical medicine, Desmosome organization, MESH: Cricetulus, MESH : CHO Cells, MESH: Animals, Cytoskeleton, Mitogen-Activated Protein Kinase 1, 0303 health sciences, Mitogen-Activated Protein Kinase 3, MESH : Gene Expression Regulation, MESH : Cricetinae, Keratinocyte activation, MESH: Transcription Factors, Cell biology, ErbB Receptors, 030220 oncology & carcinogenesis, MESH: Mitogen-Activated Protein Kinase 7, MESH: Mitogen-Activated Protein Kinase 3, MESH: Mitogen-Activated Protein Kinase 1, Slug, MESH : Cricetulus, MESH: Receptor, Epidermal Growth Factor, MESH: Cell Adhesion, 03 medical and health sciences, [SDV.CAN] Life Sciences [q-bio]/Cancer, MESH: CHO Cells, MESH: Promoter Regions, Genetic, Humans, Cell adhesion, Molecular Biology, [ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology, EGF, MESH: Humans, Epidermal Growth Factor, MESH: Phosphorylation, [ SDV.BC ] Life Sciences [q-bio]/Cellular Biology, HaCaT, MESH : Humans, MAPK, MESH : Epithelium, MESH: Wound Healing, MESH: Epithelium, Wound healing, [ SDV.MHEP.DERM ] Life Sciences [q-bio]/Human health and pathology/Dermatology, [SDV.MHEP.DERM]Life Sciences [q-bio]/Human health and pathology/Dermatology, Transcription Factors, Keratinocytes, MESH: Epidermal Growth Factor, Cytoskeleton organization, MESH : Transcription Factors, MESH: Cricetinae, Epithelium, MESH : Receptor, Epidermal Growth Factor, Cricetinae, MESH: RNA, Small Interfering, Phosphorylation, RNA, Small Interfering, Keratinocyte migration, Promoter Regions, Genetic, MESH : Epidermal Growth Factor, MESH : Mitogen-Activated Protein Kinase 1, biology, MESH : Mitogen-Activated Protein Kinase 3, ERK1/2, MESH : Keratinocytes, [ SDV.MHEP.PHY ] Life Sciences [q-bio]/Human health and pathology/Tissues and Organs [q-bio.TO], Articles, MESH: Gene Expression Regulation, MESH: Keratinocytes, MESH : Mitogen-Activated Protein Kinase 7, Motility, [SDV.CAN]Life Sciences [q-bio]/Cancer, CHO Cells, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Cricetulus, MESH : Mice, Cell Adhesion, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH: Cytoskeleton, [SDV.MHEP.PHY]Life Sciences [q-bio]/Human health and pathology/Tissues and Organs [q-bio.TO], Animals, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, MESH: Mice, Mitogen-Activated Protein Kinase 7, 030304 developmental biology, Wound Healing, Cell Biology, [SDV.MHEP.DERM] Life Sciences [q-bio]/Human health and pathology/Dermatology, biology.organism_classification, MESH : Cell Adhesion, Gene Expression Regulation, Cancer research, Snail Family Transcription Factors, MESH : Animals
Abstract

International audience; Reepithelialization during cutaneous wound healing involves numerous signals that result in basal keratinocyte activation, spreading, and migration, all linked to a loosening of cell-cell adhesion structures. The transcription factor Slug is required for this process, and EGF treatment of human keratinocytes induced activating phosphorylation of Erk5 that coincides with slug transcription. Accordingly, ectopic activation of Erk5 led to increased Slug mRNA levels and faster wound healing, whereas keratinocyte migration was totally blocked by Erk5 pathway inhibition. Expression of a shRNA specific for Erk5 strongly diminished Erk5 levels in keratinocytes and significantly decreased their motility response to EGF, along with induction of Slug expression. These Erk5-deprived keratinocytes showed an altered, more compact morphology, along with disruption of desmosome organization. Accordingly, they displayed an altered ability to form cell aggregates. These results implicate a novel EGFR/Erk5/Slug pathway in the control of cytoskeleton organization and cell motility in keratinocytes treated with EGF.

Published
2008
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12. Transforming growth factor beta controls the directional migration of hepatocyte cohorts by modulating their adhesion to fibronectin

Author

Urszula Hibner, Fabien Binamé, Patrice Lassus, Institut de Génétique Moléculaire de Montpellier ( IGMM ), Université de Montpellier ( UM ) -Centre National de la Recherche Scientifique ( CNRS ), Institut de Génétique Moléculaire de Montpellier (IGMM), and Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)

Subjects
rac1 GTP-Binding Protein, MESH : Cell Line, MESH : Protein Subunits, Apoptosis, Cell Communication, Integrin alpha5, MESH : Hepatocytes, Epithelium, Mesoderm, MESH: Hepatocytes, Mice, 0302 clinical medicine, Cell Movement, Transforming Growth Factor beta, MESH: Fibronectins, MESH : Cell Movement, MESH : Fibronectins, MESH: Animals, MESH: Cell Movement, MESH: Mesoderm, 0303 health sciences, MESH : Gene Expression Regulation, MESH: Cell Surface Extensions, MESH : Mesoderm, Articles, Adhesion, MESH: Protein Subunits, MESH: Gene Expression Regulation, Cell biology, medicine.anatomical_structure, 030220 oncology & carcinogenesis, Hepatocyte, MESH: Integrin alpha5, MESH : Cell Communication, MESH : Cell Membrane, MESH : Cell Surface Extensions, Biology, Cell Line, MESH: Cell Adhesion, MESH: Gene Expression Profiling, 03 medical and health sciences, MESH : rac1 GTP-Binding Protein, MESH : Integrin alpha5, MESH: Cell Communication, MESH : Mice, Cell Adhesion, medicine, Animals, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH: Mice, Molecular Biology, [ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH: Transforming Growth Factor beta, 030304 developmental biology, MESH: Humans, MESH: rac1 GTP-Binding Protein, Gene Expression Profiling, MESH: Apoptosis, MESH : Gene Expression Profiling, Cell Membrane, MESH : Humans, Cell Biology, MESH : Epithelium, Fibronectins, MESH: Cell Line, Fibronectin, Protein Subunits, MESH: Epithelium, MESH : Cell Adhesion, MESH : Transforming Growth Factor beta, Gene Expression Regulation, Hepatocytes, biology.protein, Cell Surface Extensions, MESH : Animals, MESH : Apoptosis, MESH: Cell Membrane, Transforming growth factor
Abstract

International audience; Transforming growth factor beta (TGF-beta) has a strong impact on liver development and physiopathology, exercised through its pleiotropic effects on growth, differentiation, survival, and migration. When exposed to TGF-beta, the mhAT3F cells, immortalized, highly differentiated hepatocytes, maintained their epithelial morphology and underwent dramatic alterations of adhesion, leading to partial or complete detachment from a culture plate, followed by readhesion and spreading. These alterations of adhesive behavior were caused by sequential changes in expression of the alpha5beta1 integrin and of its ligand, the fibronectin. The altered specificity of anchorage to the extracellular matrix gave rise to changes in cells' collective motility: cohorts adhering to fibronectin maintained a persistent, directional motility, with ezrin-rich pathfinder cells protruding from the tips of the cohorts. The absence of adhesion to fibronectin prevented the appearance of polarized pathfinders and lead to random, oscillatory motility. Our data suggest a novel role for TGF-beta in the control of collective migration of epithelial cohorts.

Published
2008
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13. Extracellular matrix-bound angiopoietin-like 4 inhibits endothelial cell adhesion, migration, and sprouting and alters actin cytoskeleton

Author

Nicolas Bréchot, Ariane Galaup, Clémence Chomel, Laurent Muller, Marine Bignon, Stéphane Germain, Catherine Monnot, Pierre Corvol, Aurélie Cazes, Holger Weber, Sébastien Le Jan, 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), Departement of Vascular Biology and Angiogenesis Research, Tumor Biology Center, Centre interdisciplinaire de recherche en biologie (CIRB), Labex MemoLife, École normale supérieure - Paris (ENS-PSL), 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-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), Mouret, Sandrine, École normale supérieure - Paris (ENS 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), Pathologie vasculaire et endocrinologie rénale, Collège de France ( CdF ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Service d'hématologie A [CHU HEGP], and Assistance publique - Hôpitaux de Paris (AP-HP)-Hôpital Européen Georges Pompidou [APHP] ( HEGP )

Subjects
MESH: Heparin, MESH : Biological Availability, Physiology, Angiogenesis, MESH: Anoxia, MESH : Actins, Extracellular matrix, MESH : Cytoskeleton, Mice, 0302 clinical medicine, Cell Movement, Ischemia, MESH: Blood Proteins, MESH : Cell Movement, MESH: Animals, MESH: Endothelial Cells, Hypoxia, Cytoskeleton, MESH: Cell Movement, Cells, Cultured, MESH: Biological Availability, Tube formation, 0303 health sciences, MESH : Neovascularization, Physiologic, MESH: Hindlimb, MESH : Extracellular Matrix, Blood Proteins, [ SDV.MHEP.CSC ] Life Sciences [q-bio]/Human health and pathology/Cardiology and cardiovascular system, MESH : Proteoglycans, Extracellular Matrix, Hindlimb, Cell biology, [SDV.MHEP.CSC] Life Sciences [q-bio]/Human health and pathology/Cardiology and cardiovascular system, Endothelial stem cell, Biochemistry, 030220 oncology & carcinogenesis, MESH: Proteoglycans, Proteoglycans, Cardiology and Cardiovascular Medicine, MESH: Neovascularization, Physiologic, MESH: Cells, Cultured, MESH : Blood Proteins, MESH : Anoxia, Biological Availability, Neovascularization, Physiologic, MESH : Mice, Inbred C57BL, Angiopoietin-like 4 Protein, In Vitro Techniques, Biology, MESH: Extracellular Matrix, MESH: Actins, Article, MESH: Cell Adhesion, Angiopoietin, 03 medical and health sciences, [SDV.MHEP.CSC]Life Sciences [q-bio]/Human health and pathology/Cardiology and cardiovascular system, MESH : Ischemia, MESH: Mice, Inbred C57BL, MESH : Mice, MESH : Cells, Cultured, Cell Adhesion, MESH: Cytoskeleton, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, Angiopoietin-Like Protein 4, Animals, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, [ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH: Mice, 030304 developmental biology, MESH : Hindlimb, MESH : Endothelial Cells, Heparin, Endothelial Cells, Actin cytoskeleton, Actins, Mice, Inbred C57BL, MESH : Cell Adhesion, MESH : Heparin, MESH : Animals, MESH: Ischemia, Angiopoietins
Abstract

Angiopoietin-like 4 (ANGPTL4) is a secreted protein that belongs to the angiopoietin family and is involved in angiogenesis and metabolism regulation. We previously reported the induction of angptl4 by hypoxia in endothelial cells and in human ischemic tissues from peripheral artery disease. We here observed in a mouse model of hindlimb ischemia that the mRNA upregulation in the vessels correlates with the accumulation of the full-length protein in ischemic tissues. We then investigated its functions in endothelial cells. In response to hypoxia, endogenous ANGPTL4 accumulates in the subendothelial extracellular matrix (ECM). Although the secreted protein undergoes proteolysis leading to truncated fragments present in the medium, only full-length ANGPTL4 interacts with the ECM. Competition and direct binding assays indicate that the strong interaction of ANGPTL4 with the ECM is heparin/heparan sulfate proteoglycan dependent. The balance between matrix-associated and soluble forms of ANGPTL4 points to the role of the ECM in the regulation of its bioavailability. The angiogenic function of the ECM-bound full-length protein was investigated using either the form associated with the conditioned ECM from ANGPTL4-transfected HEK293 cells or the purified immobilized protein. We show that matrix-associated and immobilized ANGPTL4 limit the formation of actin stress fibers and focal contacts in the adhering endothelial cells and inhibit their adhesion. Immobilized ANGPTL4 also decreases motility of endothelial cells and inhibits the sprouting and tube formation. Altogether, these findings show that hypoxic endothelial cells accumulate ANGPTL4 in the ECM, which in turn negatively regulates their angiogenic capacities through an autocrine pathway.

Published
2006
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14. ARHGAP10 is necessary for alpha-catenin recruitment at adherens junctions and for Listeria invasion

Author

Sandra Maria Zákia Lian Sousa, Didier Cabanes, Cristel Archambaud, Stéphanie Boisson-Dupuis, Marc Lecuit, Michel R. Popoff, Emmanuel Lemichez, Pascale Cossart, Edith Gouin, Pierre Legrain, Frédéric Colland, Interactions Bactéries-Cellules (UIBC), Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur [Paris]-Institut National de la Recherche Agronomique (INRA), Molecular Microbiology, Institute for Molecular and Cell Biology, Hybrigenics [Paris], Hybrigenics, Toxines bactériennes dans la relation hôtes-pathogènes, Université Nice Sophia Antipolis (... - 2019) (UNS), Université Côte d'Azur (UCA)-Université Côte d'Azur (UCA)-IFR50-Institut National de la Santé et de la Recherche Médicale (INSERM), Bactéries anaérobies et Toxines, Institut Pasteur [Paris], Centre d'infectiologie Necker-Pasteur [CHU Necker], CHU Necker - Enfants Malades [AP-HP], Service Hospitalier Frédéric Joliot (SHFJ), Université Paris-Saclay-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), Institut National de la Recherche Agronomique (INRA)-Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Nice Sophia Antipolis (1965 - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-IFR50-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Côte d'Azur (UCA), Institut Pasteur [Paris] (IP), 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), Lemichez, Emmanuel, Institut National de la Recherche Agronomique (INRA)-Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut National de la Santé et de la Recherche Médicale (INSERM)-IFR50-Université Côte d'Azur (UCA)-Université Nice Sophia Antipolis (... - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA), Institut Pasteur [Paris]-CHU Necker - Enfants Malades [AP-HP], Interactions Bactéries-Cellules ( UIBC ), Institut National de la Recherche Agronomique ( INRA ) -Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale ( INSERM ), Université Nice Sophia Antipolis ( UNS ), Université Côte d'Azur ( UCA ) -Université Côte d'Azur ( UCA ) -IFR50-Institut National de la Santé et de la Recherche Médicale ( INSERM ), CHU Necker - Enfants Malades [AP-HP]-Assistance publique - Hôpitaux de Paris (AP-HP), Département de Biologie Joliot-Curie, and Commissariat à l'énergie atomique et aux énergies alternatives ( CEA )

Subjects
MESH : Cell Line, RHOA, MESH : Hela Cells, MESH : Actins, Arp2/3 complex, MESH: GTPase-Activating Proteins, MESH: Listeria monocytogenes, Ligands, Cell junction, Cell membrane, [ SDV.MP ] Life Sciences [q-bio]/Microbiology and Parasitology, MESH: Ligands, MESH : Bacterial Proteins, MESH: Animals, MESH: Bacterial Proteins, MESH : Ligands, 0303 health sciences, biology, 030302 biochemistry & molecular biology, GTPase-Activating Proteins, MESH: Research Support, N, Adherens Junctions, Cell biology, medicine.anatomical_structure, [SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology, Intercellular Junctions, Ectodomain, MESH : Intercellular Junctions, RNA Interference, MESH: Caco-2 Cells, MESH : Cell Membrane, TWO-HYBRID METHOD, MESH: RNA Interference, ARHGAP10, Alpha catenin, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, MESH: Actins, ACTIN, MESH: Cell Adhesion, Cell Line, Adherens junction, 03 medical and health sciences, Bacterial Proteins, Two-Hybrid System Techniques, medicine, Cell Adhesion, Animals, Humans, [SDV.MP] Life Sciences [q-bio]/Microbiology and Parasitology, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, ADHERENS JUNCTION, 030304 developmental biology, MESH: Humans, [ SDV.BC ] Life Sciences [q-bio]/Cellular Biology, MESH : Humans, Cell Membrane, MESH : Caco-2 Cells, MESH : GTPase-Activating Proteins, RELATION HOTE-BACTERIE, Cell Biology, MESH : Adherens Junctions, Actin cytoskeleton, Listeria monocytogenes, Actins, MESH: Cell Line, MESH: Hela Cells, MESH : Cell Adhesion, MESH: Adherens Junctions, MICROBIAL INFECTION STRATEGY, biology.protein, MESH : Listeria monocytogenes, MESH : Animals, MESH : RNA Interference, LIGAND, Caco-2 Cells, rhoA GTP-Binding Protein, MESH : Research Support, N, alpha Catenin, MESH: Intercellular Junctions, MESH: Cell Membrane, HeLa Cells
Abstract

E-cadherin mediates the formation of adherens junctions between epithelial cells. It serves as a receptor for Listeria monocytogenes, a bacterial pathogen that enters epithelial cells. The L. monocytogenes surface protein, InlA, interacts with the extracellular domain of E-cadherin. In adherens junctions, this ectodomain is involved in homophilic interactions whereas the cytoplasmic domain binds beta-catenin, which then recruits alpha-catenin. alpha-catenin binds to actin directly, or indirectly, thus linking E-cadherin to the actin cytoskeleton. Entry of L. monocytogenes into cells and adherens junction formation are dynamic events that involve actin and membrane rearrangements. To understand these processes better, we searched for new ligands of alpha-catenin. Using a two-hybrid screen, we identified a new partner of alpha-catenin: ARHGAP10. This protein colocalized with alpha-catenin at cell-cell junctions and was recruited at L. monocytogenes entry sites. In ARHGAP10-knockdown cells, L. monocytogenes entry and alpha-catenin recruitment at cell-cell contacts were impaired. The GAP domain of ARHGAP10 has GAP activity for RhoA and Cdc42. Its overexpression disrupted actin cables, enhanced alpha-catenin and cortical actin levels at cell-cell junctions and inhibited L. monocytogenes entry. Altogether, our results show that ARHGAP10 is a new component of cell-cell junctions that controls alpha-catenin recruitment and has a key role during L. monocytogenes uptake.

Published
2005
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15. Chronology of cellular alterations during 7-ketocholesterol-induced cell death on A7R5 rat smooth muscle cells: analysis by time lapse-video microscopy and conventional fluorescence microscopy

Author

Zahm, Jean-Marie, Baconnais, Sonia, Monier, Serge, Bonnet, Noël, Bessède, Ginette, Gambert, Philippe, Puchelle, Edith, Lizard, Gérard, Dynamique cellulaire et moléculaire de la muqueuse respiratoire, Université de Reims Champagne-Ardenne (URCA)-IFR53-Institut National de la Santé et de la Recherche Médicale (INSERM), Laboratoire des Lipoprotéines humaines et interactions vasculaires, Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM), Université de Reims Champagne-Ardenne ( URCA ) -IFR53-Institut National de la Santé et de la Recherche Médicale ( INSERM ), Université de Bourgogne ( UB ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), and Birembaut, Philippe

Subjects
MESH: Cell Death, Time Factors, MESH : Microscopy, Fluorescence, MESH : Actins, Apoptosis, Cell Count, MESH: Microscopy, Fluorescence, MESH : Cell Count, [SDV.MHEP.PSR]Life Sciences [q-bio]/Human health and pathology/Pulmonology and respiratory tract, Muscle, Smooth, Vascular, Membrane Potentials, MESH : Image Cytometry, MESH : Rat, MESH : Membrane Potentials, MESH: Animals, MESH : DNA Fragmentation, Enzyme Inhibitors, Ketocholesterols, Image Cytometry, Microscopy, Video, Cell Death, MESH : Rats, MESH : Microscopy, Video, MESH: Ketocholesterols, MESH: Muscle, Smooth, Vascular, Mitochondria, MESH: Enzyme Inhibitors, MESH : Mitochondria, MESH: Rats, MESH: Mitochondria, DNA Fragmentation, MESH: Actins, MESH: Cell Adhesion, MESH: Rat, Cell Adhesion, Animals, MESH: Membrane Potentials, MESH: DNA Fragmentation, MESH : Ketocholesterols, MESH : Enzyme Inhibitors, MESH: Cell Count, MESH: Apoptosis, Rats, Inbred Strains, MESH : Muscle, Smooth, Vascular, Actins, Rats, MESH: Microscopy, Video, MESH : Cell Adhesion, Microscopy, Fluorescence, MESH : Cell Death, [SDV.MHEP.PSR] Life Sciences [q-bio]/Human health and pathology/Pulmonology and respiratory tract, MESH : Animals, MESH : Apoptosis, [ SDV.MHEP.PSR ] Life Sciences [q-bio]/Human health and pathology/Pulmonology and respiratory tract, MESH: Image Cytometry
Abstract

BACKGROUND: Time-lapse video microscopy was used to determine whether mitochondrial and nuclear changes (decrease in mitochondrial transmembrane potential, condensation, and/or fragmentation of the nuclei, morphologic features typical of apoptosis) occurring during 7-ketocholesterol-induced cell death on A7R5 rat smooth muscle cells took place before or after the loss of cell adhesion. In addition, changes in actin organization were followed by conventional fluorescence microscopy. METHODS: Morphologic, functional, and spatial changes at the mitochondrial level were investigated with 3,3'-dihexyloxacarbocyanine iodide and/or MitoTracker Red, and nuclear morphology was characterized by staining with Hoechst 33342. Actin fibers, which are major components of the filament network of the cytoskeleton, were visualized with phalloidin linked to fluorescein. The numbers of adherent and nonadherent cells were determined by cell counting. RESULTS: 7-Ketocholesterol-induced cell death was associated with a rapid alteration of actin fibers, a loss of intercellular junctions, and cell shape modifications. Analysis of mitochondrial transmembrane potential showed successively a hyperpolarization and a more or less pronounced progressive decrease followed by a dramatic drop associated with an increase in Hoechst 33342 staining, reflecting chromatin condensation and morphologic changes in the nuclei. CONCLUSIONS: During cell death induced by 7-ketocholesterol in A7R5 rat smooth muscle cells, the different methods of microscopy allowed us to establish that alterations of actin fibers and mitochondrial dysfunctions occurred before condensation and/or fragmentation of the nuclei, which preceded the loss of cell adhesion.

Published
2003
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