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1. Naturally Occurring Resistance-Associated Variants of Hepatitis C Virus Protease Inhibitors in Poor Responders to Pegylated Interferon-Ribavirin

Author: Larrat, Sylvie, Vallet, Sophie, David-Tchouda, Sandra, Caporossi, Alban, Margier, Jennifer, Ramière, Christophe, Scholtès, Caroline, Haïm-Boukobza, Stéphanie, Roque-Afonso, Anne-Marie, Besse, Bernard, André-Garnier, Elisabeth, Mohamed, Sofiane, Halfon, Philippe, Pivert, Adeline, Leguillou-Guillemette, Hélène, Abravanel, Florence, Guivarch, Matthieu, Mackiewicz, Vincent, Lada, Olivier, Mourez, Thomas, Plantier, Jean-Christophe, Baazia, Yazid, Alain, Sophie, Hantz, Sébastien, Thibault, Vincent, Gaudy-Graffin, Catherine, Bouvet, Dorine, Mirand, Audrey, Henquell, Cécile, Gozlan, Joël, Lagathu, Gisèle, Pronier, Charlotte, Velay, Aurélie, SCHVOERER, Evelyne, Trimoulet, Pascale, Fleury, Hervé, Bouvier-Alias, Magali, BROCHOT, Etienne, Duverlie, Gilles, Maylin, Sarah, Gouriou, Stéphanie, Pawlotsky, Jean-Michel, Morand, Patrice, Plantier, Christophe, Tang, Y.-W., 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), Laboratoire Universitaire de Biodiversité et Ecologie Microbienne (LUBEM), Université de Brest (UBO), Centre d'Investigation Clinique - Innovation Technologique - INSERM - CHU de Grenoble (CIC-IT Grenoble (CIT803)), Université Joseph Fourier - Grenoble 1 (UJF)-CHU Grenoble-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Grenoble Alpes - UFR Médecine (UGA UFRM), Université Grenoble Alpes (UGA), Centre Hospitalier Universitaire [Grenoble] (CHU), Biologie cellulaire des infections virales – Cell biology of viral infection, Centre International de Recherche en Infectiologie - UMR (CIRI), 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)-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 Recherche en Cancérologie de Lyon (CRCL), Centre Léon Bérard [Lyon]-Université Claude Bernard Lyon 1 (UCBL), Laboratoire de virologie, Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Hôpital Paul Brousse, Hôtel-Dieu de Nantes, Alphabio Laboratory, Hôpital Européen [Fondation Ambroise Paré - Marseille], Microbes évolution phylogénie et infections (MEPHI), Institut de Recherche pour le Développement (IRD)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Centre Hospitalier Universitaire d'Angers (CHU Angers), PRES Université Nantes Angers Le Mans (UNAM), Centre de Physiopathologie Toulouse Purpan (ex IFR 30 et IFR 150) (CPTP), Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UPS), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM), Hôpital Beaujon, Centre de recherche sur l'Inflammation (CRI), Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), CHU Rouen, Normandie Université (NU), Service de bactériologie-Virologie-Hygiène [Avicenne], Université Paris 13 (UP13)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Hôpital Avicenne, Université de Limoges (UNILIM), Service de Bactériologie, Virologie, Hygiène [CHU Limoges], CHU Limoges, CHU Pontchaillou [Rennes], Morphogénèse et antigénicité du VIH et du virus des Hépatites (MAVIVH), Centre Hospitalier Régional Universitaire de Tours (CHRU Tours)-Université de Tours-Institut National de la Santé et de la Recherche Médicale (INSERM), Epidémiologie et pathogénie des infections à entérovirus (EPIE), Université d'Auvergne - Clermont-Ferrand I (UdA), Laboratoire Microorganismes : Génome et Environnement - Clermont Auvergne (LMGE), Université Clermont Auvergne (UCA)-Centre National de la Recherche Scientifique (CNRS), CHU Saint-Antoine [APHP], Virology Laboratory, General Hospital Papageorgiou, Service de Virologie [CHRU Nancy], Centre Hospitalier Régional Universitaire de Nancy (CHRU Nancy), Stress, Immunité, Pathogènes (SIMPA), Université de Lorraine (UL), Service de virologie et d'immunologie biologique, CHU Bordeaux [Bordeaux]-Groupe hospitalier Pellegrin, Microbiologie cellulaire et moléculaire et pathogénicité (MCMP), Université Bordeaux Segalen - Bordeaux 2-Centre National de la Recherche Scientifique (CNRS), Centre National de Référence Virus des hépatites B, C et Delta, Institut National de la Transfusion Sanguine [Paris] (INTS)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), Unité de Virologie clinique et fondamentale EA 4294, CHU Amiens-Picardie, Institut de biologie de Lille - IBL (IBLI), Université de Lille, Sciences et Technologies-Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Centre National de la Recherche Scientifique (CNRS)-Université de Lille, Droit et Santé, Laboratoire de Virologie, Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], 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), Laboratoire de virologie moléculaire et structurale, CHU Grenoble, Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF), Laboratoire de Parasitologie et Mycologiede [CHRU Brest], Centre Hospitalier Régional Universitaire de Brest (CHRU Brest), Unité d'évaluation clinique et médico-économique [CHU Grenoble] (Département de la recherche clinique et de l'innovation), Techniques de l'Ingénierie Médicale et de la Complexité - Informatique, Mathématiques et Applications, Grenoble - UMR 5525 (TIMC-IMAG), VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF), Institut National de la Santé et de la Recherche Médicale (INSERM)-École normale supérieure - Lyon (ENS 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)-École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Virologie [HCL-Hôpital de la Croix Rousse], Hôpital de la Croix-Rousse [CHU - HCL], Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Service de Virologie [Villejuif], Hôpital Paul Brousse, Université Paris-Sud - Paris 11 - Faculté de médecine (UP11 UFR Médecine), Université Paris-Sud - Paris 11 (UP11), Laboratoire de Virologie [CHU Nantes], Centre hospitalier universitaire de Nantes (CHU Nantes), ALPHABIO - Laboratoire de biologie médicale, Laboratoire de virologie [CHU Angers], PRES Université Nantes Angers Le Mans (UNAM)-PRES Université Nantes Angers Le Mans (UNAM), Laboratoire de Virologie [CHU Purpan, Toulouse], Institut Fédératif de Biologie (IFB) - Hôpital Purpan, Hôpital Purpan [Toulouse], CHU Toulouse [Toulouse]-CHU Toulouse [Toulouse]-Hôpital Purpan [Toulouse], CHU Toulouse [Toulouse]-CHU Toulouse [Toulouse], Hôpital Beaujon [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Laboratoire de virologie [Rouen], Hôpital Charles Nicolle [Rouen]-CHU Rouen, Normandie Université (NU)-Normandie Université (NU)-Université de Rouen Normandie (UNIROUEN), Normandie Université (NU)-Département de microbiologie : Bactério, Virologie, Parasito, Hygiène, Université Paris 13 (UP13)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Avicenne [AP-HP], CHU Pitié-Salpêtrière [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Service de bactériologie-virologie [Tours], Hôpital Bretonneau-Centre Hospitalier Régional Universitaire de Tours (CHRU Tours), Centre national de référence du VIH INSERM U966, Université de Tours (UT), CHU Clermont-Ferrand, Laboratoire de Virologie [CHU Saint-Antoine], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Sorbonne Université (SU), Laboratoire de Virologie [Rennes] = Virology [Rennes], Institut National de la Transfusion Sanguine [Paris] (INTS)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Laboratoire de bactériologie - hygiène [CHU Amiens], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Paris Diderot - Paris 7 (UPD7)-Hopital Saint-Louis [AP-HP] (AP-HP), 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 ), Laboratoire Universitaire de Biodiversité et Ecologie Microbienne ( LUBEM ), Université de Brest ( UBO ), ThEMAS, Techniques de l'Ingénierie Médicale et de la Complexité - Informatique, Mathématiques et Applications, Grenoble ( TIMC-IMAG ), Université Joseph Fourier - Grenoble 1 ( UJF ) -Institut polytechnique de Grenoble - Grenoble Institute of Technology ( Grenoble INP ) -IMAG-Institut d'Enseignement Supérieur et de Recherche en Alimentation, Santé Animale, Sciences Agronomiques et de l'Environnement ( VetAgro Sup ) -Centre National de la Recherche Scientifique ( CNRS ) -Université Grenoble Alpes ( UGA ) -Université Joseph Fourier - Grenoble 1 ( UJF ) -Institut polytechnique de Grenoble - Grenoble Institute of Technology ( Grenoble INP ) -IMAG-Institut d'Enseignement Supérieur et de Recherche en Alimentation, Santé Animale, Sciences Agronomiques et de l'Environnement ( VetAgro Sup ) -Centre National de la Recherche Scientifique ( CNRS ) -Université Grenoble Alpes ( UGA ) -CHU Grenoble, Université Joseph Fourier - Grenoble 1 ( UJF ) -CHU Grenoble-CHU Grenoble, Pathogénèse et traitement de l'hépatite fulminante et du cancer du foie, Université Paris-Sud - Paris 11 ( UP11 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Université Grenoble Alpes - UFR Médecine ( UGA UFRM ), Université Grenoble Alpes ( UGA ), Internal Medicine and Infectious Diseases Department, European Hospital and Alphabio Laboratory, Centre Hospitalier Universitaire d'Angers ( CHU Angers ), PRES Université Nantes Angers Le Mans ( UNAM ), Laboratoire de Virologie [Toulouse], CHU Toulouse [Toulouse], Service d'hépatologie, Assistance publique - Hôpitaux de Paris (AP-HP) ( APHP ) -Université Paris Diderot - Paris 7 ( UPD7 ) -Hôpital Beaujon, Hôpital Bichat - Claude Bernard, Centre de recherche sur l'Inflammation ( CRI ), Université Paris Diderot - Paris 7 ( UPD7 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Institut de Recherche Biomédicale des Armées ( IRBA ), CHU Avicenne, Service de virologie, Université Pierre et Marie Curie - Paris 6 ( UPMC ) -Assistance publique - Hôpitaux de Paris (AP-HP) ( APHP ) -CHU Pitié-Salpêtrière [APHP], Morphogénèse et antigénicité du VIH et du virus des Hépatites ( MAVIVH ), Université de Tours-Centre Hospitalier Régional Universitaire de Tours-Institut National de la Santé et de la Recherche Médicale ( INSERM ), Centre de Calcul de l'IN2P3 ( CC-IN2P3 ), Institut National de Physique Nucléaire et de Physique des Particules du CNRS ( IN2P3 ) -Centre National de la Recherche Scientifique ( CNRS ), CHU Strasbourg, Virology Laboratory, Bordeaux University Hospital, Bordeaux, France, Microbiologie cellulaire et moléculaire et pathogénicité ( MCMP ), Université Bordeaux Segalen - Bordeaux 2-Centre National de la Recherche Scientifique ( CNRS ), Faculté de Médecine, CHU Henri Mondor, Université Joseph Fourier - Grenoble 1 (UJF)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS), Laboratoire Virologie [CHU Toulouse], Institut Fédératif de Biologie (IFB), Centre Hospitalier Universitaire de Toulouse (CHU Toulouse)-Centre Hospitalier Universitaire de Toulouse (CHU Toulouse)-Pôle Biologie [CHU Toulouse], Centre Hospitalier Universitaire de Toulouse (CHU Toulouse), Hôpital Charles Nicolle [Rouen], Normandie Université (NU)-Normandie Université (NU)-CHU Rouen, Centre Hospitalier Régional Universitaire de Tours (CHRU Tours)-Hôpital Bretonneau, SCHVOERER, Evelyne, Centre International de Recherche en Infectiologie (CIRI), École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), CHU Saint-Antoine [AP-HP], Université de Rouen Normandie (UNIROUEN), Normandie Université (NU)-Normandie Université (NU)-Département de microbiologie : Bactério, Virologie, Parasito, Hygiène-Hôpital Charles Nicolle [Rouen]-CHU Rouen, and Université de Tours
Subjects: Male, Hepacivirus, Viral Nonstructural Proteins, medicine.disease_cause, Gastroenterology, Telaprevir, chemistry.chemical_compound, Pegylated interferon, [SDV.MHEP.MI]Life Sciences [q-bio]/Human health and pathology/Infectious diseases, MESH: Hepacivirus, MESH: Treatment Outcome, MESH: Inhibitory Concentration 50, [SDV.MP.VIR] Life Sciences [q-bio]/Microbiology and Parasitology/Virology, MESH: Aged, education.field_of_study, MESH: Drug Resistance, Viral, MESH: Middle Aged, MESH: Protease Inhibitors, Hepatitis C, Middle Aged, 3. Good health, MESH: Hepatitis C, Chronic, Treatment Outcome, MESH: Oligopeptides, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, [SDV.MHEP.MI] Life Sciences [q-bio]/Human health and pathology/Infectious diseases, Female, MESH: Interferon-alpha, Viral load, Oligopeptides, medicine.drug, Microbiology (medical), MESH: Antiviral Agents, Adult, medicine.medical_specialty, Proline, Hepatitis C virus, Population, Mutation, Missense, Antiviral Agents, Inhibitory Concentration 50, MESH: Ribavirin, Internal medicine, Boceprevir, Virology, Drug Resistance, Viral, Ribavirin, medicine, Humans, Protease Inhibitors, education, Aged, Retrospective Studies, MESH: Mutation, Missense, MESH: Humans, MESH: Proline, business.industry, Interferon-alpha, MESH: Adult, MESH: Retrospective Studies, Hepatitis C, Chronic, medicine.disease, MESH: Male, chemistry, Immunology, MESH: Viral Nonstructural Proteins, business, MESH: Female, [ SDV.BBM.BS ] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM]
Abstract: The pretherapeutic presence of protease inhibitor (PI) resistance-associated variants (RAVs) has not been shown to be predictive of triple-therapy outcomes in treatment-naive patients. However, they may influence the outcome in patients with less effective pegylated interferon (pegIFN)-ribavirin (RBV) backbones. Using hepatitis C virus (HCV) population sequence analysis, we retrospectively investigated the prevalence of baseline nonstructural 3 (NS3) RAVs in a multicenter cohort of poor IFN-RBV responders (i.e., prior null responders or patients with a viral load decrease of 3-fold shift in 50% inhibitory concentration (IC 50 ) for telaprevir or boceprevir, T54S was the most frequently detected mutation (3.9%), followed by A156T, R155K (0.7%), V36M, and V55A (0.35%). Mutations were more frequently found in patients infected with genotype 1a (7.5 to 23.6%) than 1b (3.3 to 19.8%) ( P = 0.03). No other sociodemographic or viroclinical characteristic was significantly associated with a higher prevalence of RAVs. No obvious effect of baseline RAVs on viral load was observed. In this cohort of poor responders to IFN-RBV, no link was found with a sustained virological response to triple therapy, regardless of the algorithm used for the detection of mutations. Based on a cross-study comparison, baseline RAVs are not more frequent in poor IFN-RBV responders than in treatment-naive patients and, even in these difficult-to-treat patients, this study demonstrates no impact on treatment outcome, arguing against resistance analysis prior to treatment.
Published: 2015
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2. 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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3. Surveillance of HIV-1 primary infections in France from 2014 to 2016: toward stable resistance, but higher diversity, clustering and virulence?

Author: Visseaux, Benoit, Assoumou, Lambert, Grude, Maxime, Carles, Marie-Josée, Meyer, Laurence, Roussel, Catherine, Le Guillou-Guillemette, H., Ducancelle, A., Courdavault, L, Alloui, C, Honore, P, Lepiller, Q., Bettinger, D., Bellecave, P., Pinson-Recordon, P, Tumiotto, Camille, Vallet, Sophie, Payan, C., Duthe, J, Leroux, M., Dina, Julia, Vabret, A., Mirand, A., Henquell, C., Bouvier-Alias, Magali, Simohamed, A, dos Santos, G, yerly, S, Gaille, C, Caveng, W, Chapalay, S, Calmy, A., Signori-Schmuck, Anne, Morand, Patrice, Pallier, Coralie, Raho-Moussa, M, Mole, M, Dulucq, M-J, Bocket, Laurence, Alidjinou, K, Ranger-Rogez, S, Trabaud, Mary-Anne, Icard, V, Tardy, J., Tamalet, C., Delamare, C, Montes, Brigitte, Schvoerer, E., Fenaux, H., Rodallec, A., André-Garnier, E., Ferre, Virginie, de Monte, Anne, Guigon, A., Guinard, J., Descamps, Diane, Charpentier, C., Peytavin, G., Tremaux, P, Avettand-Fenoel, V., Soulie, C., Malet, I., Wirden, Marc, Marcelin, A, Calvez, V., Flandre, P., Costagliola, D, Morand-Joubert, Laurence, Lambert-Niclot, S., Fofana, D, Boukli, N., Delaugerre, C., Chaix, Marie-Laure, Mahjoub, Nadia, Amiel, Corinne, Giraudeau, G, Beby-Defaux, A., Plainchamp, D, Maillard, Anne, Alessandri-Gradt, E, Leoz, M, Plantier, Jean-Christophe, Gantner, P, Delagreverie, H, Fafi-Kremer, Samira, Fischer, P, Raymond, Stéphanie, Izopet, Jacques, Chiabrando, J, Stefic, Karl, Barin, F., Fajole, G, Burgault, O, Marque-Juillet, S, Service de Virologie [CHU Bichat], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-AP-HP - Hôpital Bichat - Claude Bernard [Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Epidémiologie, stratégies thérapeutiques et virologie cliniques dans l'infection à VIH, Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Pierre Louis d'Epidémiologie et de Santé Publique (iPLESP), Centre de recherche en épidémiologie et santé des populations (CESP), Université de Versailles Saint-Quentin-en-Yvelines (UVSQ)-Université Paris-Sud - Paris 11 (UP11)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Paul Brousse-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre Hospitalier Régional Universitaire de Tours (CHRU Tours), Centre Hospitalier Universitaire d'Angers (CHU Angers), PRES Université Nantes Angers Le Mans (UNAM), Immuno-Rhumatologie Moléculaire, Université de Strasbourg (UNISTRA)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre Hospitalier Régional Universitaire de Besançon (CHRU Besançon), Microbiologie cellulaire et moléculaire et pathogénicité (MCMP), Université Bordeaux Segalen - Bordeaux 2-Centre National de la Recherche Scientifique (CNRS), Université de Bordeaux (UB), virology, Centre Hospitalier Régional Universitaire de Brest (CHRU Brest), Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Centre de recherche sur l'hétéroepitaxie et ses applications (CRHEA), 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)-Centre National de la Recherche Scientifique (CNRS)-Université Côte d'Azur (UCA), Service de Virologie [CHU Caen], CHU Caen, Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN)-Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN), CHU Clermont-Ferrand, 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), Geneva University Hospital (HUG), Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-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), Laboratoire de virologie [CHU Lille], Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Institut Hospitalier Universitaire Méditerranée Infection (IHU Marseille), Centre Hospitalier Régional Universitaire [Montpellier] (CHRU Montpellier), Laboratoire de Chimie Physique et Microbiologie pour les Matériaux et l'Environnement (LCPME), Institut de Chimie du CNRS (INC)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Service de virologie [CHU Nantes], Centre hospitalier universitaire de Nantes (CHU Nantes), Hôtel-Dieu de Nantes, Infection, Anti-microbiens, Modélisation, Evolution (IAME (UMR_S_1137 / U1137)), Université Paris 13 (UP13)-Université Paris Diderot - Paris 7 (UPD7)-Université Sorbonne Paris Cité (USPC)-Institut National de la Santé et de la Recherche Médicale (INSERM), Service de Radiologie [CHU Rouen], CHU Rouen, Normandie Université (NU)-Normandie Université (NU), Pharmacie de l'Hôpital Bichat, Centre de Recherches Pétrographiques et Géochimiques (CRPG), Institut national des sciences de l'Univers (INSU - CNRS)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Service de Virologie [CHU Pitié-Salpêtrière], CHU Pitié-Salpêtrière [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU), CHU Saint-Antoine [AP-HP], Service de microbiologie [Saint-Louis], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Paris Diderot - Paris 7 (UPD7)-Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], Génétique et Ecologie des Virus, Génétique des Virus et Pathogénèse des Maladies Virales, Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM), Laboratoire de Virologie [CHU Saint-Louis], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], Centre d'Immunologie et des Maladies Infectieuses (CIMI), Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Centre hospitalier universitaire de Poitiers (CHU Poitiers), Unité de Rétrovirologie, Hôpital Pontchaillou, Normandie Université (NU), Laboratoire de Virologie, CHU Strasbourg, Laboratoire de Virologie [Toulouse], CHU Toulouse [Toulouse], Service de bactériologie-virologie [Tours], Centre Hospitalier Régional Universitaire de Tours (CHRU Tours)-Hôpital Bretonneau, Centre national de référence du VIH INSERM U966, Université de Tours (UT), Université de Versailles Saint-Quentin-en-Yvelines (UVSQ)-Université Paris-Sud - Paris 11 (UP11)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Hôpital Paul Brousse-Institut National de la Santé et de la Recherche Médicale (INSERM), CHU Besançon, Université de Franche-Comté (UFC), Université Grenoble Alpes (UGA), Université Nice Sophia Antipolis (... - 2019) (UNS), Université Côte d'Azur (UCA)-Université Côte d'Azur (UCA)-Centre National de la Recherche Scientifique (CNRS), 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 de biologie structurale (IBS - UMR 5075), 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)-Université Grenoble Alpes (UGA)-Centre National de la Recherche Scientifique (CNRS), Institut Hospitalier Universitaire Méditerranée Infection (IHU AMU), Stress, Immunité, Pathogènes (SIMPA), Université de Lorraine (UL), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Physiologie, Environnement et Génétique pour l'Animal et les Systèmes d'Elevage [Rennes] (PEGASE), AGROCAMPUS OUEST-Institut National de la Recherche Agronomique (INRA), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris 13 (UP13)-Université Paris Diderot - Paris 7 (UPD7)-Université Sorbonne Paris Cité (USPC), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-AP-HP - Hôpital Bichat - Claude Bernard [Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-CHU Pitié-Salpêtrière [APHP], CHU Saint-Antoine [APHP], Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Université Paris Diderot - Paris 7 (UPD7), Centre d'Immunologie et de Maladies Infectieuses (CIMI), Université de Tours, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Paris-Sud - Paris 11 (UP11)-Hôpital Paul Brousse-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Versailles Saint-Quentin-en-Yvelines (UVSQ), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12)-IFR10, 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)-Centre National de la Recherche Scientifique (CNRS), Service de virologie [CHU Pitié-Salpêtrière], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Sorbonne Université (SU)-Institut National de la Santé et de la Recherche Médicale (INSERM), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Université Paris Diderot - Paris 7 (UPD7)-Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], Hôpital Bretonneau-Centre Hospitalier Régional Universitaire de Tours (CHRU Tours), and Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Diderot - Paris 7 (UPD7)
Subjects: Male, Etravirine, HIV Infections, Men who have sex with men, Sexual and Gender Minorities, chemistry.chemical_compound, Pre-exposure prophylaxis, 0302 clinical medicine, Pharmacology (medical), 030212 general & internal medicine, Phylogeny, ComputingMilieux_MISCELLANEOUS, 0303 health sciences, education.field_of_study, Virulence, biology, [SDE.IE]Environmental Sciences/Environmental Engineering, Middle Aged, Viral Load, 3. Good health, Integrase, Infectious Diseases, Rilpivirine, Epidemiological Monitoring, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, Female, France, Viral load, medicine.drug, Adult, Microbiology (medical), Genotype, Anti-HIV Agents, [SDE.MCG]Environmental Sciences/Global Changes, Population, Evolution, Molecular, 03 medical and health sciences, Drug Resistance, Viral, medicine, Humans, education, Pharmacology, 030306 microbiology, Genetic Variation, Sequence Analysis, DNA, [SDE.ES]Environmental Sciences/Environmental and Society, Virology, chemistry, Mutation, HIV-1, biology.protein, [SDE.BE]Environmental Sciences/Biodiversity and Ecology
Abstract: ObjectivesPatients with primary HIV-1 infection (PHI) are a particular population, giving important insight about ongoing evolution of transmitted drug resistance-associated mutation (TDRAM) prevalence, HIV diversity and clustering patterns. We describe these evolutions of PHI patients diagnosed in France from 2014 to 2016.MethodsA total of 1121 PHI patients were included. TDRAMs were characterized using the 2009 Stanford list and the French ANRS algorithm. Viral subtypes and recent transmission clusters (RTCs) were also determined.ResultsPatients were mainly MSM (70%) living in the Paris area (42%). TDRAMs were identified among 10.8% of patients and rose to 18.6% when including etravirine and rilpivirine TDRAMs. Prevalences of PI-, NRTI-, first-generation NNRTI-, second-generation NNRTI- and integrase inhibitor-associated TDRAMs were 2.9%, 5.0%, 4.0%, 9.4% and 5.4%, respectively. In a multivariable analysis, age >40 years and non-R5 tropic viruses were associated with a >2-fold increased risk of TDRAMs. Regarding HIV diversity, subtype B and CRF02_AG (where CRF stands for circulating recombinant form) were the two main lineages (56% and 20%, respectively). CRF02_AG was associated with higher viral load than subtype B (5.83 versus 5.40 log10 copies/mL, P = 0.004). We identified 138 RTCs ranging from 2 to 14 patients and including overall 41% from the global population. Patients in RTCs were younger, more frequently born in France and more frequently MSM.ConclusionsSince 2007, the proportion of TDRAMs has been stable among French PHI patients. Non-B lineages are increasing and may be associated with more virulent CRF02_AG strains. The presence of large RTCs highlights the need for real-time cluster identification to trigger specific prevention action to achieve better control of the epidemic.
Published: 2019
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4. CircRNAs in Plants

Author: Xuelei Lai, Chloe Zubieta, Stuart Webb, Martin Crespi, Simon J. Conn, Jérémie Bazin, Physiologie cellulaire et végétale (LPCV), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA)-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), Institut des Sciences des Plantes de Paris-Saclay (IPS2 (UMR_9213 / UMR_1403)), Institut National de la Recherche Agronomique (INRA)-Université Paris-Sud - Paris 11 (UP11)-Université Paris Diderot - Paris 7 (UPD7)-Université d'Évry-Val-d'Essonne (UEVE)-Centre National de la Recherche Scientifique (CNRS), Flinders University of South Australia, J. Xiao, Univ Groningen, Lab Biophys Chem, Nijenborgh 7, NL-9747 AG Groningen, Netherlands, Institut National de la Recherche Agronomique (INRA)-Université d'Évry-Val-d'Essonne (UEVE)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Diderot - Paris 7 (UPD7)-Université Paris-Saclay-Université Paris-Sud - Paris 11 (UP11), European Molecular Biology Laboratory, European Molecular Biology Laboratory [Grenoble] (EMBL), 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), Université Paris-Sud - Paris 11 (UP11)-Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS)-Université d'Évry-Val-d'Essonne (UEVE)-Institut National de la Recherche Agronomique (INRA), Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Lai, Xuelei, Bazin, Jérémie, Webb, Stuart, Crespi, Martin, Zubieta, Chloe, and Conn, Simon J
Subjects: 0301 basic medicine, [SDV]Life Sciences [q-bio], Computational biology, Biology, Transcriptome, transcriptomics, 03 medical and health sciences, Transcription (biology), Arabidopsis, microRNA, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Transcriptomics, ComputingMilieux_MISCELLANEOUS, plants, Alternative splicing, food and beverages, Plants, biology.organism_classification, Genome-wide profiling, Phenotype, genome-wide profiling, 030104 developmental biology, RNA splicing, circRNAs, Biogenesis
Abstract: International audience; Circular RNAs (circRNAs) are covalently closed, single-stranded transcripts that are ubiquitously expressed in all eukaryotes and even prokaryotic archaea. Although once regarded as splicing artifacts, circRNAs are a novel class of regulatory molecules with diverse biological functions, including regulation of transcription, modulation of alternative splicing, and binding of miRNAs and proteins. The majority of studies of circRNAs have been performed in animals with a focus on the biogenesis, function, and mechanistic characterization of these molecules. In contrast, the study of circRNAs in plants is just emerging. Interestingly, recent circRNA profiling studies in model plant systems show distinct features of plant circRNAs compared with those from animals, including putative roles in stress response, differences in expression patterns, and novel biogenesis mechanisms. This provides a great opportunity to broaden our knowledge of circRNAs using plant model systems, such as Arabidopsis and rice, which are ideal for phenotypic characterization and genetic studies. In this review, we summarize current knowledge of plant circRNAs, discuss their identification and biogenesis, describe potential functions, and propose future perspectives for plant circRNA study.
Published: 2018
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5. The Non-Coding RNA Journal Club: Highlights on Recent Papers—6

Author: Hua Xiao, Patrick Shiu, Jun Shu, Gaetano Santulli, Mohammad Gheybi, Simon Conn, Baptiste Bogard, Florent Hubé, Joseph Taube, Sendurai Mani, Luo Song, George Calin, Shuxing Zhang, Columbia University Irving Medical Center (CUIMC), 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), Centre épigénétique et destin cellulaire (EDC (UMR_7216)), Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS), Université Paris Diderot - Paris 7 (UPD7), MD Anderson Cancer Center [Houston], The University of Texas Health Science Center at Houston (UTHealth), School of Pharmacy and Medical Sciences, University of South Australia [Adelaide], Centre National de la Recherche Scientifique (CNRS)-Université Paris Diderot - Paris 7 (UPD7), Xiao, Hua, Shiu, Patrick KT, Shu, Jun, Santulli, Gaetano, Gheybi, Mohammad K, Conn, Simon J, Bogard, Baptiste, Hube, Florent, Taube, Joseph H, Mani, Sendurai A, Song, Luo, Calin, George A, and Zhang, Shuxing
Subjects: 0303 health sciences, lcsh:QH426-470, [SDV]Life Sciences [q-bio], journal club, Biochemistry, 03 medical and health sciences, lcsh:Genetics, 0302 clinical medicine, n/a, Editorial, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], Genetics, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, recent papers, Molecular Biology, 030217 neurology & neurosurgery, ComputingMilieux_MISCELLANEOUS, 030304 developmental biology
Abstract: We are delighted to share with you our sixth Journal Club and highlight some of the most interesting papers published recently [...]
Published: 2018
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6. Emerging resistance mutations in PI-naive patients failing an atazanavir-based regimen (ANRS multicentre observational study)

Author: Lambert-Niclot, L, Grude, C, Chaix, L., Charpentier, L, Reigadas, C, Le Guillou-Guillemette, L, Rodallec, C, Maillard, Pascale, Dufayard, C, Mourez, C, Mirand, C, Guinard, L, Montes, S, Vallet, L, Marcelin, Marc, Descamps, L, Flandre, C, Delaugerre, L, Alloui, Chakib, Descamps, Diane, Charpentier, Charlotte, Visseaux, Benoit, Krivine, Anne, Bouviers-Alias, Ali, Pallier, Coralie, Soulié, Cathia, Wirden, Marc, Marcelin, Anne, Calvez, Vincent, Morand, Laurence, Lambert-Niclot, Sidonie, Fofana, Djeneba, Mahjoub, Nadia, Delaugerre, Constance, Chaix, Marie, Amiel, Corinne, Schneider, Veronique, Roussel, Catherine, Le Guillou-Guillemette, Hélène, Courdavault, Laurence, Reigadas, Sandrine, Recordon-Pinson, Patricia, Fleury, Hervé, Vallet, Sophie, Dina, Julia, Vabret, Astrid, Mirand, Audrey, Henquell, Cécile, Auvray, Christelle, de Rougemont, Alexis, Giraudon, Helene, Si-Mohammed, Ali, Mathez, Dominique, Signori-Schmuck, Anne, Morand, Patrice, Bocket, Laurence, Trabaud, Anne, Montes, Brigitte, Le Guen, Laura, Rodallec, Audrey, Ferré, Virginie, Jeulin, Hélène, SCHVOERER, Evelyne, Dufayard, Jacqueline, Allardet-Servent, Annick, carles, Marie, Guinard, Jérôme, Guigon, Aurélie, Giraudeau, Genevieve, Beby-Defaux, Agnès, Maillard, Anne, Plantier, Jean Christophe, Leoz, Marie, Mourez, Thomas, Bourlet, Thomas, Fafi-Kremer, Samira, Chiabrando, Julie, Raymond, Stéphanie, Izopet, Jacques, Barin, Francis, Marque-Juillet, Stéphanie, Yerly, Sabine, Institut Pierre Louis d'Epidémiologie et de Santé Publique (iPLESP), Sorbonne Université (SU)-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Pierre et Marie Curie - Paris 6 - UFR de Médecine Pierre et Marie Curie (UPMC), Université Pierre et Marie Curie - Paris 6 (UPMC), Hopital Saint-Louis [AP-HP] (AP-HP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), AP-HP - Hôpital Bichat - Claude Bernard [Paris], Hôpital Pellegrin, CHU Bordeaux [Bordeaux]-Groupe hospitalier Pellegrin, Centre Hospitalier Universitaire d'Angers (CHU Angers), PRES Université Nantes Angers Le Mans (UNAM), Centre hospitalier universitaire de Nantes (CHU Nantes), Hôpital Pontchaillou, Centre Hospitalier Universitaire de Nice (CHU Nice), CHU Rouen, Normandie Université (NU), CHU Clermont-Ferrand, Centre Hospitalier Régional d'Orléans (CHRO), Centre Hospitalier Régional Universitaire [Montpellier] (CHRU Montpellier), Hôpital de la Cavale Blanche - CHRU Brest (CHU - BREST ), CHU Pitié-Salpêtrière [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Laboratoire Bordelais de Recherche en Informatique (LaBRI), Université de Bordeaux (UB)-Centre National de la Recherche Scientifique (CNRS)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB), Infection, Anti-microbiens, Modélisation, Evolution (IAME (UMR_S_1137 / U1137)), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris 13 (UP13)-Université Paris Diderot - Paris 7 (UPD7)-Université Sorbonne Paris Cité (USPC), 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)-IFR10, Service de virologie [CHU Pitié-Salpêtrière], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Epidémiologie, stratégies thérapeutiques et virologie cliniques dans l'infection à VIH, Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM), Département de Mathématiques et Applications - ENS Paris (DMA), École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Centre National de la Recherche Scientifique (CNRS), Département d'Astrophysique, de physique des Particules, de physique Nucléaire et de l'Instrumentation Associée (DAPNIA), Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Laboratoire de Virologie [CHU Saint-Louis], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], Génétique et Ecologie des Virus, Génétique des Virus et Pathogénèse des Maladies Virales, Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM), UFR des Sciences de Santé (Université de Bourgogne), Université de Bourgogne (UB), CHU Tenon [AP-HP], Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Hémodynamique, Interaction Fibrose et Invasivité tumorales Hépatiques (HIFIH), Université d'Angers (UA), Centre Hospitalier Victor Dupouy, Microbiologie cellulaire et moléculaire et pathogénicité (MCMP), Université Bordeaux Segalen - Bordeaux 2-Centre National de la Recherche Scientifique (CNRS), CHU Bordeaux [Bordeaux], Laboratoire Universitaire de Biodiversité et Ecologie Microbienne (LUBEM), Université de Brest (UBO), Laboratoire de Virologie Humaine et Moléculaire [Caen], CHU Caen, Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN)-Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN), Groupe de Recherche sur l'Adaptation Microbienne (GRAM 2.0), Université de Rouen Normandie (UNIROUEN), Normandie Université (NU)-Normandie Université (NU)-Université de Caen Normandie (UNICAEN), Laboratoire Microorganismes : Génome et Environnement (LMGE), Université Clermont Auvergne [2017-2020] (UCA [2017-2020])-Centre National de la Recherche Scientifique (CNRS), Laboratoire de sérologie-virologie (CHU de Dijon), Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon), Hôpital Raymond Poincaré [AP-HP], 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), Département de virologie [Grenoble], Centre Hospitalier Universitaire [Grenoble] (CHU), Service de Virologie [Lille], Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Epidémiologie, Démographie et Sciences Sociales: santé reproductive, sexualité et infection à VIH (Inserm U569), Epidémiologie, sciences sociales, santé publique (IFR 69), Université Paris 1 Panthéon-Sorbonne (UP1)-Université Paris-Sud - Paris 11 (UP11)-École des hautes études en sciences sociales (EHESS)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-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 1 Panthéon-Sorbonne (UP1)-Université Paris-Sud - Paris 11 (UP11)-École des hautes études en sciences sociales (EHESS)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-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 d'études démographiques (INED)-Institut National de la Santé et de la Recherche Médicale (INSERM), Laboratoire de Chimie Physique et Microbiologie pour les Matériaux et l'Environnement (LCPME), Institut de Chimie du CNRS (INC)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Observatoire des Programmes Communautaires de Développement Rural (US ODR), Institut National de la Recherche Agronomique (INRA), Institut Pasteur de Nouvelle-Calédonie, Réseau International des Instituts Pasteur (RIIP), Centre hospitalier universitaire de Poitiers (CHU Poitiers), Ecophysiologie Végétale, Agronomie et Nutritions (EVA), Institut National de la Recherche Agronomique (INRA)-Université de Caen Normandie (UNICAEN), Normandie Université (NU)-Normandie Université (NU), Groupe Immunité des Muqueuses et Agents Pathogènes (GIMAP), Université Jean Monnet [Saint-Étienne] (UJM), Interaction virus-hôte et maladies du foie, Université de Strasbourg (UNISTRA)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Claude de Préval (ICP), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-CHU Toulouse [Toulouse]-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées, Université de Tours (UT), Centre Hospitalier de Versailles André Mignot (CHV), Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU), Université de Bordeaux (UB)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB)-Centre National de la Recherche Scientifique (CNRS), Université Paris 13 (UP13)-Université Paris Diderot - Paris 7 (UPD7)-Université Sorbonne Paris Cité (USPC)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut National de la Santé et de la Recherche Médicale (INSERM)-IFR10-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12), École normale supérieure - Paris (ENS-PSL), Microbiologie Fondamentale et Pathogénicité (MFP), Université de Caen Normandie (UNICAEN), Normandie Université (NU)-Normandie Université (NU)-Université de Rouen Normandie (UNIROUEN), 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), Normandie Université (NU)-Normandie Université (NU)-Institut National de la Recherche Agronomique (INRA), Université Jean Monnet - Saint-Étienne (UJM), Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Université de Toulouse (UT)-Centre Hospitalier Universitaire de Toulouse (CHU Toulouse)-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)-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)-Université Paris Diderot - Paris 7 (UPD7), Université de Tours, Service de Virologie [CHU Pitié-Salpêtrière], Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-CHU Toulouse [Toulouse]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Lycée agricole La Touche, Hôpital Bichat - Claude Bernard, Centre National de la Recherche Scientifique (CNRS)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB)-Université Sciences et Technologies - Bordeaux 1-Université Bordeaux Segalen - Bordeaux 2, Memo-Flu-ARDS Study Group, Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-CHU Pitié-Salpêtrière [APHP], École normale supérieure - Paris (ENS Paris)-Centre National de la Recherche Scientifique (CNRS), CHU Pitié-Salpêtrière [APHP], Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), CHU Tenon [APHP], Hôpital d'Argenteuil, Laboratoire Microorganismes : Génome et Environnement - Clermont Auvergne (LMGE), Université Clermont Auvergne (UCA)-Centre National de la Recherche Scientifique (CNRS), AP-HP Hôpital Raymond Poincaré [Garches], 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), Laboratoire de virologie moléculaire et structurale, CHU Grenoble, Laboratoire de virologie [CHU Lille], Université Panthéon-Sorbonne (UP1)-Université Paris-Sud - Paris 11 (UP11)-École des hautes études en sciences sociales (EHESS)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-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 Panthéon-Sorbonne (UP1)-Université Paris-Sud - Paris 11 (UP11)-École des hautes études en sciences sociales (EHESS)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-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)-Institut national d'études démographiques (INED), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Centre national de référence du VIH INSERM U966, Centre Hospitalier de Versailles (CHV), and Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)
Subjects: 0301 basic medicine, Microbiology (medical), Adult, Male, medicine.medical_specialty, 030106 microbiology, Atazanavir Sulfate, HIV Infections, Emtricitabine, Gastroenterology, 03 medical and health sciences, 0302 clinical medicine, Abacavir, [SDV.MHEP.MI]Life Sciences [q-bio]/Human health and pathology/Infectious diseases, Internal medicine, Drug Resistance, Viral, medicine, Humans, Pharmacology (medical), 030212 general & internal medicine, Treatment Failure, Tenofovir, ComputingMilieux_MISCELLANEOUS, Retrospective Studies, Pharmacology, business.industry, virus diseases, Lamivudine, HIV Protease Inhibitors, Middle Aged, Viral Load, Resistance mutation, Dideoxynucleosides, 3. Good health, Atazanavir, Regimen, Drug Combinations, Infectious Diseases, Mutation, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, HIV-1, Ritonavir, business, Viral load, medicine.drug
Abstract: Background Atazanavir is a PI widely used as a third agent in combination ART. We aimed to determine the prevalence and the patterns of resistance in PI-naive patients failing on an atazanavir-based regimen. Methods We analysed patients failing on an atazanavir-containing regimen used as a first line of PI therapy. We compared the sequences of reverse transcriptase and protease before the introduction of atazanavir and at failure [two consecutive viral loads (VLs) >50 copies/mL]. Resistance was defined according to the 2014 Agence Nationale de Recherche sur le SIDA et les Hepatites Virales (ANRS) algorithm. Results Among the 113 patients, atazanavir was used in the first regimen in 71 (62.8%) patients and in the first line of a PI-based regimen in 42 (37.2%). Atazanavir was boosted with ritonavir in 95 (84.1%) patients and combined with tenofovir/emtricitabine or lamivudine (n = 81) and abacavir/lamivudine or emtricitabine (n = 22). At failure, median VL was 3.05 log10 copies/mL and the median CD4+ T cell count was 436 cells/mm3. The median time on atazanavir was 21.2 months. At failure, viruses were considered resistant to atazanavir in four patients (3.5%) with the selection of the following major atazanavir-associated mutations: I50L (n = 1), I84V (n = 2) and N88S (n = 1). Other emergent PI mutations were L10V, G16E, K20I/R, L33F, M36I/L, M46I/L, G48V, F53L, I54L, D60E, I62V, A71T/V, V82I/T, L90M and I93L/M. Emergent NRTI substitutions were detected in 21 patients: M41L (n = 2), D67N (n = 3), K70R (n = 1), L74I/V (n = 3), M184V/I (n = 16), L210W (n = 1), T215Y/F (n = 3) and K219Q/E (n = 2). Conclusions Resistance to atazanavir is rare in patients failing the first line of an atazanavir-based regimen according to the ANRS. Emergent NRTI resistance-associated mutations were reported in 18% of patients.
Published: 2018
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7. Phenotypic analysis of HIV-1 E157Q integrase polymorphism and impact on virological outcome in patients initiating an integrase inhibitor-based regimen

Author: Alexandre Storto, Diane Descamps, Anne Maillard, Audrey Mirand, Corinne Amiel, Charlotte Charpentier, Vincent Calvez, Thuy Van Nguyen, Laurence Bocket, Coralie Pallier, Audrey Rodallec, Elisabeth André-Garnier, Marie Leoz, Laurence Morand-Joubert, Isabelle Malet, Stéphanie Raymond, Véronique Schneider, Catherine Roussel, Camille Tumiotto, Constance Delaugerre, Anne-Geneviève Marcelin, Julia Dina, Jean-Christophe Plantier, Brigitte Montes, Anne Signori-Schmuck, Infection, Anti-microbiens, Modélisation, Evolution (IAME (UMR_S_1137 / U1137)), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris 13 (UP13)-Université Paris Diderot - Paris 7 (UPD7)-Université Sorbonne Paris Cité (USPC), Université Paris Diderot - Paris 7 (UPD7), Institut Pierre Louis d'Epidémiologie et de Santé Publique (iPLESP), Sorbonne Université (SU)-Institut National de la Santé et de la Recherche Médicale (INSERM), Hôtel-Dieu de Nantes, Service de Virologie [Lille], Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Service de bactériologie-virologie, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Sorbonne Université (SU), Service de virologie [Hôpital Tenon], CHU Tenon [AP-HP], Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Ecophysiologie Végétale, Agronomie et Nutritions (EVA), Institut National de la Recherche Agronomique (INRA)-Université de Caen Normandie (UNICAEN), Normandie Université (NU)-Normandie Université (NU), Service de virologie [CHU Nantes], Centre hospitalier universitaire de Nantes (CHU Nantes), Laboratoire de virologie [Rouen], CHU Rouen, Normandie Université (NU)-Normandie Université (NU)-Université de Rouen Normandie (UNIROUEN), Normandie Université (NU), Centre Hospitalier Régional Universitaire [Montpellier] (CHRU Montpellier), Service de Virologie [CHU Caen], CHU Caen, Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN)-Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN), Service de Virologie [Villejuif], Hôpital Paul Brousse, Laboratoire Microorganismes : Génome et Environnement (LMGE), Université Clermont Auvergne [2017-2020] (UCA [2017-2020])-Centre National de la Recherche Scientifique (CNRS), Service de bactériologie-virologie [Tours], Hôpital Bretonneau-Centre Hospitalier Régional Universitaire de Tours (CHRU Tours), 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), Laboratoire de Virologie [Toulouse], CHU Toulouse [Toulouse], Génétique et Ecologie des Virus, Génétique des Virus et Pathogénèse des Maladies Virales, Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM), Service de Virologie [CHU Bichat], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-AP-HP - Hôpital Bichat - Claude Bernard [Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-CHU Saint-Antoine [APHP], CHU Tenon [APHP], Université de Caen Normandie (UNICAEN), Normandie Université (NU)-Normandie Université (NU)-Institut National de la Recherche Agronomique (INRA), Laboratoire Microorganismes : Génome et Environnement - Clermont Auvergne (LMGE), Université Clermont Auvergne (UCA)-Centre National de la Recherche Scientifique (CNRS), 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), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-AP-HP - Hôpital Bichat - Claude Bernard [Paris], Université Paris 13 (UP13)-Université Paris Diderot - Paris 7 (UPD7)-Université Sorbonne Paris Cité (USPC)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre Hospitalier Régional Universitaire de Tours (CHRU Tours)-Hôpital Bretonneau, Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Diderot - Paris 7 (UPD7), CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), 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), Laboratoire Virologie [CHU Toulouse], Institut Fédératif de Biologie (IFB), Centre Hospitalier Universitaire de Toulouse (CHU Toulouse)-Centre Hospitalier Universitaire de Toulouse (CHU Toulouse)-Pôle Biologie [CHU Toulouse], and Centre Hospitalier Universitaire de Toulouse (CHU Toulouse)
Subjects: 0301 basic medicine, Microbiology (medical), Genotype, 030106 microbiology, Mutation, Missense, Integrase inhibitor, HIV Infections, HIV Integrase, [SDV.MP.PRO]Life Sciences [q-bio]/Microbiology and Parasitology/Protistology, Virus, 03 medical and health sciences, chemistry.chemical_compound, Gene Frequency, medicine, Prevalence, Humans, Pharmacology (medical), HIV Integrase Inhibitors, Pharmacology, biology, Elvitegravir, business.industry, Viral Load, Raltegravir, Virology, [SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, 3. Good health, Integrase, Regimen, Infectious Diseases, Treatment Outcome, chemistry, Dolutegravir, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, biology.protein, HIV-1, France, business, Viral load, medicine.drug
Abstract: International audience; ObjectivesTo assess the phenotypic susceptibility of the E157Q polymorphism in HIV-1 integrase (IN) and the virological outcome of patients infected with E157Q-mutated virus initiating an IN inhibitor (INI)-based regimen.MethodsThis was a multicentre study assessing IN sequences from INI-naive patients among 17 French HIV clinical centres. E157Q site-directed mutants in pNL4.3 and pCRF02_AG contexts were assessed in a recombinant phenotypic assay.ResultsPrevalence of the E157Q polymorphism was 2.7% among 8528 IN sequences from INI-naive patients and its distribution was 1.7%, 5.6% and 2.2% in B, CRF02_AG and various non-B subtypes, respectively. Thirty-nine INI-naive patients with E157Q-mutated virus initiated an INI-based regimen. Among them, 15 had a viral load (VL) 50 copies/mL at the time of INI-based regimen initiation. Among them eight were receiving a first-line regimen and the only two patients who did not reach VL
Published: 2018
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8. Innovative multiplexed point-of-care immunoassay applied to hepatitis B screening

Author: Paul Kauffmann, Pn Marche, Guillaume Blaire, Sarah Delshadi, Thomas Decaens, Patrice Morand, V. Masse, O. Cugat, Sylvie Larrat, Microbiology and Infectious Diseases, Sherbrooke University Hospital, 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), Laboratoire de Génie Electrique de Grenoble (G2ELab), Université Joseph Fourier - Grenoble 1 (UJF)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP)-Institut Polytechnique de Grenoble - Grenoble Institute of Technology-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA), 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), Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Chadebec, Olivier, Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)
Subjects: medicine.diagnostic_test, business.industry, [SPI.NRJ]Engineering Sciences [physics]/Electric power, Biochemistry (medical), Clinical Biochemistry, General Medicine, Biochemistry, Virology, 3. Good health, Hepatitis B screening, Immunoassay, medicine, business, ComputingMilieux_MISCELLANEOUS, [SPI.NRJ] Engineering Sciences [physics]/Electric power, Point of care
Abstract: International audience
Published: 2019
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9. Fully automatic macromolecular crystallography: the impact of MASSIF-1 on the optimum acquisition and quality of data

Author: Didier Nurizzo, Olof Svensson, Matthew W. Bowler, European Molecular Biology Laboratory [Grenoble] (EMBL), 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), and European Synchrotron Radiation Facility (ESRF)
Subjects: 0301 basic medicine, 030103 biophysics, Computer science, [SDV]Life Sciences [q-bio], Human error, computer.software_genre, Biochemistry, 03 medical and health sciences, Software, macromolecular crystallography, Structural Biology, General Materials Science, automation, X-ray crystal centring, Focus (computing), Data collection, synchrotron instrumentation, business.industry, Robotics, General Chemistry, Condensed Matter Physics, Automation, Expert system, 030104 developmental biology, Beamline, Artificial intelligence, business, Software engineering, computer
Abstract: International audience; Automation is beginning to transform the way data are collected in almost all scientific disciplines. The combination of robotics and software now allows data to be collected consistently and reproducibly, eliminating human error and boredom. This approach has been applied to macromolecular crystallography at MASSIF-1, a fully automated beamline at the European Synchrotron Radiation Facility (ESRF). Considerable human effort is still dedicated to evaluating protein crystals in order to find the few crystals that diffract well or collecting hundreds of data sets to screen potential new drug candidates. The combination of ESRF-developed robotic sample handling and advanced software protocols now provides a new tool to structural biologists. Not only is the beamline used efficiently, running 24h a day without getting tired, data collection is also performed consistently by an expert system, often better than with a human operator. In this review, we will focus on the impact this level of automation has had on the optimum acquisition of data from crystals of biological macromolecules
Published: 2016
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10. Structural analysis of dsRNA binding to anti-viral pattern recognition receptors LGP2 and MDA5

Author: Denis Gerlier, Emiko Uchikawa, Stephen Cusack, Joanna Brunel, Hélène Malet, Mathilde Lethier, 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), European Molecular Biology Laboratory [Grenoble] (EMBL), Immunobiologie des infections virales – Immunobiology of Viral Infections (IbIV), Centre International de Recherche en Infectiologie - UMR (CIRI), 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)-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), Cusack, Stephen, Centre International de Recherche en Infectiologie (CIRI), École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-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)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF), Virologie et pathogenèse virale (VPV), Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), and Université de Lyon-Université de Lyon
Subjects: 0301 basic medicine, Models, Molecular, Interferon-Induced Helicase, IFIH1, [SDV.IMM] Life Sciences [q-bio]/Immunology, viruses, RNA-binding protein, Plasma protein binding, Transfection, [SDV.IMM.II]Life Sciences [q-bio]/Immunology/Innate immunity, Article, Cell Line, Avian Proteins, 03 medical and health sciences, Structure-Activity Relationship, Adenosine Triphosphate, Animals, Humans, Protein Interaction Domains and Motifs, Binding site, DEAD Box Protein 58, Molecular Biology, ComputingMilieux_MISCELLANEOUS, RNA, Double-Stranded, Adenosine Triphosphatases, Binding Sites, biology, Hydrolysis, LGP2, Helicase, RNA, RNA-Binding Proteins, MDA5, Cell Biology, Molecular biology, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], 3. Good health, Cell biology, Adenosine Diphosphate, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], 030104 developmental biology, Receptors, Pattern Recognition, biology.protein, Nucleic Acid Conformation, [SDV.IMM]Life Sciences [q-bio]/Immunology, Chickens, Protein Binding
Abstract: Summary RIG-I and MDA5 sense virus-derived short 5′ppp blunt-ended or long dsRNA, respectively, causing interferon production. Non-signaling LGP2 appears to positively and negatively regulate MDA5 and RIG-I signaling, respectively. Co-crystal structures of chicken (ch) LGP2 with dsRNA display a fully or semi-closed conformation depending on the presence or absence of nucleotide. LGP2 caps blunt, 3′ or 5′ overhang dsRNA ends with 1 bp longer overall footprint than RIG-I. Structures of 1:1 and 2:1 complexes of chMDA5 with short dsRNA reveal head-to-head packing rather than the polar head-to-tail orientation described for long filaments. chLGP2 and chMDA5 make filaments with a similar axial repeat, although less co-operatively for chLGP2. Overall, LGP2 resembles a chimera combining a MDA5-like helicase domain and RIG-I like CTD supporting both stem and end binding. Functionally, RNA binding is required for LGP2-mediated enhancement of MDA5 activation. We propose that LGP2 end-binding may promote nucleation of MDA5 oligomerization on dsRNA., Graphical Abstract, Highlights • chLPG2-dsRNA structures reveal RIG-I like end binding, but overhangs are possible • chMDA5-dsRNA complex structures show head-to-head packing on short dsRNAs • LGP2 also has MDA5-like behavior, coating dsRNA but with less cooperativity • Both human and chicken LGP2 enhance MDA5 signaling in an RNA-dependent manner, Uchikawa et al. reveal structural details of dsRNA recognition by MDA5 and LGP2 that synergistically sense viral RNA and activate interferon expression. LGP2 is primarily a dsRNA end binder but can also coat dsRNA, but less co-operatively than MDA5. Functional studies show that LGP2 enhancement of MDA5 signaling is RNA dependent.
Published: 2016
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11. PIWI Slicing and EXD1 Drive Biogenesis of Nuclear piRNAs from Cytosolic Targets of the Mouse piRNA Pathway

Author: Marie-Odile Fauvarque, David Homolka, Ramesh S. Pillai, Zhaolin Yang, Radha Raman Pandey, K.M. Chen, Andrew A. McCarthy, Ravi Sachidanandam, Bruno Kotska Rodino Janeiro, Michael Reuter, Laboratoire européen de biologie moléculaire - European Molecular Biology Laboratory (EMBL Grenoble), European Molecular Biology Laboratory [Grenoble] (EMBL), Department of Molecular Biology [Geneva, Switzerland], Université de Genève = University of Geneva (UNIGE), 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), Cold Spring Harbor Laboratory, Laboratoire de Biologie à Grande Échelle (BGE - UMR S1038), Institut National de la Santé et de la Recherche Médicale (INSERM)-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), Bertacchi Griffi, Nathalie, University of Geneva [Switzerland], Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF), Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])
Subjects: 0301 basic medicine, Exonucleases, Male, Models, Molecular, [SDV]Life Sciences [q-bio], Tdrd12, RNA-binding protein, Retrotransposon, piRNA, Cytosol, RNA interference, Drosophila Proteins, RNA Processing, Post-Transcriptional, RNA, Small Interfering, ComputingMilieux_MISCELLANEOUS, Genetics, RNA-Binding Proteins, Argonaute, Cell biology, [SDV] Life Sciences [q-bio], Drosophila melanogaster, Argonaute Proteins, Female, RNA Interference, endocrine system, Molecular Sequence Data, Piwi-interacting RNA, Mice, Transgenic, Inchworming, Biology, Article, 03 medical and health sciences, Slicer, RasiRNA, Animals, Protein Interaction Domains and Motifs, Exd1, Amino Acid Sequence, Molecular Biology, Cell Nucleus, PIWI, urogenital system, Cell Biology, Bombyx, 030104 developmental biology, Gene Expression Regulation, Cytoplasm, Multiprotein Complexes, Ping-pong, Carrier Proteins, Biogenesis
Abstract: Summary PIWI-interacting RNAs (piRNAs) guide PIWI proteins to suppress transposons in the cytoplasm and nucleus of animal germ cells, but how silencing in the two compartments is coordinated is not known. Here we demonstrate that endonucleolytic slicing of a transcript by the cytosolic mouse PIWI protein MILI acts as a trigger to initiate its further 5′→3′ processing into non-overlapping fragments. These fragments accumulate as new piRNAs within both cytosolic MILI and the nuclear MIWI2. We also identify Exonuclease domain-containing 1 (EXD1) as a partner of the MIWI2 piRNA biogenesis factor TDRD12. EXD1 homodimers are inactive as a nuclease but function as an RNA adaptor within a PET (PIWI-EXD1-Tdrd12) complex. Loss of Exd1 reduces sequences generated by MILI slicing, impacts biogenesis of MIWI2 piRNAs, and de-represses LINE1 retrotransposons. Thus, piRNA biogenesis triggered by PIWI slicing, and promoted by EXD1, ensures that the same guides instruct PIWI proteins in the nucleus and cytoplasm., Graphical Abstract, Highlights • MILI slicing triggers entry of target transcript into piRNA processing • Slicing generates non-overlapping, contiguous primary piRNAs in MILI and MIWI2 • EXD1 is a component of a PET (PIWI-EXD1-TDRD12) complex • RNA-binding protein EXD1 is critical for MIWI2 piRNA biogenesis, Yang et al. show how cytoplasmic slicing of a target transcript by a PIWI endonuclease triggers its conversion into piRNAs that are loaded into both cytosolic and nuclear PIWI proteins, ensuring coordinated silencing in the two compartments. The inactive nuclease EXD1 functions as an RNA-binding in this process.
Published: 2016
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12. Sequence Variation in Amplification Target Genes and Standards Influences Interlaboratory Comparison of BK Virus DNA Load Measurement

Author: Pilar Domingo-Calap, Bruno MOULIN, Valerie Giordanengo, Seiamak Bahram, Véronique Avettand-Fenoel, Sébastien Hantz, Samira Fafi-Kremer, Jerome LeGoff, ALEXANDRA DUCANCELLE, Bruno POZZETTO, Sophie Caillard, ERIC SOULIER, Peggy Perrin, Sylvie Pillet, Morgane Solis, Astrid VABRET, Immuno-Rhumatologie Moléculaire, Université de Strasbourg (UNISTRA)-Institut National de la Santé et de la Recherche Médicale (INSERM), Les Hôpitaux Universitaires de Strasbourg (HUS), French BKV Study Group, Laboratoire de Virologie [Hôpitaux universitaires de Strasbourg], Immunorhumathologie moléculaire, Institut National de la Santé et de la Recherche Médicale (INSERM), Virulence bactérienne précoce, Université de Strasbourg (EA7290), 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), CHU Strasbourg, Service de Néphrologie et Transplantation, Interaction virus-hôte et maladies du foie, Laboratoire Microorganismes : Génome et Environnement - Clermont Auvergne (LMGE), Université Clermont Auvergne (UCA)-Centre National de la Recherche Scientifique (CNRS), Virulence Bactérienne Précoce : fonctions cellulaires et contrôle de l'infection aigüe et subaigüe, Université de Strasbourg (UNISTRA), Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF), Laboratoire Microorganismes : Génome et Environnement (LMGE), Université Clermont Auvergne [2017-2020] (UCA [2017-2020])-Centre National de la Recherche Scientifique (CNRS), and BRICHEUX, Genevieve
Subjects: Microbiology (medical), Laboratory Proficiency Testing, viruses, 030230 surgery, Biology, medicine.disease_cause, [SDV.MHEP.UN]Life Sciences [q-bio]/Human health and pathology/Urology and Nephrology, Sensitivity and Specificity, Sciences du Vivant [q-bio]/Médecine humaine et pathologie, isolation & purification, 03 medical and health sciences, 0302 clinical medicine, [SDV.MHEP.MI]Life Sciences [q-bio]/Human health and pathology/Infectious diseases, Virology, Genetic variation, Genotype, medicine, Humans, Sequence variation, methods, standards, Gene, [SDV.MP.VIR] Life Sciences [q-bio]/Microbiology and Parasitology/Virology, Genetics, Sciences du Vivant [q-bio]/Ingénierie biomédicale, Polyomavirus Infections, 0303 health sciences, diagnosis, virology, 030306 microbiology, genetics, Genetic Variation, virus diseases, Viral Load, [SDV.MHEP.UN] Life Sciences [q-bio]/Human health and pathology/Urology and Nephrology, DNA extraction, Hospitals, 3. Good health, BK virus, BK Virus, DNA, Viral, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, France, Primer (molecular biology), Viral load
Abstract: International guidelines define a BK virus (BKV) load of ≥4 log 10 copies/ml as presumptive of BKV-associated nephropathy (BKVN) and a cutoff for therapeutic intervention. To investigate whether BKV DNA loads (BKVL) are comparable between laboratories, 2 panels of 15 and 8 clinical specimens (urine, whole blood, and plasma) harboring different BKV genotypes were distributed to 20 and 27 French hospital centers in 2013 and 2014, respectively. Although 68% of the reported results fell within the acceptable range of the expected result ±0.5 log 10 , the interlaboratory variation ranged from 1.32 to 5.55 log 10 . Polymorphisms specific to BKV genotypes II and IV, namely, the number and position of mutations in amplification target genes and/or deletion in standards, arose as major sources of interlaboratory disagreements. The diversity of DNA purification methods also contributed to the interlaboratory variability, in particular for urine samples. Our data strongly suggest that (i) commercial external quality controls for BKVL assessment should include all major BKV genotypes to allow a correct evaluation of BKV assays, and (ii) the BKV sequence of commercial standards should be provided to users to verify the absence of mismatches with the primers and probes of their BKV assays. Finally, the optimization of primer and probe design and standardization of DNA extraction methods may substantially decrease interlaboratory variability and allow interinstitutional studies to define a universal cutoff for presumptive BKVN and, ultimately, ensure adequate patient care.
Published: 2015
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13. Production and characterisation of Epstein–Barr virus helicase–primase complex and its accessory protein BBLF2/3

Author: Marlyse Buisson, Adam Round, Martha Brennich, Eric Thierry, Wim P. Burmeister, Stephanie Hutin, 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), European Synchrotron Radiation Facility (ESRF), European Molecular Biology Laboratory [Heidelberg] (EMBL), and Thomas, Frank
Subjects: Herpesvirus 4, Human, Insecta, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Genetic Vectors, medicine.disease_cause, Cell Line, Viral Proteins, Adenosine Triphosphate, Helicase–primase complex, Caudovirales, Virology, Scattering, Small Angle, Genetics, medicine, Animals, Molecular Biology, Polymerase, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], biology, DNA Helicases, DNA replication, Helicase, General Medicine, biology.organism_classification, Epstein–Barr virus, Molecular biology, Recombinant Proteins, Lytic cycle, Biochemistry, biology.protein, Primase, Baculoviridae
Abstract: International audience; The helicase-primase complex is part of the lytic DNA replication machinery of herpesviruses, but up to now, almost nothing is known about its structure. For Epstein-Barr virus it consists in the helicase BBLF4, the primase BSLF1 and the accessory protein BBLF2/3. The accessory protein shows only weak sequence homology within the herpesvirus family but may be related to an inactive B-family polymerase. BSLF1 belongs to the archaeo-eukaryotic primase family, whereas the helicase BBLF4 has been related either to Dda helicases of caudovirales or to Pif1 helicases. We produced the helicase-primase complex in insect cells using a baculovirus coding for all three proteins simultaneously. The soluble monomeric helicase-primase complex containing the three proteins with 1:1:1 stoichiometry showed ATPase activity, which is strongly stimulated in the presence of ssDNA oligomers. Furthermore, we expressed BBLF2/3 as soluble monomeric protein and performed small-angle X-ray scattering experiments which yielded an envelope whose shape is compatible with B-family polymerases.
Published: 2015
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14. Structural Insights into Bunyavirus Replication and Its Regulation by the vRNA Promoter

Author: Stephen Cusack, Hélène Malet, Piotr Gerlach, Juan Reguera, European Molecular Biology Laboratory [Heidelberg] (EMBL), 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), European Molecular Biology Laboratory [Grenoble] (EMBL), and Université Joseph Fourier - Grenoble 1 (UJF)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Centre National de la Recherche Scientifique (CNRS)
Subjects: Models, Molecular, Orthobunyavirus, viruses, MESH: Sequence Alignment, MESH: Catalytic Domain, MESH: Orthobunyavirus, RNA-dependent RNA polymerase, Sequence alignment, MESH: Amino Acid Sequence, Crystallography, X-Ray, Article, General Biochemistry, Genetics and Molecular Biology, 03 medical and health sciences, Catalytic Domain, MESH: Promoter Regions, Genetic, Amino Acid Sequence, Promoter Regions, Genetic, Peptide sequence, Polymerase, 030304 developmental biology, Genetics, 0303 health sciences, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], biology, 030306 microbiology, Biochemistry, Genetics and Molecular Biology(all), Ribonucleoprotein particle, RNA, RNA virus, MESH: Crystallography, X-Ray, RNA-Dependent RNA Polymerase, biology.organism_classification, 3. Good health, MESH: Ribonucleoproteins, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Ribonucleoproteins, MESH: RNA, Viral, biology.protein, RNA, Viral, MESH: RNA Replicase, Sequence Alignment, MESH: Models, Molecular
Abstract: Summary Segmented negative-strand RNA virus (sNSV) polymerases transcribe and replicate the viral RNA (vRNA) within a ribonucleoprotein particle (RNP). We present cryo-EM and X-ray structures of, respectively, apo- and vRNA bound La Crosse orthobunyavirus (LACV) polymerase that give atomic-resolution insight into how such RNPs perform RNA synthesis. The complementary 3′ and 5′ vRNA extremities are sequence specifically bound in separate sites on the polymerase. The 5′ end binds as a stem-loop, allosterically structuring functionally important polymerase active site loops. Identification of distinct template and product exit tunnels allows proposal of a detailed model for template-directed replication with minimal disruption to the circularised RNP. The similar overall architecture and vRNA binding of monomeric LACV to heterotrimeric influenza polymerase, despite high sequence divergence, suggests that all sNSV polymerases have a common evolutionary origin and mechanism of RNA synthesis. These results will aid development of replication inhibitors of diverse, serious human pathogenic viruses., Graphical Abstract, Highlights • Bunyavirus polymerase binds its complementary 3′ and 5′ vRNA ends in distinct sites • 5′ vRNA binding allosterically structures and activates the polymerase catalytic site • Distinct template/product exit tunnels explain RNA synthesis in a circularized RNP • Monomeric bunyavirus and trimeric influenza polymerases are structurally similar, The structure of the monomeric bunyavirus polymerase reveals that divergent segmented negative-strand RNA virus polymerases have a common overall architecture, explains how viral RNA binding allosterically regulates polymerase activity, and suggests a replication model that could apply to all related RNA viruses.
Published: 2015
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15. Ultradeep Pyrosequencing of NS3 To Predict Response to Triple Therapy with Protease Inhibitors in Previously Treated Chronic Hepatitis C Patients

Author: Olivier François, Katia Fusillier, Michael G. B. Blum, Jean-Pierre Zarski, Sylvie Larrat, Pauline Tremeaux, Patrice Morand, Réjane Beugnot, Alice Marlu, Jean-Baptiste Claude, Agnès Plages, Om Kulkarni, Anne Signori-Schmuck, Julien Lupo, Hervé Duborjal, Vincent Leroy, 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), Biologie Computationnelle et Mathématique (TIMC-IMAG-BCM), Techniques de l'Ingénierie Médicale et de la Complexité - Informatique, Mathématiques et Applications, Grenoble - UMR 5525 (TIMC-IMAG), VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF)-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF), Genostar, GENOME Express (GENOME Express), GENOME Express, Earth and Environmental, AMEC, Biochimie et Génétique Moléculaire, CHU Grenoble-Hôpital Michallon, Pôle Digidune, Pôle DigiDune, Unité de Biophysique, CRSSA, Département d'hépato-gastroentérologie, Université Grenoble Alpes (UGA)-CHU Grenoble, Matière et Systèmes Complexes (MSC (UMR_7057)), Centre National de la Recherche Scientifique (CNRS)-Université Paris Diderot - Paris 7 (UPD7), CHU Grenoble-Université Grenoble Alpes (UGA), Université Joseph Fourier - Grenoble 1 (UJF)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Centre National de la Recherche Scientifique (CNRS), Université Joseph Fourier - Grenoble 1 (UJF)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP)-IMAG-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Université Joseph Fourier - Grenoble 1 (UJF)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP)-IMAG-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA), Université Joseph Fourier - Grenoble 1 (UJF)-CHU Grenoble, and Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS)
Subjects: Adult, Male, Microbiology (medical), NS3, medicine.medical_specialty, Entropy, medicine.medical_treatment, Mutation, Missense, Salvage therapy, Viral quasispecies, Viral Nonstructural Proteins, Biology, Antiviral Agents, Peripheral blood mononuclear cell, Gastroenterology, Young Adult, 03 medical and health sciences, 0302 clinical medicine, Virology, Internal medicine, Drug Resistance, Viral, Genotype, medicine, Humans, Protease Inhibitors, Aged, 030304 developmental biology, Aged, 80 and over, Salvage Therapy, 0303 health sciences, Protease, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Genetic Variation, High-Throughput Nucleotide Sequencing, virus diseases, Odds ratio, Hepatitis C, Chronic, Middle Aged, Prognosis, digestive system diseases, Confidence interval, 3. Good health, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], HCV, Immunology, Drug Therapy, Combination, Female, 030211 gastroenterology & hepatology
Abstract: Despite the gain in sustained virological responses (SVR) provided by protease inhibitors (PIs), failures still occur. The aim of this study was to determine if a baseline analysis of the NS3 region using ultradeep pyrosequencing (UDPS) can help to predict an SVR. Serum samples from 40 patients with previously nonresponding genotype 1 chronic hepatitis C who were retreated with triple therapy, including a PI, were analyzed. Baseline UDPS of the NS3 gene was performed on plasma and peripheral blood mononuclear cells (PBMC). Mutations conferring resistance to PIs were sought. The overall diversity of the quasispecies was evaluated by calculating the Shannon entropy (SE). Resistance mutations were found in plasma and PBMC but were not discriminating enough to predict an SVR. NS3 quasispecies heterogeneity was significantly lower at baseline in patients achieving an SVR than in those not achieving an SVR (SE of 26.98 ±16.64 x10^3 versus 44.93 ±19.58 x10^3, P =0.0047). With multivariate analysis, the independent predictors of an SVR were fibrosis of stage F <2 (odds ratio [OR], 13.3; 95% confidence interval [CI], 1.25 to 141.096; P < 0.03) and SE below the median (OR, 5.4; 95% CI, 1.22 to 23.87; P < 0.03). More than the presence of minor mutations at the baseline in plasma or in PBMC, the NS3 viral heterogeneity determined by UDPS is an independent factor for an SVR in previously treated patients receiving triple therapy that includes a PI.&nbsp
Published: 2015
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16. A Fusion Intermediate gp41 Immunogen Elicits Neutralizing Antibodies to HIV-1

Author: Jonathan L. Heeney, Winfried Weissenhorn, Nancy Tumba, Lynn Morris, Hanna Dreja, David C. Montefiori, Jens Radzimanowski, Alexander Kliche, Susan W. Barnett, Ralf Wagner, Grégory Effantin, Rachel P. J. Lai, Miriam Hock, Celia C. LaBranche, Michael S. Seaman, David J. Seilly, David Lutje Hulsik, Paul Tonks, Thomas, Frank, Tonks, Paul [0000-0003-3512-5631], Heeney, Jonathan [0000-0003-2702-1621], Apollo - University of Cambridge Repository, Dept. of Computer Science and Technology, Peking University [Beijing], 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), Beth Israel Deaconess Medical Center, Beth Israel Deaconess Medical Center [Boston] (BIDMC), Harvard Medical School [Boston] (HMS)-Harvard Medical School [Boston] (HMS), 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), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)
Subjects: Immunogen, HIV-1 gp41, Protein Conformation, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], viruses, Proteolipids, Guinea Pigs, Molecular Sequence Data, Antibody Affinity, HIV Antibodies, Gp41, Membrane Fusion, Microbiology, Biochemistry, Epitope, Immunoglobulin G, Neutralization, Vaccine Development, Scattering, Small Angle, Animals, Humans, Amino Acid Sequence, Molecular Biology, chemistry.chemical_classification, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], biology, Immune Sera, virus diseases, Lipid bilayer fusion, Cell Biology, Antibodies, Neutralizing, Virology, Molecular biology, HIV Envelope Protein gp41, Protein Structure, Tertiary, 3. Good health, AIDS, Neutron Diffraction, chemistry, HIV-1, biology.protein, Female, Immunization, Antibody, Glycoprotein, X-ray Scattering
Abstract: International audience; The membrane-proximal external region (MPER) of the human immunodeficiency virus, type 1 (HIV-1) envelope glycoprotein subunit gp41 is targeted by potent broadly neutralizing antibodies 2F5, 4E10, and 10E8. These antibodies recognize linear epitopes and have been suggested to target the fusion intermediate conformation of gp41 that bridges viral and cellular membranes. Anti-MPER antibodies exert different degrees of membrane interaction, which is considered to be the limiting factor for the generation of such antibodies by immunization. Here we characterize a fusion intermediate conformation of gp41 (gp41int-Cys) and show that it folds into an elongated ∼12-nm-long extended structure based on small angle x-ray scattering data. Gp41int-Cys was covalently linked to liposomes via its C-terminal cysteine and used as immunogen. The gp41int-Cys proteoliposomes were administered alone or in prime-boost regimen with trimeric envelope gp140CA018 in guinea pigs and elicited high anti-gp41 IgG titers. The sera interacted with a peptide spanning the MPER region, demonstrated competition with broadly neutralizing antibodies 2F5 and 4E10, and exerted modest lipid binding, indicating the presence of MPER-specific antibodies. Although the neutralization potency generated solely by gp140CA018 was higher than that induced by gp41int-Cys, the majority of animals immunized with gp41int-Cys proteoliposomes induced modest breadth and potency in neutralizing tier 1 pseudoviruses and replication-competent simian/human immunodeficiency viruses in the TZM-bl assay as well as responses against tier 2 HIV-1 in the A3R5 neutralization assay. Our data thus demonstrate that liposomal gp41 MPER formulation can induce neutralization activity, and the strategy serves to improve breadth and potency of such antibodies by improved vaccination protocols.
Published: 2014
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17. Intrinsically disordered proteins implicated in paramyxoviral replication machinery

Author: Rob W. H. Ruigrok, Guillaume Communie, Malene Ringkjøbing Jensen, Martin Blackledge, 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), 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), 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), Université Joseph Fourier - Grenoble 1 (UJF)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Centre National de la Recherche Scientifique (CNRS), and Thomas, Frank
Subjects: Paramyxoviridae Infections, Protein Conformation, [SDV]Life Sciences [q-bio], Intrinsic protein, Computational biology, Biology, Virus Replication, Bioinformatics, Intrinsically disordered proteins, Replication (computing), Nucleoprotein, [SDV] Life Sciences [q-bio], Intrinsically Disordered Proteins, Viral Proteins, Order (biology), Virology, Paramyxoviridae, Animals, Humans, Function (biology)
Abstract: International audience; The development of mechanistic insight into the molecular basis of how intrinsically disordered proteins function is a key challenge for contemporary molecular biology. Intrinsic protein disorder is abundant in the replication machinery of paramyxoviruses. In order to study this kind of protein, new methods are required that specifically take account of the highly dynamic nature of the chain, and describe this disorder in quantitative terms. Here we review recent studies of conformational disorder in paramyxoviral phosphoproteins and nucleoproteins using solution-based approaches such as nuclear magnetic resonance.
Published: 2014
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18. Structural and Functional Studies on the Interaction of Adenovirus Fiber Knobs and Desmoglein 2

Author: Pascal Fender, Hua Cao, Liang Ran, Julie Gralow, Chloe Zubieta, Forrest Epstein, Maximilian Richter, Hongjie Wang, Roma Yumul, André Lieber, Xiaolong Fan, Thomas, Frank, Division of Medical Genetics [Seattle], University of Washington [Seattle], Unit for Virus Host-Cell Interactions [Grenoble] (UVHCI), and Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF)
Subjects: Protein Conformation, viruses, DNA Mutational Analysis, Mutant, Plasma protein binding, Crystallography, X-Ray, medicine.disease_cause, law.invention, MESH: Recombinant Proteins, MESH: Mutant Proteins, MESH: Protein Conformation, Protein structure, law, Protein Interaction Mapping, MESH: DNA Mutational Analysis, Receptor, MESH: Capsid Proteins, Desmoglein 2, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Recombinant Proteins, Host-Pathogen Interactions, Recombinant DNA, Receptors, Virus, Protein Binding, MESH: Adenoviruses, Human, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Viral protein, Immunology, Biology, Microbiology, Cell Line, Virology, medicine, Humans, MESH: Protein Binding, Protein Interaction Domains and Motifs, MESH: Protein Interaction Domains and Motifs, MESH: Humans, CD46, Adenoviruses, Human, Structure and Assembly, MESH: Protein Interaction Mapping, MESH: Host-Pathogen Interactions, MESH: Crystallography, X-Ray, MESH: Receptors, Virus, Molecular biology, MESH: Cell Line, Cell culture, Insect Science, Capsid Proteins, Mutant Proteins, MESH: Desmoglein 2
Abstract: Human adenovirus (Ad) serotypes Ad3, Ad7, Ad11, and Ad14, as well as a recently emerged strain of Ad14 (Ad14p1), use the epithelial junction protein desmoglein 2 (DSG2) as a receptor for infection. Unlike Ad interaction with CAR and CD46, structural details for Ad binding to DSG2 are still elusive. Using an approach based on Escherichia coli expression libraries of random Ad3 and Ad14p1 fiber knob mutants, we identified amino acid residues that, when mutated individually, ablated or reduced Ad knob binding to DSG2. These residues formed three clusters inside one groove at the extreme distal end of the fiber knob. The Ad3 fiber knob mutant library was also used to identify variants with increased affinity to DSG2. We found a number of mutations within or near the EF loop of the Ad3 knob that resulted in affinities to DSG2 that were several orders of magnitude higher than those to the wild-type Ad3 knob. Crystal structure analysis of one of the mutants showed that the introduced mutations make the EF loop more flexible, which might facilitate the interaction with DSG2. Our findings have practical relevance for cancer therapy. We have recently reported that an Ad3 fiber knob-containing recombinant protein (JO-1) is able to trigger opening of junctions between epithelial cancer cells which, in turn, greatly improved the intratumoral penetration and efficacy of therapeutic agents (I. Beyer, et al., Clin. Cancer Res. 18:3340–3351, 2012; I. Beyer, et al., Cancer Res. 71:7080–7090, 2011). Here, we show that affinity-enhanced versions of JO-1 are therapeutically more potent than the parental protein in a series of cancer models.
Published: 2013
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19. Natural non-homologous recombination led to the emergence of a duplicated V3-NS5A region in HCV-1b strains associated with hepatocellular carcinoma

Author: Françoise Stoll-Keller, Henia Saoudin, Arielle A. Rosenberg, Dominique Bettinger, Pascal Veillon, Odile Petsaris, Cécile Henquell, Bruno Pozzetto, Catherine Gaudy-Graffin, Jean-Baptiste Bour, Sophie Minjolle-Cha, Jacques Izopet, Christopher Payan, E. Bouthry, Alexandra Ducancelle, Adeline Pivert, Hélène Le Guillou-Guillemette, Gisèle Lagathu, Sylvie Larrat, P. André, Anne Signori-Schmuck, Sophie Vallet, Françoise Lunel-Fabiani, Jean-Michel Pawlotsky, Yazid Baazia, Elisabeth André-Garnier, Christophe Lemaire, Florence Abravanel, Pascale Trimoulet, Sophie Alain, Vincent Thibault, Hémodynamique, Interaction Fibrose et Invasivité tumorales Hépatiques (HIFIH), Université d'Angers (UA), Centre Hospitalier Universitaire d'Angers (CHU Angers), PRES Université Nantes Angers Le Mans (UNAM), CHU Clermont-Ferrand, Laboratoire Microorganismes : Génome et Environnement (LMGE), Université Clermont Auvergne [2017-2020] (UCA [2017-2020])-Centre National de la Recherche Scientifique (CNRS), Laboratoire Universitaire de Biodiversité et Ecologie Microbienne (LUBEM), Université de Brest (UBO), Service de Bactériologie, Virologie, Hygiène [CHU Limoges], CHU Limoges, Anti-infectieux : supports moléculaires des résistances et innovations thérapeutiques (RESINFIT), CHU Limoges-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Génomique, Environnement, Immunité, Santé, Thérapeutique (GEIST), Université de Limoges (UNILIM)-Université de Limoges (UNILIM), CHU Pontchaillou [Rennes], Centre de Physiopathologie Toulouse Purpan (CPTP), 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)-Centre National de la Recherche Scientifique (CNRS), Virologie de l'hépatite C (EA 4474), Université Paris Descartes - Paris 5 (UPD5), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Hôpital Cochin [AP-HP], Hôtel-Dieu de Nantes, Laboratoire de sérologie-virologie (CHU de Dijon), Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon), Service de bactériologie-Virologie-Hygiène [Avicenne], Université Paris 13 (UP13)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Avicenne [AP-HP], Service de virologie et d'immunologie biologique, CHU Bordeaux [Bordeaux]-Groupe hospitalier Pellegrin, Hôpital de la Croix-Rousse [CHU - HCL], Hospices Civils de Lyon (HCL), Morphogénèse et antigénicité du VIH et du virus des Hépatites (MAVIVH - U1259 Inserm - CHRU Tours ), Centre Hospitalier Régional Universitaire de Tours (CHRU Tours)-Université de Tours (UT)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre Hospitalier Régional Universitaire de Besançon (CHRU Besançon), Université Grenoble Alpes - UFR Pharmacie (UGA UFRP), Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-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), Groupe Immunité des Muqueuses et Agents Pathogènes (GIMAP), Université Jean Monnet - Saint-Étienne (UJM), Interaction virus-hôte et maladies du foie, Université de Strasbourg (UNISTRA)-Institut National de la Santé et de la Recherche Médicale (INSERM), 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), Institut de Recherche en Horticulture et Semences (IRHS), Université d'Angers (UA)-Institut National de la Recherche Agronomique (INRA)-AGROCAMPUS OUEST, Laboratoire Microorganismes : Génome et Environnement - Clermont Auvergne (LMGE), Université Clermont Auvergne (UCA)-Centre National de la Recherche Scientifique (CNRS), Centre de Physiopathologie Toulouse Purpan ex IFR 30 et IFR 150 (CPTP), 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-Centre National de la Recherche Scientifique (CNRS), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), CHU Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Université Paris 13 (UP13)-Hôpital Avicenne, Morphogénèse et antigénicité du VIH et du virus des Hépatites (MAVIVH), Centre Hospitalier Régional Universitaire de Tours (CHRU Tours)-Université de Tours-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre Hospitalier Régional Universitaire [Besançon] (CHRU Besançon), Université Grenoble Alpes (UGA), Institut de biologie structurale (IBS - UMR 5075), 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)-Université Grenoble Alpes (UGA)-Centre National de la Recherche Scientifique (CNRS), 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), Université Jean Monnet [Saint-Étienne] (UJM), Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Génomique, Environnement, Immunité, Santé, Thérapeutique (GEIST), Université de Limoges (UNILIM)-Université de Limoges (UNILIM)-CHU Limoges, 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 de Tours-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre Hospitalier Régional Universitaire de Tours (CHRU TOURS), 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)-Centre National de la Recherche Scientifique (CNRS), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12)-IFR10, AGROCAMPUS OUEST, Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut National de la Recherche Agronomique (INRA)-Université d'Angers (UA), Université Paris 13 (UP13)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Hôpital Avicenne, Université de Tours-Centre Hospitalier Régional Universitaire de Tours (CHRU Tours)-Institut National de la Santé et de la Recherche Médicale (INSERM), and AGROCAMPUS OUEST-Institut National de la Recherche Agronomique (INRA)-Université d'Angers (UA)
Subjects: RNA viruses, 0301 basic medicine, Mutant, lcsh:Medicine, Hepacivirus, Viral Nonstructural Proteins, Biochemistry, Cohort Studies, Database and Informatics Methods, Viral Envelope Proteins, Gene Duplication, Gene duplication, Evolutionary Emergence, lcsh:Science, Homologous Recombination, Pathology and laboratory medicine, Phylogeny, Data Management, Recombination, Genetic, Genetics, Multidisciplinary, Hepatitis C virus, Liver Neoplasms, Phylogenetic Analysis, Medical microbiology, Hepatitis C, 3. Good health, Nucleic acids, Phylogenetics, Viruses, Host-Pathogen Interactions, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, Pathogens, Sequence Analysis, Recombination, Research Article, Computer and Information Sciences, Evolutionary Processes, Carcinoma, Hepatocellular, DNA recombination, Bioinformatics, 030106 microbiology, Viral quasispecies, Biology, Research and Analysis Methods, Microbiology, Viral Evolution, Evolution, Molecular, 03 medical and health sciences, Virology, Humans, Evolutionary Systematics, Gene, DNA sequence analysis, Taxonomy, Medicine and health sciences, Cloning, Evolutionary Biology, Polymorphism, Genetic, Biology and life sciences, Flaviviruses, lcsh:R, Organisms, Viral pathogens, RNA, Bayes Theorem, DNA, Hepatitis viruses, Organismal Evolution, Microbial pathogens, 030104 developmental biology, Genetic Loci, Microbial Evolution, Mutation, lcsh:Q, Homologous recombination, Sequence Alignment
Abstract: International audience; Background: The emergence of new strains in RNA viruses is mainly due to mutations or intra and inter-genotype homologous recombination. Non-homologous recombinations may be deleterious and are rarely detected. In previous studies, we identified HCV-1b strains bearing two tandemly repeated V3 regions in the NS5A gene without ORF disruption. This polymorphism may be associated with an unfavorable course of liver disease and possibly involved in liver carcinogenesis. Here we aimed at characterizing the origin of these mutant strains and identifying the evolutionary mechanism on which the V3 duplication relies.Methods: Direct sequencing of the entire NS5A and E1 genes was performed on 27 mutant strains. Quasispecies analyses in consecutive samples were also performed by cloning and sequencing the NS5A gene for all mutant and wild strains. We analyzed the mutant and wild-type sequence polymorphisms using Bayesian methods to infer the evolutionary history of and the molecular mechanism leading to the duplication-like event.Results: Quasispecies were entirely composed of exclusively mutant or wild-type strains respectively. Mutant quasispecies were found to have been present since contamination and had persisted for at least 10 years. This V3 duplication-like event appears to have resulted from non-homologous recombination between HCV-1b wild-type strains around 100 years ago. The association between increased liver disease severity and these HCV-1b mutants may explain their persistence in chronically infected patients.Conclusions: These results emphasize the possible consequences of non-homologous recombination in the emergence and severity of new viral diseases.
Published: 2017
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20. Structure and function of an insect α-carboxylesterase (αEsterase7) associated with insecticide resistance

Author: Paul D. Carr, Faisal Younus, Jian-Wei Liu, Robyn J. Russell, Chris M. Coppin, Tamara Meirelles, John G. Oakeshott, David L. Ollis, Mathilde Lethier, Martin Weik, Gunjan Pandey, Colin J. Jackson, Research School of Chemistry, Australian National University (ANU), 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), CSIRO - Land & Water National Research Flagship, 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), Commonwealth Scientific and Industrial Research Organisation [Canberra] (CSIRO), 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), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)
Subjects: 0106 biological sciences, Insecticides, Drug Resistance, MESH: Protein Structure, Secondary, MESH: Catalytic Domain, Genes, Insect, MESH: Genes, Insect, Crystallography, X-Ray, 01 natural sciences, Protein Structure, Secondary, Carboxylesterase, Substrate Specificity, chemistry.chemical_compound, Catalytic Domain, MESH: Animals, MESH: Carboxylesterase, Phosphorylation, directed evolution, MESH: Diptera, chemistry.chemical_classification, 0303 health sciences, Multidisciplinary, biology, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], MESH: Sheep Diseases, Organophosphate, Biological Sciences, Directed evolution, Acetylcholinesterase, Biochemistry, Lucilia cuprina, MESH: Drug Resistance, ali-esterase, MESH: Australia, Sheep Diseases, MESH: Sheep, 03 medical and health sciences, Animals, 030304 developmental biology, Enzyme substrate complex, Sheep, MESH: Phosphorylation, Diptera, Australia, Active site, protein engineering, MESH: Acetylcholinesterase, biology.organism_classification, MESH: Crystallography, X-Ray, MESH: Insecticides, 010602 entomology, Enzyme, chemistry, biology.protein, MESH: Substrate Specificity
Abstract: Insect carboxylesterases from the α Esterase gene cluster, such as αE7 (also known as E3) from the Australian sheep blowfly Lucilia cuprina ( Lc αE7), play an important physiological role in lipid metabolism and are implicated in the detoxification of organophosphate (OP) insecticides. Despite the importance of OPs to agriculture and the spread of insect-borne diseases, the molecular basis for the ability of α-carboxylesterases to confer OP resistance to insects is poorly understood. In this work, we used laboratory evolution to increase the thermal stability of Lc αE7, allowing its overexpression in Escherichia coli and structure determination. The crystal structure reveals a canonical α/β-hydrolase fold that is very similar to the primary target of OPs (acetylcholinesterase) and a unique N-terminal α-helix that serves as a membrane anchor. Soaking of Lc αE7 crystals in OPs led to the capture of a crystallographic snapshot of Lc αE7 in its phosphorylated state, which allowed comparison with acetylcholinesterase and rationalization of its ability to protect insects against the effects of OPs. Finally, inspection of the active site of Lc αE7 reveals an asymmetric and hydrophobic substrate binding cavity that is well-suited to fatty acid methyl esters, which are hydrolyzed by the enzyme with specificity constants (∼10 6 M −1 s −1 ) indicative of a natural substrate.
Published: 2013
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21. Misidentification of a Hepatitis C Virus (HCV) recombinant form leading to treatment failure with new direct acting antivirals

Author: Decroos, Amandine, de Oliveira, Fabienne, Leoz, Marie, Larrat, Sylvie, Riachi, Ghassan, Mourez, Thomas, Groupe de Recherche sur l'Adaptation Microbienne (GRAM 2.0), Université de Rouen Normandie (UNIROUEN), Normandie Université (NU)-Normandie Université (NU)-Université de Caen Normandie (UNICAEN), Normandie Université (NU), 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), Service d'Hépato-Gastroentérologie [CHU Rouen], Hôpital Charles Nicolle [Rouen]-CHU Rouen, Normandie Université (NU)-Normandie Université (NU)-Université de Rouen Normandie (UNIROUEN), and Mzembaba, Sandy
Subjects: [SDV.MP.VIR] Life Sciences [q-bio]/Microbiology and Parasitology/Virology, [SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, [SDV.MP] Life Sciences [q-bio]/Microbiology and Parasitology, ComputingMilieux_MISCELLANEOUS
Abstract: International audience
Published: 2016

22. Fusion to a homo-oligomeric scaffold allows cryo-EM analysis of a small protein

Author: Francesca Coscia, Hélène Malet, Guy Schoehn, Leandro F. Estrozi, Marjolaine Noirclerc-Savoye, Fabienne Hans, Carlo Petosa, 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), 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), 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 Joseph Fourier - Grenoble 1 (UJF)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Centre National de la Recherche Scientifique (CNRS)
Subjects: 0301 basic medicine, Scaffold protein, Scaffold, Cryo-electron microscopy, Recombinant Fusion Proteins, [SDV]Life Sciences [q-bio], macromolecular substances, Biology, Maltose-Binding Proteins, Article, 03 medical and health sciences, chemistry.chemical_compound, 0302 clinical medicine, Glutamate-Ammonia Ligase, chemistry.chemical_classification, DNA ligase, Multidisciplinary, Resolution (electron density), Cryoelectron Microscopy, 030104 developmental biology, Dodecameric protein, Monomer, chemistry, Biochemistry, Biophysics, Target protein, Protein Multimerization, 030217 neurology & neurosurgery
Abstract: Recent technical advances have revolutionized the field of cryo-electron microscopy (cryo-EM). However, most monomeric proteins remain too small (
Published: 2016
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23. The Myb domain of LUX ARRHYTHMO in complex with DNA: expression, purification and crystallization

Author: Chloe Zubieta, Catarina S. Silva, Xuelei Lai, Max H. Nanao, Physiologie cellulaire et végétale (LPCV), Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-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), European Synchrotron Radiation Facility (ESRF), 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), ATIP-Avenir, European Project: 283570,EC:FP7:INFRA,FP7-INFRASTRUCTURES-2011-1,BIOSTRUCT-X(2011), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF), and Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG)
Subjects: 0301 basic medicine, animal structures, Arabidopsis thaliana, Circadian clock, Biophysics, Arabidopsis, Protein-DNA complex, Electrophoretic Mobility Shift Assay, Crystallography, X-Ray, Genes, Plant, Biochemistry, Myb domain, Research Communications, 03 medical and health sciences, chemistry.chemical_compound, Proto-Oncogene Proteins c-myb, Structural Biology, Genetics, Electrophoretic mobility shift assay, MYB, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, DNA binding, LUX ARRHYTHMO, Transcription factor, 030102 biochemistry & molecular biology, biology, Arabidopsis Proteins, fungi, food and beverages, DNA, Plant, Condensed Matter Physics, biology.organism_classification, Molecular biology, Cell biology, protein–DNA complex, 030104 developmental biology, chemistry, sense organs, Crystallization
Abstract: LUX ARRHYTHMO (LUX) is a Myb-domain transcription factor that plays an important role in regulating the circadian clock.Luxmutations cause severe clock defects and arrhythmia in constant light and dark. In order to examine the molecular mechanisms underlying the function of LUX, the DNA-binding Myb domain was cloned, expressed and purified. The DNA-binding activity of the Myb domain was confirmed using electrophoretic mobility shift assays (EMSAs), demonstrating that the LUX Myb domain is able to bind to DNA with nanomolar affinity. In order to investigate the specificity determinants of protein–DNA interactions, the protein was co-crystallized with a 10-mer cognate DNA. Initial crystallization results for the selenomethionine-derivatized protein and data-set collection statistics are reported. Data collection was performed using theMeshAndCollectworkflow available at the ESRF.
Published: 2016
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24. Protein Surface and Core Dynamics Show Concerted Hydration-Dependent Activation

Author: Wood, Kathleen, Gallat, François-Xavier, Otten, Renee, van Heel, Auke J, Lethier, Mathilde, van Eijck, Lambert, Moulin, Martin, Haertlein, Michael, Weik, Martin, Mulder, Frans a A, Van heel, Auke J., Van eijck, Lambert, Mulder, Frans A. A., Australian Nuclear Science and Technology Organisation (ANSTO), Australian Nuclear Science and Technology Organisation, Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-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), 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 Laue-Langevin (ILL), Groningen University, Gröningen, 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)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF), ILL, and Groningen Biomolecular Sciences and Biotechnology
Subjects: Models, Molecular, Neutron scattering, 01 natural sciences, MYOGLOBIN, Isotopic labeling, chemistry.chemical_compound, Molecular dynamics, Side chain, MESH: Proteins, ENZYME-ACTIVITY, Physics::Chemical Physics, TEMPERATURE, Physics::Biological Physics, Quantitative Biology::Biomolecules, 0303 health sciences, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], biology, BACTERIORHODOPSIN, Quantitative Biology::Molecular Networks, General Medicine, Myoglobin, Chemical physics, THERMAL MOTIONS, INELASTIC NEUTRON-SCATTERING, protein dynamical transitions, NMR neutron scattering protein dynamics, Glass transition, hydration, MESH: Models, Molecular, Surface Properties, GLASS-TRANSITION, 010402 general chemistry, Catalysis, 03 medical and health sciences, MESH: Water, isotopic labeling, 030304 developmental biology, MESH: Surface Properties, Proteins, Water, Bacteriorhodopsin, General Chemistry, Nanosecond, 0104 chemical sciences, neutron spectroscopy, Crystallography, LYSOZYME, chemistry, MOLECULAR-DYNAMICS, biological sciences, biology.protein
Abstract: International audience; By specifically labeling leucine/valine methyl groups and lysine side chains "inside" and "outside" dynamics of proteins on the nanosecond timescale are compared using neutron scattering. Surprisingly, both groups display similar dynamics as a function of temperature, and the buried hydrophobic core is sensitive to hydration and undergoes a dynamical transition.
Published: 2012
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25. CC2D1A Is a Regulator of ESCRT-III CHMP4B

Author: Heinrich G. Göttlinger, Charles Sabin, Bettina Hartlieb, Yoshiko Usami, Nolwenn Miguet, Sergiy V. Avilov, Winfried Weissenhorn, Nicolas Martinelli, Aurelien Dordor, Euripedes A. Ribeiro, 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), 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 Thomas, Frank
Subjects: Models, Molecular, Regulator, MESH: Protein Structure, Secondary, Plasma protein binding, Protein Structure, Secondary, MESH: Endosomal Sorting Complexes Required for Transport, MESH: HIV-1, MESH: Protein Structure, Tertiary, 0302 clinical medicine, Protein structure, Membrane fission, Structural Biology, Cell Line, Transformed, 0303 health sciences, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], 3. Good health, Transport protein, DNA-Binding Proteins, Protein Transport, Biochemistry, MESH: Repressor Proteins, MESH: HEK293 Cells, MESH: Models, Molecular, Protein Binding, MESH: Protein Transport, MESH: Mutation, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Endosome, Endosomes, Biology, Article, ESCRT, 03 medical and health sciences, Humans, MESH: Protein Binding, MESH: Cell Line, Transformed, Binding site, Molecular Biology, 030304 developmental biology, Binding Sites, MESH: Humans, Endosomal Sorting Complexes Required for Transport, Protein Structure, Tertiary, Repressor Proteins, HEK293 Cells, MESH: Binding Sites, MESH: Endosomes, Mutation, HIV-1, Biophysics, MESH: DNA-Binding Proteins, 030217 neurology & neurosurgery
Abstract: International audience; Endosomal sorting complexes required for transport (ESCRTs) regulate diverse processes ranging from receptor sorting at endosomes to distinct steps in cell division and budding of some enveloped viruses. Common to all processes is the membrane recruitment of ESCRT-III that leads to membrane fission. Here, we show that CC2D1A is a novel regulator of ESCRT-III CHMP4B function. We demonstrate that CHMP4B interacts directly with CC2D1A and CC2D1B with nanomolar affinity by forming a 1:1 complex. Deletion mapping revealed a minimal CC2D1A-CHMP4B binding construct, which includes a short linear sequence within the third DM14 domain of CC2D1A. The CC2D1A binding site on CHMP4B was mapped to the N-terminal helical hairpin. Based on a crystal structure of the CHMP4B helical hairpin, two surface patches were identified that interfere with CC2D1A interaction as determined by surface plasmon resonance. Introducing these mutations into a C-terminal truncation of CHMP4B that exerts a potent dominant negative effect on human immunodeficiency virus type 1 budding revealed that one of the mutants lost this effect completely. This suggests that the identified CC2D1A binding surface might be required for CHMP4B polymerization, which is consistent with the finding that CC2D1A binding to CHMP4B prevents CHMP4B polymerization in vitro. Thus, CC2D1A might act as a negative regulator of CHMP4B function.
Published: 2012
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26. Architecture of a Dodecameric Bacterial Replicative Helicase

Author: Ulrike Kapp, Meike Stelter, Alexandre Bazin, Marc Jamin, Goran Bajic, Gael Goret, Laurent Terradot, Irina Gutsche, Joanna Timmins, Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-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), Laboratoire de Virologie Moléculaire et Structurale, Centre National de la Recherche Scientifique (CNRS), 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 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), 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)-Centre National de la Recherche Scientifique (CNRS), and Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF)
Subjects: Models, Molecular, Protein Conformation, [SDV]Life Sciences [q-bio], Replication Origin, MESH: Microscopy, Electron, medicine.disease_cause, Polymerization, 03 medical and health sciences, Protein structure, MESH: Protein Conformation, Structural Biology, medicine, Escherichia coli, MESH: Replication Origin, MESH: Protein Binding, Molecular Biology, dnaB helicase, 030304 developmental biology, Genetics, 0303 health sciences, MESH: DnaB Helicases, biology, Helicobacter pylori, MESH: Escherichia coli, 030302 biochemistry & molecular biology, DNA replication, Helicase, RNA Helicase A, DnaA, Cell biology, Microscopy, Electron, MESH: Polymerization, biology.protein, bacteria, MESH: Helicobacter pylori, dnaC, DnaB Helicases, MESH: Models, Molecular, Protein Binding
Abstract: International audience; Hexameric DnaB helicases are often loaded at DNA replication forks by interacting with the initiator protein DnaA and/or a helicase loader (DnaC in Escherichia coli). These loaders are not universally required, and DnaB from Helicobacter pylori was found to bypass DnaC when expressed in E. coli cells. The crystal structure of Helicobacter pylori DnaB C-terminal domain (HpDnaB-CTD) reveals a large two-helix insertion (named HPI) in the ATPase domain that protrudes away from the RecA fold. Biophysical characterization and electron microscopy (EM) analysis of the full-length protein show that HpDnaB forms head-to-head double hexamers remarkably similar to helicases found in some eukaryotes, archaea, and viruses. The docking of the HpDnaB-CTD structure into EM reconstruction of HpDnaB provides a model that shows how hexamerization of the CTD is facilitated by HPI-HPI interactions. The HpDnaB double-hexamer architecture supports an alternative strategy to load bacterial helicases onto forks in the absence of helicase loaders.
Published: 2012
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27. Extracellular Complexes of the Hematopoietic Human and Mouse CSF-1 Receptor Are Driven by Common Assembly Principles

Author: Alexander V. Shkumatov, Bjorn Vergauwen, Jonathan Elegheert, Kenneth Verstraete, Irina Gutsche, Xiongwu Wu, Ambroise Desfosses, Nathalie Bracke, Kathleen Van Craenenbroeck, Dmitri I. Svergun, Savvas N. Savvides, Bernard R. Brooks, Unit Struct Biol, Lab Prot Biochem & Biomol Engn L ProBE, Structural and Computational Biology Unit, European Molecular Biology Laboratory [Grenoble] (EMBL), 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), and Thomas, Frank
Subjects: MESH: Receptor, Macrophage Colony-Stimulating Factor, Macrophage colony-stimulating factor, MESH: Macrophage Colony-Stimulating Factor, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Hematopoietic System, Receptor, Macrophage Colony-Stimulating Factor, Stem cell factor, MESH: Microscopy, Electron, Biology, Ligands, Article, Mice, 03 medical and health sciences, 0302 clinical medicine, Structural Biology, MESH: Ligands, Extracellular, Animals, Humans, MESH: Animals, MESH: Hematopoietic System, Binding site, Cytokine binding, Receptor, MESH: Mice, Molecular Biology, Ternary complex, 030304 developmental biology, 0303 health sciences, Binding Sites, MESH: Humans, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Macrophage Colony-Stimulating Factor, Cell biology, Microscopy, Electron, MESH: Binding Sites, Thermodynamics, MESH: Thermodynamics, Tyrosine kinase, 030217 neurology & neurosurgery
Abstract: International audience; The hematopoietic colony stimulating factor-1 receptor (CSF-1R or FMS) is essential for the cellular repertoire of the mammalian immune system. Here, we report a structural and mechanistic consensus for the assembly of human and mouse CSF-1:CSF-1R complexes. The EM structure of the complete extracellular assembly of the human CSF-1:CSF-1R complex reveals how receptor dimerization by CSF-1 invokes a ternary complex featuring extensive homotypic receptor contacts and striking structural plasticity at the extremities of the complex. Studies by small-angle X-ray scattering of unliganded hCSF-1R point to large domain rearrangements upon CSF-1 binding, and provide structural evidence for the relevance of receptor predimerization at the cell surface. Comparative structural and binding studies aiming to dissect the assembly principles of human and mouse CSF-1R complexes, including a quantification of the CSF-1/CSF-1R species cross-reactivity, show that bivalent cytokine binding to receptor coupled to ensuing receptor-receptor interactions are common denominators in extracellular complex formation.
Published: 2011
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28. Radiation-damage-induced phasing: a case study using UV irradiation with light-emitting diodes

Author: Chloe Zubieta, Max H. Nanao, Hugo Caserotto, Franck Felisaz, Daniele de Sanctis, European Synchrotron Radiation Facility (ESRF), Laboratoire de physiologie cellulaire végétale (LPCV), Université Joseph Fourier - Grenoble 1 (UJF)-Institut National de la Recherche Agronomique (INRA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), European Molecular Biology Laboratory, 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), Physiologie cellulaire et végétale (LPCV), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA)-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), European Molecular Biology Laboratory [Grenoble] (EMBL), Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF), Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG)
Subjects: Models, Molecular, 0301 basic medicine, 030103 biophysics, Protein Conformation, Ultraviolet Rays, [SDV]Life Sciences [q-bio], Quantitative Biology::Tissues and Organs, light-emitting diodes, ultraviolet light, Nanotechnology, disulfide bonds, Crystallography, X-Ray, medicine.disease_cause, law.invention, 03 medical and health sciences, Structural Biology, law, Ultraviolet light, Radiation damage, medicine, Insulin, Disulfides, Irradiation, Diode, phasing, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], business.industry, Chemistry, Equipment Design, Laser, Research Papers, UV, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], 030104 developmental biology, RIP, radiation damage, SIR, Optoelectronics, Crystallization, business, Ultraviolet, Light-emitting diode, Visible spectrum
Abstract: A case study of radiation-damage-induced phasing is discussed using ultraviolet light-emitting diodes to induce specific radiation damage., Exposure to X-rays, high-intensity visible light or ultraviolet radiation results in alterations to protein structure such as the breakage of disulfide bonds, the loss of electron density at electron-rich centres and the movement of side chains. These specific changes can be exploited in order to obtain phase information. Here, a case study using insulin to illustrate each step of the radiation-damage-induced phasing (RIP) method is presented. Unlike a traditional X-ray-induced damage step, specific damage is introduced via ultraviolet light-emitting diodes (UV-LEDs). In contrast to UV lasers, UV-LEDs have the advantages of small size, low cost and relative ease of use.
Published: 2016
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29. Cryo-electron Microscopy Structure of the Native Prototype Foamy Virus Glycoprotein and Virus Architecture

Author: Winfried Weissenhorn, Nick Aschman, Patricia Renesto, Guy Schoehn, Grégory Effantin, Dirk Lindemann, Leandro F. Estrozi, Nicole Stanke, Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-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), 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), Thérapeutique Recombinante Expérimentale (TIMC-IMAG-TheREx), Techniques de l'Ingénierie Médicale et de la Complexité - Informatique, Mathématiques et Applications, Grenoble - UMR 5525 (TIMC-IMAG), Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Institute of Virology [Dresden], Technische Universität Dresden = Dresden University of Technology (TU Dresden), ANR-10-INBS-0005,FRISBI,Infrastructure Française pour la Biologie Structurale Intégrée(2010), ANR-10-LABX-0049,GRAL,Grenoble Alliance for Integrated Structural Cell Biology(2010), ANR-14-CE09-0003,ESCRTfission,Mécanique de fission membranaire induite par ESCRT-III(2014), 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)-Centre National de la Recherche Scientifique (CNRS), and Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF)
Subjects: 0301 basic medicine, Cryo-electron microscopy, Protein Conformation, viruses, Cell Membranes, Glycobiology, Cytoplasmic part, Biochemistry, Membrane Fusion, Viral Packaging, Virions, Protein structure, Image Processing, Computer-Assisted, Electron Microscopy, Biology (General), Spumavirus, Microscopy, biology, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Transmembrane domain, Capsid, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, Cellular Structures and Organelles, Research Article, Signal peptide, QH301-705.5, Immunology, Blotting, Western, Viral Structure, Transfection, Research and Analysis Methods, Microbiology, Cell Line, 03 medical and health sciences, Virology, Genetics, Humans, Molecular Biology, Glycoproteins, Virus Glycoproteins, Cryoelectron Microscopy, Gene Products, env, Biology and Life Sciences, Electron Cryo-Microscopy, Cell Biology, Viral membrane, RC581-607, biology.organism_classification, Molecular biology, [SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, Viral Replication, 030104 developmental biology, Biophysics, Parasitology, Immunologic diseases. Allergy
Abstract: Foamy viruses (FV) belong to the genus Spumavirus, which forms a distinct lineage in the Retroviridae family. Although the infection in natural hosts and zoonotic transmission to humans is asymptomatic, FVs can replicate well in human cells making it an attractive gene therapy vector candidate. Here we present cryo-electron microscopy and (cryo-)electron tomography ultrastructural data on purified prototype FV (PFV) and PFV infected cells. Mature PFV particles have a distinct morphology with a capsid of constant dimension as well as a less ordered shell of density between the capsid and the membrane likely formed by the Gag N-terminal domain and the cytoplasmic part of the Env leader peptide gp18LP. The viral membrane contains trimeric Env glycoproteins partly arranged in interlocked hexagonal assemblies. In situ 3D reconstruction by subtomogram averaging of wild type Env and of a Env gp48TM- gp80SU cleavage site mutant showed a similar spike architecture as well as stabilization of the hexagonal lattice by clear connections between lower densities of neighboring trimers. Cryo-EM was employed to obtain a 9 Å resolution map of the glycoprotein in its pre-fusion state, which revealed extensive trimer interactions by the receptor binding subunit gp80SU at the top of the spike and three central helices derived from the fusion protein subunit gp48TM. The lower part of Env, presumably composed of interlaced parts of gp48TM, gp80SU and gp18LP anchors the spike at the membrane. We propose that the gp48TM density continues into three central transmembrane helices, which interact with three outer transmembrane helices derived from gp18LP. Our ultrastructural data and 9 Å resolution glycoprotein structure provide important new insights into the molecular architecture of PFV and its distinct evolutionary relationship with other members of the Retroviridae., Author Summary Foamy viruses (FVs), which belong to the retroviral genus Spumavirus, are endemic to non-human primates and can be transmitted to humans. They are considered as potential vectors for gene therapy due to their broad cell tropism and their apparent apathogenicity in natural hosts and humans. In order to gain more insight into the ultrastructure of the prototype FV (PFV) we performed (cryo-)electron tomography and microscopy of infected cells and of isolated virions. We find that PFV contains a nucleocapsid of constant dimensions at its center, an intermediate shell of protein positioned between the core capsid and the viral membrane and glycoprotein that arranges into regular hexagonal lattices on the virus membrane. Structural analysis of the glycoprotein was performed in situ to a resolution of 9Å, which shows regular helical features such as a trimeric coiled coil of the fusion protein subunit, a hallmark of class I fusion proteins, spacer arms between the glycoprotein trimers and the arrangement of six transmembrane helices, a characteristic feature of the PFV Env glycoprotein. We discuss our results in light of the evolutionary relationship of PFV with other retroviruses as well as the role of the unique glycoprotein architecture on the virus life cycle.
Published: 2016
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30. Structural insights and membrane binding properties of MGD1, the major galactolipid synthase in plants

Author: Magali Audry, Christelle Breton, Agnès Girard-Egrot, Joana Rocha, Joe Sarkis, Monica M. Palcic, Maryse A. Block, Laurence Pitou, Daniele de Sanctis, J. Radzimanowski, Valérie Chazalet, Eric Maréchal, Aline Thomas, Centre de Recherches sur les Macromolécules Végétales (CERMAV ), Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Génie Enzymatique, Membrane Biomimétique et Assemblages Supramoléculaires (GEMBAS), Institut de Chimie et Biochimie Moléculaires et Supramoléculaires (ICBMS), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-École Supérieure Chimie Physique Électronique de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-École Supérieure Chimie Physique Électronique de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), 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), European Synchrotron Radiation Facility (ESRF), Carlsberg Laboratory, Carlsberg Group, Carlsberg Laboratory-Carlsberg Laboratory, Physiologie cellulaire et végétale (LPCV), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA)-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), Région Rhône-Alpes, ANR-10-BLAN-1524,ReGal,Régulation moléculaire de la biosynthèse des Galactolipides chloroplastiques(2010), Université de Lyon-Université de Lyon-École Supérieure de Chimie Physique Électronique de Lyon (CPE)-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-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), Carlsberg Fondation = Carlsbergfondet [Copenhague], Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Centre de Recherches sur les Macromolécules Végétales (CERMAV), Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-École Supérieure Chimie Physique Électronique de Lyon-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-École Supérieure Chimie Physique Électronique de Lyon-Centre National de la Recherche Scientifique (CNRS), Laboratoire de physiologie cellulaire végétale (LPCV), and Université Joseph Fourier - Grenoble 1 (UJF)-Institut National de la Recherche Agronomique (INRA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)
Subjects: 0106 biological sciences, 0301 basic medicine, Models, Molecular, Arabidopsis thaliana, Arabidopsis, Plant Science, Crystallography, X-Ray, 01 natural sciences, Chloroplast membrane, Thylakoids, Protein Structure, Secondary, MGDG, X-Ray Diffraction, Catalytic Domain, Langmuir monolayer, ComputingMilieux_MISCELLANEOUS, ATP synthase, biology, At4g31780, Galactosyltransferases, Lipids, Chloroplast, Membrane, Biochemistry, Thylakoid, Electrophoresis, Polyacrylamide Gel, lipids (amino acids, peptides, and proteins), Protein Binding, Galactolipid, Galactolipids biosynthesis, Uridine Diphosphate, Diglycerides, 03 medical and health sciences, Protein Domains, Scattering, Small Angle, Genetics, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Amino Acid Sequence, Diacylglycerol kinase, Sequence Homology, Amino Acid, Arabidopsis Proteins, Galactolipids, Crystal structure, Envelope membrane, Galactose, Cell Biology, Intracellular Membranes, Plant, Biosynthetic Pathways, 030104 developmental biology, Metabolism, Mutation, biology.protein, Biocatalysis, Biogenesis, 010606 plant biology & botany
Abstract: International audience; Monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG) are the major lipid components of photosynthetic membranes, and hence the most abundant lipids in the biosphere. They are essential for assembly and function of the photosynthetic apparatus. In Arabidopsis, the first step of galactolipid synthesis is catalyzed by MGDG synthase 1 (MGD1), which transfers a galactosyl residue from UDP-galactose to diacylglycerol (DAG). MGD1 is a monotopic protein that is embedded in the inner envelope membrane of chloroplasts. Once produced, MGDG is transferred to the outer envelope membrane, where DGDG synthesis occurs, and to thylakoids. Here we present two crystal structures of MGD1: one unliganded and one complexed with UDP. MGD1 has a long and flexible region (approximately 50 amino acids) that is required for DAG binding. The structures reveal critical features of the MGD1 catalytic mechanism and its membrane binding mode, tested on biomimetic Langmuir monolayers, giving insights into chloroplast membrane biogenesis. The structural plasticity of MGD1, ensuring very rapid capture and utilization of DAG, and its interaction with anionic lipids, possibly driving the construction of lipoproteic clusters, are consistent with the role of this enzyme, not only in expansion of the inner envelope membrane, but also in supplying MGDG to the outer envelope and nascent thylakoid membranes.
Published: 2016
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31. From Molecules to Living Organisms: An Interplay Between Biology and Physics

Author: Pebay-Peyroula, Eva, Nury, Hugues, Parcy, François, Ruigrok, Rob W. H., Ziegler, Christine, Cugliandolo, Leticia F., Thomas, Frank, Eva Pebay-Peyroula, Institut de biologie structurale (IBS - UMR 5075), 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 (UGA)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de physiologie cellulaire végétale (LPCV), Université Joseph Fourier - Grenoble 1 (UJF)-Institut National de la Recherche Agronomique (INRA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), 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 für Biophysik und physicalische Biochemie, Universität Regensburg (UR), Laboratoire de Physique Théorique et Hautes Energies (LPTHE), Université Pierre et Marie Curie - Paris 6 (UPMC)-Centre National de la Recherche Scientifique (CNRS), Institut de biologie structurale (IBS - UMR 5075 ), 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)-Centre National de la Recherche Scientifique (CNRS), Physiologie cellulaire et végétale (LPCV), Institut National de la Recherche Agronomique (INRA)-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), Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF), 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), Centre National de la Recherche Scientifique (CNRS), Université Grenoble Alpes (UGA), Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes (UGA)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), UIMI 3265 UVHCI Unit of Virus Host Interactions, European Molecular Biology Laboratory, Institute of Biophysics and Physical Biochemistry, and Centre National de la Recherche Scientifique (CNRS)-Université Pierre et Marie Curie - Paris 6 (UPMC)
Subjects: [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], education, membrane fusion, surface proteins, modelling, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], développement de la plante, interaction virus cellule, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, molecular model, biologie intégrée, protéine membranaire, fusion membranaire, modèle moléculaire, modélisation
Abstract: Ecole d'été organisée par :Eva Pebay-Peyroula, IBS, Grenoble, FRChristine Ziegler, Univ. Regensburg et MPI Frankfurt, AllemagneYves Gaudin, VMS, Gif-sur-Yvette, FRHugues Nury, EPFL Lausanne, Suisse et IBS, Grenoble, FRFrançois Parcy, iRTSV/PCV, Grenoble, FRRob Ruigrok, UVHCI, Grenoble, FR; The book gathers lecture notes of courses given at the 2014 summer school on integrated biology in Les Houches, France, Session CII. It addresses an emerging field ranging from molecules to cells and to organisms. Through examples it presents a new way of thinking using a combination of interdisciplinary and cutting-edge methods, bridging physics and biology beyond current biophysics. Important novel developments are expected in the coming years that may well introduce paradigm shifts in biological science. The school had the ambition to prepare participants to become major actors in these breakthroughs.The power of integrated approaches is illustrated through two cases: interactions between viruses and host cells, and flower development. The role of forces in biology, as well as their mathematical modeling, is illustrated in both processes: how they allow flower organs to emerge or how they control membrane fusion during virus budding. The book also underlines the importance of conformational changes and dynamics of proteins particularly during membrane processes. It explains how membrane proteins can be handled and studied by molecular simulations. Finally, the book also contains concepts in cell biology, in thermodynamics and several novel approaches such as in-cell NMR. Altogether, the chapters show how examining a biological system from different viewpoints based on multidisciplinary aspects often leads to enriching controversial arguments.
Published: 2016

32. The solution configurations of inactive and activated DntR have implications for the sliding dimer mechanism of LysR transcription factors

Author: Cyril Dian, Michael Lerche, Rosa Lönneborg, Peter Brzezinski, Adam Round, Gordon A. Leonard, European Synchrotron Radiation Facility (ESRF), Institut des sciences du végétal (ISV), Centre National de la Recherche Scientifique (CNRS), European Molecular Biology Laboratory [Grenoble] (EMBL), 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), KTH Royal Inst Technol, Sch Educ & Commun Engn Sci ECE, SE-10044 Stockholm, Sweden, and Stockholm Univ, Arrhenius Labs Nat Sci, Dept Biochem & Biophys, SE-10691 Stockholm, Sweden
Subjects: 0301 basic medicine, Models, Molecular, Conformational change, Protein Conformation, [SDV]Life Sciences [q-bio], 030106 microbiology, Mutant, Plasma protein binding, Biology, Crystallography, X-Ray, Article, 03 medical and health sciences, chemistry.chemical_compound, Protein structure, Bacterial Proteins, X-Ray Diffraction, Transcription (biology), Scattering, Small Angle, Promoter Regions, Genetic, Transcription factor, Multidisciplinary, Base Sequence, DNA, DNA binding site, Solutions, 030104 developmental biology, chemistry, Biochemistry, Biophysics, Protein Multimerization, Protein Binding, Transcription Factors
Abstract: LysR Type Transcriptional Regulators (LTTRs) regulate basic metabolic pathways or virulence gene expression in prokaryotes. Evidence suggests that the activation of LTTRs involves a conformational change from an inactive compact apo- configuration that represses transcription to an active, expanded holo- form that promotes it. However, no LTTR has yet been observed to adopt both configurations. Here, we report the results of structural studies of various forms of the LTTR DntR. Crystal structures of apo-DntR and of a partially autoinducing mutant H169T-DntR suggest that active and inactive DntR maintain a compact homotetrameric configuration. However, Small Angle X-ray Scattering (SAXS) studies on solutions of apo-, H169T- and inducer-bound holo-DntR indicate a different behaviour, suggesting that while apo-DntR maintains a compact configuration in solution both H169T- and holo-DntR adopt an expanded conformation. Models of the SAXS-obtained solution conformations of apo- andholo-DntR homotetramers in complex with promoter-operator region DNA are consistent with previous observations of a shifting of LTTR DNA binding sites upon activation and a consequent relaxation in the bend of the promoter-operator region DNA. Our results thus provide clear evidence at the molecular level which strongly supports the ‘sliding dimer’ hypothesis concerning LTTR activation mechanisms.
Published: 2016
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33. RoboDiff: combining a sample changer and goniometer for highly automated macromolecular crystallography experiments

Author: Christoph Mueller-Dieckmann, Nicolas Guichard, Hugo Caserotto, Florent Cipriani, Gordon A. Leonard, Didier Nurizzo, Fabien Dobias, David Flot, Sean McSweeney, John Surr, Gergely Papp, Pascal Theveneau, Matias Guijarro, Matthew W. Bowler, Thierry Giraud, European Synchrotron Radiation Facility (ESRF), European Molecular Biology Laboratory [Grenoble] (EMBL), 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), Brookhaven National Laboratory [Upton, NY] (BNL), U.S. Department of Energy [Washington] (DOE)-UT-Battelle, LLC-Stony Brook University [SUNY] (SBU), and State University of New York (SUNY)-State University of New York (SUNY)
Subjects: 0301 basic medicine, 030103 biophysics, Computer science, RoboDiff, [SDV]Life Sciences [q-bio], Thermolysin, Bacillus, high throughput, Crystallography, X-Ray, Bacterial protein, 03 medical and health sciences, Software, Bacterial Proteins, Structural Biology, Animals, Trypsin, automation, robotics, goniometers, business.industry, Macromolecular crystallography, Proteins, Equipment Design, Automation, Sample (graphics), Research Papers, Crystallography, 030104 developmental biology, Fully automated, Beamline, Goniometer, Cattle, business, Computer hardware
Abstract: An industrial six-axis robot has been combined with a high-accuracy air-bearing rotation axis to create a single device with the capabilities of both transferring cryocooled protein crystals from a sample-containing dewar and collecting complete X-ray diffraction data sets., Automation of the mounting of cryocooled samples is now a feature of the majority of beamlines dedicated to macromolecular crystallography (MX). Robotic sample changers have been developed over many years, with the latest designs increasing capacity, reliability and speed. Here, the development of a new sample changer deployed at the ESRF beamline MASSIF-1 (ID30A-1), based on an industrial six-axis robot, is described. The device, named RoboDiff, includes a high-capacity dewar, acts as both a sample changer and a high-accuracy goniometer, and has been designed for completely unattended sample mounting and diffraction data collection. This aim has been achieved using a high level of diagnostics at all steps of the process from mounting and characterization to data collection. The RoboDiff has been in service on the fully automated endstation MASSIF-1 at the ESRF since September 2014 and, at the time of writing, has processed more than 20 000 samples completely automatically.
Published: 2016
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34. Kinetic discrimination of self/non-self RNA by the ATPase activity of RIG-I and MDA5

Author: Louber, Jade, Brunel, Joanna, Uchikawa, Emiko, Cusack, Stephen, Gerlier, Denis, Immunobiologie des infections virales – Immunobiology of Viral Infections (IbIV), Centre International de Recherche en Infectiologie - UMR (CIRI), 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)-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), 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), Centre International de Recherche en Infectiologie (CIRI), École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-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), and BMC, BMC
Subjects: Interferon-Induced Helicase, IFIH1, MDA5, viruses, [SDV]Life Sciences [q-bio], dsRNA, self from non-self, Cell Line, DEAD-box RNA Helicases, RIG-I, Adenosine Triphosphate, Humans, Point Mutation, ATPase, Receptors, Immunologic, RNA, Double-Stranded, Adenosine Triphosphatases, Binding Sites, Agricultural and Biological Sciences(all), Biochemistry, Genetics and Molecular Biology(all), Hydrolysis, Keratin-8, Protein Structure, Tertiary, [SDV] Life Sciences [q-bio], Kinetics, DEAD Box Protein 58, Research Article, Protein Binding
Abstract: Background The cytoplasmic RIG-like receptors are responsible for the early detection of viruses and other intracellular microbes by activating the innate immune response mediated by type I interferons (IFNs). RIG-I and MDA5 detect virus-specific RNA motifs with short 5′-tri/diphosphorylated, blunt-end double-stranded RNA (dsRNA) and >0.5–2 kb long dsRNA as canonical agonists, respectively. However, in vitro, they can bind to many RNA species, while in cells there is an activation threshold. As SF2 helicase/ATPase family members, ATP hydrolysis is dependent on co-operative RNA and ATP binding. Whereas simultaneous ATP and cognate RNA binding is sufficient to activate RIG-I by releasing autoinhibition of the signaling domains, the physiological role of the ATPase activity of RIG-I and MDA5 remains controversial. Results A cross-analysis of a rationally designed panel of RNA binding and ATPase mutants and truncated receptors, using type I IFN promoter activation as readout, allows us to refine our understanding of the structure-function relationships of RIG-I and MDA5. RNA activation of RIG-I depends on multiple critical RNA binding sites in its helicase domain as confirmed by functional evidence using novel mutations. We found that RIG-I or MDA5 mutants with low ATP hydrolysis activity exhibit constitutive activity but this was fully reverted when associated with mutations preventing RNA binding to the helicase domain. We propose that the turnover kinetics of the ATPase domain enables the discrimination of self/non-self RNA by both RIG-I and MDA5. Non-cognate, possibly self, RNA binding would lead to fast ATP turnover and RNA disassociation and thus insufficient time for the caspase activation and recruitment domains (CARDs) to promote downstream signaling, whereas tighter cognate RNA binding provides a longer time window for downstream events to be engaged. Conclusions The exquisite fine-tuning of RIG-I and MDA5 RNA-dependent ATPase activity coupled to CARD release allows a robust IFN response from a minor subset of non-self RNAs within a sea of cellular self RNAs. This avoids the eventuality of deleterious autoimmunity effects as have been recently described to arise from natural gain-of-function alleles of RIG-I and MDA5. Electronic supplementary material The online version of this article (doi:10.1186/s12915-015-0166-9) contains supplementary material, which is available to authorized users.
Published: 2015
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35. [The keys to success in French Medical National Ranking Examination: Integrated training activities in teaching hospital and medical school]

Author: Marie Fourcot, Jean-Luc Bosson, Céline Genty, Patrice Morand, Pierre Gillois, Techniques pour l'Evaluation et la Modélisation des Actions de la Santé (TIMC-IMAG-ThEMAS), Techniques de l'Ingénierie Médicale et de la Complexité - Informatique, Mathématiques et Applications, Grenoble - UMR 5525 (TIMC-IMAG), VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF)-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF), Division de Biostatistiques, Université Joseph Fourier - Grenoble 1 (UJF)-CHU Grenoble-Hôpital Michallon, 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), Ingénieries santé médicament, Unit for Virus Host-Cell Interactions [Grenoble] (UVHCI), and Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF)
Subjects: MESH: Education, Medical, education, Specialty, Library science, [SCCO.COMP]Cognitive science/Computer science, MESH: Logistic Models, Ranking (information retrieval), MESH: Career Choice, [STAT.ML]Statistics [stat]/Machine Learning [stat.ML], MESH: Internship and Residency, [MATH.MATH-ST]Mathematics [math]/Statistics [math.ST], Internship, ComputingMilieux_COMPUTERSANDEDUCATION, Medicine, ComputingMilieux_MISCELLANEOUS, [STAT.AP]Statistics [stat]/Applications [stat.AP], MESH: Humans, MESH: Schools, Medical, MESH: Curriculum, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], ComputingMilieux_THECOMPUTINGPROFESSION, business.industry, 4. Education, MESH: Achievement, Success factors, MESH: Adult, MESH: Medicine, General Medicine, MESH: Hospitals, Teaching, [SDV.BIBS]Life Sciences [q-bio]/Quantitative Methods [q-bio.QM], MESH: Male, 3. Good health, MESH: France, MESH: Educational Measurement, business, MESH: Female, [STAT.ME]Statistics [stat]/Methodology [stat.ME]
Abstract: International audience; The National Ranking Examination (NRE) is the key to the choice of career and specialty for future physicians; it lets them choose their place of employment in a specialty and an hospital for their internship. It seems interesting to model the success factors to this exam for the medical students from Grenoble University.
Published: 2015
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36. Intravacuolar Membranes Regulate CD8 T Cell Recognition of Membrane-Bound Toxoplasma gondii Protective Antigen

Author: Lopez, Jodie, Bittame, Amina, Massera, Céline, Vasseur, Virginie, Effantin, Grégory, Valat, Anne, Buaillon, Célia, Allart, Sophie, Fox, Barbara A, Rommereim, Leah M, Bzik, David J, Schoehn, Guy, Weissenhorn, Winfried, Dubremetz, Jean-François, Gagnon, Jean, Mercier, Corinne, Cesbron-Delauw, Marie-France, Blanchard, Nicolas, Centre de Physiopathologie Toulouse Purpan (CPTP), 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)-Centre National de la Recherche Scientifique (CNRS), Université Joseph Fourier - Grenoble 1 (UJF), Laboratoire Adaptation et pathogénie des micro-organismes [Grenoble] (LAPM), Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS), Barrière Naturelle et Infectiosité (TIMC-IMAG-BNI), Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS), 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 de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-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), Department of Microbiology and Immunology, Dartmouth Medical School (DMS), Dartmouth College [Hanover]-Dartmouth College [Hanover], Dynamique des interactions membranaires normales et pathologiques (DIMNP), Université Montpellier 1 (UM1)-Université Montpellier 2 - Sciences et Techniques (UM2)-Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Bisanz, Cordelia, 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), Dartmouth Medical School, Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF), Techniques de l'Ingénierie Médicale et de la Complexité - Informatique, Mathématiques et Applications, Grenoble - UMR 5525 (TIMC-IMAG), VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF)-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Institut polytechnique de Grenoble - Grenoble Institute of Technology (Grenoble INP )-Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF), Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and 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)
Subjects: Male, Blotting, Western, Antigen presentation, Protozoan Proteins, Fluorescent Antibody Technique, Antigens, Protozoan, Vacuole, CD8-Positive T-Lymphocytes, Lymphocyte Activation, General Biochemistry, Genetics and Molecular Biology, Host-Parasite Interactions, Mice, Antigen, MHC class I, parasitic diseases, Animals, Cytotoxic T cell, lcsh:QH301-705.5, [SDV.MP] Life Sciences [q-bio]/Microbiology and Parasitology, Antigen Presentation, biology, Effector, Toxoplasma gondii, biology.organism_classification, Cell biology, Mice, Inbred C57BL, Disease Models, Animal, [SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology, lcsh:Biology (General), Vacuoles, biology.protein, Female, Toxoplasmosis, CD8
Abstract: International audience; Apicomplexa parasites such asToxoplasma gondiitarget effectors to and across the boundary of theirparasitophorous vacuole (PV), resulting in host cellsubversion and potential presentation by MHC classI molecules for CD8 T cell recognition. The host-para-site interface comprises the PV limiting membraneand a highly curved, membranous intravacuolarnetwork (IVN) of uncertain function. Here, using acell-free minimal system, we dissect how membranetubules are shaped by the parasite effectors GRA2and GRA6. We show that membrane associationregulates access of the GRA6 protective antigen tothe MHC I pathway in infected cells. Although inser-tion of GRA6 in the PV membrane is key for immuno-genicity, association of GRA6 with the IVN limitspresentation and curtails GRA6-specific CD8 re-sponses in mice. Thus, membrane deformations ofthe PV regulate access of antigens to the MHC classI pathway, and the IVN may play a role in immunemodulation.
Published: 2015
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37. Liaison à l'ARN et Assemblage des ARN-Polymérases des virus Humains de la Grippe A

Author: Swale, Christopher, 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), Université Grenoble Alpes, Rob W. H. Ruigrok, Thibaut Crépin, and STAR, ABES
Subjects: [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], RNA polymerase, viruses, Grippe, ARN viral, Viral RNA, ARN polymérase, Influenza, Rnp
Abstract: Influenza A virus is a negative-strand RNA virus belonging to the Orthomyxoviriadea family whose replication occurs in the nucleus of infected cells. The genome organisation of influenza virus is segmented in eight vRNA segments of negative polarity coding for at least 16 different viral proteins. Each vRNA is bound to multiple copies of nucleoprotein (NP) and to the heterotrimeric RNA-dependent RNA-polymerase complex (PA, PB1 and PB2) through its 5' and 3' extremities. This macromolecular assembly (vRNA/polymerase/NP) forms the ribonucleoprotein (RNP) particle, which acts as a separate genomic entity within the virion. The RNP complex is at the core of viral replication and in the context of RNPs, the polymerase performs both transcription and replication of the vRNA genome. As such, the polymerase constitutes a major antiviral drug target. The research work presented within this thesis focuses on the underlying determinants of the RNA polymerase assembly process and its interaction with its vRNA genome. To fulfill these goals, our lab, in collaboration with other groups, has set up a novel polyprotein expression system to express the polymerase but also to reconstitute polymerase and cellular partner complexes, notably RanBP5, which belongs to the importin-β family., Le virus de la grippe A est un virus à ARN négatif appartenant à la famille des Orthomyxoviriadea dont la réplication se produit dans le noyau des cellules infectées. L'organisation du génome est segmentée en huit segments d'ARNv de polarité négative, codant pour un minimum de 16 protéines virales différentes. Ces ARN viraux (ARNv) sont en complexe avec de nombreuses copies de nucléoprotéines et liés par leurs extrémités 5' et 3' au complexe hétérotrimérique de l'ARN-polymérase ARN-dépendante composé des sous unités PA, PB1 et PB2. Cet assemblage macromoléculaire (ARNv / polymérase / NP) nommée Ribonucléoprotéine (RNP) constitue une entité génomique indépendante. Dans le contexte de la RNP, l'ARN-polymérase assure à la fois la transcription et la réplication du génome ARNv. En assurant ces deux fonctions, l'ARN-polymérase joue un rôle majeur dans la réplication virale et constitue une cible antivirale privilégiée. Les travaux de recherche présentés dans cette thèse se concentrent sur les éléments structuraux participants à l'assemblage de l'ARN polymérase et son interaction avec les avec les ARNv. Pour atteindre ces objectifs, notre laboratoire, en collaboration avec d'autres groupes, a mis en place un système d'expression en polyprotéines permettant d'exprimer la polymérase. Plus encore, cette méthode a aussi permis de reconstituer des complexes entre l'ARN-polymérase et des partenaires cellulaires, notamment RanBP5 qui appartient à la famille des importines-β.
Published: 2015

38. Large-Scale Conformational Dynamics Control H5N1 Influenza Polymerase PB2 Binding to Importin α

Author: Nicola Salvi, Adam Round, Elise Delaforge, Martin Blackledge, Sigrid Milles, Stephane Boivin, Darren J. Hart, Anne Martel, Malene Ringkjøbing Jensen, Guillaume Bouvignies, Denis Bouvier, Damien Maurin, Edward A. Lemke, 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), Integrated Structural Biology Grenoble, Unit of Virus Host Cell Interactions, Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Institut Laue-Langevin (ILL), ILL, European Molecular Biology Laboratory [Heidelberg] (EMBL), Structural and Computational Biology Unit, EMBL, 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), University of Toronto, 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), Agence Nationale de Recherche under ComplexDynamics [SIMI7], EMBO long-term fellowship, Swiss National Science Foundation [P2ELP2\₁48858], Region Rhone-Alpes, FINOVI foundation, FRISBI [ANR-10-INSB-05-02], GRAL within the Grenoble Partnership for Structural Biology (PSB) [ANR-10-LABX-49-01], Emmy Noether program of the DFG, and Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF)
Subjects: Protein Conformation, Population, Importin, Karyopherins, 010402 general chemistry, Crystallography, X-Ray, 01 natural sciences, Biochemistry, environment and public health, Catalysis, 03 medical and health sciences, Viral Proteins, Colloid and Surface Chemistry, Protein structure, Importin-alpha, Fluorescence Resonance Energy Transfer, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, education, Nuclear Magnetic Resonance, Biomolecular, Dynamic equilibrium, Polymerase, ComputingMilieux_MISCELLANEOUS, 030304 developmental biology, 0303 health sciences, education.field_of_study, biology, Influenza A Virus, H5N1 Subtype, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Chemistry, General Chemistry, 0104 chemical sciences, Solutions, Crystallography, Förster resonance energy transfer, biology.protein, Salt bridge, Protein Binding
Abstract: International audience; Influenza A RNA polymerase complex is formed from three components, PA, PB1, and PB2. PB2 is independently imported into the nucleus prior to polymerase reconstitution. All crystallographic structures of the PB2 C-terminus (residues 536-759) reveal two globular domains, 627 and NLS, that form a tightly packed heterodimer. The molecular basis of the affinity of 627-NLS for importins remained unclear from these structures, apparently requiring large-scale conformational changes prior to importin binding. Using a combination of solution-state NMR, small-angle neutron scattering, small-angle X-ray scattering (SAXS), and Forster resonance energy transfer (FRET), we show that 627-NLS populates a temperature-dependent dynamic equilibrium between closed and open states. The closed state is stabilized by a tripartite salt bridge involving the 627-NLS interface and the linker, that becomes flexible in the open state, with 627 and NLS dislocating into a highly dynamic ensemble. Activation enthalpies and entropies associated with the rupture of this interface were derived from simultaneous analysis of temperature-dependent chemical exchange saturation transfer measurements, revealing a strong temperature dependence of both open-state population and exchange rate. Single-molecule FRET and SAXS demonstrate that only the open-form is capable of binding to importin alpha and that, upon binding, the 627 domain samples a dynamic conformational equilibrium in the vicinity of the C-terminus of importin alpha. This intrinsic large-scale conformational flexibility therefore enables 627-NLS to bind importin through conformational selection from a temperature-dependent equilibrium comprising both functional forms of the protein.
Published: 2015
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39. Hidden Selection of Bacterial Resistance to Fluoroquinolones In Vivo: The Case of Legionella pneumophila and Humans

Author: Sophie Jarraud, Lubana Shadoud, Dominique Schneider, Jerome Etienne, Iyad Almahmoud, Sylvie Larrat, Jean-François Timsit, Max Maurin, Carole Schwebel, Laboratoire Adaptation et pathogénie des micro-organismes [Grenoble] (LAPM), Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS), Centre Nationale de Référence des Légionelles, Laboratoire de Virologie moléculaire et structurale, CHU Grenoble, Service de Réanimation Médicale [Grenoble], CHU Grenoble-Hôpital Michallon, Epidémiologie pronostique des cancers et affections graves, Université Joseph Fourier - Grenoble 1 (UJF)-Institut National de la Santé et de la Recherche Médicale (INSERM), Génomique et Évolution des Microorganismes [?-2015] (TIMC-GEM [?-2015]), Techniques de l'Ingénierie Médicale et de la Complexité - Informatique, Mathématiques et Applications, Grenoble - UMR 5525 [2011-2015] (TIMC [2011-2015]), Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS)-IMAG-Institut polytechnique de Grenoble - Grenoble Institute of Technology [2007-2019] (Grenoble INP [2007-2019])-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS)-IMAG-Institut polytechnique de Grenoble - Grenoble Institute of Technology [2007-2019] (Grenoble INP [2007-2019])-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS), Thérapeutique Recombinante Expérimentale [?-2015] (TIMC-TheREx [?-2015]), Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF), Pathogenèse des légionelles- Legionella pathogenesis (LegioPath), Centre International de Recherche en Infectiologie - UMR (CIRI), Institut National de la Santé et de la Recherche Médicale (INSERM)-École normale supérieure - Lyon (ENS 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)-École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Pathogénie des Staphylocoques – Staphylococcal Pathogenesis (StaPath), 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), Service de Réanimation Médicale [CHU Grenoble], 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), Centre International de Recherche en Infectiologie (CIRI), École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), and Université Joseph Fourier - Grenoble 1 (UJF)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Centre National de la Recherche Scientifique (CNRS)
Subjects: Male, Antibiotics, Colony Count, Microbial, Drug Resistance, lcsh:Medicine, Fluoroquinolone resistance, Legionella pneumophila, In vivo selection, Cohort Studies, Microbial resistance, 80 and over, ComputingMilieux_MISCELLANEOUS, Aged, 80 and over, lcsh:R5-920, Cross Infection, biology, [SDV.BID.EVO]Life Sciences [q-bio]/Biodiversity/Populations and Evolution [q-bio.PE], Bacterial, Drug Resistance, Microbial, General Medicine, Bacterial Infections, Middle Aged, 3. Good health, Human morbidity, Anti-Bacterial Agents, Hospitalization, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, [SDV.IMM]Life Sciences [q-bio]/Immunology, Female, lcsh:Medicine (General), Research Paper, Fluoroquinolones, DNA, Bacterial, Adult, medicine.drug_class, Microbial Sensitivity Tests, General Biochemistry, Genetics and Molecular Biology, Microbiology, Antibiotic resistance, In vivo, Next generation sequencing, Drug Resistance, Bacterial, medicine, Escherichia coli, Humans, Aged, Legionellosis, Bacteria, lcsh:R, DNA, Resistance detection, biology.organism_classification, Virology, [SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, Bacterial Load, Genes, Genes, Bacterial, Mutation, Commentary
Abstract: Background Infectious diseases are the leading cause of human morbidity and mortality worldwide. One dramatic issue is the emergence of microbial resistance to antibiotics which is a major public health concern. Surprisingly however, such in vivo adaptive ability has not been reported yet for many intracellular human bacterial pathogens such as Legionella pneumophila. Methods We examined 82 unrelated patients with Legionnaire's disease from which 139 respiratory specimens were sampled during hospitalization and antibiotic therapy. We both developed a real time PCR assay and used deep-sequencing approaches to detect antibiotic resistance mutations in L. pneumophila and follow their selection and fate in these samples. Findings We identified the in vivo selection of fluoroquinolone resistance mutations in L. pneumophila in two infected patients treated with these antibiotics. By investigating the mutational dynamics in patients, we showed that antibiotic resistance occurred during hospitalization most likely after fluoroquinolone treatment. Interpretation In vivo selection of antibiotic resistances in L. pneumophila may be associated with treatment failures and poor prognosis. This hidden resistance must be carefully considered in the therapeutic management of legionellosis patients and in the control of the gradual loss of effectiveness of antibiotics., Highlights • Legionellosis is a pneumonia caused by the inhalation of aerosols containing Legionella, mainly L. pneumophila. • Its average mortality rate is 10%, despite availability of effective antibiotics such as the macrolides and the fluoroquinolones. • Using modern molecular approaches, we identified the selection of fluoroquinolone resistance in L. pneumophila in patients under fluoroquinolone therapy. • This may lead to reduction of treatment efficacy and prognosis worsening. • Our findings should lead to revised guidelines for therapeutic management and prognosis evaluation of legionellosis.
Published: 2015
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40. Structure and primase-mediated activation of a bacterial dodecameric replicative helicase

Author: Alexandre Bazin, Laurent Terradot, MickaÃ«l V. Cherrier, Joanna Timmins, Irina Gutsche, Bases moléculaires et structurales des systèmes infectieux (BMSSI), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), 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), 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), ATIP-Avenir, 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 Joseph Fourier - Grenoble 1 (UJF)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Centre National de la Recherche Scientifique (CNRS)
Subjects: Models, Molecular, [SDV]Life Sciences [q-bio], DNA, Single-Stranded, DNA Primase, Biology, Primosome, 03 medical and health sciences, chemistry.chemical_compound, 0302 clinical medicine, Bacterial Proteins, ATP hydrolysis, Structural Biology, Genetics, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], dnaB helicase, 030304 developmental biology, Adenosine Triphosphatases, 0303 health sciences, Helicobacter pylori, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], DNA replication, Helicase, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, DNA, Protein Structure, Tertiary, Enzyme Activation, [SDV.BBM.BP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biophysics, Dodecameric protein, Biochemistry, chemistry, biology.protein, Biophysics, Primase, Protein Multimerization, DnaB Helicases, 030217 neurology & neurosurgery
Abstract: International audience; Replicative helicases are essential ATPases that unwind DNA to initiate chromosomal replication. While bacterial replicative DnaB helicases are hexameric, Helicobacter pylori DnaB (HpDnaB) was found to form double hexamers, similar to some archaeal and eukaryotic replicative helicases. Here we present a structural and functional analysis of HpDnaB protein during primosome formation. The crystal structure of the HpDnaB at 6.7 ˚ A resolution reveals a dode-cameric organization consisting of two hexamers assembled via their N-terminal rings in a stack-twisted mode. Using fluorescence anisotropy we show that HpDnaB dodecamer interacts with single-stranded DNA in the presence of ATP but has a low DNA unwinding activity. Multi-angle light scattering and small angle X-ray scattering demonstrate that interaction with the DnaG primase helicase-binding domain dissociates the helicase dodecamer into single ringed primosomes. Functional assays on the proteins and associated complexes indicate that these single ringed primosomes are the most active form of the helicase for ATP hydrolysis, DNA binding and unwinding. These findings shed light onto an activation mechanism of HpDnaB by the primase that might be relevant in other bacteria and possibly other organisms exploiting dodecameric helicases for DNA replication.
Published: 2015
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41. Hepatitis E and neuralgic amyotrophy: Five cases and review of literature

Author: Sylvie Larrat, Françoise Sarrot-Reynauld, Anaïs Dartevel, Emmeline Lagrange, B. Colombe, Laurence Bouillet, A. Bosseray, Aude Belbézier, Université Grenoble Alpes - UFR Médecine (UGA UFRM), Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Clinique de médecine interne, Université Joseph Fourier - Grenoble 1 (UJF)-CHU Grenoble, 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), Département de neurologie, and Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF)
Subjects: Parsonage–Turner syndrome, Adult, Male, Pediatrics, medicine.medical_specialty, Usually asymptomatic, Sex Factors, MESH: Sex Factors, Virology, medicine, Brachial Plexus Neuritis, Humans, Parsonage turner syndrome, MESH: Brachial Plexus Neuritis, MESH: Developed Countries, Neuralgic amyotrophy, MESH: Humans, MESH: Middle Aged, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Acute hepatitis E, business.industry, Developed Countries, MESH: Hepatitis E, Mean age, Strong hepatitis E, MESH: Adult, Middle Aged, Hepatitis E, medicine.disease, Nervous system diseases, MESH: Immunoglobulin M, MESH: Male, 3. Good health, Surgery, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Infectious Diseases, Immunoglobulin M, Female, business, Brachial plexus, MESH: Female, Hepatitis E virus infection
Abstract: International audience; Hepatitis E virus infection - mainly genotype 3 - is increasingly common in industrialized countries. Infection is usually asymptomatic, but cases of central or peripheral neurological symptoms with hepatitis E have been described. The most frequent is Guillain-Barre but somes cases of neuralgic amyotrophy have been described. In our center, since 2010, we have identified five cases of neuralgic amyotrophy associated with acute hepatitis E in immunocompetent patients. For all these patients, neuralgic amyotrophy was diagnosed with electromyogram and positive IgM for hepatitis E, and detectable HEV RNA in 4 of the cases. Including our patients, we count 26 cases in literature. The mean age of the patients was 44 years old, with a large predominance of males (88%). The disorder is bilateral and asymmetric in 69% of cases. Peripheral nerves other than the brachial plexus were affected in 6 patients (23%). In industrialized countries, any neuralgic amyotrophy, particularly if there is bilateral, asymmetric associated with involvement of nerves outside the brachial plexus, should lead physicians to consider a diagnosis of acute hepatitis E.
Published: 2015
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42. Assessment of Residual Moisture and Maintenance of Sterility in Surgical Instrument Sets after Sterilization

Author: Camille Fayard, Marion Levast, Christophe Lambert, Raphaelle Germi, Catherine Guimier-Pingault, 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), and Thomas, Frank
Subjects: Microbiology (medical), medicine.medical_specialty, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Epidemiology, Sterility, Residual, complex mixtures, medicine, Humans, Cross Infection, business.industry, Residual moisture, Sterilization, Water, food and beverages, Sterilization (microbiology), Surgical Instruments, Steam sterilization, humanities, Surgery, Steam, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Infectious Diseases, Surgical instrument, Equipment Contamination, business
Abstract: Good sterilization practices include discarding items containing residual moisture after steam sterilization. In this small laboratory study, however, the presence of residual water did not appear to compromise the sterility of surgical instruments in 2 commonly used types of packaging during routine storage after steam sterilization.Infect Control Hosp Epidemiol 2015;36(8):990–992
Published: 2015

43. Crystal Structure of the Vaccinia Virus Uracil-DNA Glycosylase in Complex with DNA

Author: Christophe N. Peyrefitte, Céline Contesto-Richefeu, Wim P. Burmeister, Pascal Fender, Frédéric Iseni, Nicolas Tarbouriech, 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 de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-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), Virologie et pathogenèse virale (VPV), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF), 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)-Centre National de la Recherche Scientifique (CNRS), and Thomas, Frank
Subjects: [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Protein Conformation, DNA polymerase, viruses, DNA polymerase II, Molecular Sequence Data, MESH: Sequence Alignment, Vaccinia virus, MESH: Amino Acid Sequence, Crystallography, X-Ray, Biochemistry, DNA polymerase delta, 03 medical and health sciences, MESH: Protein Conformation, MESH: Vaccinia virus, Vaccinia, MESH: Molecular Docking Simulation, Humans, Amino Acid Sequence, Uracil-DNA Glycosidase, Molecular Biology, Replication protein A, MESH: Uracil-DNA Glycosidase, 030304 developmental biology, 0303 health sciences, DNA clamp, MESH: Humans, MESH: Molecular Sequence Data, biology, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], 030302 biochemistry & molecular biology, MESH: DNA, DNA, Cell Biology, MESH: Crystallography, X-Ray, Molecular biology, MESH: Vaccinia, 3. Good health, Molecular Docking Simulation, MESH: Nucleic Acid Conformation, DNA glycosylase, Uracil-DNA glycosylase, Protein Structure and Folding, biology.protein, Nucleic Acid Conformation, DNA polymerase I, Sequence Alignment
Abstract: International audience; Vaccinia virus polymerase holoenzyme is composed of the DNA polymerase catalytic subunit E9 associated with its heterodimeric co-factor A20·D4 required for processive genome synthesis. Although A20 has no known enzymatic activity, D4 is an active uracil-DNA glycosylase (UNG). The presence of a repair enzyme as a component of the viral replication machinery suggests that, for poxviruses, DNA synthesis and base excision repair is coupled. We present the 2.7 Å crystal structure of the complex formed by D4 and the first 50 amino acids of A20 (D4·A201-50) bound to a 10-mer DNA duplex containing an abasic site resulting from the cleavage of a uracil base. Comparison of the viral complex with its human counterpart revealed major divergences in the contacts between protein and DNA and in the enzyme orientation on the DNA. However, the conformation of the dsDNA within both structures is very similar, suggesting a dominant role of the DNA conformation for UNG function. In contrast to human UNG, D4 appears rigid, and we do not observe a conformational change upon DNA binding. We also studied the interaction of D4·A201-50 with different DNA oligomers by surface plasmon resonance. D4 binds weakly to nonspecific DNA and to uracil-containing substrates but binds abasic sites with a Kd of
Published: 2015

44. Structural virology. Near-atomic cryo-EM structure of the helical measles virus nucleocapsid

Author: Gutsche, Irina, Desfosses, Ambroise, Effantin, Grégory, Ling, Wai-Li, Haupt, Melina, Ruigrok, Rob W H, Sachse, Carsten, Schoehn, Guy, 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), Structural and Computational Biology Unit, European Molecular Biology Laboratory [Grenoble] (EMBL), Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-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), Institut Laue-Langevin (ILL), Centre National de la Recherche Scientifique (CNRS)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF), 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)-Centre National de la Recherche Scientifique (CNRS), and ILL
Subjects: [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], viruses, Cryoelectron Microscopy, Molecular Sequence Data, Nucleocapsid Proteins, Protein Structure, Secondary, Viral Proteins, Nucleoproteins, Measles virus, Humans, Nucleic Acid Conformation, RNA, Viral, Amino Acid Sequence, Nucleocapsid, Measles
Abstract: International audience; Measles is a highly contagious human disease. We used cryo-electron microscopy and single particle-based helical image analysis to determine the structure of the helical nucleocapsid formed by the folded domain of the measles virus nucleoprotein encapsidating an RNA at a resolution of 4.3 angstroms. The resulting pseudoatomic model of the measles virus nucleocapsid offers important insights into the mechanism of the helical polymerization of nucleocapsids of negative-strand RNA viruses, in particular via the exchange subdomains of the nucleoprotein. The structure reveals the mode of the nucleoprotein-RNA interaction and explains why each nucleoprotein of measles virus binds six nucleotides, whereas the respiratory syncytial virus nucleoprotein binds seven. It provides a rational basis for further analysis of measles virus replication and transcription, and reveals potential targets for drug design.
Published: 2015

45. The status of the macromolecular crystallography beamlines at the European Synchrotron Radiation Facility

Author: Andrew A. McCarthy, David von Stetten, Max H. Nanao, Daniele de Sanctis, Antoine Royant, David Flot, Petra Pernot, Adam Round, Alexander Popov, Christoph Mueller-Dieckmann, Matthew W. Bowler, Gordon A. Leonard, Didier Nurizzo, Philippe Carpentier, European Synchrotron Radiation Facility (ESRF), 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), European Molecular Biology Laboratory [Grenoble] (EMBL), European Molecular Biology Laboratory [Hamburg] (EMBL), 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), Cryobench, 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), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)
Subjects: Focus (computing), [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], business.industry, Computer science, Detector, Electrical engineering, General Physics and Astronomy, Synchrotron radiation, Tracking system, Tracking (particle physics), Synchrotron, law.invention, Beamline, law, business, ComputingMilieux_MISCELLANEOUS, Graphical user interface
Abstract: The European Synchrotron Radiation Facility (ESRF) is the oldest and most powerful 3rd generation synchrotron in Europe, providing X-rays to more than 40 experimental stations welcoming several thousand researchers per year. A major success story has been the ESRF's facilities for macromolecular crystallography (MX). These are grouped around 3 straight sections: On ID23 canted undulators accommodate ID23-1, a mini-focus tuneable energy end station and ID23-2, the world's first micro-focus beamline dedicated to MX; ID29 houses a single, mini-focus, tuneable energy end station; ID30 will provide three end stations for MX due in operation from mid-2014 to early 2015. Here, one branch of a canted X-ray source feeds two fixed-energy end stations (MASSIF-1, MASSIF-3). The second feeds ID30B, a variable focus, tuneable energy beamline. MASSIF-1 is optimised for automatic high-throughput experiments requiring a relatively large beam size at the sample position, MASSIF-3 is a high-intensity, micro-focus facility designed to complement ID23-2. All end stations are highly automated, equipped with sample mounting robots and large area, fast-readout photon-counting detectors. Experiment control and tracking is achieved via a combination of the MXCuBE2 graphical user interface and the ISPyB database, the former allowing user-friendly control of all beamline components, the latter providing data tracking before, after and during experiments.
Published: 2015
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46. Structure determination of feline calicivirus virus-like particles in the context of a pseudo-octahedral arrangement

Author: Wim P Burmeister, Marlyse Buisson, Leandro F Estrozi, Guy Schoehn, Olivier Billet, Zahia Hannas, Cécile Sigoillot, Hervé Poulet, 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), Mérial [Lyon], Mérial, Thomas, Frank, 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), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)
Subjects: Models, Molecular, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Protein Conformation, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], viruses, lcsh:R, Molecular Sequence Data, Virion, lcsh:Medicine, virus diseases, biochemical phenomena, metabolism, and nutrition, Cats, Animals, lcsh:Q, Capsid Proteins, Amino Acid Sequence, lcsh:Science, Sequence Alignment, Research Article, Calicivirus, Feline
Abstract: International audience; The vesivirus feline calicivirus (FCV) is a positive strand RNA virus encapsidated by an icosahedral T=3 shell formed by the viral VP1 protein. Upon its expression in the insect cell - baculovirus system in the context of vaccine development, two types of virus-like particles (VLPs) were formed, a majority built of 60 subunits (T=1) and a minority probably built of 180 subunits (T=3). The structure of the small particles was determined by x-ray crystallography at 0.8 nm resolution helped by cryo-electron microscopy in order to understand their formation. Cubic crystals belonged to space group P213. Their self-rotation function showed the presence of an octahedral pseudo-symmetry similar to the one described previously by Agerbandje and co-workers for human parvovirus VLPs. The crystal structure could be solved starting from the published VP1 structure in the context of the T=3 viral capsid. In contrast to viral capsids, where the capsomers are interlocked by the exchange of the N-terminal arm (NTA) domain, this domain is disordered in the T=1 capsid of the VLPs. Furthermore it is prone to proteolytic cleavage. The relative orientation of P (protrusion) and S (shell) domains is alerted so as to fit VP1 to the smaller T=1 particle whereas the intermolecular contacts around 2-fold, 3-fold and 5-fold axes are conserved. By consequence the surface of the VLP is very similar compared to the viral capsid and suggests a similar antigenicity. The knowledge of the structure of the VLPs will help to improve their stability, in respect to a use for vaccination.
Published: 2015

47. Visualizing the molecular recognition trajectory of an intrinsically disordered protein using multinuclear relaxation dispersion NMR

Author: Guillaume Communie, Malene Ringkjøbing Jensen, Martin Blackledge, Rob W. H. Ruigrok, Damien Maurin, Jaka Kragelj, D. Flemming Hansen, Robert Schneider, Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-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), 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), University of Grenoble Alpes, France, 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)-Centre National de la Recherche Scientifique (CNRS), 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), 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)-Université Grenoble Alpes (UGA)-Centre National de la Recherche Scientifique (CNRS), Unit of Virus Host Cell Interactions (UVHCI), and Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS)
Subjects: Models, Molecular, 010402 general chemistry, Intrinsically disordered proteins, 01 natural sciences, Biochemistry, Sendai virus, Catalysis, 03 medical and health sciences, Colloid and Surface Chemistry, Molecular recognition, Binding site, Nuclear Magnetic Resonance, Biomolecular, 030304 developmental biology, 0303 health sciences, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Chemistry, Relaxation (NMR), General Chemistry, Phosphoproteins, Receptor–ligand kinetics, 0104 chemical sciences, Folding (chemistry), Crystallography, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Nucleoproteins, Phosphoprotein, Helix, Biophysics
Abstract: International audience; Despite playing important roles throughout biology, molecular recognition mechanisms in intrinsically disordered proteins remain poorly understood. We present a combination of (1)H(N), (13)C', and (15)N relaxation dispersion NMR, measured at multiple titration points, to map the interaction between the disordered domain of Sendai virus nucleoprotein (NT) and the C-terminal domain of the phosphoprotein (PX). Interaction with PX funnels the free-state equilibrium of NT by stabilizing one of the previously identified helical substates present in the prerecognition ensemble in a nonspecific and dynamic encounter complex on the surface of PX. This helix then locates into the binding site at a rate coincident with intrinsic breathing motions of the helical groove on the surface of PX. The binding kinetics of complex formation are thus regulated by the intrinsic free-state conformational dynamics of both proteins. This approach, providing high-resolution structural and kinetic information about a complex folding and binding interaction trajectory, can be applied to a number of experimental systems to provide a general framework for understanding conformational disorder in biomolecular function
Published: 2015
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48. MeshAndCollect: an automated multi-crystal data-collection workflow for synchrotron macromolecular crystallography beamlines

Author: Alexander Popov, Gleb Bourenkov, Andrew A. McCarthy, Ulrich Zander, Christoph Mueller-Dieckmann, Gordon A. Leonard, Daniele de Sanctis, Ekaterina Round, Olof Svensson, Valentin Gordeliy, European Synchrotron Radiation Facility (ESRF), DESY, European Mol Biol Lab, D-22607 Hamburg, Germany, European Molecular Biology Laboratory [Grenoble] (EMBL), 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), Moscow Inst Phys & Technol, Lab Adv Studies Membrane Proteins, Dolgoprudnyi 141700, Russia, Res Ctr Juelich, ICS, ICS Mol Biophys 6, D-52425 Julich, Germany, Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-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)-European Molecular Biology Laboratory [Grenoble] (EMBL)-Université Joseph Fourier - Grenoble 1 (UJF), 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)-Centre National de la Recherche Scientifique (CNRS), European Molecular Biology Laboratory [Hamburg] (EMBL), and Institut de biologie structurale [1992-2019] (IBS - UMR 5075 [1992-2019])
Subjects: Halobacterium salinarum, Models, Molecular, Diffraction, Thermolysin, Analytical chemistry, Bacillus, Crystallography, X-Ray, Workflow, law.invention, Computational science, hierarchical cluster analysis, Set (abstract data type), macromolecular crystallography, Structural Biology, law, Proton transport, Animals, Cluster Analysis, Plant Proteins, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Chemistry, Proteins, General Medicine, multi-crystal data collection, Plants, Bombyx, Research Papers, Synchrotron, Hierarchical clustering, Data set, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Plant protein, Bacteriorhodopsins, Insect Proteins, synchrotron serial crystallography, Muramidase, Crystallization, Synchrotrons
Abstract: The fully automated collection and merging of partial data sets from a series of cryocooled crystals of biological macromolecules contained on the same support is presented, as are the results of test experiments carried out on various systems., Here, an automated procedure is described to identify the positions of many cryocooled crystals mounted on the same sample holder, to rapidly predict and rank their relative diffraction strengths and to collect partial X-ray diffraction data sets from as many of the crystals as desired. Subsequent hierarchical cluster analysis then allows the best combination of partial data sets, optimizing the quality of the final data set obtained. The results of applying the method developed to various systems and scenarios including the compilation of a complete data set from tiny crystals of the membrane protein bacteriorhodopsin and the collection of data sets for successful structure determination using the single-wavelength anomalous dispersion technique are also presented.
Published: 2015
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49. Polyproteins in structural biology

Author: Maxime Chaillet, Frederic Garzoni, Thibaut Crépin, Alexandre Monod, Christopher Swale, Imre Berger, Thomas, Frank, 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), European Molecular Biology Laboratory [Grenoble] (EMBL), Unit of Virus Host Cell Interactions (UVHCI), and Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS)
Subjects: Models, Molecular, Protein Folding, Polyproteins, Protein Conformation, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Protein subunit, viruses, Molecular Sequence Data, 02 engineering and technology, Crystallography, X-Ray, Microscopy, Atomic Force, Article, 03 medical and health sciences, Synthetic biology, Viral Proteins, Protein structure, Structural Biology, Animals, Humans, Amino Acid Sequence, Molecular Biology, Polymerase, 030304 developmental biology, ComputingMethodologies_COMPUTERGRAPHICS, 0303 health sciences, biology, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], 021001 nanoscience & nanotechnology, Recombinant Proteins, 3. Good health, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Structural biology, Biochemistry, Proteome, Viruses, biology.protein, Protein folding, 0210 nano-technology
Abstract: Graphical abstract, Highlights • Structures have been determined for natural and recombinant polyproteins. • Native HIV Gag polyprotein architecture was revealed by cryo-EM of immature capsids. • Recombinant polyprotein technology has resolved sample preparation bottlenecks. • The high-resolution structure of influenza polymerase has been solved. • Single-molecule analysis of polyproteins revealed their folding characteristics., Polyproteins are chains of covalently conjoined smaller proteins that occur in nature as versatile means to organize the proteome of viruses including HIV. During maturation, viral polyproteins are typically cleaved into the constituent proteins with different biological functions by highly specific proteases, and structural analyses at defined stages of this maturation process can provide clues for antiviral intervention strategies. Recombinant polyproteins that use similar mechanisms are emerging as powerful tools for producing hitherto inaccessible protein targets such as the influenza polymerase, for high-resolution structure determination by X-ray crystallography. Conversely, covalent linking of individual protein subunits into single polypeptide chains are exploited to overcome sample preparation bottlenecks. Moreover, synthetic polyproteins provide a promising tool to dissect dynamic folding of polypeptide chains into three-dimensional architectures in single-molecule structure analysis by atomic force microscopy (AFM). The recent use of natural and synthetic polyproteins in structural biology and major achievements are highlighted in this contribution.
Published: 2015

50. Disseminated rhinovirus C8 infection with infectious virus in blood and fatal outcome in a child with repeated episodes of bronchiolitis

Author: Bruno Lina, Patrice Morand, Nathalie Lin-Marq, Samuel Constant, Laurent Kaiser, Julie Bardet, Christine Morel-Baccard, Isabelle Schuffenecker, Johannes Alexander Lobrinus, Caroline Tapparel, Julien Lupo, Valérie Payen, 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), French National Enterovirus/Parechovirus Reference centre, Hospices Civils de Lyon (HCL), Centre hospitalier universitaire de Grenoble (CHU de Grenoble), CHU Grenoble, and University of Geneva [Switzerland]
Subjects: Microbiology (medical), Virus Cultivation, Fatal outcome, Rhinovirus, Respiratory System, Viremia, Case Reports, medicine.disease_cause, Feces, 03 medical and health sciences, Fatal Outcome, 0302 clinical medicine, Cerebrospinal fluid, stomatognathic system, medicine, Humans, 030212 general & internal medicine, Respiratory system, Cerebrospinal Fluid, 030304 developmental biology, ddc:616, 0303 health sciences, Picornaviridae Infections, business.industry, Infant, virus diseases, medicine.disease, Virology, 3. Good health, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Blood, Bronchiolitis, Immunology, Female, Airway, business
Abstract: We report a fatal case of acute lower respiratory tract disease with human rhinovirus C (HRV-C) as the unique cause in a 19-month-old girl with a history of repeated episodes of bronchiolitis. HRV-C type 8 nucleic acids were observed in respiratory, stool, and cerebrospinal fluid samples, and infectious virions were isolated from patient serum after inoculation onto reconstituted airway epithelia.
Published: 2015

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