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5 results on '"Sylviane Hoos"'

1. A High‐Affinity Calmodulin‐Binding Site in the CyaA Toxin Translocation Domain is Essential for Invasion of Eukaryotic Cells

Author

Alexis Voegele, Mirko Sadi, Darragh Patrick O'Brien, Pauline Gehan, Dorothée Raoux‐Barbot, Maryline Davi, Sylviane Hoos, Sébastien Brûlé, Bertrand Raynal, Patrick Weber, Ariel Mechaly, Ahmed Haouz, Nicolas Rodriguez, Patrice Vachette, Dominique Durand, Sébastien Brier, Daniel Ladant, and Alexandre Chenal

Subjects
adenylate cyclase, Bordetella pertussis, calmodulin, CyaA toxin, membrane translocation, protein membrane interaction, Science
Abstract

Abstract The molecular mechanisms and forces involved in the translocation of bacterial toxins into host cells are still a matter of intense research. The adenylate cyclase (CyaA) toxin from Bordetella pertussis displays a unique intoxication pathway in which its catalytic domain is directly translocated across target cell membranes. The CyaA translocation region contains a segment, P454 (residues 454–484), which exhibits membrane‐active properties related to antimicrobial peptides. Herein, the results show that this peptide is able to translocate across membranes and to interact with calmodulin (CaM). Structural and biophysical analyses reveal the key residues of P454 involved in membrane destabilization and calmodulin binding. Mutational analysis demonstrates that these residues play a crucial role in CyaA translocation into target cells. In addition, calmidazolium, a calmodulin inhibitor, efficiently blocks CyaA internalization. It is proposed that after CyaA binding to target cells, the P454 segment destabilizes the plasma membrane, translocates across the lipid bilayer and binds calmodulin. Trapping of CyaA by the CaM:P454 interaction in the cytosol may assist the entry of the N‐terminal catalytic domain by converting the stochastic motion of the polypeptide chain through the membrane into an efficient vectorial chain translocation into host cells.

Published
2021
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2. The c‐di‐ <scp>AMP</scp> ‐binding protein <scp>CbpB</scp> modulates the level of <scp>ppGpp</scp> alarmone in Streptococcus agalactiae

Author

Giovanni Covaleda‐Cortés, Ariel Mechaly, Terry Brissac, Heike Baehre, Laura Devaux, Patrick England, Bertrand Raynal, Sylviane Hoos, Myriam Gominet, Arnaud Firon, Patrick Trieu‐Cuot, Pierre Alexandre Kaminski, Biologie des Bactéries pathogènes à Gram-positif - Biology of Gram-Positive Pathogens, Université Paris Cité (UPCité)-Microbiologie Intégrative et Moléculaire (UMR6047), Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Cristallographie (Plateforme) - Crystallography (Platform), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité), Hannover Medical School [Hannover] (MHH), Biophysique Moléculaire (plateforme) - Molecular Biophysics (platform), This work was supported by grants from the French Government ‘Laboratory of Excellence - Integrative Biology of Emerging Infectious Diseases’ (LabEx IBEID, grant number ANR-10-LABX-62-IBEID to PTC), the Fondation pour la Recherche Médicale (FRM grant number DEQ20181039599 to PTC)., We acknowledge SOLEIL for provision of synchrotron radiation facilities, and we thank the staff of beamlines PROXIMA-1 and PROXIMA-2A for assistance. The authors are grateful to the Staff of the Crystallography platform at the Institut Pasteur for robot-driven crystallization screenings. We acknowledge Eduard Baquero Salazar for his help during the early stage of this work. We also acknowledge the help of Mariette Matondo and Thibault Chaze from the Proteomics facility of Institut Pasteur., and ANR-10-LABX-0062,IBEID,Integrative Biology of Emerging Infectious Diseases(2010)

Subjects
CbpB protein, Cyclic diadenosine monophosphate (c‐di‐AMP), [SDV]Life Sciences [q-bio], X‐ray crystallography & binding, Streptococcus, Cell Biology, Potassium homeostasis, CBS domain, Molecular Biology, Biochemistry
Abstract

International audience; Cyclic di-AMP is an essential signaling molecule in Gram-positive bacteria. This second messenger regulates the osmotic pressure of the cell by interacting directly with the regulatory domains, either RCK_C or CBS domains, of several potassium and osmolyte uptake membrane protein systems. Cyclic di-AMP also targets stand-alone CBS domain proteins such as DarB in Bacillus subtilis and CbpB in Listeria monocytogenes. We show here that the CbpB protein of Group B Streptococcus binds c-di-AMP with a very high affinity. Crystal structures of CbpB reveal the determinants of binding specificity and significant conformational changes occurring upon c-di-AMP binding. Deletion of the cbpB gene alters bacterial growth in low potassium conditions most likely due to a decrease in the amount of ppGpp caused by a loss of interaction between CbpB and Rel, the GTP/GDP pyrophosphokinase.

Published
2023

4. A High‐Affinity Calmodulin‐Binding Site in the CyaA Toxin Translocation Domain is Essential for Invasion of Eukaryotic Cells

Author

Nicolas Rodriguez, Daniel Ladant, Maryline Davi, Ahmed Haouz, Bertrand Raynal, Darragh P. O'Brien, Alexis Voegele, Ariel E. Mechaly, Patrick Weber, Sébastien Brûlé, Pauline Gehan, Dominique Durand, Mirko Sadi, Sylviane Hoos, Sébastien Brier, Alexandre Chenal, Dorothée Raoux-Barbot, Patrice Vachette, Biochimie des Interactions Macromoléculaires / Biochemistry of Macromolecular Interactions, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Université de Paris (UP), Laboratoire des biomolécules (LBM UMR 7203), Centre National de la Recherche Scientifique (CNRS)-Sorbonne Université (SU)-Département de Chimie - ENS Paris, École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Chimie Moléculaire de Paris Centre (FR 2769), Institut de Chimie du CNRS (INC)-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)-Ecole Nationale Supérieure de Chimie de Paris - Chimie ParisTech-PSL (ENSCP), Université Paris sciences et lettres (PSL)-Ecole Superieure de Physique et de Chimie Industrielles de la Ville de Paris (ESPCI Paris), Université Paris sciences et lettres (PSL)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Ecole Nationale Supérieure de Chimie de Paris - Chimie ParisTech-PSL (ENSCP), Université Paris sciences et lettres (PSL)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM), Biophysique Moléculaire (plateforme) - Molecular Biophysics (platform), Cristallographie (Plateforme) - Crystallography (Platform), Institut de Biologie Intégrative de la Cellule (I2BC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS), Plateforme technologique de RMN biologique - Biological NMR Technological Platform, A.V. was supported by a DIM MalInf grant from the region Ile‐de‐France. M.S. was supported by the Pasteur – Paris University (PPU) International PhD Program. D.P.O.B. was supported by Institut Pasteur (grants PasteurInnoV15006‐01A and PTR451). P.G. was supported by Sorbonne Université. Funding is acknowledged from Agence Nationale de la Recherche (grant number CACSICE Equipex ANR‐11‐EQPX‐0008). Region Ile de France (grant number DIM MalInf 2016), CNRS (UMR 3528), Institut Pasteur (grant numbers PasteurInnoV15006‐01A, PTR451 and PTR166‐19, PPUIP program). The funders have no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript., The authors acknowledge SOLEIL and ESRF for provision of synchrotron radiation facilities. The authors thank the staffs of the DISCO, PROXIMA‐1, and SWING beamlines for constant support and help during data collection (Synchrotron SOLEIL, St Aubin, France) and MASSIF (Synchrotron ESRF, Grenoble, France) beamlines for assistance during the X‐ray diffraction data collection., ANR-11-EQPX-0008,CACSICE,Centre d'analyse de systèmes complexes dans les environnements complexes(2011), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), Université Paris Cité (UPCité), Chimie Moléculaire de Paris Centre (FR 2769), École normale supérieure - Paris (ENS-PSL), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Ecole Nationale Supérieure de Chimie de Paris - Chimie ParisTech-PSL (ENSCP), Université Paris sciences et lettres (PSL)-Institut de Chimie du CNRS (INC)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Paris (ENS-PSL), Université Paris sciences et lettres (PSL)-Institut de Chimie du CNRS (INC)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS)-Département de Chimie - ENS Paris, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Ecole Nationale Supérieure de Chimie de Paris - Chimie ParisTech-PSL (ENSCP), Université Paris sciences et lettres (PSL)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Ecole Nationale Supérieure de Chimie de Paris - Chimie ParisTech-PSL (ENSCP), Université Paris sciences et lettres (PSL)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM), Chenal, Alexandre, and Equipements d'excellence - Centre d'analyse de systèmes complexes dans les environnements complexes - - CACSICE2011 - ANR-11-EQPX-0008 - EQPX - VALID

Subjects
calmodulin, General Chemical Engineering, General Physics and Astronomy, Medicine (miscellaneous), 02 engineering and technology, [SDV.BBM.BM] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, 01 natural sciences, Bordetella pertussis, protein membrane interaction, General Materials Science, Internalization, Lipid bilayer, [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], media_common, biology, Full Paper, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Chemistry, General Engineering, Full Papers, 021001 nanoscience & nanotechnology, [SDV.BBM.BP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biophysics, Protein Transport, Eukaryotic Cells, Adenylate Cyclase Toxin, 0210 nano-technology, Protein Binding, adenylate cyclase, Calmodulin, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], CyaA toxin, media_common.quotation_subject, Science, Antimicrobial peptides, [SDV.BBM.BP] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biophysics, protein–protein interactions, 010402 general chemistry, Biochemistry, Genetics and Molecular Biology (miscellaneous), Protein–protein interaction, Protein Domains, membrane translocation, Binding site, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Binding Sites, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, cyaA, 0104 chemical sciences, Cytosol, Biophysics, biology.protein
Abstract

The molecular mechanisms and forces involved in the translocation of bacterial toxins into host cells are still a matter of intense research. The adenylate cyclase (CyaA) toxin from Bordetella pertussis displays a unique intoxication pathway in which its catalytic domain is directly translocated across target cell membranes. The CyaA translocation region contains a segment, P454 (residues 454–484), which exhibits membrane‐active properties related to antimicrobial peptides. Herein, the results show that this peptide is able to translocate across membranes and to interact with calmodulin (CaM). Structural and biophysical analyses reveal the key residues of P454 involved in membrane destabilization and calmodulin binding. Mutational analysis demonstrates that these residues play a crucial role in CyaA translocation into target cells. In addition, calmidazolium, a calmodulin inhibitor, efficiently blocks CyaA internalization. It is proposed that after CyaA binding to target cells, the P454 segment destabilizes the plasma membrane, translocates across the lipid bilayer and binds calmodulin. Trapping of CyaA by the CaM:P454 interaction in the cytosol may assist the entry of the N‐terminal catalytic domain by converting the stochastic motion of the polypeptide chain through the membrane into an efficient vectorial chain translocation into host cells., The mechanism of cell invasion by bacterial toxins is still poorly understood. The adenylate cyclase (CyaA) toxin from Bordetella pertussis, the causative agent of whooping cough, directly translocates its catalytic domain across plasma membranes. The membrane‐permeabilizing segment (yellow) of CyaA translocates across the plasma membrane and binds calmodulin (red), which assists the entry of the catalytic domain (blue) into host cells while the hydrophobic and acylation domains (green) interact with the membrane and the C‐terminal Repeat‐in‐Toxin domain (grey) remains in the extra‐cellular milieu.

Published
2021

5. Characterization of the elongasome core PBP2 : MreC complex of Helicobacter pylori

Author

Sylviane Hoos, Christine Ebel, Alexandre Martins, Christine Schmitt, Frank Gabel, Chantal Ecobichon, Patrick England, Andréa Dessen, Pierre-Jean Matteï, Ivo G. Boneca, Meriem El Ghachi, Marie-Christine Prévost, and Frédéric Colland

Subjects
chemistry.chemical_classification, 0303 health sciences, 030306 microbiology, Cell, Periplasmic space, biochemical phenomena, metabolism, and nutrition, Biology, Cell morphology, Microbiology, Yeast, Bacterial cell structure, Amino acid, law.invention, 03 medical and health sciences, medicine.anatomical_structure, chemistry, Biochemistry, law, polycyclic compounds, Recombinant DNA, Biophysics, medicine, Surface plasmon resonance, Molecular Biology, 030304 developmental biology
Abstract

Summary The definition of bacterial cell shape is a complex process requiring the participation of multiple components of an intricate macromolecular machinery. We aimed at characterizing the determinants involved in cell shape of the helical bacterium Helicobacter pylori. Using a yeast two-hybrid screen with the key cell elongation protein PBP2 as bait, we identified an interaction between PBP2 and MreC. The minimal region of MreC required for this interaction ranges from amino acids 116 to 226. Using recombinant proteins, we showed by affinity and size exclusion chromatographies and surface plasmon resonance that PBP2 and MreC form a stable complex. In vivo, the two proteins display a similar spatial localization and their complex has an apparent 1:1 stoichiometry; these results were confirmed in vitro by analytical ultracentrifugation and chemical cross-linking. Small angle X-ray scattering analyses of the PBP2 : MreC complex suggest that MreC interacts directly with the C-terminal region of PBP2. Depletion of either PBP2 or MreC leads to transition into spherical cells that lose viability. Finally, the specific expression in trans of the minimal interacting domain of MreC with PBP2 in the periplasmic space leads to cell rounding, suggesting that the PBP2/MreC complex formation in vivo is essential for cell morphology.

Published
2011

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