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1. The Sec61 translocon is a therapeutic vulnerability in multiple myeloma

Author: Antoine Domenger, Caroline Choisy, Ludivine Baron, Véronique Mayau, Emeline Perthame, Ludovic Deriano, Bertrand Arnulf, Jean‐Christophe Bories, Gilles Dadaglio, Caroline Demangel, Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), Université Paris Cité (UPCité), Université Sorbonne Paris Cité (USPC), Immunologie humaine, physiopathologie & immunothérapie (HIPI (UMR_S_976 / U976)), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), Hub Bioinformatique et Biostatistique - Bioinformatics and Biostatistics HUB, Institut Pasteur [Paris] (IP)-Université Paris Cité (UPCité), Intégrité du génome, immunité et cancer - Genome integrity, Immunity and Cancer, Hopital Saint-Louis [AP-HP] (AP-HP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), This work was supported by the Cancéropôle Ile de France (CD and J-CB, Emergence 2019), the Fondation Française pour la Recherche contre le Myélome et les Gammapathies (CD, FFRMG), and the Sanofi iAwards program (CD, 2020). CD also acknowledges core support from Institut Pasteur and INSERM (U1221). LB was recipient of a Roux-Cantarini post-doctoral fellowship (Institut Pasteur). AD is a BioSPC-Université de Paris PhD student, recipient of a doctoral fellowship from the Ministère français de l’Enseignement Supérieur, de la Recherche et de l’Innovation., and Kop, Marie-Luce
Subjects: Sec61 translocon, Proteasome Endopeptidase Complex, [SDV.IMM] Life Sciences [q-bio]/Immunology, proteostatic stress, [SDV]Life Sciences [q-bio], Antineoplastic Agents, Endoplasmic Reticulum Stress, [SDV] Life Sciences [q-bio], Mice, Protein Transport, Cell Line, Tumor, [SDV.IMM]Life Sciences [q-bio]/Immunology, Animals, Humans, Molecular Medicine, Multiple Myeloma, Proteasome Inhibitors, SEC Translocation Channels
Abstract: International audience; Multiple myeloma (MM) is an incurable malignancy characterized by the uncontrolled expansion of plasma cells in the bone marrow. While proteasome inhibitors like bortezomib efficiently halt MM progression, drug resistance inevitably develop, and novel therapeutic approaches are needed. Here, we used a recently discovered Sec61 inhibitor, mycolactone, to assess the interest of disrupting MM proteostasis via protein translocation blockade. In human MM cell lines, mycolactone caused rapid defects in secretion of immunoglobulins and expression of pro-survival interleukin (IL)-6 receptor and CD40, whose activation stimulates IL-6 production. Mycolactone also triggered pro-apoptotic endoplasmic reticulum stress responses synergizing with bortezomib for induction of MM cell death and overriding acquired resistance to the proteasome inhibitor. Notably, the mycolactone-bortezomib combination rapidly killed patient-derived MM cells ex vivo, but not normal mononuclear cells. In immunodeficient mice engrafted with MM cells, it demonstrated superior therapeutic efficacy over single drug treatments, without inducing toxic side effects. Collectively, these findings establish Sec61 blockers as novel anti-MM agents and reveal the interest of targeting both the translocon and the proteasome in proteostasis-addicted tumors.
Published: 2022

2. Early antibiotic exposure and adverse outcomes in very preterm infants at low risk of early-onset sepsis: the EPIPAGE-2 cohort study

Author: Mathilde Letouzey, Elsa Lorthe, Laetitia Marchand-Martin, Gilles Kayem, Caroline Charlier, Marine Butin, Ayoub Mitha, Monique Kaminski, Valerie Benhammou, Pierre-Yves Ancel, Pascal Boileau, Laurence Foix-L'Hélias, Christèle Gras-Le Guen, Pierre Kuhn, Emeline Maisonneuve, Jeanne Sibiude, Héloïse Torchin, Equipe 1 : EPOPé - Épidémiologie Obstétricale, Périnatale et Pédiatrique (CRESS - U1153), Centre de Recherche Épidémiologie et Statistique Sorbonne Paris Cité (CRESS (U1153 / UMR_A_1125 / UMR_S_1153)), Conservatoire National des Arts et Métiers [CNAM] (CNAM), HESAM Université - Communauté d'universités et d'établissements Hautes écoles Sorbonne Arts et métiers université (HESAM)-HESAM Université - Communauté d'universités et d'établissements Hautes écoles Sorbonne Arts et métiers université (HESAM)-Université Sorbonne Paris Cité (USPC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Conservatoire National des Arts et Métiers [CNAM] (CNAM), HESAM Université - Communauté d'universités et d'établissements Hautes écoles Sorbonne Arts et métiers université (HESAM)-HESAM Université - Communauté d'universités et d'établissements Hautes écoles Sorbonne Arts et métiers université (HESAM)-Université Sorbonne Paris Cité (USPC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Centre hospitalier intercommunal de Poissy/Saint-Germain-en-Laye - CHIPS [Poissy], Geneva University Hospitals and Geneva University, CHU Trousseau [APHP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), Centre collaborateur de l'OMS Listeria / WHO Collaborating Centre Listeria (CC-OMS / WHO-CC), Institut Pasteur [Paris] (IP)-Organisation Mondiale de la Santé / World Health Organization Office (OMS / WHO)-Université Paris Cité (UPCité), Biologie des Infections - Biology of Infection, Centre National de Référence Listeria - National Reference Center Listeria (CNRL), Hospices Civils de Lyon (HCL), Hôpital Jeanne de Flandres, Université de Lille, Droit et Santé-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), CIC - Mère Enfant Necker Cochin Paris Centre (CIC 1419), Hôpital Cochin [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)-Université Paris Cité (UPCité), Université de Versailles Saint-Quentin-en-Yvelines - UFR Sciences de la santé Simone Veil (UVSQ Santé), Université de Versailles Saint-Quentin-en-Yvelines (UVSQ), he EPIPAGE-2 study was funded by the French Institute of Public Health Research/Institute of Public Health and its partners: the French Health Ministry, National Institute of Health and Medical Research (Inserm), National Institute of Cancer, and National Solidarity Fund for Autonomy (CNSA). Additional support for this study was provided by the National Research Agency through the French EquipEx 'Investments in the Future' Program (ANR-11-EQPX-0038 and ANR-19-COHO-001), the PREMUP Foundation, Fondation de France (11779), and Fondation pour la Recherche M edicale (SPF20160936356). The funding sources had no role in the design and conduct of the study, collection, management, analysis, and interpretation of the data, preparation, review, or approval of the manuscript, or the decision to submit the manuscript for publication. The authors declare no conflicts of interest., ANR-11-EQPX-0038,RE-CO-NAI,Plateforme de REcherche sur les COhortes d'enfants suivis depuis la NAIssance(2011), ANR-19-COHO-0001,RE-CO-NAI (COHORTES),Resarch platform on cohorts of children followed from birth(2019), LETOUZEY, Mathilde, Equipements d'excellence - Plateforme de REcherche sur les COhortes d'enfants suivis depuis la NAIssance - - RE-CO-NAI2011 - ANR-11-EQPX-0038 - EQPX - VALID, Resarch platform on cohorts of children followed from birth - - RE-CO-NAI (COHORTES)2019 - ANR-19-COHO-0001 - COHO - VALID, and Centre National de Référence Listeria - National Reference Center Listeria (CNR)
Subjects: neonatal death, [SDV.MHEP.PED]Life Sciences [q-bio]/Human health and pathology/Pediatrics, necrotizing enterocolitis, prematurity, preterm birth, severe cerebral lesions, empirical antibiotics, neonatal outcome, [SDV.MHEP.PED] Life Sciences [q-bio]/Human health and pathology/Pediatrics, [SDV.SPEE] Life Sciences [q-bio]/Santé publique et épidémiologie, Pediatrics, Perinatology and Child Health, late-onset sepsis, bronchopulmonary dysplasia, [SDV.SPEE]Life Sciences [q-bio]/Santé publique et épidémiologie
Abstract: International audience; Objective: To assess the association between early empirical antibiotics and neonatal adverse outcomes in very preterm infants without risk factors of early-onset sepsis.Study design: This is a secondary analysis of the EPIPAGE-2 study, a prospective national population-based cohort that included all liveborn infants at 22-31 completed weeks of gestation in France in 2011. Infants at high risk of early-onset sepsis (born after preterm labor or preterm premature rupture of membranes, or from mother who had clinical chorioamnionitis or received antibiotics during the last 72 hours) were excluded. Early antibiotic exposure was defined as antibiotics started at Day 0 or Day 1 of life, regardless of the duration and type of antibiotics. We compared treated and untreated patients by using inverse probability of treatment weighting based on estimated propensity score. Results: Among 648 very preterm infants at low risk of early-onset sepsis, 173 (26.2%) had early antibiotics treatment. Early antibiotic exposure was not associated with death or late-onset sepsis or necrotizing enterocolitis (OR=1.04 [95%CI=0.72-1.50]). However, it was associated with higher odds of severe cerebral lesions (OR=2.71 [95%CI=1.25-5.86]) and moderate-to-severe bronchopulmonary dysplasia (OR=2.30 [95%CI=1.21-4.38]). Conclusion: Early empirical antibiotics administrated in very preterm infants at low risk of early-onset sepsis were associated with a higher risk of severe cerebral lesions and moderate-to-severe bronchopulmonary dysplasia.
Published: 2022

3. De novo synthesized polyunsaturated fatty acids operate as both host immunomodulators and nutrients for Mycobacterium tuberculosis

Author: Thomas Laval, Laura Pedró-Cos, Wladimir Malaga, Laure Guenin-Macé, Alexandre Pawlik, Véronique Mayau, Hanane Yahia-Cherbal, Océane Delos, Wafa Frigui, Justine Bertrand-Michel, Christophe Guilhot, Caroline Demangel, Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), Université Paris Cité (UPCité), Université Sorbonne Paris Cité (USPC), Institut de pharmacologie et de biologie structurale (IPBS), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS), Pathogénomique mycobactérienne intégrée - Integrated Mycobacterial Pathogenomics, 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), Immunorégulation - Immunoregulation, Institut Pasteur [Paris] (IP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Paris Cité (UPCité), MetaboHUB-MetaToul, Génopole Toulouse Midi-Pyrénées [Auzeville] (GENOTOUL), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Ecole Nationale Vétérinaire de Toulouse (ENVT), Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Institut des Maladies Métaboliques et Casdiovasculaires (UPS/Inserm U1297 - I2MC), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM), This study was supported by the Institut Pasteur (TL, CD) and INSERM (U1224, CD)., We thank the Cytometry and Biomarkers (UTechS CB) Technological Platform for help with nCounter FLEX NanoString use and the Image Analysis Hub of Institut Pasteur for confocal microscopy data analysis. We gratefully acknowledge the MetaToul (Toulouse metabolomics and fluxomics facilities, https://www.metatoul.fr), part of the French National Infrastructure for Metabolomics and Fluxomics MetaboHUB-ANR-11-INBS-0010, for lipid analysis. We also thank Prof. Luke Chamberlain (University of Strathclyde, Glasgow, UK) for sharing his expertise of FA labeling by click-chemistry., Kop, Marie-Luce, Université de Toulouse (UT)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS), MetaboHUB-Génopole Toulouse Midi-Pyrénées [Auzeville] (GENOTOUL), Université de Toulouse (UT)-Université de Toulouse (UT)-Ecole Nationale Vétérinaire de Toulouse (ENVT), Université de Toulouse (UT)-Université de Toulouse (UT)-Institut National Polytechnique (Toulouse) (Toulouse INP), Université de Toulouse (UT)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Université de Toulouse (UT)-Université de Toulouse (UT)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Pasteur [Paris], Université Paris Cité (UPC)-Université Paris Cité (UPC)-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Paris Cité (UPC), Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées, Centre National de la Recherche Scientifique (CNRS)-Institut Pasteur [Paris], Université Paris Cité (UPC)-Université Paris Cité (UPC), and MetaToul-MetaboHUB
Subjects: Male, [SDV.IMM] Life Sciences [q-bio]/Immunology, Mouse, QH301-705.5, Science, infectious disease, [SDV]Life Sciences [q-bio], immunomodulation, fatty acids, Cell Line, immunology, Mice, Immunology and Inflammation, Animals, Humans, Immunologic Factors, Biology (General), innate immunity, Microbiology and Infectious Disease, nutrient, microbiology, Mycobacterium tuberculosis, Nutrients, respiratory system, Immunity, Innate, macrophages, [SDV] Life Sciences [q-bio], inflammation, Fatty Acids, Unsaturated, Medicine, [SDV.IMM]Life Sciences [q-bio]/Immunology, Female, Research Article
Abstract: International audience; Successful control of Mycobacterium tuberculosis (Mtb) infection by macrophages relies on immunometabolic reprogramming, where the role of fatty acids (FAs) remains poorly understood. Recent studies unraveled Mtb’s capacity to acquire saturated and monounsaturated FAs via the Mce1 importer. However, upon activation, macrophages produce polyunsaturated fatty acids (PUFAs), mammal-specific FAs mediating the generation of immunomodulatory eicosanoids. Here, we asked how Mtb modulates de novo synthesis of PUFAs in primary mouse macrophages and whether this benefits host or pathogen. Quantitative lipidomics revealed that Mtb infection selectively activates the biosynthesis of ω6 PUFAs upstream of the eicosanoid precursor arachidonic acid (AA) via transcriptional activation of Fads2 . Inhibiting FADS2 in infected macrophages impaired their inflammatory and antimicrobial responses but had no effect on Mtb growth in host cells nor mice. Using a click-chemistry approach, we found that Mtb efficiently imports ω6 PUFAs via Mce1 in axenic culture, including AA. Further, Mtb preferentially internalized AA over all other FAs within infected macrophages by mechanisms partially depending on Mce1 and supporting intracellular persistence. Notably, IFNγ repressed de novo synthesis of AA by infected mouse macrophages and restricted AA import by intracellular Mtb. Together, these findings identify AA as a major FA substrate for intracellular Mtb, whose mobilization by innate immune responses is opportunistically hijacked by the pathogen and downregulated by IFNγ.
Published: 2021

4. Biochemical and Biological Assays of Mycolactone-Mediated Inhibition of Sec61

Author: Caroline Demangel, Sarah O'Keefe, Stephen High, University of Manchester [Manchester], Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), and Kop, Marie-Luce
Subjects: Sec61, [SDV.IMM] Life Sciences [q-bio]/Immunology, [SDV]Life Sciences [q-bio], Cell, MESH: Biological Assay, chemistry.chemical_compound, In vitro transcription and translation, Ubiquitin, MESH: Buruli Ulcer, Microsomes, medicine, Humans, Endoplasmic reticulum (ER), Flow cytometry, Mycolactone, Buruli Ulcer, Sec61 translocon, MESH: Humans, biology, Chemistry, Endoplasmic reticulum, Enzyme-linked immunosorbent assay (ELISA), Membrane Proteins, Cell biology, [SDV] Life Sciences [q-bio], Cytosol, medicine.anatomical_structure, Membrane protein, Proteasome, biology.protein, [SDV.IMM]Life Sciences [q-bio]/Immunology, Biological Assay, Co-translational translocation, MESH: Membrane Proteins, Macrolides, MESH: Macrolides, MESH: SEC Translocation Channels, SEC Translocation Channels
Abstract: International audience; Mycobacterium ulcerans, the causative agent of Buruli ulcer disease, is unique among human pathogens in its capacity to produce mycolactone, a diffusible macrolide with immunosuppressive and cytotoxic properties. Recent studies have shown that mycolactone operates by inhibiting the host membrane translocation complex (Sec61), with an unprecedented potency compared to previously identified Sec61 blockers. Mycolactone binding to the pore-forming subunit of Sec61 inhibits its capacity to transport nascent secretory and membrane proteins into the endoplasmic reticulum, leading to their cytosolic degradation by the ubiquitin:proteasome system. In T lymphocytes, Sec61 blockade by mycolactone manifests as a sharp decrease in the cell's ability to express homing receptors and release cytokines following activation. Sustained exposure of human cells to mycolactone typically generates proteotoxic stress responses in their cytosol and endoplasmic reticulum (ER), ultimately inducing apoptosis. Here we describe cell-free systems for studying Sec61-mediated protein translocation that allow the impact of mycolactone on the biogenesis of secretory and membrane proteins to be probed. We also describe biological assays of mycolactone-driven inhibition of Sec61 providing rapid and sensitive means to quantitatively assess the presence of the toxin in biological samples.
Published: 2021

5. Molecular Mechanisms Underpinning the Circulation and Cellular Uptake of Mycobacterium ulcerans Toxin Mycolactone

Author: Bruno Tello Rubio, Florence Bugault, Blandine Baudon, Bertrand Raynal, Sébastien Brûlé, Jean-David Morel, Sarah Saint-Auret, Nicolas Blanchard, Caroline Demangel, Laure Guenin-Macé, Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), Biophysique Moléculaire (plateforme) - Molecular Biophysics (platform), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité), Laboratoire d'innovation moléculaire et applications (LIMA), Université de Strasbourg (UNISTRA)-Université de Haute-Alsace (UHA) Mulhouse - Colmar (Université de Haute-Alsace (UHA))-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), This work was supported by the Fondation Raoul Follereau., Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), and Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS)
Subjects: Pharmacology, SR-B1, Mycobacterium ulcerans, lipid carriers, uptake, [SDV.SP.PHARMA]Life Sciences [q-bio]/Pharmaceutical sciences/Pharmacology, mycolactone, Therapeutics. Pharmacology, RM1-950, Original Research
Abstract: International audience; Mycolactone is a diffusible lipid toxin produced by Mycobacterium ulcerans, the causative agent of Buruli ulcer disease. Altough bacterially derived mycolactone has been shown to traffic from cutaneous foci of infection to the bloodstream, the mechanisms underpinning its access to systemic circulation and import by host cells remain largely unknown. Using biophysical and cell-based approaches, we demonstrate that mycolactone specific association to serum albumin and lipoproteins is necessary for its solubilization and is a major mechanism to regulate its bioavailability. We also demonstrate that Scavenger Receptor (SR)-B1 contributes to the cellular uptake of mycolactone. Overall, we suggest a new mechanism of transport and cell entry, challenging the dogma that the toxin enters host cells via passive diffusion.
Published: 2021

6. Insights into the ancestry evolution of the Mycobacterium tuberculosis complex from analysis of Mycobacterium riyadhense

Author: Talal AlMutairi, Shamsudeen F. Fagbo, Muhammad Amin urRahman, Albel Singh, Caroline Demangel, Arnab Pain, Stephen V. Gordon, Alicia Smyth, Thomas Laval, Yasuhiko Suzuki, Sara Mfarrej, John A. Browne, Anwar Hoosen, Faisal A. Alasmari, Chie Nakajima, Apoorva Bhatt, Musa A. Garbati, Alya Alruwaili, Qingtian Guan, Conor J. Meehan, King Abdullah University of Science and Technology (KAUST), King Fahad Medical City [Riyadh] (KFMC), Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), Université Paris Diderot - Paris 7 (UPD7), University of Birmingham [Birmingham], Saudi Center for Disease Prevention and Control [Riyadh, Saudi Arabia], Nigerian Institute of Medical Research [Yaba, Lagos, Nigeria] (NIMR), University College Dublin [Dublin] (UCD), University of Bradford, Hokkaido University [Sapporo, Japan], KAUST [BAS/1/1020-01- 01], BBSRC [BB/N004574/1 to S.G., A.B.], SFI [SFI/15/IA/3154 to S.G.], We wish to thank members of the Bioscience Core Lab (BCL) of King Abdullah University of Science and Technology for sequencing the bacterial genomic DNAs on the Illumina HiSeq 4000 and PacBio RSII platforms., and Kop, Marie-Luce
Subjects: Genetics, Tuberculosis, [SDV.IMM] Life Sciences [q-bio]/Immunology, [SDV]Life Sciences [q-bio], Virulence, Biology, biology.organism_classification, medicine.disease, Genome, Transcriptome, Mycobacterium tuberculosis, [SDV] Life Sciences [q-bio], Mycobacterium tuberculosis complex, medicine, [SDV.IMM]Life Sciences [q-bio]/Immunology, Antitoxin, Gene
Abstract: Current evolutionary scenarios posit the emergence of Mycobacterium tuberculosis from an environmental saprophyte through a cumulative process of genome adaptation. Mycobacterium riyadhense, a related bacillus, is being increasingly isolated from human clinical cases with tuberculosis-like symptoms in various parts of the world. To elucidate the evolutionary relationship between M. riyadhense and other mycobacterial species, including members of the M. tuberculosis complex (MTBC), eight clinical isolates of M. riyadhense were sequenced and analyzed. We show, among other features, that M. riyadhense shares a large number of conserved orthologs with M. tuberculosis and shows the expansion of toxin/antitoxin pairs, PE/PPE family proteins compared with other non-tuberculous mycobacteria. We observed M. riyadhense lacks wecE gene which may result in the absence of lipooligosaccharides (LOS) IV. Comparative transcriptomic analysis of infected macrophages reveals genes encoding inducers of Type I IFN responses, such as cytosolic DNA sensors, were relatively less expressed by macrophages infected with M. riyadhense or M. kansasii, compared to BCG or M. tuberculosis. Overall, our work sheds new light on the evolution of M. riyadhense, its relationship to the MTBC, and its potential as a system for the study of mycobacterial virulence and pathogenesis.
Published: 2021

7. The impact of gut microbiota on depressive-like behaviors and adult hippocampal neurogenesis requires the endocannabinoid system

Author: Ivo G. Boneca, Gabriel Lepousez, Laure Guenin-Macé, Aline Rifflet, Thomas Laval, Gérard Eberl, Eleni Siopi, Pierre-Marie Lledo, Caroline Demangel, Maud Pascal, Grégoire Chevalier, François Leulier, Microenvironnement et Immunité - Microenvironment and Immunity, Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur [Paris], Perception et Mémoire / Perception and Memory, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Biologie et Génétique de la Paroi bactérienne - Biology and Genetics of Bacterial Cell Wall (BGPB), Institut Pasteur [Paris], Institut de Génomique Fonctionnelle de Lyon (IGFL), 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 Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), Institut Pasteur [Paris] (IP), École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), and lepousez, gabriel
Subjects: 0303 health sciences, medicine.medical_specialty, [SCCO.NEUR]Cognitive science/Neuroscience, [SCCO.NEUR] Cognitive science/Neuroscience, Neurogenesis, Biology, Hippocampal formation, Gut flora, medicine.disease, biology.organism_classification, Endocannabinoid system, 03 medical and health sciences, chemistry.chemical_compound, 0302 clinical medicine, Endocrinology, chemistry, Mood disorders, Internal medicine, medicine, Arachidonic acid, Chronic stress, Dysbiosis, 030217 neurology & neurosurgery, 030304 developmental biology
Abstract: SUMMARYDepression is the leading cause of disability worldwide. Recent observations have revealed an association between mood disorders and alterations of the intestinal microbiota, but causality remains yet to be established. Here, using unpredictable chronic mild stress (UCMS) as a mouse model of depression, we show that the UCMS mice display phenotypic alterations — characterized by an altered gut microbiota composition, a reduced adult hippocampal neurogenesis and a depressive-like behaviors — which could be transferred from UCMS donors to naïve recipient mice by fecal microbiota transplantation. The cellular and behavioral alterations observed in recipient mice were accompanied by a decrease in the endocannabinoid (eCB) signaling due to lower peripheral levels of fatty acid precursors of eCB ligands. The adverse effects of UCMS-transferred microbiota on adult neurogenesis and behavior in naïve recipient mice were alleviated by selectively enhancing the central eCB tone or by adding arachidonic acid, a fatty acid precursor of eCB ligands, to the diet. In the gut of both UCMS donors and recipients, the microbiota composition was characterized by a relative decrease in Lactobacilli abundance, and complementation of the UCMS recipient microbiota with a strain of the Lactobacilli genus was sufficient to restore normal eCB brain levels, hippocampal neurogenesis and to alleviate depressive-like behaviors. Our findings provide a mechanistic scenario for how chronic stress, diet and gut microbiota dysbiosis generate a pathological feed-forward loop that contributes to despair behavior via the central eCB system.
Published: 2021

8. Not too fat to fight -The emerging role of macrophage fatty acid metabolism in immunity to Mycobacterium tuberculosis

Author: Lise Chaumont, Thomas Laval, Caroline Demangel, Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Université de Paris (UP), C.D. acknowledges core support from Institut Pasteur and INSERM (U1221). T.L. was a BioSPC- Université de Paris PhD student, recipient of doctoral fellowships from the Ministère français de l’Enseignement Supérieur, de la Recherche et de l’Innovation (2016-2019) and Fondation pour la Recherche Médicale (FDT201904008040)., Kop, Marie-Luce, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), and Université Paris Cité (UPCité)
Subjects: 0301 basic medicine, [SDV.IMM] Life Sciences [q-bio]/Immunology, Immunology, cell lineages and subsets, Virulence, Biology, infectious diseases, Microbiology, lung, 03 medical and health sciences, 0302 clinical medicine, Lipid droplet, Humans, Tuberculosis, Immunology and Allergy, Macrophage, bacterial, Pathogen, Phagosome, Fatty Acids, Lipid metabolism, Lipid signaling, Mycobacterium tuberculosis, Lipid Metabolism, 3. Good health, macrophages, 030104 developmental biology, fatty acid metabolism, Host-Pathogen Interactions, [SDV.IMM]Life Sciences [q-bio]/Immunology, monocytes, Intracellular, 030215 immunology
Abstract: International audience; While the existence of a special relationship between Mycobacterium tuberculosis (Mtb) and host lipids has long been known, it remains a challenging enigma. It was clearly established that Mtb requires host fatty acids (FAs) and cholesterol to produce energy, build its distinctive lipid-rich cell wall and produce lipid virulence factors. It was also observed that in infected hosts, Mtb constantly resides in a FA-rich environment that the pathogen contributes to generate by inducing a lipid-laden “foamy” phenotype in host macrophages. These observations, and the proximity between lipid droplets and phagosomes containing bacteria within infected macrophages gave rise to the hypothesis that Mtb reprograms host cell lipid metabolism to ensure a continuous supply of essential nutrients and its long-term persistence in vivo. However, recent studies question this principle by indicating that in Mtb-infected macrophages, lipid droplet formation prevents bacterial acquisition of host FAs while supporting the production of FA-derived protective lipid mediators. Further, in vivo investigations reveal discrete macrophage phenotypes linking the FA metabolisms of host cell and intracellular pathogen. Notably, FA storage within lipid droplets characterizes both macrophages controlling Mtb infection and dormant intracellular Mtb. In this review, we integrate findings from immunological and microbiological studies illustrating the new concept that cytoplasmic accumulation of FAs is a metabolic adaptation of macrophages to Mtb infection, which potentiates their antimycobacterial responses and forces the intracellular pathogen to shift into fat-saving, survival mode.
Published: 2021

9. Immunity against Mycobacterium ulcerans - The subversive role of mycolactone

Author: Caroline Demangel, Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut PasteurU1221/Institut National de la Santé et de la Recherche Médicale, Kop, Marie-Luce, and Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM)
Subjects: 0301 basic medicine, Buruli ulcer, [SDV.IMM] Life Sciences [q-bio]/Immunology, Immunology, Biology, 03 medical and health sciences, chemistry.chemical_compound, 0302 clinical medicine, Immune system, Immunity, Immune suppression, medicine, Humans, Immunology and Allergy, Cytotoxic T cell, Tissue Distribution, Mycolactone, Buruli Ulcer, Secretory pathway, Sec61 translocon, Bacterial < Infectious Diseases, Mycobacterium ulcerans, medicine.disease, biology.organism_classification, 3. Good health, Blockade, Skin < Tissues, 030104 developmental biology, chemistry, [SDV.IMM]Life Sciences [q-bio]/Immunology, Macrolides, 030215 immunology
Abstract: International audience; Mycobacterium ulcerans causes Buruli ulcer, a neglected tropical skin disease manifesting as chronic wounds that can leave victims with major, life-long deformity and disability. Differently from other mycobacterial pathogens, M. ulcerans produces mycolactone, a diffusible lipid factor with unique cytotoxic and immunomodulatory properties. Both traits result from mycolactone targeting Sec61, the entry point of the secretory pathway in eukaryotic cells. By inhibiting Sec61, mycolactone prevents the host cell’s production of secreted proteins, and most of its transmembrane proteins. This molecular blockade dramatically alters the functions of immune cells, thereby the generation of protective immunity. Moreover, sustained inhibition of Sec61 triggers proteotoxic stress responses leading to apoptotic cell death, which can stimulate vigorous immune responses. The dynamics of bacterial production of mycolactone and elimination by infected hosts thus critically determine the balance between its immuno-stimulatory and -suppressive effects. Following an introduction summarizing the essential information on Buruli ulcer disease, this review focuses on the current state of knowledge regarding mycolactone’s regulation and biodistribution. We then detail the consequences of mycolactone-mediated Sec61 blockade on initiation and maintenance of innate and adaptive immune responses. Finally, we discuss the key questions to address in order to improve immunity to M. ulcerans, and how increased knowledge of mycolactone biology may pave the way to innovative therapeutics.
Published: 2021

10. Antigen-Specific In Vivo Killing Assay

Author: Nada Chaoul, Gilles Dadaglio, University of Bari Aldo Moro (UNIBA), Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Margherita Gigante, Elena Ranieri, Università degli studi di Bari Aldo Moro = University of Bari Aldo Moro (UNIBA), and Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM)
Subjects: 0301 basic medicine, Lysis, biology, Chemistry, Molecular biology, Epitope, Mouse model, Antigen-specific lysis of target cells, 03 medical and health sciences, 030104 developmental biology, 0302 clinical medicine, Antigen, [SDV.IMM.IA]Life Sciences [q-bio]/Immunology/Adaptive immunology, In vivo, Antigen specific, MHC class I, biology.protein, Cytotoxic CD8+ T lymphocytes, Cytotoxic T cell, Vaccine testing, CD8, 030215 immunology, Flow cytometry-based In vivo killing assay
Abstract: International audience; The in vivo killing assay allows the quantification of the antigen-specific killing capacity of Cytotoxic CD8+ T Lymphocytes (CTLs) in mice. CTLs are indeed known for the lysis of cells expressing foreign or modified antigen peptides on their MHC class I molecules. Here we describe the detailed protocol used for the in vivo specific lysis of cells expressing the H-2 Kb immunodominant CD8+ T-cell epitope of the OVA protein: an 8 amino acid peptide corresponding to the 257-264 region of OVA (SIINFEKL).
Published: 2021

11. Mycolactone Purification from M. ulcerans Cultures and HPLC-Based Approaches for Mycolactone Quantification in Biological Samples

Author: Aline Rifflet, Caroline Demangel, Laure Guenin-Macé, Kop, Marie-Luce, Biologie et Génétique de la Paroi bactérienne - Biology and Genetics of Bacterial Cell Wall, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-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), Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), Institut Pasteur [Paris] (IP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité)-Microbiologie Intégrative et Moléculaire (UMR6047), and Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)
Subjects: Folch’s extraction, [SDV.IMM] Life Sciences [q-bio]/Immunology, [SDV]Life Sciences [q-bio], TLC, Membrane translocation, Human pathogen, MESH: Mycobacterium ulcerans, High-performance liquid chromatography, Microbiology, chemistry.chemical_compound, Polyketide synthase, Lc ms ms, Quantitative assessment, Humans, LC-MS/MS, Mycolactone, MESH: Chromatography, High Pressure Liquid, Chromatography, High Pressure Liquid, MESH: Humans, biology, Mycobacterium ulcerans, Chemistry, biology.organism_classification, [SDV] Life Sciences [q-bio], MESH: Polyketide Synthases, biology.protein, [SDV.IMM]Life Sciences [q-bio]/Immunology, Macrolides, MESH: Macrolides, Polyketide Synthases
Abstract: International audience; Mycolactones are a family of polyketide synthase products made by the human pathogen Mycobacterium ulcerans that were recently identified as novel inhibitors of the host membrane translocation complex (Sec61). Here, we provide protocols for the purification of mycolactones from bacterial cultures, and for their quantitative assessment in biological samples.
Published: 2021

12. Spatiotemporal analysis of mycolactone distribution in vivo reveals partial diffusion in the central nervous system

Author: Thomas Laval, Sarah Saint-Auret, Ivo G. Boneca, Laurent Boucontet, Caroline Demangel, Jean-Pierre Levraud, Laure Guenin-Macé, Christophe Prehaud, Anaelle da Costa, Emma Colucci-Guyon, Aline Rifflet, Nicolas Blanchard, Macrophages et Développement de l’Immunité, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Biologie et Génétique de la Paroi bactérienne - Biology and Genetics of Bacterial Cell Wall, Laboratoire d'innovation moléculaire et applications (LIMA), Université de Strasbourg (UNISTRA)-Université de Haute-Alsace (UHA) Mulhouse - Colmar (Université de Haute-Alsace (UHA))-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Neuro-Immunologie Virale - Viral Neuro-immunology, Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Paris Diderot, Sorbonne Paris Cité, Paris, France, Université Paris Diderot - Paris 7 (UPD7), This work was supported by the Institut Pasteur 'Microbes & Brain' collaborative project (LGM) and the Fondation Raoul Follereau (CD), core support from Institut Pasteur and INSERM (U1221). This study has received funding from the French Government’s Investissement d’Avenir program, Laboratoire d’Excellence 'Integrative Biology of Emerging Infectious Diseases' (grant n °ANR-10-LABX-62-IBEID) (IGB). AR was support by a post-doctoral fellowship from the Laboratoire d’Excellence 'Integrative Biology of Emerging Infectious Diseases. T.L. was a BioSPC-Université Paris Diderot PhD student, recipient of doctoral fellowships from the Ministère français de l’Enseignement Supérieur, de la Recherche et de l’Innovation (2016–2019) and Fondation pour la Recherche Médicale (FDT201904008040)., We thank the Image Analysis Hub of the Institut Pasteur for help with image analysis. We are grateful to Laurent Marsollier for the malaysian human isolate M. ulcerans 1615 and to Tim Stinear for the bioluminescent strain of M.ulcerans., ANR-10-LABX-0062,IBEID,Integrative Biology of Emerging Infectious Diseases(2010), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), and Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM)
Subjects: Central Nervous System, Bacterial Diseases, 0301 basic medicine, Buruli ulcer, Life Cycles, [SDV]Life Sciences [q-bio], RC955-962, Nervous System, chemistry.chemical_compound, Larvae, Medical Conditions, 0302 clinical medicine, Animal Cells, Arctic medicine. Tropical medicine, Medicine and Health Sciences, Mycolactone, Buruli Ulcer, Zebrafish, biology, Optical Imaging, Eukaryota, Animal Models, 3. Good health, Cell biology, Actinobacteria, Infectious Diseases, medicine.anatomical_structure, Experimental Organism Systems, Osteichthyes, Blood-Brain Barrier, Larva, Mycobacterium ulcerans, Vertebrates, Physical Sciences, [SDV.IMM]Life Sciences [q-bio]/Immunology, Macrolides, Cellular Types, Anatomy, Public aspects of medicine, RA1-1270, Research Article, Neglected Tropical Diseases, Imaging Techniques, Materials Science, Material Properties, Bacterial Toxins, Central nervous system, Glial Cells, Research and Analysis Methods, Blood–brain barrier, Permeability, Cell Line, 03 medical and health sciences, Model Organisms, Spatio-Temporal Analysis, In vivo, Fluorescence Imaging, medicine, Animals, Humans, Microglial Cells, Bacteria, Organisms, Public Health, Environmental and Occupational Health, Biology and Life Sciences, Endothelial Cells, Cell Biology, Tropical Diseases, biology.organism_classification, medicine.disease, In vitro, Fish, 030104 developmental biology, chemistry, Astrocytes, Animal Studies, Zoology, 030217 neurology & neurosurgery, Developmental Biology
Abstract: Mycobacterium ulcerans, the causative agent of Buruli ulcer (BU) disease, is unique amongst human pathogens in its capacity to produce a lipid toxin called mycolactone. While previous studies have demonstrated that bacterially-released mycolactone diffuses beyond infection foci, the spatiotemporal distribution of mycolactone remained largely unknown. Here, we used the zebrafish model to provide the first global kinetic analysis of mycolactone’s diffusion in vivo, and multicellular co-culture systems to address the critical question of the toxin’s access to the brain. Zebrafish larvae were injected with a fluorescent-derivative of mycolactone to visualize the in vivo diffusion of the toxin from the peripheral circulation. A rapid, body-wide distribution of mycolactone was observed, with selective accumulation in tissues near the injection site and brain, together with an important excretion through the gastro-intestinal tract. Our conclusion that mycolactone reached the central nervous system was reinforced by an in cellulo model of human blood brain barrier and a mouse model of M. ulcerans-infection. Here we show that mycolactone has a broad but heterogenous profile of distribution in vivo. Our investigations in vitro and in vivo support the view that a fraction of bacterially-produced mycolactone gains access to the central nervous system. The relative persistence of mycolactone in the bloodstream suggests that assays of circulating mycolactone are relevant for BU disease monitoring and treatment optimization., Author summary Mycolactone is the major virulence factor of Mycobacterium ulcerans, the human pathogen causing Buruli ulcer (BU) disease. While it is now established that mycolactone is able to diffuse from infected tissues to exert immunomodulatory and analgesic effects at the systemic level, the in vivo spatiotemporal distribution of mycolactone remained largely unknown. Here, using the zebrafish larva, we describe the spatiotemporal distribution of a fluorescent derivative of mycolactone in vivo. We show that fluorescent mycolactone quickly diffuses from the blood circulation into various organs, and accumulates in muscles and brain. Mycolactone’s diffusion into the central nervous system was further confirmed in the mouse model of M. ulcerans infection. We reported previously that mycolactone suppresses the production of inflammatory mediators by primary microglia at nanomolar concentrations in vitro [1]. In the present study, we provide evidence suggesting that bacterially-produced mycolactone is able to reach the brain. While additional in vivo investigations will be required, we can speculate that the amount of mycolactone reaching the brain in the context of M. ulcerans infection is sufficient to modulate inflammation and pain transmission.
Published: 2020

13. Repair of G1 induced DNA double-strand breaks in S-G2/M by alternative NHEJ

Author: Loelia Babin, Caroline Demangel, Hélène Lenden-Hasse, Ludovic Deriano, Ludivine Baron, Erika Brunet, José Yélamos, Marie Bedora-Faure, Wei Yu, Chloé Lescale, Intégrité du génome, immunité et cancer - Genome integrity, Immunity and Cancer, Institut Pasteur [Paris], Genome dynamics in the immune system (Equipe Inserm U1163), Imagine - Institut des maladies génétiques (IMAGINE - U1163), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), IMIM-Hospital del Mar, Generalitat de Catalunya, This project is funded by the Institut Pasteur (L.D. and C.D. labs), the Institut National du Cancer (INCa Grant # PLBIO16-181 to L.D. and E.B. labs), the Ligue Nationale Contre le Cancer (Équipe Labellisée 2019 L.D. lab and Équipe Labellisée 2017 and 2020 E.B. lab) the Cancéropôle IdF-INCa (Emergence 2016 grant to L.D. and C.D.), the Spanish Ministerio de Economía, Industria y Competitividad (Grant SAF2017-83565-R to J.Y.) as well as by the Fundación Científica de la Asociación Española Contra el Cáncer (AECC) (Grant PROYEI6018YÉLA to J.Y.)., We thank the Institut Pasteur genomics and cytometry platforms for help with sequencing and cell sorting, Frederick Alt and members of his lab for tremendous help with LAM-HTGTS experiments, Barry Sleckman for providing the RAG2-Thy1.1 complementation vector, Joy Bianchi for providing the Rag2−/− p53−/− v-Abl pro-B cell lines, Carine Giovannangeli and Anne de Cian for providing the Cas9 protein and Thomas Mercher for helpful discussions and suggestions., Institut Pasteur [Paris] (IP), Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Demangel, Caroline, and Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur [Paris]
Subjects: 0301 basic medicine, Genome instability, DNA End-Joining Repair, MESH: DNA Breaks, Double-Stranded, Poly (ADP-Ribose) Polymerase-1, General Physics and Astronomy, MESH: Cell Cycle, MESH: Poly (ADP-Ribose) Polymerase-1, Genome, Càncer--Tractament, chemistry.chemical_compound, Mice, 0302 clinical medicine, [SDV.MHEP.MI]Life Sciences [q-bio]/Human health and pathology/Infectious diseases, DNA Breaks, Double-Stranded, MESH: Animals, lcsh:Science, Double strand, Multidisciplinary, Cell Cycle, DNA repair protein XRCC4, Cell cycle, 3. Good health, Cell biology, DNA-Binding Proteins, 030220 oncology & carcinogenesis, [SDV.MHEP.MI] Life Sciences [q-bio]/Human health and pathology/Infectious diseases, Cell division, DNA repair, ADN--Dany, DNA recombination, Science, Double-strand DNA breaks, MESH: G1 Phase, General Biochemistry, Genetics and Molecular Biology, Article, 03 medical and health sciences, Targeted therapies, Animals, MESH: Mice, B cells, fungi, G1 Phase, General Chemistry, MESH: DNA End-Joining Repair, enzymes and coenzymes (carbohydrates), 030104 developmental biology, chemistry, lcsh:Q, Homologous recombination, DNA, MESH: DNA-Binding Proteins
Abstract: The alternative non-homologous end-joining (NHEJ) pathway promotes DNA double-strand break (DSB) repair in cells deficient for NHEJ or homologous recombination, suggesting that it operates at all stages of the cell cycle. Here, we use an approach in which DNA breaks can be induced in G1 cells and their repair tracked, enabling us to show that joining of DSBs is not functional in G1-arrested XRCC4-deficient cells. Cell cycle entry into S-G2/M restores DSB repair by Pol θ-dependent and PARP1-independent alternative NHEJ with repair products bearing kilo-base long DNA end resection, micro-homologies and chromosome translocations. We identify a synthetic lethal interaction between XRCC4 and Pol θ under conditions of G1 DSBs, associated with accumulation of unresolved DNA ends in S-G2/M. Collectively, our results support the conclusion that the repair of G1 DSBs progressing to S-G2/M by alternative NHEJ drives genomic instability and represent an attractive target for future DNA repair-based cancer therapies., Depending on the cell cycle stage, cells can repair their genome via different pathways. Here the authors reveal mechanistic insights into repair of double strand breaks induced during G1 in an error-prone manner by Pol θ-dependent and PARP1-independent alt NHEJ during the SG2/M phases of the cell cycle
Published: 2020

14. Mycolactone induces cell death by SETD1B-dependent degradation of glutathione

Author: Thorsten Thye, Caroline Demangel, Birgit Förster, Bernhard Nocht Institute for Tropical Medicine - Bernhard-Nocht-Institut für Tropenmedizin [Hamburg, Germany] (BNITM), Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), We would like to thank T. Brummelkamp for providing us with the KBM-7 wild-type cells. The modified pGT vector was generously provided by Lidia Duncan from Cambridge Institute for Medical Research, Addenbrooke's Hospital, Cambridge, United Kingdom., Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), and Bodescot, Myriam
Subjects: 0301 basic medicine, Buruli ulcer, Genetic Screens, RC955-962, Gene Identification and Analysis, Gene Expression, Apoptosis, [SDV.BC.IC] Life Sciences [q-bio]/Cellular Biology/Cell Behavior [q-bio.CB], Toxicology, Pathology and Laboratory Medicine, medicine.disease_cause, Biochemistry, chemistry.chemical_compound, 0302 clinical medicine, [SDV.MHEP.MI]Life Sciences [q-bio]/Human health and pathology/Infectious diseases, Arctic medicine. Tropical medicine, [SDV.BC.IC]Life Sciences [q-bio]/Cellular Biology/Cell Behavior [q-bio.CB], Medicine and Health Sciences, Toxins, Cytotoxic T cell, Mycolactone, Cell Death, Chemistry, Genomics, Glutathione, 3. Good health, Cell biology, Infectious Diseases, Cell Processes, [SDV.MHEP.MI] Life Sciences [q-bio]/Human health and pathology/Infectious diseases, Macrolides, Public aspects of medicine, RA1-1270, Transcriptome Analysis, Research Article, Programmed cell death, Toxic Agents, 030231 tropical medicine, Necrotic Cell Death, 03 medical and health sciences, Downregulation and upregulation, Cell Line, Tumor, Genetics, medicine, Humans, Endoplasmic reticulum, Public Health, Environmental and Occupational Health, Biology and Life Sciences, Computational Biology, Cell Biology, Histone-Lysine N-Methyltransferase, Genome Analysis, medicine.disease, [SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, Acetylcysteine, 030104 developmental biology, Gene Expression Regulation, Mutagenesis, [SDV.MP.BAC] Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, Peptides, Gene Deletion, Exotoxin
Abstract: Mycobacterium ulcerans is a human pathogen that causes a necrotizing skin disease known as Buruli ulcer. Necrosis of infected skin is driven by bacterial production of mycolactone, a diffusible exotoxin targeting the host translocon (Sec61). By blocking Sec61, mycolactone prevents the transport of nascent secretory proteins into the endoplasmic reticulum of host cells. This triggers pro-apoptotic stress responses partially depending on activation of the ATF4 transcription factor. To gain further insight into the molecular pathways mediating the cytotoxic effects of mycolactone we conducted the first haploid genetic screen with the M. ulcerans toxin in KBM-7 cells. This approach allowed us to identify the histone methyltransferase SETD1B as a novel mediator of mycolactone-induced cell death. CRISPR/Cas9-based inactivation of SETD1B rendered cells resistant to lethal doses of the toxin, highlighting the critical importance of this gene’s expression. To understand how SETD1B contributes to mycolactone cytotoxicity, we compared the transcriptomes of wild-type (WT) and SETD1B knockout KBM-7 cells upon exposure to the toxin. While ATF4 effectors were upregulated by mycolactone in both WT and SETD1B knockout cells, mycolactone selectively induced the expression of pro-apoptotic genes in WT cells. Among those genes we identified CHAC1, which codes for a major glutathione (GSH)-degrading enzyme, and whose strong upregulation in mycolactone-treated WT cells correlated with a marked reduction in GSH protein level. Moreover, GSH supplementation conferred cells with substantial protection against the toxic effects of mycolactone. Our data thus identify SETD1B/CHAC1/GSH as a novel, epigenetic mechanism connecting Sec61 blockade with apoptotic cell death. They suggest that GSH-based treatments might have the capacity to limit skin necrosis in Buruli ulcer., Author summary The human pathogen Mycobacterium ulcerans causes a necrotizing skin disease known as Buruli ulcer. The major toxin of the mycobacteria, mycolactone, prevents the transport of secretory proteins into the endoplasmic reticulum, and thereby triggers a deadly stress response. We conducted the first haploid genetic screen to identify host factors with impact on mycolactone toxicity. This enabled us to identify the histone methyltransferase SETD1B as a novel mediator of mycolactone-induced cell death. RNA analyses of wild-type cells and resistant SETD1B knockout cells treated with mycolactone then showed a selective induction of genes implicated in programmed cell-death only in wild-type cells. This was accompanied by a marked reduction of the antioxidant glutathione, which might cause the mycolactone induced cell death.
Published: 2020

15. Dysregulation of tryptophan catabolism at the host-skin microbiota interface in Hidradenitis Suppurativa

Author: Jean-Marc Doisne, Pedro Gonçalves, Lysiane Boulet, Marie-Noëlle Ungeheuer, Laure Guenin-Macé, Sabine Duchatelet, Vincent Bondet, James P. Di Santo, Alain Hovnanian, Aude Nassif, Darragh Duffy, Maïa Delage, Angèle Schiavo, Véronique Mayau, J Morel, Caroline Demangel, Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Sorbonne Paris Cité (USPC), École normale supérieure de Lyon (ENS de Lyon), Immunité Innée - Innate Immunity, Biochimie Hormonale et Nutritionnelle, Université Joseph Fourier - Grenoble 1 (UJF)-CHU Grenoble, Genetic skin diseases : from disease mechanism to therapies (Equipe Inserm U1163), Imagine - Institut des maladies génétiques (IHU) (Imagine - U1163), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), Service de Génétique Médicale [CHU Necker], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Immunobiologie des Cellules dendritiques, Investigation Clinique et d’Accès aux Ressources Biologiques (Plate-forme) - Clinical Investigation and Access to BioResources (ICAReB), Institut Pasteur [Paris] (IP), Centre Médical de l'Institut Pasteur (CMIP), This study was supported by Institut Pasteur (to CD and JPDS) via the Center for Translational Science, INSERM (U1221, to CD, U1223, to JPDS), and 'Société Française de Dermatologie' (to AN). The Biomics Platform, C2RT, Institut Pasteur is supported by France Génomique (ANR-10-INBS-09-09) and IBISA., We thank Armanda Casrouge for assistance in performing the immunostaining of skin biopsies, Olivier Join-Lambert for fruitful discussions on the skin microbiota of patients with HS, and Harry Sokol for his critical reading of this manuscript. We also thank Valérie Briolat and Laurence Motreff of the Biomics Wet-Lab Platform for 16S rRNA sequencing and the Image Analysis Hub for immunofluorescence data processing., ANR-10-INBS-0009,France-Génomique,Organisation et montée en puissance d'une Infrastructure Nationale de Génomique(2010), Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur [Paris], École normale supérieure - Lyon (ENS Lyon), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Paris (UP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Paris (UP), Institut Pasteur [Paris], Centre Médical de l'Institut Pasteur, Vougny, Marie-Christine, Organisation et montée en puissance d'une Infrastructure Nationale de Génomique - - France-Génomique2010 - ANR-10-INBS-0009 - INBS - VALID, and Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM)
Subjects: Male, 0301 basic medicine, Kynurenine pathway, chemistry.chemical_compound, 030207 dermatology & venereal diseases, 0302 clinical medicine, Hidradenitis suppurativa, Immune homeostasis, Kynurenine, Skin, 0303 health sciences, integumentary system, biology, Tryptophan, General Medicine, Middle Aged, Hidradenitis Suppurativa, 3. Good health, 030220 oncology & carcinogenesis, Medicine, [SDV.IMM]Life Sciences [q-bio]/Immunology, Female, medicine.symptom, Research Article, Adult, [SDV.IMM] Life Sciences [q-bio]/Immunology, Inflammation, Dermatology, Amino acid metabolism, Tryptophan catabolism, 03 medical and health sciences, medicine, Humans, 030304 developmental biology, Host Microbial Interactions, business.industry, Cellular immune response, Fibroblasts, medicine.disease, Aryl hydrocarbon receptor, Metabolism, 030104 developmental biology, Receptors, Aryl Hydrocarbon, chemistry, Axilla, Immunology, biology.protein, business, Dysbiosis
Abstract: Hidradenitis suppurativa (HS) is a chronic skin disorder of unknown etiology that manifests as recurrent, painful lesions. Cutaneous dysbiosis and unresolved inflammation are hallmarks of active HS, but their origin and interplay remain unclear. Our metabolomic profiling of HS skin revealed an abnormal induction of the kynurenine pathway of tryptophan catabolism in dermal fibroblasts, correlating with the release of kynurenine pathway–inducing cytokines by inflammatory cell infiltrates. Notably, overactivation of the kynurenine pathway in lesional skin was associated with local and systemic depletion in tryptophan. Yet the skin microbiota normally degrades host tryptophan into indoles regulating tissue inflammation via engagement of the aryl hydrocarbon receptor (AHR). In HS skin lesions, we detected contextual defects in AHR activation coinciding with impaired production of bacteria-derived AHR agonists and decreased incidence of AHR ligand-producing bacteria in the resident flora. Dysregulation of tryptophan catabolism at the skin-microbiota interface thus provides a mechanism linking the immunological and microbiological features of HS lesions. In addition to revealing metabolic alterations in patients with HS, our study suggests that correcting AHR signaling would help restore immune homeostasis in HS skin., Loss of homeostasis of tryptophan metabolism at the host-microbiota interface may contribute to Hidradenitis Suppurativa.
Published: 2020

16. Metabolic alterations in Hurley stage 1 Hidradenitis Suppurativa

Author: Delage, Maïa, Guenin-Macé, Laure, Morel, Jean-David, Doisne, Jean-Marc, Schiavo, Angèle, Ungeheuer, Marie-Noëlle, Lam, Thi, Join-Lambert, Olivier, Santo, James, Nassif, Aude, Demangel, Caroline, Centre Médical de l'Institut Pasteur, Institut Pasteur [Paris], Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Immunité Innée - Innate Immunity, Investigation Clinique et d’Accès aux Ressources Biologiques (Plate-forme) - Clinical Investigation and Access to BioResources (ICAReB), Service de Microbiologie [CHU Caen], Université de Caen Normandie (UNICAEN), Normandie Université (NU)-Normandie Université (NU)-CHU Caen, Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN)-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), Normandie Université (NU), Centre Médical de l'Institut Pasteur (CMIP), Institut Pasteur [Paris] (IP), Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Normandie Université (NU)-Normandie Université (NU)-Université de Rouen Normandie (UNIROUEN), Mzembaba, Sandy, and Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur [Paris]
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.BAC] Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, [SDV.MP] Life Sciences [q-bio]/Microbiology and Parasitology, [SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, ComputingMilieux_MISCELLANEOUS
Abstract: International audience
Published: 2020

17. Effect of gut microbiota on depressive-like behaviors in mice is mediated by the endocannabinoid system

Author: Grégoire Chevalier, Gabriel Lepousez, Alain Pruvost, Thomas Laval, Aline Rifflet, Emeline Chu-Van, Maud Pascal, Ivo G. Boneca, Laure Guenin-Macé, Caroline Demangel, Gérard Eberl, Eleni Siopi, Benoit Colsch, Pierre-Marie Lledo, François Leulier, Aurélie Messager, Microenvironnement et Immunité - Microenvironment and Immunity, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Perception et Mémoire / Perception and Memory, Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), Immunobiologie de l'Infection - Immunobiology of Infection, Biologie et Génétique de la Paroi bactérienne - Biology and Genetics of Bacterial Cell Wall, Médicaments et Technologies pour la Santé (MTS), Université Paris-Saclay-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Inflammasome NLRP3 – NLRP3 Inflammasome, 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), This work was supported by the Major Federating Program 'Microbes & Brain' of the Institut Pasteur, Agence Nationale de la Recherche Grant ANR-16-CE15-0021-02-PG-Brain and a grant from the Fédération pour la Recherche sur le Cerveau (FRC). Lledo’s laboratory is supported by Agence Nationale de la Recherche Grants ANR-16-CE37-0010-ORUPS and ANR-15-NEUC-0004-02 'Circuit-OPL,' Laboratory for Excellence Programme 'Revive' Grant ANR-10-LABX-73, and the Life Insurance Company AG2R-La Mondiale. Boneca’s laboratory and A.R. were supported by the French Government’s Investissement d’Avenir program, Laboratoire d’Excellence 'Integrative Biology of Emerging Infectious Diseases' (grant no. ANR-10-LABX-62-IBEID)., ANR-16-CE15-0021,PG-Brain,Modulation de l'activité du cerveau par le peptidoglycan bactérien(2016), ANR-16-CE37-0010,ORUPS,Représentation sensorielle lors d'états psychotiques(2016), ANR-15-NEUC-0004,CIRCUIT-OPL,US-French Research Proposal- Neural Circuits and Plasticity- Olfactory Perception and Learning(2015), ANR-10-LABX-0073,REVIVE,Stem Cells in Regenerative Biology and Medicine(2010), ANR-10-LABX-0062,IBEID,Integrative Biology of Emerging Infectious Diseases(2010), Institut de Génomique Fonctionnelle de Lyon (IGFL), 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 Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), HUGOT, Bérengère, Modulation de l'activité du cerveau par le peptidoglycan bactérien - - PG-Brain2016 - ANR-16-CE15-0021 - AAPG2016 - VALID, Représentation sensorielle lors d'états psychotiques - - ORUPS2016 - ANR-16-CE37-0010 - AAPG2016 - VALID, US-French Research Proposal- Neural Circuits and Plasticity- Olfactory Perception and Learning - - CIRCUIT-OPL2015 - ANR-15-NEUC-0004 - CRCNS - VALID, Laboratoires d'excellence - Stem Cells in Regenerative Biology and Medicine - - REVIVE2010 - ANR-10-LABX-0073 - LABX - VALID, Integrative Biology of Emerging Infectious Diseases - - IBEID2010 - ANR-10-LABX-0062 - LABX - VALID, Centre International de Recherche en Infectiologie - UMR (CIRI), 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), Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Pasteur [Paris]-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)-Institut National de la Santé et de la Recherche Médicale (INSERM)-École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), and Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)
Subjects: 0301 basic medicine, Male, Science, Neurogenesis, [SDV]Life Sciences [q-bio], [SDV.NEU.NB]Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC]/Neurobiology, General Physics and Astronomy, Biology, Gut flora, Microbiology, Neural circuits, General Biochemistry, Genetics and Molecular Biology, Article, 03 medical and health sciences, Mice, 0302 clinical medicine, medicine, Animals, Chronic stress, Pathological, Multidisciplinary, Behavior, Animal, Depression, Fatty Acids, [SDV.NEU.NB] Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC]/Neurobiology, General Chemistry, Fecal Microbiota Transplantation, medicine.disease, biology.organism_classification, Phenotype, Endocannabinoid system, 3. Good health, Gastrointestinal Microbiome, Complementation, Mice, Inbred C57BL, Disease Models, Animal, Lactobacillus, 030104 developmental biology, Mood disorders, Immunology, Stress and resilience, 030217 neurology & neurosurgery, Stress, Psychological, Endocannabinoids
Abstract: Depression is the leading cause of disability worldwide. Recent observations have revealed an association between mood disorders and alterations of the intestinal microbiota. Here, using unpredictable chronic mild stress (UCMS) as a mouse model of depression, we show that UCMS mice display phenotypic alterations, which could be transferred from UCMS donors to naïve recipient mice by fecal microbiota transplantation. The cellular and behavioral alterations observed in recipient mice were accompanied by a decrease in the endocannabinoid (eCB) signaling due to lower peripheral levels of fatty acid precursors of eCB ligands. The adverse effects of UCMS-transferred microbiota were alleviated by selectively enhancing the central eCB or by complementation with a strain of the Lactobacilli genus. Our findings provide a mechanistic scenario for how chronic stress, diet and gut microbiota generate a pathological feed-forward loop that contributes to despair behavior via the central eCB system., The gut microbiota may contribute to depression, but the underlying mechanism is not well understood. Here the authors use a mouse model of stress induced depression to demonstrate that behavioural changes conferred by fecal transplant from stressed to naïve mice require the endocannabinoid system.
Published: 2020

18. CR3 Engaged by PGL-I Triggers Syk-Calcineurin-NFATc to Rewire the Innate Immune Response in Leprosy

Author: Doz-Deblauwe, Emilie, Carreras, Florence, Arbues, Ainhoa, Remot, Aude, Epardaud, Mathieu, Malaga, Wladimir, Mayau, Véronique, Prandi, Jacques, Astarie-Dequeker, Catherine, Guilhot, Christophe, Demangel, Caroline, Winter, Nathalie, ISP, Infectiologie et Santé Publique, INRA, Université de Tours, Nouzilly, Infectiologie et Santé Publique (UMR ISP), Institut National de la Recherche Agronomique (INRA)-Université de Tours (UT), Institut de pharmacologie et de biologie structurale (IPBS), Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées, Immunobiologie de l'Infection - Immunobiology of Infection, Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur [Paris], ANR-2011-BSV3-0001 PGLECT, Région Centre Val de Loire (No. 32000584 Inflammation et Infection), bourse du programme France Futur Elevage, ANR-11-BSV3-0001,PGLECT,Interaction PGL/lectines et modulation de la réponse immunitaire par les mycobactéries pathogènes(2011), Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS), Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), and Institut National de la Recherche Agronomique (INRA)-Université de Tours
Subjects: Male, lcsh:Immunologic diseases. Allergy, NFAT, Neutrophils, dendritic cell, Immunology, Macrophage-1 Antigen, phenol glycolipid-1, chemical and pharmacologic phenomena, macrophage, [SDV.IMM.II]Life Sciences [q-bio]/Immunology/Innate immunity, Mice, Phagocytosis, Leprosy, Animals, Syk Kinase, Syk, ComputingMilieux_MISCELLANEOUS, Original Research, CR3, Mice, Knockout, Antigens, Bacterial, NFATC Transcription Factors, Calcineurin, neutrophil, hemic and immune systems, Dendritic Cells, Immunity, Innate, Mycobacterium leprae, [SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology, Cytokines, [SDV.IMM]Life Sciences [q-bio]/Immunology, Glycolipids, lcsh:RC581-607, Signal Transduction
Abstract: International audience; Mycobacterium leprae, the causative agent of leprosy, is unique amongst human pathogens in its capacity to produce the virulence factor phenolic glycolipid (PGL)-I. In addition to mediating bacterial tropism for neurons, PGL-I interacts with Complement Receptor (CR)3 on macrophages (MPs) to promote infection. We demonstrate here that PGL-I binding to CR3 also enhances bacterial invasion of both polymorphonuclear neutrophils (PMNs) and dendritic cells (DCs). Moreover, in all cell types CR3 engagement by PGL-I activates the Syk tyrosine kinase, inducing calcineurin-dependent nuclear translocation of the transcription factor NFATc. This selectively augments the production of IL-2 by DCs, IL-10 by PMNs and IL-1β by MPs. In intranasally-infected mice PGL-I binding to CR3 heightens mycobacterial phagocytosis by lung PMNs and MPs, and stimulates NFATc-controlled production of Syk-dependent cytokines. Our study thus identifies the CR3-Syk-NFATc axis as a novel signaling pathway activated by PGL-I in innate immune cells, rewiring host cytokine responses to M. leprae.
Published: 2019

19. Recombinant Antibodies against Mycolactone

Author: M. Frank Erasmus, Fortunato Ferrara, Caroline Demangel, Leslie Naranjo, Sara D'Angelo, Andre A. Teixera, Nicolas Blanchard, Andrew Bradbury, Specifica Inc.[Santa Fe], Laboratoire d'innovation moléculaire et applications (LIMA), Université de Strasbourg (UNISTRA)-Université de Haute-Alsace (UHA) Mulhouse - Colmar (Université de Haute-Alsace (UHA))-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), This research was funded by NIH, R01-AI-113266 from the National Institutes of Health., and Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM)
Subjects: Buruli ulcer, single chain Fv, Phage display, Health, Toxicology and Mutagenesis, lcsh:Medicine, mycolactone, Yeast display, Biology, Toxicology, 03 medical and health sciences, chemistry.chemical_compound, 0302 clinical medicine, Antigen, Yeasts, medicine, Humans, recombinant antibody, Biotinylation, yeast display, Mycolactone, 030304 developmental biology, 0303 health sciences, Communication, lcsh:R, Antibodies, Monoclonal, High-Throughput Nucleotide Sequencing, medicine.disease, biology.organism_classification, Virology, Recombinant Proteins, 3. Good health, Immunization, chemistry, 030220 oncology & carcinogenesis, Mycobacterium ulcerans, biology.protein, [SDV.IMM]Life Sciences [q-bio]/Immunology, Macrolides, Antibody, phage display
Abstract: International audience; In the past, it has proved challenging to generate antibodies against mycolactone, the primary lipidic toxin A of Mycobacterium ulcerans causing Buruli ulcer, due to its immunosuppressive properties. Here we show that in vitro display, comprising both phage and yeast display, can be used to select antibodies recognizing mycolactone from a large human naïve phage antibody library. Ten different antibodies were isolated, and hundreds more identified by next generation sequencing. These results indicate the value of in vitro display methods to generate antibodies against difficult antigenic targets such as toxins, which cannot be used for immunization unless inactivated by structural modification. The possibility to easily generate anti-mycolactone antibodies is an exciting prospect for the development of rapid and simple diagnostic/detection methods.
Published: 2019

20. Ipomoeassin F Binds Sec61α to Inhibit Protein Translocation

Author: Wei Shi, Guanghui Zong, Zhijian Hu, Sarah O’Keefe, Dale Tranter, Ludivine Baron, Michael Iannotti, Belinda Hall, Katherine Corfield, Anja Paatero, Mark Henderson, Peristera Roboti, Jianhong Zhou, Xianwei Sun, Mugunthan Govindarajan, Jason M. Rohde, Nicolas Blanchard, Rachel Simmonds, James Inglese, Yuchun Du, Caroline Demangel, Stephen High, Ville Paavilainen, University of Arkansas [Fayetteville], Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Laboratoire d'innovation moléculaire et applications (LIMA), Université de Strasbourg (UNISTRA)-Université de Haute-Alsace (UHA) Mulhouse - Colmar (Université de Haute-Alsace (UHA))-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Faculty of Life Sciences [Manchester], University of Manchester [Manchester], Institute of Biotechnology, Biosciences, Doctoral Programme in Biomedicine, Doctoral Programme in Integrative Life Science, and Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM)
Subjects: Binding Sites, ANALOGS, Molecular Structure, BLOCKADE, [CHIM.ORGA]Chemical Sciences/Organic chemistry, RESIN GLYCOSIDES, [CHIM.THER]Chemical Sciences/Medicinal Chemistry, SQUAMOSA, 3. Good health, Protein Transport, INSIGHTS, NATURAL-PRODUCTS, MYCOLACTONE, REVEALS, Humans, 1182 Biochemistry, cell and molecular biology, BIOLOGICAL EVALUATION, APRATOXIN, Glycoconjugates, SEC Translocation Channels
Abstract: Funding Information: We thank the Arkansas Nano & Bio Materials Characterization Facility at the Institute for Nano Sciences & Engineering for our imaging studies, and Prof Yoshito Kishi (Harvard University) for the kind gift of synthetic mycolactone A/B used by S.H. and R.S. W.S. is supported by Grant No. R15GM116032 from the National Institute of General Medical Sciences of the National Institutes of Health (NIH) and startup funds from the University of Arkansas. This work was also supported in part by Grant No. P30 GM103450 from the National Institute of General Medical Sciences of the NIH and by seed money from the Arkansas Biosciences Institute (ABI). S.O’K. is the recipient of a Biotechnology and Biological Sciences Research Council (BBSRC) Doctoral Training Programme Award (BB/J014478/ 1), and S.H. holds a Welcome Trust Investigator Award in Science (204957/Z/16/Z). The alpha-1 antitrypsin work was supported by the Alpha-1 Foundation (J.I. and M.J.I.). J.I. and M.J.H. were supported by the intramural program of NCATS, National Institutes of Health, projects 1ZIATR000048-03 (J.I.) and ZIATR000063-04 (M.J.H.). R.S. holds a Welcome Trust Investigator Award in Science (202843/Z/16/Z). C.D. received funding from the Institut Pasteur, the Institut National de la Santé et de la Recherche Med́ icale, and the Fondation Raoul Follereau. N.B.’s synthesis and chemical biology studies of mycolactone were supported by CNRS, Université de Strasbourg, Fondations Potier et Follereau, and the Investisse-ment d’Avenir (Idex Unistra). V.O.P. is supported by the Academy of Finland (Grants 289737 and 314672) and the Sigrid Juselius Foundation. Funding Information: We thank the Arkansas Nano & Bio Materials Characterization Facility at the Institute for Nano Sciences & Engineering for our imaging studies, and Prof Yoshito Kishi (Harvard University) for the kind gift of synthetic mycolactone A/B used by S.H. and R.S. W.S. is supported by Grant No. R15GM116032 from the National Institute of General Medical Sciences of the National Institutes of Health (NIH) and startup funds from the University of Arkansas. This work was also supported in part by Grant No. P30 GM103450 from the National Institute of General Medical Sciences of the NIH and by seed money from the Arkansas Biosciences Institute (ABI). S.O'K. is the recipient of a Biotechnology and Biological Sciences Research Council (BBSRC) Doctoral Training Programme Award (BB/J014478/1), and S.H. holds a Welcome Trust Investigator Award in Science (204957/Z/16/Z). The alpha-1 antitrypsin work was supported by the Alpha-1 Foundation (J.I. and M.J.I.). J.I. and M.J.H. were supported by the intramural program of NCATS, National Institutes of Health, projects 1ZIATR000048-03 (J.I.) and ZIATR000063-04 (M.J.H.). R.S. holds a Welcome Trust Investigator Award in Science (202843/Z/16/Z). C.D. received funding from the Institut Pasteur, the Institut National de la Sante et de la Recherche Medicale, and the Fondation Raoul Follereau. N.B.'s synthesis and chemical biology studies of mycolactone were supported by CNRS, Universite de Strasbourg, Fondations Potier et Follereau and the Investissement d'Avenir (Idex Unistra). V.O.P. is supported by the Academy of Finland (Grants 289737 and 314672) and the Sigrid Juselius Foundation. Publisher Copyright: © 2019 American Chemical Society. Ipomoeassin F is a potent natural cytotoxin that inhibits growth of many tumor cell lines with single-digit nanomolar potency. However, its biological and pharmacological properties have remained largely unexplored. Building upon our earlier achievements in total synthesis and medicinal chemistry, we used chemical proteomics to identify Sec61 alpha (protein transport protein Sec61 subunit alpha isoform 1), the pore-forming subunit of the Sec61 protein translocon, as a direct binding partner of ipomoeassin F in living cells. The interaction is specific and strong enough to survive lysis conditions, enabling a biotin analogue of ipomoeassin F to pull down Sec61 alpha from live cells, yet it is also reversible, as judged by several experiments including fluorescent streptavidin staining, delayed competition in affinity pulldown, and inhibition of TNF biogenesis after washout. Sec61 alpha forms the central subunit of the ER protein translocation complex, and the binding of ipomoeassin F results in a substantial, yet selective, inhibition of protein translocation in vitro and a broad ranging inhibition of protein secretion in live cells. Lastly, the unique resistance profile demonstrated by specific amino acid single-point mutations in Sec61 alpha provides compelling evidence that Sec61 alpha is the primary molecular target of ipomoeassin F and strongly suggests that the binding of this natural product to Sec61 alpha is distinctive. Therefore, ipomoeassin F represents the first plant-derived, carbohydrate-based member of a novel structural class that offers new opportunities to explore Sec61 alpha function and to further investigate its potential as a therapeutic target for drug discovery.
Published: 2019

21. Mycolactone: More than Just a Cytotoxin

Author: Laure Guenin-Macé, Caroline Demangel, Gerd Pluschke, Marie-Thérèse Ruf, Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Swiss Tropical and Public Health Institute [Basel], University of Basel (Unibas), and Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM)
Subjects: 0303 health sciences, Toxin, [SDV]Life Sciences [q-bio], 030231 tropical medicine, Mycobacterial culture, Biology, medicine.disease_cause, 3. Good health, Microbiology, 03 medical and health sciences, chemistry.chemical_compound, 0302 clinical medicine, chemistry, Focal necrosis, medicine, %22">Fish , Mycolactone, Cytotoxicity, 030304 developmental biology
Abstract: International audience; From their observation of necrotic areas around bacterial foci in Buruli ulcers (BUs), Connor and co-workers were the first, back in 1965, to suggest that M. ulcerans may produce a diffusible cytotoxin [1]. This hypothesis was later confirmed by injecting mycobacterial culture filtrates into the skin of guinea pigs, showing that this causes focal necrosis resembling that of naturally occurring human infections [1–3]. In 1999, George et al. succeeded in isolating a cytotoxic factor from M. ulcerans lipid extracts, and deciphered its chemical nature [4]. The M. ulcerans toxin was named mycolactone, based on its mycobacterial origin and macrolactone structure: a 12-membered lactone ring, to which a C5-O-linked polyunsaturated acyl side chain and a C-linked upper side chain comprising C12–C20 are appended (Fig. 1). Follow-up studies showed that M. ulcerans-derived mycolactone was in reality a mixture of two stereoisomers called A and B [6, 7] (Fig. 1). Since the initial discovery of mycolactone A/B, eight additional mycolactone congeners have been identified that are either produced by M. ulcerans strains of different geographical origins or genetically related fish and frog pathogens, which were initially given different species designations such as M. pseudoshottsii and M. liflandii. Comparative genome analysis later revealed that all mycolactone-producing mycobacteria evolved from a common M. marinum-like progenitor, and are therefore ecovars of a single M. ulcerans species [8]. While the macrolide core structure and upper side chain are conserved, mycolactone populations from different M. ulcerans sub-lineages vary in the length, number and localization of hydroxyl groups and number of double bonds of the lower side chain. These modifications of the lower polyunsaturated acyl side chain cause pronounced changes in cytotoxicity [9–11]. The origin and chemistry of natural mycolactones, structure-activity relationships and the various approaches that were developed to generate synthetic mycolactones have been reviewed recently [12, 13]. These aspects will therefore not be further discussed here. Instead, this chapter provides an update on our understanding of mycolactone (A/B) biology, and discusses its proposed mechanisms of action in relation with BU pathogenesis.
Published: 2019

22. Mécanisme responsable des effets immunosuppresseurs de la mycolactone, toxine de M. ulcerans

Author: Morel, Jean-David, Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Sorbonne Paris Cité, and Caroline Demangel
Subjects: [SDV.IMM]Life Sciences [q-bio]/Immunology, Immunosuppression, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology, Mycolactone
Abstract: Mycolactone is a diffusible lipid produced by the human pathogen Mycobacteriumulcerans, the causative agent of a tropical skin disease called Buruli ulcer. Bacterial production of mycolactone in infected skin causes local tissue necrosis, while inducing immunosuppressive defects at the systemic level. When I started my PhD, the molecular mechanism(s) underpinning these effects were unknown. Over the course of my thesis, I contributed to demonstrate that mycolactoneis a novel inhibitor of the Sec61 translocon, a channel regulating the biogenesis of most secretedand membrane proteins in eukaryotic cells. Indeed, a single point mutation in the alpha subunit ofSec61 protected cells from the cytotoxic and immunosuppressive effects of mycolactone. I showed that mycolactone-mediated blockade of the Sec61 translocon efficiently prevents the synthesis ofkey immune receptors and signaling molecules, impeding the communication between immunecells that is required for the development of anti-mycobacterial immunity. Through a series of larges caleproteomic studies, I demonstrated that mycolactone is a broad-acting inhibitor of Sec61 and identified the Sec61 clients that are primarily down regulated by mycolactone in physiologicallyrelevant cell types. These analyses also allowed me to describe a unique stress response,encompassing elements of the unfolded protein response and integrated stress response, that isinduced upon protein translocation blockade and ultimately causes cell apoptosis. The Sec61 translocon has been proposed to play a role in other cell functions that require the retrograde transport of proteins across membranes, namely Endoplasmic Reticulum-Associated Degradation(ERAD), an essential process in protein quality control, and antigen export to the cytosol during cross-presentation, a pathway essential to the activation of adaptive immunity to intracellular pathogens and cancer. Using mycolactone, I showed that Sec61 blockade does not affect protein export to the cytosol in either of these pathways, arguing against Sec61 operating as are trotranslocon. Altogether, my work provided a molecular mechanism for the diverse effects of mycolactone in Buruli Ulcer patients, and thus for M. ulcerans virulence. Mycolactone representing the most potent Sec61 blocker identified to date, my studies also revealed the key importance of Sec61-mediated protein translocation in the regulation of immune responses and protein homeostasis.; La mycolactone est un lipide diffusible produit par Mycobacterium ulcerans, la bactérie responsable d'une maladie tropicale de la peau appelée ulcère de Buruli. La production de mycolactone dans la peau infectée par M. ulcerans provoque une nécrose tissulaire locale, tout en induisant des anomalies immunosuppressives au niveau systémique. Lorsque j'ai commencé mon doctorat, les mécanismes moléculaires à l'origine de ces effets étaient inconnus. Au cours de ma thèse, j'ai contribué à démontrer que la mycolactone est un nouvel inhibiteur du translocon Sec61,le canal régulant la biogenèse de la plupart des protéines sécrétées et membranaires dans les cellules eucaryotes. J’ai démontré qu’une mutation ponctuelle dans la sous-unité alpha de Sec61 protège les cellules des effets cytotoxiques et immunosuppresseurs de la mycolactone. J'ai montré que le blocage du translocon Sec61 par la mycolactone empêche efficacement la synthèse des principaux récepteurs immunitaires et des molécules de signalisation du système immunitaire,blockant la communication entre les cellules immunitaires et inhibant l'immunité antimycobactérienne. Par une série d'études protéomiques à grande échelle, j'ai démontré que la mycolactone est un inhibiteur à large action de Sec61 et j’ai identifié les substrats de Sec61 les plus impactés dans différents types cellulaires. Ces analyses m'ont également permis de décrire la réponse au stress induite par le blocage de la translocation des protéines, qui inclut des éléments de la réponse au stress protéostatique (UPR) et de la réponse intégrée au stress (ISR), provoquant finalement l'apoptose cellulaire. Plusieurs études ont impliqué le translocon Sec61 dans des processus qui requièrent le transport rétrograde de protéines à travers les membranes : la Dégradation Associée au Réticulum Endoplasmique (ERAD), processus essentiel du contrôle de la qualité des protéines et la cross-présentation, une voie essentielle à l'activation de l'immunité adaptative aux pathogènes intracellulaires et au cancer. J'ai montré que le blocage de Sec61 par la mycolactone n'affecte pas l'export de protéines vers le cytosol dans ces deux voies, suggérant que Sec61 ne peut pas fonctionner comme un rétrotranslocon. Mes travaux ont permis d’élucider le moléculaire responsable des divers effets de la mycolactone observés chez les patients atteints d’ulcère de Buruli et de révéler l’importance majeure de la translocation des protéines dans la régulation des réponses immunitaires et de l’homéostasie des protéines.
Published: 2018

23. Mécanisme responsable des effets immunosuppresseurs de la mycolactone, toxine de M. ulcerans

Author: Morel, Jean-David, Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Sorbonne Paris Cité, Caroline Demangel, STAR, ABES, and Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM)
Subjects: [SDV.MHEP] Life Sciences [q-bio]/Human health and pathology, [SDV.IMM] Life Sciences [q-bio]/Immunology, [SDV.IMM]Life Sciences [q-bio]/Immunology, Immunosuppression, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology, Mycolactone
Abstract: Mycolactone is a diffusible lipid produced by the human pathogen Mycobacteriumulcerans, the causative agent of a tropical skin disease called Buruli ulcer. Bacterial production of mycolactone in infected skin causes local tissue necrosis, while inducing immunosuppressive defects at the systemic level. When I started my PhD, the molecular mechanism(s) underpinning these effects were unknown. Over the course of my thesis, I contributed to demonstrate that mycolactoneis a novel inhibitor of the Sec61 translocon, a channel regulating the biogenesis of most secretedand membrane proteins in eukaryotic cells. Indeed, a single point mutation in the alpha subunit ofSec61 protected cells from the cytotoxic and immunosuppressive effects of mycolactone. I showed that mycolactone-mediated blockade of the Sec61 translocon efficiently prevents the synthesis ofkey immune receptors and signaling molecules, impeding the communication between immunecells that is required for the development of anti-mycobacterial immunity. Through a series of larges caleproteomic studies, I demonstrated that mycolactone is a broad-acting inhibitor of Sec61 and identified the Sec61 clients that are primarily down regulated by mycolactone in physiologicallyrelevant cell types. These analyses also allowed me to describe a unique stress response,encompassing elements of the unfolded protein response and integrated stress response, that isinduced upon protein translocation blockade and ultimately causes cell apoptosis. The Sec61 translocon has been proposed to play a role in other cell functions that require the retrograde transport of proteins across membranes, namely Endoplasmic Reticulum-Associated Degradation(ERAD), an essential process in protein quality control, and antigen export to the cytosol during cross-presentation, a pathway essential to the activation of adaptive immunity to intracellular pathogens and cancer. Using mycolactone, I showed that Sec61 blockade does not affect protein export to the cytosol in either of these pathways, arguing against Sec61 operating as are trotranslocon. Altogether, my work provided a molecular mechanism for the diverse effects of mycolactone in Buruli Ulcer patients, and thus for M. ulcerans virulence. Mycolactone representing the most potent Sec61 blocker identified to date, my studies also revealed the key importance of Sec61-mediated protein translocation in the regulation of immune responses and protein homeostasis., La mycolactone est un lipide diffusible produit par Mycobacterium ulcerans, la bactérie responsable d'une maladie tropicale de la peau appelée ulcère de Buruli. La production de mycolactone dans la peau infectée par M. ulcerans provoque une nécrose tissulaire locale, tout en induisant des anomalies immunosuppressives au niveau systémique. Lorsque j'ai commencé mon doctorat, les mécanismes moléculaires à l'origine de ces effets étaient inconnus. Au cours de ma thèse, j'ai contribué à démontrer que la mycolactone est un nouvel inhibiteur du translocon Sec61,le canal régulant la biogenèse de la plupart des protéines sécrétées et membranaires dans les cellules eucaryotes. J’ai démontré qu’une mutation ponctuelle dans la sous-unité alpha de Sec61 protège les cellules des effets cytotoxiques et immunosuppresseurs de la mycolactone. J'ai montré que le blocage du translocon Sec61 par la mycolactone empêche efficacement la synthèse des principaux récepteurs immunitaires et des molécules de signalisation du système immunitaire,blockant la communication entre les cellules immunitaires et inhibant l'immunité antimycobactérienne. Par une série d'études protéomiques à grande échelle, j'ai démontré que la mycolactone est un inhibiteur à large action de Sec61 et j’ai identifié les substrats de Sec61 les plus impactés dans différents types cellulaires. Ces analyses m'ont également permis de décrire la réponse au stress induite par le blocage de la translocation des protéines, qui inclut des éléments de la réponse au stress protéostatique (UPR) et de la réponse intégrée au stress (ISR), provoquant finalement l'apoptose cellulaire. Plusieurs études ont impliqué le translocon Sec61 dans des processus qui requièrent le transport rétrograde de protéines à travers les membranes : la Dégradation Associée au Réticulum Endoplasmique (ERAD), processus essentiel du contrôle de la qualité des protéines et la cross-présentation, une voie essentielle à l'activation de l'immunité adaptative aux pathogènes intracellulaires et au cancer. J'ai montré que le blocage de Sec61 par la mycolactone n'affecte pas l'export de protéines vers le cytosol dans ces deux voies, suggérant que Sec61 ne peut pas fonctionner comme un rétrotranslocon. Mes travaux ont permis d’élucider le moléculaire responsable des divers effets de la mycolactone observés chez les patients atteints d’ulcère de Buruli et de révéler l’importance majeure de la translocation des protéines dans la régulation des réponses immunitaires et de l’homéostasie des protéines.
Published: 2018

24. Proteomics Reveals Scope of Mycolactone-mediated Sec61 Blockade and Distinctive Stress Signature

Author: Morel, Jean-David, Paatero, Anja O., Wei, Jiajie, Yewdell, Jonathan W., Guenin-Mace, Laure, Van Haver, Delphi, Impens, Francis, Pietrosemoli, Natalia, Paavilainen, Ville O., Demangel, Caroline, Immunobiologie de l'Infection - Immunobiology of Infection, Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur [Paris], University of Helsinki, National Institute of Allergy and Infectious Diseases [Bethesda] (NIAID-NIH), National Institutes of Health [Bethesda] (NIH), Universiteit Gent = Ghent University [Belgium] (UGENT), Hub Bioinformatique et Biostatistique - Bioinformatics and Biostatistics HUB, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Helsingin yliopisto = Helsingfors universitet = University of Helsinki, Universiteit Gent = Ghent University (UGENT), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), Institute of Biotechnology, Doctoral Programme in Integrative Life Science, and University Management
Subjects: Proteomics, Transcription, Genetic, Research, Membrane Proteins, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, QUANTIFICATION, Endoplasmic Reticulum Stress, ER-STRESS, [SDV.IMM.II]Life Sciences [q-bio]/Immunology/Innate immunity, TOXIN MYCOLACTONE, BURULI ULCER, Substrate Specificity, Up-Regulation, Mice, Viral Proteins, [SDV.IMM.IA]Life Sciences [q-bio]/Immunology/Adaptive immunology, Stress, Physiological, [SDV.MHEP.MI]Life Sciences [q-bio]/Human health and pathology/Infectious diseases, MYCOBACTERIUM-ULCERANS, 1182 Biochemistry, cell and molecular biology, Animals, Macrolides, PROTEIN TRANSLOCATION, SEC Translocation Channels, Transcription Factors
Abstract: International audience; Mycolactone is a bacteria-derived macrolide that blocks the biogenesis of a large array of secretory and integral transmembrane proteins (TMP) through potent inhibition of the Sec61 translocon. Here, we used quantitative pro-teomics to delineate the direct and indirect effects of mycolactone-mediated Sec61 blockade in living cells. In T lymphocytes, dendritic cells and sensory neurons, Sec61 substrates downregulated by mycolactone were in order of incidence: secretory proteins (with a signal peptide but no transmembrane domain), TMPs with a signal peptide (Type I) and TMPs without signal peptide and a cytosolic N terminus (Type II). TMPs without a signal peptide and the opposite N terminus topology (Type III) were refractory to mycolactone inhibition. This rule applied comparably to single-and multi-pass TMPs, and extended to exogenous viral proteins. Parallel to its broad-spectrum inhibition of Sec61-mediated protein translocation, mycolactone rapidly induced cytosolic chaperones Hsp70/Hsp90. Moreover , it activated an atypical endoplasmic reticulum stress response, differing from conventional unfolded protein response by the down-regulation of Bip. In addition to refining our mechanistic understanding of Sec61 inhibition by mycolactone, our findings thus reveal that Sec61 blockade induces proteostatic stress in the cytosol and the endoplas-mic reticulum.
Published: 2018

25. Sec61 blockade by mycolactone: A central mechanism in Buruli ulcer disease

Author: Caroline, Demangel, Stephen, High, Kop, Marie-Luce, Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), University of Manchester [Manchester], Research in our laboratories is supported by the Institut Pasteur, the Institut National de la Santé et de la Recherche Médicale and the Fondation Raoul Follereau (C.D.), and and the Wellcome Trust (204957/Z/16/Z) (S.H.).
Subjects: MESH: Protein Transport, [SDV.IMM] Life Sciences [q-bio]/Immunology, [SDV]Life Sciences [q-bio], MESH: Mycobacterium ulcerans, Endoplasmic Reticulum, Stress, Models, Biological, Immunomodulation, MESH: Buruli Ulcer, MESH: Endoplasmic Reticulum, Animals, Humans, MESH: Animals, MESH: Humans, Mycobacterium ulcerans, MESH: Models, Biological, Sec61, [SDV] Life Sciences [q-bio], Protein Transport, [SDV.IMM]Life Sciences [q-bio]/Immunology, Macrolides, MESH: Macrolides, SEC Translocation Channels, Buruli ulcer, MESH: SEC Translocation Channels, Mycolactone
Abstract: International audience; Infection with Mycobacterium ulcerans results in a necrotising skin disease known as a Buruli ulcer, the pathology of which is directly linked to the bacterial production of the toxin mycolactone. Recent studies have identified the protein translocation machinery of the endoplasmic reticulum (ER) membrane as the primary cellular target of mycolactone, and shown that the toxin binds to the core subunit of the Sec61 complex. Mycolactone binding strongly inhibits the capacity of the Sec61 translocon to transport newly synthesised membrane and secretory proteins into and across the ER membrane. Since the ER acts as the entry point for the mammalian secretory pathway, and hence regulates initial access to the entire endomembrane system, mycolactone-treated cells have a reduced ability to produce a range of proteins including secretory cytokines and plasma membrane receptors. The global effect of this molecular blockade of protein translocation at the ER is that the host is unable to mount an effective immune response to the underlying mycobacterial infection. Prolonged exposure to mycolactone is normally cytotoxic, since it triggers stress responses activating the transcription factor ATF4 and ultimately inducing apoptosis.
Published: 2018

26. Mycobacterial Phenolic Glycolipids Selectively Disable TRIF-Dependent TLR4 Signaling in Macrophages

Author: Reid Oldenburg, Caroline Demangel, Jacques Prandi, Catherine Werts, Christophe Guilhot, Véronique Mayau, Nathalie Winter, Catherine Astarie-Dequeker, Ainhoa Arbues, Immunobiologie de l'Infection, Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut de pharmacologie et de biologie structurale (IPBS), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS), Biologie et Génétique de la Paroi bactérienne - Biology and Genetics of Bacterial Cell Wall (BGPB), Institut Pasteur [Paris], Infectiologie Animale et Santé Publique - IASP (Nouzilly, France), Institut National de la Recherche Agronomique (INRA), Université Francois Rabelais [Tours], This study was supported by the 'Marie Skłodowska-Curie Actions' of the European Union’s Seventh Framework Programme FP7/2007-2013, REA grant agreement no. 317057, the French National Research Agency (ANR-11-BSV3-001), Institut Pasteur and Institut National de la Santé et de la Recherche Médicale (INSERM)., European Project: 317057,EC:FP7:PEOPLE,FP7-PEOPLE-2012-ITN,HOMIN(2013), Immunobiologie de l'Infection - Immunobiology of Infection, Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées, Infectiologie et Santé Publique (UMR ISP), Institut National de la Recherche Agronomique (INRA)-Université de Tours, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Régulation des Infections Rétrovirales, Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UPS), Biologie et Génétique de la Paroi bactérienne, Génétique mycobactérienne, GUILHOT, Christophe, Host-microbe interactions in health and disease. Interface with the immune system - HOMIN - - EC:FP7:PEOPLE2013-04-01 - 2017-03-31 - 317057 - VALID, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Pasteur [Paris] (IP), Institut National de la Recherche Agronomique (INRA)-Université de Tours (UT), Université de Toulouse (UT)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur [Paris], and Demangel, Caroline
Subjects: 0301 basic medicine, [SDV]Life Sciences [q-bio], Nitric Oxide Synthase Type II, Complement receptor, [SDV.IMM.II]Life Sciences [q-bio]/Immunology/Innate immunity, Mice, glycolipide, Cell Wall, Immunologie, mycobactérie, Immunology and Allergy, TLR4, TRIF, ComputingMilieux_MISCELLANEOUS, Cells, Cultured, Original Research, Mice, Knockout, biology, Chemistry, Microbiology and Parasitology, Microbiologie et Parasitologie, Cell biology, Nitric oxide synthase, iNOS, Mycobacterium leprae, [SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology, [SDV.IMM]Life Sciences [q-bio]/Immunology, phenolic glycolipids, Signal transduction, Tyrosine kinase, réponse inflammatoire, Signal Transduction, lcsh:Immunologic diseases. Allergy, macrophages, mycobacteria, Immunology, macrophage, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, 03 medical and health sciences, Downregulation and upregulation, Leprosy, CXCL10, Animals, [SDV.IMM.II] Life Sciences [q-bio]/Immunology/Innate immunity, Antigens, Bacterial, Macrophages, Interferon-beta, Chemokine CXCL10, Mice, Inbred C57BL, Toll-Like Receptor 4, Adaptor Proteins, Vesicular Transport, 030104 developmental biology, biology.protein, Glycolipids, lcsh:RC581-607
Abstract: Phenolic glycolipids (PGLs) are cell wall components of a subset of pathogenic mycobacteria, with immunomodulatory properties. Here we show that in addition, PGLs exert anti-bactericidal activity by limiting the production of nitric oxide synthase (iNOS) in mycobacteria-infected macrophages. PGL-mediated downregulation of iNOS was complement receptor (CR)3-dependent, and comparably induced by bacterially-displayed and purified PGLs. Using Mycobacterium leprae PGL-1 as a model, we found that PGLs dampen the Toll-like receptor (TLR)4 signaling pathway, with macrophage exposure to PGLs leading to significant reduction in TIR-domain-containing adapter-inducing interferon-beta (TRIF) protein level. PGL-driven decrease in TRIF operated post-transcriptionally and independently of Src-family tyrosine kinases, lysosomal and proteasomal degradation. It resulted in the defective production of TRIF-dependent IFN-beta and CXCL10 in TLR4-stimulated macrophages, in addition to iNOS. Our results unravel a mechanism by which PGLs hijack both the bactericidal and inflammatory responses of host macrophages. Moreover, they identify TRIF as a critical node in the crosstalk between CR3 and TLR4.
Published: 2018

27. Modular total syntheses of mycolactone A/B and its [ 2 H]- isotopologue

Author: Caroline Demangel, Nicolas Blanchard, Sarah Saint-Auret, Didier Le Nouen, Philippe Bisseret, Hajer Abdelkafi, Laure Guenin-Macé, Laboratoire de chimie moléculaire (LCM), Université de Strasbourg (UNISTRA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Chimie organique et bioorganique (COB), Université de Haute-Alsace (UHA) Mulhouse - Colmar (Université de Haute-Alsace (UHA))-Centre National de la Recherche Scientifique (CNRS), Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), The authors thank the Université de Strasbourg, Investissement d'Avenir (Idex Unistra), CNRS, Fondation 'Raoul Follereau', Fondation 'Pour Le Développement De La Chimie Des Substances Naturelles Et Ses Applications' and Roche for a generous gift of methyl 3-hydroxy-2-methylpropionate., ANR-10-IDEX-0002,UNISTRA,Par-delà les frontières, l'Université de Strasbourg(2010), Chimie organique et bioorganique (COB), Université de Haute-Alsace (UHA) Mulhouse - Colmar (Université de Haute-Alsace (UHA))-Ecole Nationale Supérieure de Chimie de Mulhouse-Centre National de la Recherche Scientifique (CNRS), Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), and Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)
Subjects: Natural product, biology, 010405 organic chemistry, Stereochemistry, Organic Chemistry, Total synthesis, Biological activity, 010402 general chemistry, medicine.disease_cause, biology.organism_classification, 01 natural sciences, Biochemistry, 0104 chemical sciences, chemistry.chemical_compound, chemistry, Mycobacterium ulcerans, [SDV.TOX]Life Sciences [q-bio]/Toxicology, medicine, Epimer, Isotopologue, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Physical and Theoretical Chemistry, Mycolactone, Exotoxin
Abstract: A modular total synthesis of mycolactone A/B, the exotoxin produced by Mycobacterium ulcerans, has been achieved through the orchestration of several Pd-catalyzed key steps. While this route leads to a mixture of the natural product and its C12 epimer (4 : 1 ratio), this was inconsequential from the biological activity standpoint. Compared to the previously reported routes, this synthetic blueprint allows the late-stage modification of the toxin, as exemplified by the preparation of [22,22,22-2H3]-mycolactone A/B.
Published: 2017

28. Zika virus induces massive cytoplasmic vacuolization and paraptosis‐like death in infected cells

Author: Pierre Génin, Nicoletta Casartelli, Laurent Meertens, Julien Burlaud-Gaillard, Olivier Schwartz, Laura Sinigaglia, Etienne Simon-Loriere, Alex A. Compton, Nicolas Roy, Caroline Demangel, Philippe Roingeard, Nolwenn Jouvenet, Timothée Bruel, Ali Amara, Sonia Amraoui, Françoise Porrot, Arnaud Moris, Blandine Monel, Florence Guivel-Benhassine, Robert Weil, Virus et Immunité - Virus and immunity (CNRS-UMR3569), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), Plateforme IBISA de Microscopie Electronique [CHRU de Tours] (UNIV Tours), Centre Hospitalier Régional Universitaire de Tours (CHRU Tours)-Université de Tours (UT), Institut National de la Santé et de la Recherche Médicale (INSERM), Signalisation et Pathogénèse, Pathologie cellulaire : aspects moléculaires et viraux / Pathologie et Virologie Moléculaire, Institut Universitaire d'Hématologie (IUH), Université Paris Diderot - Paris 7 (UPD7)-Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Centre National de la Recherche Scientifique (CNRS), Génomique virale et vaccination, Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Génétique fonctionnelle des Maladies infectieuses - Functional Genetics of Infectious Diseases, Centre d'Immunologie et de Maladies Infectieuses (CIMI), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), AC was supported from postdoctoral fellowships from the Pasteur Foundation and ANRS. Work was supported by grants from the 'CHIKV-Viro-Immuno' ANR-14-CE14-0015-01 and 'TIMTAMDEN' ANR-14-CE14-0029 projects, the Labex IBEID program ANR-10-IHUB-0002, the Labex VRI program ANR-10-LABX-77 and Institut Pasteur., We thank members of the Virus & Immunity Unit for discussions. We thank Van-Mai Cao-Lormeau, Institut Louis Malardé (French Polynesia) and Myrielle Dupont-Rouzeyrol (Institut Pasteur Nouvelle Calédonie) for the gift of reagents. We thank Stephane Petres (Institut Pasteur) for preparation of the 4G2 antibody and the Imagopole (Institut Pasteur) for help in image acquisition., ANR-14-CE14-0015,CHIKV-Viro-Immuno,Multiplication et Relation avec l'hôte du virus Chikungunya(2014), ANR-14-CE14-0029,TIMTAMDEN,Rôle des récepteurs TIM et TAM dans l'infection des cellules cibles par le virus de la dengue(2014), ANR-10-LABX-0062,IBEID,Integrative Biology of Emerging Infectious Diseases(2010), ANR-10-LABX-0077,VRI,Initiative for the creation of a Vaccine Research Institute(2010), Virus et Immunité - Virus and immunity, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Institut Pasteur [Paris], Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur [Paris], Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Pierre et Marie Curie - Paris 6 (UPMC), and Centre Hospitalier Régional Universitaire de Tours (CHRU TOURS)-Université de Tours
Subjects: 0301 basic medicine, MESH: Cell Death, MESH: Signal Transduction, Vacuole, Endoplasmic Reticulum, Paraptosis, IFITM, Phosphatidylinositol 3-Kinases, 0302 clinical medicine, Cytopathogenic Effect, Viral, Cells, Cultured, Cytopathic effect, ZIKA virus, General Neuroscience, RNA-Binding Proteins, Articles, 3. Good health, Cell biology, cell death, MESH: Epithelial Cells, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, MESH: Membrane Proteins, MESH: SEC Translocation Channels, Signal Transduction, MESH: Cells, Cultured, Programmed cell death, MESH: Zika Virus, Biology, Virus, General Biochemistry, Genetics and Molecular Biology, cytopathic effect, MESH: Vacuoles, 03 medical and health sciences, MESH: Endoplasmic Reticulum, Humans, Molecular Biology, MESH: Humans, General Immunology and Microbiology, MESH: Proto-Oncogene Proteins c-akt, Endoplasmic reticulum, Membrane Proteins, Epithelial Cells, Fibroblasts, MESH: Cytopathogenic Effect, Viral, Virology, MESH: Astrocytes, 030104 developmental biology, MESH: RNA-Binding Proteins, Viral replication, Vacuolization, MESH: Fibroblasts, MESH: Phosphatidylinositol 3-Kinases, Astrocytes, Vacuoles, paraptosis, Proto-Oncogene Proteins c-akt, SEC Translocation Channels, 030217 neurology & neurosurgery
Abstract: International audience; The cytopathic effects of Zika virus (ZIKV) are poorly characterized. Innate immunity controls ZIKV infection and disease in most infected patients through mechanisms that remain to be understood. Here, we studied the morphological cellular changes induced by ZIKV and addressed the role of interferon-induced transmembrane proteins (IFITM), a family of broad-spectrum antiviral factors, during viral replication. We report that ZIKV induces massive vacuolization followed by "implosive" cell death in human epithelial cells, primary skin fibroblasts and astrocytes, a phenomenon which is exacerbated when IFITM3 levels are low. It is reminiscent of paraptosis, a caspase-independent, non-apoptotic form of cell death associated with the formation of large cytoplasmic vacuoles. We further show that ZIKV-induced vacuoles are derived from the endoplasmic reticulum (ER) and dependent on the PI3K/Akt signaling axis. Inhibiting the Sec61 ER translocon in ZIKV-infected cells blocked vacuole formation and viral production. Our results provide mechanistic insight behind the ZIKV-induced cytopathic effect and indicate that IFITM3, by acting as a gatekeeper for incoming virus, restricts virus takeover of the ER and subsequent cell death.
Published: 2017

29. Pathogenic and immunosuppressive properties of mycobacterial phenolic glycolipids

Author: Reid Oldenburg, Caroline Demangel, Université Paris Diderot - Paris 7 (UPD7), Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), This study was supported by the 'Marie Skłodowska Curie Actions' of the European Union's Seventh Framework Programme FP7/2007-2013, REA grant agreement n°317057., European Project: 317057,EC:FP7:PEOPLE,FP7-PEOPLE-2012-ITN,HOMIN(2013), Kop, Marie-Luce, Host-microbe interactions in health and disease. Interface with the immune system - HOMIN - - EC:FP7:PEOPLE2013-04-01 - 2017-03-31 - 317057 - VALID, and Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM)
Subjects: 0301 basic medicine, [SDV.IMM] Life Sciences [q-bio]/Immunology, [SDV]Life Sciences [q-bio], Virulence, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Biochemistry, Bacterial cell structure, Microbiology, Mycobacterium, Immunomodulation, 03 medical and health sciences, 0302 clinical medicine, Immune system, Glycolipid, Polyketide synthase, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, Animals, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, biology, Host (biology), Phenolic glycolipids, Mycobacteria, General Medicine, [SDV] Life Sciences [q-bio], 030104 developmental biology, Host cell invasion, biology.protein, [SDV.IMM]Life Sciences [q-bio]/Immunology, Glycolipids, Intracellular, Immunosuppressive Agents, 030215 immunology
Abstract: International audience; Phenolic glycolipids (PGL) are polyketide synthase products that are uniquely produced by a subset of pathogenic mycobacteria and are displayed at the bacterial cell surface, in a strategic position to interfere with host immune cells. Their expression has been associated with enhanced mycobacterial virulence in vivo, and suppression of the inflammatory responses of host phagocytes in vitro. In this review, we will present our current understanding of the mode of operation of PGL, along with functional evidence that demonstrates the evolutionary advantage conferred by PGL production for host cell invasion, intracellular persistence and evasion of host immune and bactericidal responses.
Published: 2017

30. Mycolactone displays anti-inflammatory effects on the nervous system

Author: Isaac, Caroline, Mauborgne, Annie, Grimaldi, Alfonso, Ade, Kemy, Pohl, Michel, Limatola, Cristina, Boucher, Yves, Demangel, Caroline, Guenin-Macé, Laure, Kop, Marie-Luce, Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut de psychiatrie et neurosciences (U894 / UMS 1266), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Department of Physiology and Pharmacology [Rome], Università degli Studi di Roma 'La Sapienza' = Sapienza University [Rome] (UNIROMA), This work was supported by the Ministero dell'Istruzione, dell'Universita e della Ricerca with the grant PRIN 2015E8EMCM to CL. It also recieved funding from Institut Pasteur, Institut National de la Santé et de la Recherche Médicale (INSERM) and Association Raoul Follereau (CD). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript., Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Università degli Studi di Roma 'La Sapienza' = Sapienza University [Rome], and Centre de Psychiatrie et Neurosciences (U894)
Subjects: Physiology, Cytotoxicity, [SDV]Life Sciences [q-bio], MESH: Neuralgia, Anti-Inflammatory Agents, microglia, mycolactone, MESH: Mycobacterium ulcerans, Pathology and Laboratory Medicine, Toxicology, Nervous System, Mice, Animal Cells, Immune Physiology, Medicine and Health Sciences, MESH: Animals, Immune Response, Cells, Cultured, Neurons, Innate Immune System, lcsh:Public aspects of medicine, [SDV] Life Sciences [q-bio], Spinal Cord, MESH: Cell Survival, Cytokines, [SDV.IMM]Life Sciences [q-bio]/Immunology, Macrolides, Cellular Types, Anatomy, MESH: Macrolides, Neuronal Tuning, Research Article, MESH: Cells, Cultured, lcsh:Arctic medicine. Tropical medicine, [SDV.IMM] Life Sciences [q-bio]/Immunology, MESH: Rats, lcsh:RC955-962, Cell Survival, Immunology, Pain, Glial Cells, Signs and Symptoms, Diagnostic Medicine, Animals, Microglial Cells, MESH: Mice, Neuropathic Pain, Inflammation, MESH: Nervous System, Mycobacterium ulcerans, Biology and Life Sciences, lcsh:RA1-1270, Cell Biology, Molecular Development, Rats, Neuroanatomy, Disease Models, Animal, Cellular Neuroscience, Immune System, MESH: Anti-Inflammatory Agents, Neuralgia, MESH: Disease Models, Animal, Neuroscience, Developmental Biology
Abstract: Background Mycolactone is a macrolide produced by the skin pathogen Mycobacterium ulcerans, with cytotoxic, analgesic and immunomodulatory properties. The latter were recently shown to result from mycolactone blocking the Sec61-dependent production of pro-inflammatory mediators by immune cells. Here we investigated whether mycolactone similarly affects the inflammatory responses of the nervous cell subsets involved in pain perception, transmission and maintenance. We also investigated the effects of mycolactone on the neuroinflammation that is associated with chronic pain in vivo. Methodology/ Principle findings Sensory neurons, Schwann cells and microglia were isolated from mice for ex vivo assessment of mycolactone cytotoxicity and immunomodulatory activity by measuring the production of proalgesic cytokines and chemokines. In all cell types studied, prolonged (>48h) exposure to mycolactone induced significant cell death at concentrations >10 ng/ml. Within the first 24h treatment, nanomolar concentrations of mycolactone efficiently suppressed the cell production of pro-inflammatory mediators, without affecting their viability. Notably, mycolactone also prevented the pro-inflammatory polarization of cortical microglia. Since these cells critically contribute to neuroinflammation, we next tested if mycolactone impacts this pathogenic process in vivo. We used a rat model of neuropathic pain induced by chronic constriction of the sciatic nerve. Here, mycolactone was injected daily for 3 days in the spinal canal, to ensure its proper delivery to spinal cord. While this treatment failed to prevent injury-induced neuroinflammation, it decreased significantly the local production of inflammatory cytokines without inducing detectable cytotoxicity. Conclusion/ Significance The present study provides in vitro and in vivo evidence that mycolactone suppresses the inflammatory responses of sensory neurons, Schwann cells and microglia, without affecting the cell viability. Together with previous studies using peripheral blood leukocytes, our work implies that mycolactone-mediated analgesia may, at least partially, be explained by its anti-inflammatory properties., Author summary Mycolactone is a complex macrolidic polyketide produced by the skin pathogen Mycobacterium ulcerans, with cytotoxic, analgesic and anti-inflammatory properties. Peripheral nerve destruction and activation of type 2 angiotensin II receptors on sensory neurons have been proposed to mediate bacteria-induced hypoesthesia in infected skin. In addition, mycolactone was recently shown to block the co-translational translocation of secretory proteins into the endoplasmic reticulum in host cells, leading to defective inflammatory responses. Here we examined if this last mechanism may also contribute to inhibit neuro-inflammation and particularly in the context of neuropathic pain. Using a representative panel of primary cells from the central and peripheral nervous systems, we found that indeed, mycolactone potently inhibits the production of pro-inflammatory mediators at non-cytotoxic concentrations. Notably, mycolactone prevented the polarization and pro-inflammatory functions of cortical microglia, which are critical inducers of neuroinflammation. Consistent with our in vitro findings, mycolactone had potent anti-inflammatory effects on the spinal cord of rats injected in the spinal canal, with no apparent side effects. Our data show that mycolactone suppresses inflammatory responses in the nervous system similarly as in the immune system, suggesting that mycolactone-mediated analgesia may, at least partially, be explained by its anti-inflammatory properties.
Published: 2017

31. Mycolactone subverts immunity by selectively blocking the Sec61 translocon

Author: Fatoumata Niang, Caroline Demangel, Ville O. Paavilainen, Jack Taunton, Sarah Saint-Auret, J Morel, Francis Impens, Anja O. Paatero, Laure Guenin-Macé, Ludivine Baron, Sandra Pellegrini, Rabea Dillmann, Nicolas Blanchard, Institute of Biotechnology, Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), University of Helsinki, Interactions Bactéries-Cellules (UIBC), Institut National de la Recherche Agronomique (INRA)-Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Laboratoire de chimie moléculaire (LCM), Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA), Signalisation des Cytokines - Cytokine Signaling, University of California [San Francisco] (UCSF), University of California, This work was supported by the Fondation pour la Recherche Médicale (FRM 2012 DEQ20120323704 to C. Demangel), the Association Raoul Follereau (C. Demangel), the Région Ile de France (dim130027 to C. Demangel), the Academy of Finland (289737 to V.O. Paavilainen), the Sigrid Juselius Foundation (V.O. Paavilainen), and Biocentrum Helsinki (V.O. Paavilainen). F. Impens received financial support from a Pasteur-Roux Fellowship., Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Helsingin yliopisto = Helsingfors universitet = University of Helsinki, Institut National de la Recherche Agronomique (INRA)-Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Université de Strasbourg (UNISTRA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), University of California [San Francisco] (UC San Francisco), University of California (UC), Kop, Marie-Luce, Immunobiologie de l'Infection, Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur [Paris]-Institut National de la Recherche Agronomique (INRA), Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS), Signalisation des Cytokines, Paavilainen, Ville O., and Demangel, Caroline
Subjects: 0301 basic medicine, T-Lymphocytes, [SDV]Life Sciences [q-bio], MEMBRANES, Jurkat cells, Jurkat Cells, chemistry.chemical_compound, INFECTION, Immunology and Allergy, Mycolactone, Integral membrane protein, Research Articles, Receptors, Interferon, 3. Good health, Cell biology, [SDV] Life Sciences [q-bio], medicine.anatomical_structure, Biochemistry, [SDV.IMM]Life Sciences [q-bio]/Immunology, MYCOBACTERIUM-ULCERANS, Macrolides, Signal transduction, Signal Transduction, [SDV.IMM] Life Sciences [q-bio]/Immunology, T cell, Immunology, ENDOPLASMIC-RETICULUM, INHIBITION, Biology, PERIPHERAL-BLOOD, BURULI ULCER, 03 medical and health sciences, Cell Adhesion, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, medicine, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Cell adhesion, 030102 biochemistry & molecular biology, Brief Definitive Report, SEC61 Translocon, 030104 developmental biology, Secretory protein, chemistry, VIRULENCE, 1182 Biochemistry, cell and molecular biology, PROTEIN TRANSLOCATION, COTRANSLATIONAL TRANSLOCATION, SEC Translocation Channels
Abstract: Baron et al. show that mycolactone, an immunosuppressive macrolide produced by the pathogen Mycobacterium ulcerans, operates by targeting the Sec61 translocon. This identifies the most potent Sec61 inhibitor reported to date and the potential of inhibiting Sec61 for immune modulation., Mycolactone, an immunosuppressive macrolide released by the human pathogen Mycobacterium ulcerans, was previously shown to impair Sec61-dependent protein translocation, but the underlying molecular mechanism was not identified. In this study, we show that mycolactone directly targets the α subunit of the Sec61 translocon to block the production of secreted and integral membrane proteins with high potency. We identify a single–amino acid mutation conferring resistance to mycolactone, which localizes its interaction site near the lumenal plug of Sec61α. Quantitative proteomics reveals that during T cell activation, mycolactone-mediated Sec61 blockade affects a selective subset of secretory proteins including key signal-transmitting receptors and adhesion molecules. Expression of mutant Sec61α in mycolactone-treated T cells rescued their homing potential and effector functions. Furthermore, when expressed in macrophages, the mycolactone-resistant mutant restored IFN-γ receptor–mediated antimicrobial responses. Thus, our data provide definitive genetic evidence that Sec61 is the host receptor mediating the diverse immunomodulatory effects of mycolactone and identify Sec61 as a novel regulator of immune cell functions.
Published: 2016

32. Functional analysis via standardized whole-blood stimulation systems defines the boundaries of a healthy immune response to complex stimuli

Author: Ivo Gomperts-Boneca, Ludovic Deriano, Spencer L. Shorte, Annick Dubois, Anavaj Sakuntabhai, Claude Leclerc, Caroline Demangel, Ana Cumano, Lluis Quintana-Murci, Matthew L. Albert, Antonio A. Freitas, Antoine Toubert, Lars Rogge, Manfred Schmolz, Gérard Eberl, Vincent Rouilly, Valentina Libri, Alejandra Urrutia, James P. Di Santo, Eric Tartour, Marie-Noëlle Ungeheuer, Françoise Dromer, Hugo Mouquet, Olivier Schwartz, Benoit Beitz, Mikael David, Olivier Lantz, Stéphanie Thomas, Ivo G. Boneca, Philippe Bousso, Laurent Abel, Vassili Soumelis, Cécile Delval, Darragh Duffy, Benno Schwikowski, Frédéric Tangy, Serge Hercberg, Jost Enninga, Milena Hasan, Samuel S.T. LaBrie, Sandra Pellegrini, Aurélie Bisiaux, Stanislas Pol, Marc Daëron, Andrés Alcover, Immunobiologie des Cellules dendritiques, Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre de Recherche Translationnelle - Center for Translational Science (CRT), Institut Pasteur [Paris] (IP), Centre d'Immunologie Humaine (CIH), Substances Lichéniques et Photoprotection, Université de Rennes (UR), Centre d'Immunologie et de Maladies Infectieuses (CIMI), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Immunobiologie des Cellules Dendritiques, Myriad rules based medicine, Chambre Régionale d'Agriculture des Pays de la Loire, Biologie et Génétique de la Paroi bactérienne - Biology and Genetics of Bacterial Cell Wall (BGPB), Laboratoire de géographie physique : Environnements Quaternaires et Actuels (LGP), Université Paris 1 Panthéon-Sorbonne (UP1)-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12)-Centre National de la Recherche Scientifique (CNRS), Département d'Immunologie - Department of Immunology, Immunorégulation, Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), Génétique Evolutive Humaine - Human Evolutionary Genetics, Laboratory of Cellular Physiology and Immunology and Chris Browne Center, Rockefeller University [New York], St. Giles Laboratory of Human Genetics of Infectious Diseases, Laboratory of Human Genetics of Infectious Diseases, Institut National de la Santé et de la Recherche Médicale (INSERM), Imagine - Institut des maladies génétiques (IMAGINE - U1163), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Biologie Cellulaire des Lymphocytes - Lymphocyte Cell Biology, Dynamiques des Réponses Immunes, Lymphopoïèse (Lymphopoïèse (UMR_1223 / U1223 / U-Pasteur_4)), Institut Pasteur [Paris] (IP)-Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM), Cellule Pasteur, Université Paris Diderot - Paris 7 (UPD7)-PRES Sorbonne Paris Cité, Centre d'Immunologie de Marseille - Luminy (CIML), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Immunobiologie de l'Infection - Immunobiology of Infection, Département de Génomes et Génétique - Department of Genomes and Genetics, Mycologie moléculaire, Centre National de Référence Mycologie et Antifongiques-Mycologie Moléculaire (CNRMA), Microenvironnement et Immunité - Microenvironment and Immunity, Dynamique des Interactions Hôte-Pathogène - Dynamics of Host-Pathogen Interactions, Equipe 3: EREN- Equipe de Recherche en Epidémiologie Nutritionnelle (CRESS - U1153), Université Paris 13 (UP13)-Institut National de la Recherche Agronomique (INRA)-Conservatoire National des Arts et Métiers [CNAM] (CNAM), HESAM Université - Communauté d'universités et d'établissements Hautes écoles Sorbonne Arts et métiers université (HESAM)-HESAM Université - Communauté d'universités et d'établissements Hautes écoles Sorbonne Arts et métiers université (HESAM)-Centre de Recherche Épidémiologie et Statistique Sorbonne Paris Cité (CRESS (U1153 / UMR_A_1125 / UMR_S_1153)), Institut National de la Recherche Agronomique (INRA)-Université Paris Diderot - Paris 7 (UPD7)-Université Paris Descartes - Paris 5 (UPD5)-Université Sorbonne Paris Cité (USPC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de la Recherche Agronomique (INRA)-Université Paris Diderot - Paris 7 (UPD7)-Université Paris Descartes - Paris 5 (UPD5)-Université Sorbonne Paris Cité (USPC)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut de formation en masso-kinésithérapie (IFMK Brest), Université de Brest (UBO)-Centre Hospitalier Régional Universitaire de Brest (CHRU Brest), Réponse humorale aux pathogènes, Signalisation des Cytokines - Cytokine Signaling, Service d'hépatologie médicale [CHU Cochin], Hôpital Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Génétique fonctionnelle des Maladies infectieuses - Functional Genetics of Infectious Diseases, Centre de recherche sur les liens sociaux (CERLIS - UMR 8070), Université Sorbonne Nouvelle - Paris 3-Université Paris Descartes - Paris 5 (UPD5)-Centre National de la Recherche Scientifique (CNRS), Biologie systémique - Systems Biology, Imagopole (CITECH), Laboratoire d'Immunologie Clinique, Institut Curie [Paris], Génomique virale et vaccination, Cytokines et Immunologie des Tumeurs Humaines (U753), Université Paris-Sud - Paris 11 (UP11)-Institut Gustave Roussy (IGR)-Institut National de la Santé et de la Recherche Médicale (INSERM), Hôpital Saint-Louis, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Paris Diderot - Paris 7 (UPD7), Investigation Clinique et d’Accès aux Ressources Biologiques (Plate-forme) - Clinical Investigation and Access to BioResources (ICAReB), This work benefited from support of the French government’s Invest in the Future Program, managed by the Agence Nationale de la Recherche (ANR, reference 10-LABX-69-01). We also acknowledge A. Pugsley for his support in obtaining funding and the logistical aspects of initiating the project, Milieu Intérieur Consortium (32 collaborateurs) : Abel L, Alcover A, Bousso P, Cumano A, Daëron M, Delval C, Demangel C, Deriano L, Di Santo J, Dromer F, Eberl G, Enninga J, Freitas A, Gomperts-Boneca I, Hercberg S, Lantz O, Leclerc C, Mouquet H, Pellegrini S, Pol S, Rogge L, Sakuntabhai A, Schwartz O, Schwikowski B, Shorte S, Soumelis V, Tangy F, Tartour E, Toubert A, Ungeheuer MN, Quintana-Murci L, Albert ML., ANR-10-LABX-0069,MILIEU INTERIEUR,GENETIC & ENVIRONMENTAL CONTROL OF IMMUNE PHENOTYPE VARIANCE: ESTABLISHING A PATH TOWARDS PERSONALIZED MEDICINE(2010), Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Pasteur [Paris], Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES), Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Institut Pasteur [Paris]-Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU), Centre National de la Recherche Scientifique (CNRS)-Institut Pasteur [Paris], Institut National de la Recherche Agronomique (INRA)-Conservatoire National des Arts et Métiers [CNAM] (CNAM)-Université Paris 13 (UP13)-Centre de Recherche Épidémiologie et Statistique Sorbonne Paris Cité (CRESS (U1153 / UMR_A_1125 / UMR_S_1153)), Université Paris Diderot - Paris 7 (UPD7)-Université Sorbonne Paris Cité (USPC)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de la Recherche Agronomique (INRA)-Université Paris Diderot - Paris 7 (UPD7)-Université Sorbonne Paris Cité (USPC)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de la Recherche Agronomique (INRA), Centre Hospitalier Régional Universitaire de Brest (CHRU Brest)-Université de Brest (UBO), Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale ( INSERM ), Centre de Recherche Translationnelle ( CRT ), Centre d'Immunologie Humaine ( CIH ), Université de Rennes 1 ( UR1 ), Université de Rennes ( UNIV-RENNES ) -Université de Rennes ( UNIV-RENNES ), Centre d'Immunologie et de Maladies Infectieuses ( CIMI ), Université Pierre et Marie Curie - Paris 6 ( UPMC ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Biologie et Génétique de la Paroi bactérienne, Laboratoire de géographie physique ( LGP ), Université Panthéon-Sorbonne ( UP1 ) -Université Paris-Est Créteil Val-de-Marne - Paris 12 ( UPEC UP12 ) -Centre National de la Recherche Scientifique ( CNRS ), Département d'Immunologie, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique ( CNRS ), rockefeller university, St. Giles Laboratory of Human Genetics of Infectious Diseases, Rockefeller Branch, Institut National de la Santé et de la Recherche Médicale ( INSERM ), Imagine - Institut des maladies génétiques ( IMAGINE - U1163 ), Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Biologie Cellulaire des Lymphocytes, Lymphopoïèse, Université Paris Diderot - Paris 7 ( UPD7 ) -PRES Sorbonne Paris Cité, Centre d'Immunologie de Marseille - Luminy ( CIML ), Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Aix Marseille Université ( AMU ) -Centre National de la Recherche Scientifique ( CNRS ), Immunobiologie de l'Infection, Département de Génomes et Génétique, Centre National de Référence Mycologie et Antifongiques-Mycologie Moléculaire ( CNRMA ), Microenvironnement et Immunité, Dynamique des Interactions Hôte-Pathogène, Equipe 3: EREN- Equipe de Recherche en Epidémiologie Nutritionnelle ( CRESS - U1153 ), Université Paris 13 ( UP13 ) -Institut National de la Recherche Agronomique ( INRA ) -Conservatoire National des Arts et Métiers [CNAM] ( CNAM ) -Centre de Recherche Épidémiologie et Statistique Sorbonne Paris Cité ( CRESS (U1153 / UMR_A 1125) ), Institut National de la Recherche Agronomique ( INRA ) -Université Sorbonne Paris Cité ( USPC ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Institut National de la Recherche Agronomique ( INRA ) -Université Sorbonne Paris Cité ( USPC ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Institut de formation en masso-kinésithérapie ( IFMK Brest ), Université de Brest ( UBO ) -Centre Hospitalier Régional Universitaire de Brest ( CHRU Brest ), Signalisation des Cytokines, Assistance publique - Hôpitaux de Paris (AP-HP)-CHU Cochin [AP-HP], Génétique fonctionnelle des Maladies infectieuses, CERLIS - Centre de recherche sur les liens sociaux - UMR 8070 ( CERLIS - UMR 8070 ), Centre National de la Recherche Scientifique ( CNRS ) -Université Paris Descartes - Paris 5 ( UPD5 ) -Université Sorbonne Nouvelle - Paris 3, Biologie systémique, Imagopole ( CITECH ), Institut Curie, Institut Curie-Institut Curie, Cytokines et Immunologie des Tumeurs Humaines ( U753 ), Université Paris-Sud - Paris 11 ( UP11 ) -Institut Gustave Roussy ( IGR ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Assistance publique - Hôpitaux de Paris (AP-HP)-Université Paris Diderot - Paris 7 ( UPD7 ), Investigation Clinique et d’Accès aux Ressources Biologiques (Plate-forme) - Clinical Investigation and Access to BioResources ( ICAReB ), ANR : 10-LABX-0069,MILIEU INTERIEUR,GENETIC & ENVIRONMENTAL CONTROL OF IMMUNE PHENOTYPE VARIANCE: ESTABLISHING A PATH TOWARDS PERSONALIZED MEDICINE ( 2010 ), Centre de Recherche Translationnelle (CRT), Centre National de la Recherche Scientifique (CNRS)-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12)-Université Panthéon-Sorbonne (UP1), Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Paris 13 (UP13)-Institut National de la Recherche Agronomique (INRA)-Conservatoire National des Arts et Métiers [CNAM] (CNAM)-Centre de Recherche Épidémiologie et Statistique Sorbonne Paris Cité (CRESS (U1153 / UMR_A_1125 / UMR_S_1153)), CHU Cochin [AP-HP]-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), Université Paris Diderot - Paris 7 (UPD7)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), and ANR: 10-LABX-0069,MILIEU INTERIEUR,GENETIC & ENVIRONMENTAL CONTROL OF IMMUNE PHENOTYPE VARIANCE: ESTABLISHING A PATH TOWARDS PERSONALIZED MEDICINE(2010)
Subjects: Chemokine, MESH : Cytokines, MESH : Antigens, Disease, MESH : Monitoring, Immunologic, Adaptive Immunity, 0302 clinical medicine, Immunophenotyping, Reference Values, [ SDV.IMM ] Life Sciences [q-bio]/Immunology, Immunology and Allergy, MESH: Monitoring, Immunologic, MESH: Healthy Volunteers, 0303 health sciences, MESH: Cytokines, biology, MESH: Reference Values, MESH : Immunity, Innate, Healthy Volunteers, 3. Good health, MESH: Reproducibility of Results, Infectious Diseases, Cytokines, [SDV.IMM]Life Sciences [q-bio]/Immunology, MESH: Antigens, MESH: Immunity, Innate, Inflammation Mediators, Functional analysis (psychology), Whole blood stimulation, MESH : Adaptive Immunity, Immunology, MESH: Inflammation Mediators, MESH : Reference Values, MESH : Inflammation Mediators, 03 medical and health sciences, Immune system, Immunity, Monitoring, Immunologic, Humans, Antigens, 030304 developmental biology, MESH : Healthy Volunteers, MESH: Humans, MESH : Reproducibility of Results, MESH : Humans, Reproducibility of Results, Immunity, Innate, Reference values, biology.protein, MESH: Adaptive Immunity, 030215 immunology
Abstract: International audience; Standardization of immunophenotyping procedures has become a high priority. We have developed a suite of whole-blood, syringe-based assay systems that can be used to reproducibly assess induced innate or adaptive immune responses. By eliminating preanalytical errors associated with immune monitoring, we have defined the protein signatures induced by (1) medically relevant bacteria, fungi, and viruses; (2) agonists specific for defined host sensors; (3) clinically employed cytokines; and (4) activators of T cell immunity. Our results provide an initial assessment of healthy donor reference values for induced cytokines and chemokines and we report the failure to release interleukin-1α as a common immunological phenotype. The observed naturally occurring variation of the immune response may help to explain differential susceptibility to disease or response to therapeutic intervention. The implementation of a general solution for assessment of functional immune responses will help support harmonization of clinical studies and data sharing.
Published: 2014

33. Mycolactone activation of Wiskott-Aldrich syndrome proteins underpins Buruli ulcer formation

Author: Romain Veyron-Churlet, Philippe Bousso, Maria-Isabel Thoulouze, Chiara Zurzolo, Marie-France Carlier, Serge Mostowy, Laure Guenin-Macé, Marie-Thérèse Ruf, Hui Hong, Peter F. Leadlay, Anne Danckaert, Fabrice Chrétien, Caroline Demangel, Guillaume Romet-Lemonne, Immunobiologie de l'Infection - Immunobiology of Infection, Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Biologie Cellulaire des Lymphocytes (BIOCELLY), Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Laboratoire d'Enzymologie et Biochimie Structurales, Centre National de la Recherche Scientifique (CNRS), Cambridge Systems Biology Centre & Department of Biochemistry (CSBC), University of Cambridge [UK] (CAM), Department of Biochemistry, Imagopole (CITECH), Institut Pasteur [Paris], University of Basel (Unibas), Interactions Bactéries-Cellules (UIBC), Institut National de la Recherche Agronomique (INRA)-Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Trafic membranaire et Pathogénèse, Dynamiques des Réponses Immunes, Histopathologie humaine et Modèles animaux, Université de Versailles Saint-Quentin-en-Yvelines - UFR Sciences de la santé Simone Veil (UVSQ Santé), Université de Versailles Saint-Quentin-en-Yvelines (UVSQ), Financial support from the 'Association Raoul Follereau,' the ANR (project PCV 2006-2010), the European Community's Seventh Framework Programme (FP7, nos. 241500 and 241548), an ERC advanced grant (no. 229982 Forcefulactin), the 'Ligue contre le Cancer,' and the 'Region Ile de France,' which awarded R. Veyron-Churlet the postdoctoral fellowship 'DIM Maladies Infectieuses, Parasitaires, et Nosocomiales Emergentes.', Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), Institut Pasteur [Paris] (IP), Institut National de la Recherche Agronomique (INRA)-Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Immunobiologie de l'Infection, Molecular Immunology, Swiss Tropical and Public Health Institute [Basel], Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur [Paris]-Institut National de la Recherche Agronomique (INRA), UVSQ - UFR des sciences de la santé Simone Veil (UVSQ Santé), Service d'Anatomie Pathologique, and AP-HP, Hôpital R. Poincarré, Service d'Anatomie Pathologique, Garches
Subjects: Keratinocytes, Buruli ulcer, Wiskott–Aldrich syndrome, Wiskott-Aldrich Syndrome Protein, Neuronal, MESH: Mycobacterium ulcerans, MESH: Amino Acid Sequence, Propanolamines, Pathogenesis, Mice, chemistry.chemical_compound, Cell Movement, MESH: Buruli Ulcer, MESH: Animals, Mycolactone, Buruli Ulcer, MESH: Cell Movement, MESH: Wiskott-Aldrich Syndrome Protein, Neuronal, 0303 health sciences, biology, MESH: Protein Multimerization, 030302 biochemistry & molecular biology, General Medicine, MESH: Keratinocytes, Cell biology, Actin Cytoskeleton, Protein Transport, Mycobacterium ulcerans, Macrolides, MESH: Macrolides, Research Article, MESH: Cell Nucleus, MESH: Protein Transport, Bacterial Toxins, Molecular Sequence Data, Carbazoles, macromolecular substances, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, MESH: Actins, Actin-Related Protein 2-3 Complex, MESH: Cell Adhesion, 03 medical and health sciences, MESH: Mice, Inbred C57BL, Cell Adhesion, medicine, Animals, Humans, Amino Acid Sequence, Cell adhesion, MESH: Mice, MESH: Propanolamines, Actin, 030304 developmental biology, Cell Nucleus, MESH: Humans, MESH: Molecular Sequence Data, medicine.disease, biology.organism_classification, Actin cytoskeleton, Actins, Wiskott-Aldrich Syndrome Protein Family, Mice, Inbred C57BL, MESH: Actin-Related Protein 2-3 Complex, chemistry, MESH: Bacterial Toxins, MESH: Wiskott-Aldrich Syndrome Protein Family, Immunology, MESH: HeLa Cells, Epidermis, Protein Multimerization, MESH: Epidermis, MESH: Actin Cytoskeleton, MESH: Carbazoles, HeLa Cells
Abstract: Mycolactone is a diffusible lipid secreted by the human pathogen Mycobacterium ulcerans, which induces the formation of open skin lesions referred to as Buruli ulcers. Here, we show that mycolactone operates by hijacking the Wiskott-Aldrich syndrome protein (WASP) family of actin-nucleating factors. By disrupting WASP autoinhibition, mycolactone leads to uncontrolled activation of ARP2/3-mediated assembly of actin in the cytoplasm. In epithelial cells, mycolactone-induced stimulation of ARP2/3 concentrated in the perinuclear region, resulting in defective cell adhesion and directional migration. In vivo injection of mycolactone into mouse ears consistently altered the junctional organization and stratification of keratinocytes, leading to epidermal thinning, followed by rupture. This degradation process was efficiently suppressed by coadministration of the N-WASP inhibitor wiskostatin. These results elucidate the molecular basis of mycolactone activity and provide a mechanism for Buruli ulcer pathogenesis. Our findings should allow for the rationale design of competitive inhibitors of mycolactone binding to N-WASP, with anti-Buruli ulcer therapeutic potential.
Published: 2013

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33 results on '"Immunobiologie de l'Infection - Immunobiology of Infection"'

1. The Sec61 translocon is a therapeutic vulnerability in multiple myeloma

2. Early antibiotic exposure and adverse outcomes in very preterm infants at low risk of early-onset sepsis: the EPIPAGE-2 cohort study

3. De novo synthesized polyunsaturated fatty acids operate as both host immunomodulators and nutrients for Mycobacterium tuberculosis

4. Biochemical and Biological Assays of Mycolactone-Mediated Inhibition of Sec61

5. Molecular Mechanisms Underpinning the Circulation and Cellular Uptake of Mycobacterium ulcerans Toxin Mycolactone

6. Insights into the ancestry evolution of the Mycobacterium tuberculosis complex from analysis of Mycobacterium riyadhense

7. The impact of gut microbiota on depressive-like behaviors and adult hippocampal neurogenesis requires the endocannabinoid system

8. Not too fat to fight -The emerging role of macrophage fatty acid metabolism in immunity to Mycobacterium tuberculosis

9. Immunity against Mycobacterium ulcerans - The subversive role of mycolactone

10. Antigen-Specific In Vivo Killing Assay

11. Mycolactone Purification from M. ulcerans Cultures and HPLC-Based Approaches for Mycolactone Quantification in Biological Samples

12. Spatiotemporal analysis of mycolactone distribution in vivo reveals partial diffusion in the central nervous system

13. Repair of G1 induced DNA double-strand breaks in S-G2/M by alternative NHEJ

14. Mycolactone induces cell death by SETD1B-dependent degradation of glutathione

15. Dysregulation of tryptophan catabolism at the host-skin microbiota interface in Hidradenitis Suppurativa

16. Metabolic alterations in Hurley stage 1 Hidradenitis Suppurativa

17. Effect of gut microbiota on depressive-like behaviors in mice is mediated by the endocannabinoid system

18. CR3 Engaged by PGL-I Triggers Syk-Calcineurin-NFATc to Rewire the Innate Immune Response in Leprosy

19. Recombinant Antibodies against Mycolactone

20. Ipomoeassin F Binds Sec61α to Inhibit Protein Translocation

21. Mycolactone: More than Just a Cytotoxin

22. Mécanisme responsable des effets immunosuppresseurs de la mycolactone, toxine de M. ulcerans

23. Mécanisme responsable des effets immunosuppresseurs de la mycolactone, toxine de M. ulcerans

24. Proteomics Reveals Scope of Mycolactone-mediated Sec61 Blockade and Distinctive Stress Signature

25. Sec61 blockade by mycolactone: A central mechanism in Buruli ulcer disease

26. Mycobacterial Phenolic Glycolipids Selectively Disable TRIF-Dependent TLR4 Signaling in Macrophages

27. Modular total syntheses of mycolactone A/B and its [ 2 H]- isotopologue

28. Zika virus induces massive cytoplasmic vacuolization and paraptosis‐like death in infected cells

29. Pathogenic and immunosuppressive properties of mycobacterial phenolic glycolipids

30. Mycolactone displays anti-inflammatory effects on the nervous system

31. Mycolactone subverts immunity by selectively blocking the Sec61 translocon

32. Functional analysis via standardized whole-blood stimulation systems defines the boundaries of a healthy immune response to complex stimuli

33. Mycolactone activation of Wiskott-Aldrich syndrome proteins underpins Buruli ulcer formation

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33 results on '"Immunobiologie de l'Infection - Immunobiology of Infection"'

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