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Host cell invasion by Lassa virus

English title Host cell invasion by Lassa virus
Applicant Kunz Stefan
Number 149746
Funding scheme Project funding
Research institution Institut de Microbiologie - CHUV Faculté de Biologie et Médecine Université de Lausanne
Institution of higher education University of Lausanne - LA
Main discipline Medical Microbiology
Start/End 01.01.2014 - 31.12.2016
Approved amount 623'818.00
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Keywords (5)

Dendritic cells; Receptor; Cell entry; hemorrhagic fever; Lassa fever

Lay Summary (German)

Lead
Unser Projekt erforscht den Eintritt von Lassavirus in die menschliche Zelle. Mit einer Kombination von biochemischen und zellbiologischen Methoden werden wir zelluläre Faktoren identifizieren welche für die Invasion der Wirtszelle essentiell sind. Solche Faktoren werden dann als mögliche Ziele für antivirale Medikamente evaluiert. Unser Projekt wird bisher unbekannte Aspekte der Virus-Zell Interaktion beleuchten und zur Entwicklung neuer wirksamer antiviraler Medikamente beitragen.
Lay summary
Das Lassavirus ist endemisch in Westafrika und verursacht Lassafieber, eine schwere Erkrankung mit einer Sterberate von bis zu 30% bei hospitalisierten Patienten. Lassavirus ist ein grosses Gesundheitsproblem in betroffenen Regionen. Es existiert derzeit keine zugelassene Impfung und die therapeutischen Möglichkeiten sind limitiert. Die Entwicklung neuer und wirksamer Medikamente gegen Lassavirus hat deshalb hohe Priorität. Das Andocken von Viren an zelluläre Rezeptoren und die folgende Internalisierung sind die ersten Schritte jeder viralen Infektion. Als obligatorische zelluläre Parasiten sind Viren dabei weitgehend von der Wirtszelle abhängig. Unser Projekt zielt darauf ab zelluläre Faktoren zu identifizieren welche für die Invasion der Wirtszelle durch Lassavirus essentiell sind. Mit einer Kombination von biochemischen und zellbiologischen Methoden werden wir zelluläre Proteine identifizieren welche Bindung und Eintritt des Virus ermöglichen. Mittels hochauflösender Mikroskopie werden wir dann die Reise des Virus innerhalb menschlicher Zellen verfolgen. Zelluläre Proteine welche für den Vireneintritt essentiell sind, werden in einem nächsten Schritt als mögliche Ziele für antivirale Medikamente evaluiert. Unser Projekt wird unser derzeit limitiertes Verständnis der Interaktion des Lassavirus mit menschlichen Wirtszellen vertiefen. Gleichzeitig werden unsere Resultate neue Wege der therapeutischen Intervention aufzeigen. Vom wissenschaftlichen Standpunkt wird unser Projekt neue Einblicke in derzeit noch unbekannte Aspekte eines wichtigen menschlichen Krankheitserregers liefern. Eine mögliche Verwertbarkeit unserer Resultate für die Entwicklung neuer wirksamer antiviraler Medikamente würde einen Nutzen für die gesamte Gesellschaft darstellen.      
Direct link to Lay Summary Last update: 08.10.2013

Responsible applicant and co-applicants

Employees

Publications

Publication
Lassa Virus Cell Entry via Dystroglycan Involves an Unusual Pathway of Macropinocytosis.
Oppliger Joel, Torriani Giulia, Herrador Antonio, Kunz Stefan (2016), Lassa Virus Cell Entry via Dystroglycan Involves an Unusual Pathway of Macropinocytosis., in Journal of virology, 90(14), 6412-29.
A Molecular Sensor To Characterize Arenavirus Envelope Glycoprotein Cleavage by Subtilisin Kexin Isozyme 1/Site 1 Protease.
Oppliger Joel, da Palma Joel Ramos, Burri Dominique J, Bergeron Eric, Khatib Abdel-Majid, Spiropoulou Christina F, Pasquato Antonella, Kunz Stefan (2015), A Molecular Sensor To Characterize Arenavirus Envelope Glycoprotein Cleavage by Subtilisin Kexin Isozyme 1/Site 1 Protease., in Journal of virology, 90(2), 705-14.
Lymphocytic Choriomeningitis Virus Differentially Affects the Virus-Induced Type I Interferon Response and Mitochondrial Apoptosis Mediated by RIG-I/MAVS.
Pythoud Christelle, Rothenberger Sylvia, Martínez-Sobrido Luis, de la Torre Juan Carlos, Kunz Stefan (2015), Lymphocytic Choriomeningitis Virus Differentially Affects the Virus-Induced Type I Interferon Response and Mitochondrial Apoptosis Mediated by RIG-I/MAVS., in Journal of virology, 89(12), 6240-50.
Lassa virus cell entry reveals new aspects of virus-host cell interaction.
Torriani Giulia, Galan-Navarro Clara, Kunz Stefan, Lassa virus cell entry reveals new aspects of virus-host cell interaction., in Journal of virology.

Collaboration

Group / person Country
Types of collaboration
Dr. Christina Spiroupoulou, Centers of Disase Control and Prevention, Atlanta United States of America (North America)
- Research Infrastructure
Prof. Pierre-Yves Lozach, Universite de Quebec, Laval Canada (North America)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
- Research Infrastructure
Prof. Kevin P. Campbell, head Howard Hughes Medical Institute, University of Iowa United States of America (North America)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
Prof. Juan-Carlos de la Torre, Scripps Research Institute United States of America (North America)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
- Exchange of personnel
Dr. Gisa Gerold, TWINCORE Center, Hannover Germany (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Research Infrastructure
- Exchange of personnel
Prof. Gisou F. van der Goot, EPFL, Lausanne Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
- Research Infrastructure

Scientific events

Active participation

Title Type of contribution Title of article or contribution Date Place Persons involved
Weekly Seminar Department of Immunology and Microbial Science Scripps Research Institute Individual talk Novel Strategies to Combat Pathogenic Arenaviruses 09.12.2016 La Jolla, California, United States of America Kunz Stefan;
Keystone Conference Hemorrhagic Fever Viruses Talk given at a conference Molecular Mechanisms underlying Lassa Virus Invasion of Vascular Endothelial Cells 04.12.2016 Santa Fe, United States of America Kunz Stefan;
Weekly Seminar Institute of Virology University of Heidelberg, Germany Individual talk Novel Strategies to Combat Human Pathogenic Arenaviruses 24.09.2015 Heidelberg, Germany Kunz Stefan;
Conference on Negative Strand Viruses Talk given at a conference Lymphocytic choriomeningitis virus differentially affects virus-induced type I IFN response and mitochondrial apoptosis mediated by RIG-I/MAVS 14.09.2015 Siena, Italy Kunz Stefan;
IUMS 16th International Congress of Virology Individual talk State-of-the-art lecture on arenaviruses 27.08.2014 Montreal, Canada Kunz Stefan;
Symposium Arenaviruses, Institute of Virology Individual talk Arednavirus entry 17.03.2014 Marburg, Germany Kunz Stefan;


Associated projects

Number Title Start Funding scheme
135536 Targeting the processing of arenavirus glycoprotein for anti-viral therapy 01.04.2011 Project funding
132844 Host cell invasion by Lassa virus 01.01.2011 Project funding
160780 Viral Plasticity Underlying Tropism And Pathogenesis/ Innate Immune Evasion Of Emerging Viruses 01.11.2015 Sinergia
143754 Immunoengineering a synthetic vaccine against emerging human pathogenic viruses 01.10.2012 Interdisciplinary projects
132844 Host cell invasion by Lassa virus 01.01.2011 Project funding
170108 Host cell invasion by Lassa virus 01.01.2017 Project funding
170108 Host cell invasion by Lassa virus 01.01.2017 Project funding

Abstract

The Old Word arenavirus Lassa (LASV) is the causative agent of a severe hemorrhagic fever with high mortality in humans. In Western Africa, LASV causes several hundred thousand infections per year with thousands of deaths and represents a serious public health problem. Considering the lack of a licensed vaccine for LASV and the limited therapeutic options, the development of novel and efficacious strategies for anti-viral intervention is an urgent need. The development of novel therapeutics against LASV is hampered by the currently limited knowledge on the viral structure, its life cycle, and its interaction with the host cell. The present project aims at an in-depth understanding of the molecular mechanisms underlying the first and most fundamental steps of LASV infection in human cells, receptor binding and entry. During cell invasion, the virus heavily depends on the molecular machinery of the host cell and cellular factors implicated in viral entry represent promising targets for therapeutic intervention.During the first funding period of this project, we were able to show that LASV binding to its principal cellular receptor dystroglycan (DG) induces receptor-mediated signaling, resulting in post-translational modification of the receptor. These virus-induced signaling events were accompanied by changes in the composition and dynamics of the receptor complex. Upon internalization by a yet unknown endocytotic pathway, the virus was delivered to the multivesicular body (MVB) of the endosome. At the MVB, the virus was able to hijack the endosomal sorting complex required for transport (ESCRT), allowing rapid delivery to late endosomal compartments, where low pH-induced fusion occurred. In the present project renewal, we will continue and extend these studies addressing major unresolved questions about the mechanisms underlying LASV cell invasion. In a first aim, we will investigate the receptor-associated protein-protein interactions involved in LASV attachment and entry using state-of-the art quantitative proteomics. Using stable isotope labeling of amino acids in culture (SILAC), we will identify DG-associated cellular proteins whose interactions change in presence of the virus. Such cellular factors will then be evaluated for their functions in the basic cell biology of DG and their specific role in LASV cell entry assessed. These studies will reveal yet unknown cellular factors involved in LASV infection and also provide novel insights into the biological function of DG in the host cell.A second aim will employ microscopic and imaging techniques to follow the virus’ journey from the cell surface to fusion in the late endosome and beyond. Applying established confocal microscopy techniques, combined with biochemical methods, we will for the first time try to visualize receptor attachment, clustering, and sorting at the plasma membrane. We will then track the virus during endosomal transport and try to identify the yet elusive initial compartment reached after internalization. Upon fusion at the late endosome, the viral ribonucleoprotein (RNP) core enters the cytoplasm, where distinctive replication-transcription complexes are formed. Here we will investigate the role of the cytoskeleton in cytosolic transport of the viral RNP and in early replication. The data at hand indicate that LASV cell attachment and entry involve virus-induced signaling events. The identification of the signaling molecules and pathways involved represents another major goal of our research. To this end, we will employ a two-pronged approach. First, we will use phosphoproteine antibody arrays to identify signaling pathways activated by the virus. This will be complemented by a targeted RNA interference (RNAi) silencing screen for kinases implicated in endocytosis. Candidates will be tested for their role in viral entry in an infection model.In human Lassa fever, dendritic cells (DCs) represent crucial targets of the virus. Our studies in this project revealed that LASV entry in human DCs is facilitated by the C-type lectin DC-SIGN, and likely involves yet unknown entry factors. In the present renewal, we will clarify the specific role of DC-SIGN in LASV entry. In addition, we will try to identify the yet unknown LASV entry factors in DCs using SILAC-based proteomics. In sum, our studies will deepen our understanding of LASV host cell invasion and the discovery of cellular factors involved may provide novel drug targets that will then be evaluated for therapeutic intervention.
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