The Experts below are selected from a list of 495 Experts worldwide ranked by ideXlab platform
Stefan Kunz - One of the best experts on this subject based on the ideXlab platform.
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Lassa Virus Cell Entry Reveals New Aspects of Virus-Host Cell Interaction.
Journal of virology, 2017Co-Authors: Giulia Torriani, Clara Galan-navarro, Stefan KunzAbstract:ABSTRACT Viral entry represents the first step of every viral infection and is a determinant for the host range and disease potential of a virus. Here, we review the latest developments on cell entry of the highly pathogenic Old World Arenavirus Lassa virus, providing novel insights into the complex virus-host cell interaction of this important human pathogen. We will cover new discoveries on the molecular mechanisms of receptor recognition, endocytosis, and the use of late endosomal entry factors.
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Lassa virus cell entry via dystroglycan involves an unusual pathway of macropinocytosis
Journal of virology, 2016Co-Authors: Joël Oppliger, Giulia Torriani, Antonio Herrador, Stefan KunzAbstract:ABSTRACT The pathogenic Old World Arenavirus Lassa virus (LASV) causes a severe hemorrhagic fever with a high rate of mortality in humans. Several LASV receptors, including dystroglycan (DG), TAM receptor tyrosine kinases, and C-type lectins, have been identified, suggesting complex receptor use. Upon receptor binding, LASV enters the host cell via an unknown clathrin- and dynamin-independent pathway that delivers the virus to late endosomes, where fusion occurs. Here we investigated the mechanisms underlying LASV endocytosis in human cells in the context of productive Arenavirus infection, using recombinant lymphocytic choriomeningitis virus (rLCMV) expressing the LASV glycoprotein (rLCMV-LASVGP). We found that rLCMV-LASVGP entered human epithelial cells via DG using a macropinocytosis-related pathway independently of alternative receptors. Dystroglycan-mediated entry of rLCMV-LASVGP required sodium hydrogen exchangers, actin, and the GTPase Cdc42 and its downstream targets, p21-activating kinase-1 (PAK1) and Wiskott-Aldrich syndrome protein (N-Wasp). Unlike other viruses that enter cells via macropinocytosis, rLCMV-LASVGP entry did not induce overt changes in cellular morphology and hardly affected actin dynamics or fluid uptake. Screening of kinase inhibitors identified protein kinase C, phosphoinositide 3-kinase, and the receptor tyrosine kinase human hepatocyte growth factor receptor (HGFR) to be regulators of rLCMV-LASVGP entry. The HGFR inhibitor EMD 1214063, a candidate anticancer drug, showed antiviral activity against rLCMV-LASVGP at the level of entry. When combined with ribavirin, which is currently used to treat human Arenavirus infection, EMD 1214063 showed additive antiviral effects. In sum, our study reveals that DG can link LASV to an unusual pathway of macropinocytosis that causes only minimal perturbation of the host cell and identifies cellular kinases to be possible novel targets for therapeutic intervention. IMPORTANCE Lassa virus (LASV) causes several hundred thousand infections per year in Western Africa, with the mortality rate among hospitalized patients being high. The current lack of a vaccine and the limited therapeutic options at hand make the development of new drugs against LASV a high priority. In the present study, we uncover that LASV entry into human cells via its major receptor, dystroglycan, involves an unusual pathway of macropinocytosis and define a set of cellular factors implicated in the regulation of LASV entry. A screen of kinase inhibitors revealed HGFR to be a possible candidate target for antiviral drugs against LASV. An HGFR candidate inhibitor currently being evaluated for cancer treatment showed potent antiviral activity and additive drug effects with ribavirin, which is used in the clinic to treat human LASV infection. In sum, our study reveals novel fundamental aspects of the LASV-host cell interaction and highlights a possible candidate drug target for therapeutic intervention.
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Lymphocytic Choriomeningitis Virus Differentially Affects the Virus-Induced Type I Interferon Response and Mitochondrial Apoptosis Mediated by RIG-I/MAVS
Journal of virology, 2015Co-Authors: Christelle Pythoud, Juan-carlos De La Torre, Sylvia Rothenberger, Luis Martínez-sobrido, Stefan KunzAbstract:Arenaviruses are important emerging human pathogens maintained by noncytolytic persistent infection in their rodent reservoir hosts. Despite high levels of viral replication, persistently infected carrier hosts show only mildly elevated levels of type I interferon (IFN-I). Accordingly, the Arenavirus nucleoprotein (NP) has been identified as a potent IFN-I antagonist capable of blocking activation of interferon regulatory factor 3 (IRF3) via the retinoic acid inducible gene (RIG)-I/mitochondrial antiviral signaling (MAVS) pathway. Another important mechanism of host innate antiviral defense is represented by virus-induced mitochondrial apoptosis via RIG-I/MAVS and IRF3. In the present study, we investigated the ability of the prototypic Old World Arenavirus lymphocytic choriomeningitis virus (LCMV) to interfere with RIG-I/MAVS-dependent apoptosis. We found that LCMV does not induce apoptosis at any time during infection. While LCMV efficiently blocked induction of IFN-I via RIG-I/MAVS in response to superinfection with cytopathic RNA viruses, virus-induced mitochondrial apoptosis remained fully active in LCMV-infected cells. Notably, in LCMV-infected cells, RIG-I was dispensable for virus-induced apoptosis via MAVS. Our study reveals that LCMV infection efficiently suppresses induction of IFN-I but does not interfere with the cell's ability to undergo virus-induced mitochondrial apoptosis as a strategy of innate antiviral defense. The RIG-I independence of mitochondrial apoptosis in LCMV-infected cells provides the first evidence that Arenaviruses can reshape apoptotic signaling according to their needs. IMPORTANCE Arenaviruses are important emerging human pathogens that are maintained in their rodent hosts by persistent infection. Persistent virus is able to subvert the cellular interferon response, a powerful branch of the innate antiviral defense. Here, we investigated the ability of the prototypic Arenavirus lymphocytic choriomeningitis virus (LCMV) to interfere with the induction of programmed cell death, or apoptosis, in response to superinfection with cytopathic RNA viruses. Upon viral challenge, persistent LCMV efficiently blocked induction of interferons, whereas virus-induced apoptosis remained fully active in LCMV-infected cells. Our studies reveal that the persistent virus is able to reshape innate apoptotic signaling in order to prevent interferon production while maintaining programmed cell death as a strategy for innate defense. The differential effect of persistent virus on the interferon response versus its effect on apoptosis appears as a subtle strategy to guarantee sufficiently high viral loads for efficient transmission while maintaining apoptosis as a mechanism of defense.
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Old World Arenaviruses Enter the Host Cell via the Multivesicular Body and Depend on the Endosomal Sorting Complex Required for Transport
2013Co-Authors: Giulia Pasqual, Jillian M Rojek, Mark Masin, Jean Yves Chatton, Stefan KunzAbstract:The highly pathogenic Old World Arenavirus Lassa virus (LASV) and the prototypic Arenavirus lymphocytic choriomeningitis virus (LCMV) use a-dystroglycan as a cellular receptor and enter the host cell by an unusual endocytotic pathway independent of clathrin, caveolin, dynamin, and actin. Upon internalization, the viruses are delivered to acidified endosomes in a Rab5-independent manner bypassing classical routes of incoming vesicular trafficking. Here we sought to identify cellular factors involved in the unusual and largely unknown entry pathway of LASV and LCMV. Cell entry of LASV and LCMV required microtubular transport to late endosomes, consistent with the low fusion pH of the viral envelope glycoproteins. Productive infection with recombinant LCMV expressing LASV envelope glycoprotein (rLCMV-LASVGP) and LCMV depended on phosphatidyl inositol 3-kinase (PI3K) as well as lysobisphosphatidic acid (LBPA), an unusual phospholipid that is involved in the formation of intraluminal vesicles (ILV) of the multivesicular body (MVB) of the late endosome. We provide evidence for a role of the endosomal sorting complex required for transport (ESCRT) in LASV and LCMV cell entry, in particular the ESCRT components Hrs, Tsg101, Vps22, and Vps24, as well as the ESCRT-associated ATPase Vps4 involved in fission of ILV. Productive infection with rLCMV-LASVGP and LCMV also critically depended on the ESCRT-associated protein Alix, which is implicated in membrane dynamics of the MVB/late endosomes. Our study identifies crucial cellular factors implicated in Old World Arenavirus cell entry and indicates that LASV and LCMV invade the host cell passing via the MVB/late endosome. Ou
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The role of proteolytic processing and the stable signal peptide in expression of the Old World Arenavirus envelope glycoprotein ectodomain.
Virology, 2012Co-Authors: Dominique J. Burri, Michael B. A. Oldstone, Antonella Pasquato, Joel Ramos Da Palma, Sébastien Igonet, Stefan KunzAbstract:Maturation of the Arenavirus GP precursor (GPC) involves proteolytic processing by cellular signal peptidase and the proprotein convertase subtilisin kexin isozyme 1 (SKI-1)/site 1 protease (S1P), yielding a tripartite complex comprised of a stable signal peptide (SSP), the receptor-binding GP1, and the fusion-active transmembrane GP2. Here we investigated the roles of SKI-1/S1P processing and SSP in the biosynthesis of the recombinant GP ectodomains of lymphocytic choriomeningitis virus (LCMV) and Lassa virus (LASV). When expressed in mammalian cells, the LCMV and LASV GP ectodomains underwent processing by SKI-1/S1P, followed by dissociation of GP1 from GP2. The GP2 ectodomain spontaneously formed trimers as revealed by chemical cross-linking. The endogenous SSP, known to be crucial for maturation and transport of full-length Arenavirus GPC was dispensable for processing and secretion of the soluble GP ectodomain, suggesting a specific role of SSP in the stable prefusion conformation and transport of full-length GPC.
Tomas Kirchhausen - One of the best experts on this subject based on the ideXlab platform.
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nrp2 and cd63 are host factors for lujo virus cell entry
Cell Host & Microbe, 2017Co-Authors: Matthijs Raaben, Andrew S Herbert, Ana I Kuehne, Sarah H Stubbs, Yiying Chou, Vincent A Blomen, Tomas Kirchhausen, Thijn R BrummelkampAbstract:Summary Arenaviruses cause fatal hemorrhagic disease in humans. Old World Arenavirus glycoproteins (GPs) mainly engage α-dystroglycan as a cell-surface receptor, while New World Arenaviruses hijack transferrin receptor. However, the Lujo virus (LUJV) GP does not cluster with New or Old World Arenaviruses. Using a recombinant vesicular stomatitis virus containing LUJV GP as its sole attachment and fusion protein (VSV-LUJV), we demonstrate that infection is independent of known Arenavirus receptor genes. A genome-wide haploid genetic screen identified the transmembrane protein neuropilin 2 (NRP2) and tetraspanin CD63 as factors for LUJV GP-mediated infection. LUJV GP binds the N-terminal domain of NRP2, while CD63 stimulates pH-activated LUJV GP-mediated membrane fusion. Overexpression of NRP2 or its N-terminal domain enhances VSV-LUJV infection, and cells lacking NRP2 are deficient in wild-type LUJV infection. These findings uncover this distinct set of host cell entry factors in LUJV infection and are attractive focus points for therapeutic intervention.
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Superinfection exclusion is absent during acute Junin virus infection of Vero and A549 cells
Scientific Reports, 2015Co-Authors: Raphaël Gaudin, Tomas KirchhausenAbstract:Many viruses have evolved strategies of so-called “superinfection exclusion” to prevent re-infection of a cell that the same virus has already infected. Although Old World Arenavirus infection results in down-regulation of its viral receptor and thus superinfection exclusion, whether New World Arenaviruses have evolved such a mechanism remains unclear. Here we show that acute infection by the New World Junin virus (JUNV) failed to down-regulate the transferrin receptor and did not induce superinfection exclusion. We observed that Vero cells infected by a first round of JUNV ( Candid1 strain) preserve an ability to internalize new incoming JUNV particles that is comparable to that of non-infected cells. Moreover, we developed a dual infection assay with the wild-type Candid1 JUNV and a recombinant JUNV-GFP virus to discriminate between first and second infections at the transcriptional and translational levels. We found that Vero and A549 cells already infected by JUNV were fully competent to transcribe viral RNA from a second round of infection. Furthermore, flow cytometry analysis of viral protein expression indicated that viral translation was normal, regardless of whether cells were previously infected or not. We conclude that in acutely infected cells, Junin virus lacks a superinfection exclusion mechanism.
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superinfection exclusion is absent during acute junin virus infection of vero and a549 cells
Scientific Reports, 2015Co-Authors: Raphaël Gaudin, Tomas KirchhausenAbstract:Many viruses have evolved strategies of so-called “superinfection exclusion” to prevent re-infection of a cell that the same virus has already infected. Although Old World Arenavirus infection results in down-regulation of its viral receptor and thus superinfection exclusion, whether New World Arenaviruses have evolved such a mechanism remains unclear. Here we show that acute infection by the New World Junin virus (JUNV) failed to down-regulate the transferrin receptor and did not induce superinfection exclusion. We observed that Vero cells infected by a first round of JUNV (Candid1 strain) preserve an ability to internalize new incoming JUNV particles that is comparable to that of non-infected cells. Moreover, we developed a dual infection assay with the wild-type Candid1 JUNV and a recombinant JUNV-GFP virus to discriminate between first and second infections at the transcriptional and translational levels. We found that Vero and A549 cells already infected by JUNV were fully competent to transcribe viral RNA from a second round of infection. Furthermore, flow cytometry analysis of viral protein expression indicated that viral translation was normal, regardless of whether cells were previously infected or not. We conclude that in acutely infected cells, Junin virus lacks a superinfection exclusion mechanism. Arenaviruses are enveloped viruses with two segments of an ambisense single-stranded RNA genome. Some of these viruses cause hemorrhagic fever with poor prognoses in humans, including the New World (NW) Arenavirus (clade B) Junin virus (JUNV), which is responsible for Argentine hemorrhagic
Jillian M Rojek - One of the best experts on this subject based on the ideXlab platform.
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Old World Arenaviruses Enter the Host Cell via the Multivesicular Body and Depend on the Endosomal Sorting Complex Required for Transport
2013Co-Authors: Giulia Pasqual, Jillian M Rojek, Mark Masin, Jean Yves Chatton, Stefan KunzAbstract:The highly pathogenic Old World Arenavirus Lassa virus (LASV) and the prototypic Arenavirus lymphocytic choriomeningitis virus (LCMV) use a-dystroglycan as a cellular receptor and enter the host cell by an unusual endocytotic pathway independent of clathrin, caveolin, dynamin, and actin. Upon internalization, the viruses are delivered to acidified endosomes in a Rab5-independent manner bypassing classical routes of incoming vesicular trafficking. Here we sought to identify cellular factors involved in the unusual and largely unknown entry pathway of LASV and LCMV. Cell entry of LASV and LCMV required microtubular transport to late endosomes, consistent with the low fusion pH of the viral envelope glycoproteins. Productive infection with recombinant LCMV expressing LASV envelope glycoprotein (rLCMV-LASVGP) and LCMV depended on phosphatidyl inositol 3-kinase (PI3K) as well as lysobisphosphatidic acid (LBPA), an unusual phospholipid that is involved in the formation of intraluminal vesicles (ILV) of the multivesicular body (MVB) of the late endosome. We provide evidence for a role of the endosomal sorting complex required for transport (ESCRT) in LASV and LCMV cell entry, in particular the ESCRT components Hrs, Tsg101, Vps22, and Vps24, as well as the ESCRT-associated ATPase Vps4 involved in fission of ILV. Productive infection with rLCMV-LASVGP and LCMV also critically depended on the ESCRT-associated protein Alix, which is implicated in membrane dynamics of the MVB/late endosomes. Our study identifies crucial cellular factors implicated in Old World Arenavirus cell entry and indicates that LASV and LCMV invade the host cell passing via the MVB/late endosome. Ou
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Old World Arenaviruses enter the host cell via the multivesicular body and depend on the endosomal sorting complex required for transport
PLOS Pathogens, 2011Co-Authors: Giulia Pasqual, Jillian M Rojek, Mark Masin, Jean Yves Chatton, Stefan KunzAbstract:The highly pathogenic Old World Arenavirus Lassa virus (LASV) and the prototypic Arenavirus lymphocytic choriomeningitis virus (LCMV) use α-dystroglycan as a cellular receptor and enter the host cell by an unusual endocytotic pathway independent of clathrin, caveolin, dynamin, and actin. Upon internalization, the viruses are delivered to acidified endosomes in a Rab5-independent manner bypassing classical routes of incoming vesicular trafficking. Here we sought to identify cellular factors involved in the unusual and largely unknown entry pathway of LASV and LCMV. Cell entry of LASV and LCMV required microtubular transport to late endosomes, consistent with the low fusion pH of the viral envelope glycoproteins. Productive infection with recombinant LCMV expressing LASV envelope glycoprotein (rLCMV-LASVGP) and LCMV depended on phosphatidyl inositol 3-kinase (PI3K) as well as lysobisphosphatidic acid (LBPA), an unusual phospholipid that is involved in the formation of intraluminal vesicles (ILV) of the multivesicular body (MVB) of the late endosome. We provide evidence for a role of the endosomal sorting complex required for transport (ESCRT) in LASV and LCMV cell entry, in particular the ESCRT components Hrs, Tsg101, Vps22, and Vps24, as well as the ESCRT-associated ATPase Vps4 involved in fission of ILV. Productive infection with rLCMV-LASVGP and LCMV also critically depended on the ESCRT-associated protein Alix, which is implicated in membrane dynamics of the MVB/late endosomes. Our study identifies crucial cellular factors implicated in Old World Arenavirus cell entry and indicates that LASV and LCMV invade the host cell passing via the MVB/late endosome. Our data further suggest that the virus-receptor complexes undergo sorting into ILV of the MVB mediated by the ESCRT, possibly using a pathway that may be linked to the cellular trafficking and degradation of the cellular receptor.
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Different Mechanisms of Cell Entry by Human-Pathogenic Old World and New World Arenaviruses
Journal of virology, 2008Co-Authors: Jillian M Rojek, Ana B. Sanchez, Ngoc Thao Nguyen, Juan-carlos De La Torre, Stefan KunzAbstract:The Old World Arenavirus Lassa virus (LASV) is the causative agent of severe viral hemorrhagic fever (VHF) in humans and is the most prevalent human pathogen among Arenaviruses. The present study investigated the largely unknown mechanisms of cell entry of LASV, a process know to be mediated solely by the virus envelope glycoprotein (GP). To circumvent biosafety restrictions associated with the use of live LASV, we used reverse genetics to generate a recombinant variant of the prototypic Arenavirus lymphocytic choriomeningitis virus (LCMV) expressing the LASV GP (rLCMV-LASVGP). The rescued rLCMV-LASVGP grew to titers comparable to that of LCMV and showed the receptor binding characteristics of LASV. We used rLCMV-LASVGP to characterize the cellular mechanisms of LASV entry in the context of a productive Arenavirus infection. The kinetics of pH-dependent membrane fusion of rLCMV-LASVGP resembled those of the human-pathogenic New World Arenavirus Junin virus (JUNV) and other enveloped viruses that use clathrin-mediated endocytosis for entry. However, rLCMV-LASVGP entered cells predominantly via a clathrin-, caveolin-, and dynamin-independent endocytotic pathway similar to the one recently described for LCMV. Productive infection of rLCMV-LASVGP was only mildly affected by a dominant negative mutant of Rab5 and was independent of Rab7, suggesting an unusual mechanism of delivery to endosomes. In addition, rLCMV-LASVGP infection was independent of actin but required intact microtubules. Our data indicate that LASV enters cells via a pathway distinct from the one used by human-pathogenic New World Arenaviruses.
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Cell entry by human pathogenic Arenaviruses.
Cellular microbiology, 2007Co-Authors: Jillian M Rojek, Stefan KunzAbstract:The Arenaviruses Lassa virus (LASV) in Africa and Machupo (MACV), Guanarito (GTOV) and Junin viruses (JUNV) in South America cause severe haemorrhagic fevers in humans with fatality rates of 15-35%. The present review focuses on the first steps of infection with human pathogenic Arenaviruses, the interaction with their cellular receptor molecules and subsequent entry into the host cell. While similarities exist in genomic organization, structure and clinical disease caused by pathogenic Old World and New World Arenaviruses these pathogens use different primary receptors. The Old World Arenaviruses employ alpha-dystroglycan, a cellular receptor for proteins of the extracellular matrix, and the human pathogenic New World Arenaviruses use the cellular cargo receptor transferrin receptor 1. While the New World Arenavirus JUNV enters cells via clathrin-dependent endocytosis, evidence occurred for clathrin-independent entry of the prototypic Old World Arenavirus lymphocytic choriomeningitis virus. Upon internalization, Arenaviruses are delivered to the endosome, where pH-dependent membrane fusion is mediated by the envelope glycoprotein (GP). While Arenavirus GPs share characteristics with class I fusion GPs of other enveloped viruses, unusual mechanistic features of GP-mediated membrane fusion have recently been discovered for Arenaviruses with important implications for viral entry.
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Cellular Entry of Lymphocytic Choriomeningitis Virus
Journal of virology, 2007Co-Authors: Jillian M Rojek, Mar Perez, Stefan KunzAbstract:In contrast to most enveloped viruses that enter the host cell via clathrin-dependent endocytosis, the Old World Arenavirus lymphocytic choriomeningitis virus (LCMV) enters cells via noncoated vesicles that deliver the virus to endosomes, where pH-dependent membrane fusion occurs. Here, we investigated the initial steps of LCMV infection. We found that the attachment of LCMV to its cellular receptor α-dystroglycan occurs rapidly and is not dependent on membrane cholesterol. However, subsequent virus internalization is sensitive to cholesterol depletion, indicating the involvement of a cholesterol-dependent pathway. We provide evidence that LCMV entry involves an endocytotic pathway that is independent of clathrin and caveolin and that does not require the GTPase dynamin. In addition, neither the structural integrity nor the dynamics of the actin cytoskeleton are required for infection. These findings indicate that the prototypic Old World Arenavirus LCMV uses a mechanism of entry that is different from clathrin-mediated endocytosis, which is used by the New World Arenavirus Junin virus, and pathways used by other enveloped viruses.
Michael J. Buchmeier - One of the best experts on this subject based on the ideXlab platform.
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Single Nucleoprotein Residue Modulates Arenavirus Replication Complex Formation
mBio, 2015Co-Authors: Kristeene A. Knopp, Tuan Ngo, Paul D. Gershon, Michael J. BuchmeierAbstract:TheArenaviridae are enveloped, negative-sense RNA viruses with several family members that cause hemorrhagic fevers. This work provides immunofluorescence evidence that, unlike those of New World Arenaviruses, the replication and tran- scription complexes (RTC) of lymphocytic choriomeningitis virus (LCMV) colocalize with eukaryotic initiation factor 4E (eIF4E) and that eIF4E may participate in the translation of LCMV mRNA. Additionally, we identify two residues in the LCMV nucleoprotein (NP) that are conserved in every mammalian Arenavirus and are required for recombinant LCMV recovery. One of these sites, Y125, was confirmed to be phosphorylated by using liquid chromatography-tandem mass spectrometry (LC-MS/ MS). NP Y125 is located in the N-terminal region of NP that is disordered when RNA is bound. The other site, NP T206, was pre- dicted to be a phosphorylation site. Immunofluorescence analysis demonstrated that NP T206 is required for the formation of the punctate RTC that are typically observed during LCMV infection. A minigenome reporter assay using NP mutants, as well as Northern blot analysis, demonstrated that although NP T206A does not form punctate RTC, it can transcribe and replicate a minigenome. However, in the presence of matrix protein (Z) and glycoprotein (GP), translation of the minigenome message with NP T206A was inhibited, suggesting that punctate RTC formation is required to regulate viral replication. Together, these results highlight a significant difference between New and Old World Arenaviruses and demonstrate the importance of RTC formation and translation priming in RTC for Old World Arenaviruses. IMPORTANCE Several members of the Arenaviridae cause hemorrhagic fevers and are classified as category A pathogens. Arena- virus replication-transcription complexes (RTC) are nucleated by the viral nucleoprotein. This study demonstrates that the for- mation of these complexes is required for virus viability and suggests that RTC nucleation is regulated by the phosphorylation of a single nucleoprotein residue. This work adds to the body of knowledge about how these key viral structures are formed and participate in virus replication. Additionally, the fact that Old World Arenavirus complexes colocalize with the eukaryotic initia- tion factor 4E, while New World Arenaviruses do not, is only the second notable difference observed between New and Old World Arenaviruses, thefirst being the difference in the glycoprotein receptor.
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A multivalent vaccination strategy for the prevention of Old World Arenavirus infection in humans.
Journal of Virology, 2010Co-Authors: Jason Botten, J. Lindsay Whitton, Polly Barrowman, John Sidney, Jason K. Whitmire, Jeff Alexander, Maya F. Kotturi, Alessandro Sette, Michael J. BuchmeierAbstract:Arenaviruses cause severe human disease ranging from aseptic meningitis following lymphocytic choriomeningitis virus (LCMV) infection to hemorrhagic fever syndromes following infection with Guanarito virus (GTOV), Junin virus (JUNV), Lassa virus (LASV), Machupo virus (MACV), Sabia virus (SABV), or Whitewater Arroyo virus (WWAV). Cellular immunity, chiefly the CD8 T-cell response, plays a critical role in providing protective immunity following infection with the Old World Arenaviruses LASV and LCMV. In the current study, we evaluated whether HLA class I-restricted epitopes that are cross-reactive among pathogenic Arenaviruses could be identified for the purpose of developing an epitope-based vaccination approach that would cross-protect against multiple Arenaviruses. We were able to identify a panel of HLA-A*0201-restricted peptides derived from the same region of the glycoprotein precursor (GPC) of LASV (GPC spanning residues 441 to 449 [GPC441-449]), LCMV (GPC447-455), JUNV (GPC429-437), MACV (GPC444-452), GTOV (GPC427-435), and WWAV (GPC428-436) that displayed high-affinity binding to HLA-A*0201 and were recognized by CD8 T cells in a cross-reactive manner following LCMV infection or peptide immunization of HLA-A*0201 transgenic mice. Immunization of HLA-A*0201 mice with the Old World peptide LASV GPC441-449 or LCMV GPC447-455 induced high-avidity CD8 T-cell responses that were able to kill syngeneic target cells pulsed with either LASV GPC441-449 or LCMV GPC447-455 in vivo and provided significant protection against viral challenge with LCMV. Through this study, we have demonstrated that HLA class I-restricted, cross-reactive epitopes exist among diverse Arenaviruses and that individual epitopes can be utilized as effective vaccine determinants for multiple pathogenic Arenaviruses. Arenaviruses are a family of rodent-borne viruses that are associated with severe disease in humans. Phylogenetically, the Arenaviruses are organized into Old World or New World groups, with a subdivision of the New World viruses falling into three distinct lineages (A to C) (17). At least eight Arenaviruses are known to cause human disease. Of the New World viruses, Junin virus (JUNV), Machupo virus (MACV), Guanarito virus (GTOV), and Sabia virus (SABV) (all members of lineage B) are etiologic agents of hemorrhagic fever syndromes in South
Sylvain Baize - One of the best experts on this subject based on the ideXlab platform.
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The exonuclease domain of Lassa virus nucleoprotein is involved in antigen-presenting-cell-mediated NK cell responses
Journal of Virology, 2014Co-Authors: Marion Russier, Xavier Carnec, Stéphanie Reynard, Sylvain BaizeAbstract:Lassa virus is an Old World Arenavirus which causes Lassa hemorrhagic fever in humans, mostly in West Africa. Lassa fever is an important public health problem, and a safe and effective vaccine is urgently needed. The infection causes immunosuppression, probably due to the absence of activation of antigen-presenting cells (dendritic cells and macrophages), low type I interferon (IFN) production, and deficient NK cell function. However, a recombinant Lassa virus carrying D389A and G392A substitutions in the nucleoprotein that abolish the exonuclease activity and IFN activation loses its inhibitory activity and induces strong type I IFN production by dendritic cells and macrophages. We show here that during infection by this mutant Lassa virus, antigen-presenting cells trigger efficient human NK cell responses in vitro, including production of IFN-γ and cytotoxicity. NK cell activation involves close contact with both antigen-presenting cells and soluble factors. We report that infected dendritic cells and macrophages express the NKG2D ligands major histocompatibility complex (MHC) class I-related chains A and B and that they may produce interleukin-12 (IL-12), IL-15, and IL-18, all involved in NK cell functions. NK cell degranulation is significantly increased in cocultures, suggesting that NK cells seem to kill infected dendritic cells and macrophages. This work confirms the inhibitory function of Lassa virus nucleoprotein. Importantly, we demonstrate for the first time that Lassa virus nucleoprotein is involved in the inhibition of antigen-presenting cell-mediated NK cell responses. IMPORTANCE: The pathogenesis and immune responses induced by Lassa virus are poorly known. Recently, an exonuclease domain contained in the viral nucleoprotein has been shown to be able to inhibit the type I IFN response by avoiding the recognition of viral RNA by cell sensors. Here, we studied the responses of NK cells to dendritic cells and macrophages infected with a recombinant Lassa virus in which the exonuclease functions have been abolished and demonstrated that NK cells are strongly activated and presented effective functions. These results show that the strategy developed by Lassa virus to evade innate immunity is also effective on NK cells, explaining the weak NK cell activation observed with the wild-type virus. By providing a better understanding of the interactions between Lassa virus and the host immune system, these results are important for the field of Arenavirus biology and may be useful for a vaccine approach against Lassa fever.
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Immune responses and Lassa virus infection.
Viruses, 2012Co-Authors: Marion Russier, Delphine Pannetier, Sylvain BaizeAbstract:Lassa fever is a hemorrhagic fever endemic to West Africa and caused by Lassa virus, an Old World Arenavirus. It may be fatal, but most patients recover from acute disease and some experience asymptomatic infection. The immune mechanisms associated with these different outcomes have not yet been fully elucidated, but considerable progress has recently been made, through the use of in vitro human models and nonhuman primates, the only relevant animal model that mimics the pathophysiology and immune responses induced in patients. We discuss here the roles of the various components of the innate and adaptive immune systems in Lassa virus infection and in the control of viral replication and pathogenesis.
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Lassa Virus Nucleoprotein Mutants Generated by Reverse Genetics Induce a Robust Type I Interferon Response in Human Dendritic Cells and Macrophages
Journal of virology, 2011Co-Authors: Xavier Carnec, Sylvain Baize, Noel Tordo, Stéphanie Reynard, Laure Diancourt, Valérie Caro, Michèle BouloyAbstract:Lassa virus (LASV; Arenaviridae) is responsible for severe hemorrhagic fevers in Africa. LASV nucleoprotein (NP) plays important roles in regulating viral transcription and replication and in inhibiting type I interferon (IFN) production. The NP C-terminal domain contains a 3-to-5 exonuclease activity involved in suppressing IFN induction. We have established a murine polymerase (Pol) I reverse genetics system for LASV, showing that residues D389 and G392 of NP were critical for LASV viability, while the D389A/G392A and D389T/392A double mutants were severely altered in the ability to suppress IFN in macrophages and dendritic cells. Assessing their attenuation in vivo may open new perspectives in vaccinology. Lassa fever is a viral hemorrhagic fever caused by an Old World Arenavirus, Lassa virus (LASV; family Arenaviridae) (45) transmitted by infected Mastomys natalensis, a peridomestic rodent (35). It is a major public health concern in regions of endemicity in West Africa and a threat for importation and misuse as a bioterrorism agent in industrial countries (9). The severity of the disease varies from asymptomatic infection to fatal hemorrhagic fever (19, 33, 34). Whether infection leads to death seems to depend on host immune responses, although the mechanisms involved remain to be clarified. LASV tropism for antigen-presenting cells (APC), such as dendritic cells (DC) and macrophages (MP), in the early stages of infection probably plays a key role in the defective cellular responses observed for severe cases (5, 28, 29, 51). DC and MP massively release LASV but are not activated and do not produce cytokines, except for a modest type I interferon (IFN) production (5, 7).
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Lassa Virus Infection of Human Dendritic Cells and Macrophages Is Productive but Fails to Activate Cells
Journal of Immunology, 2004Co-Authors: Sylvain Baize, Delphine Pannetier, Jérôme Kaplon, Caroline Faure, Marie-claude Georges-courbot, Vincent DeubelAbstract:Lassa fever is a hemorrhagic fever caused by Lassa virus (LV), an Old-World Arenavirus. Little is known about the immune responses that occur during the disease, but protection seems to be linked to the induction of cellular responses specific for viral glycoproteins. Conversely, severe Lassa fever may be associated with immunosuppression. We studied the infection of human dendritic cells (DC) and macrophages (MP) by LV. Both these cell types are susceptible to LV infection. Viral nucleoprotein was detected in DC and MP, and high and moderate viral titers were obtained with culture supernatants of DC and MP, respectively. LV did not induce apoptosis in DC and MP. These cells were not activated by LV infection. No change was observed in the expression of surface molecules involved in activation, costimulation, adhesion, and Ag presentation following LV infection, or in the functional properties of DC. Inflammatory cytokine production was not detected at the mRNA or protein level after LV infection of DC and MP. Thus, MP, and particularly DC, are crucial targets for LV and are probably involved in the early replication of LV from the initial site of infection. The lack of activation and maturation of cells following infection may be associated with the immunosuppression observed in severe LV infection.
