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Pascale Cossart - One of the best experts on this subject based on the ideXlab platform.
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role in virulence of phospholipases Listeriolysin o and Listeriolysin s from epidemic listeria monocytogenes using the chicken embryo infection model
Veterinary Research, 2018Co-Authors: Juan J Quereda, Pascale Cossart, Christopher Andersson, Jorgen Johansson, Javier PizarrocerdaAbstract:Most human listeriosis outbreaks are caused by Listeria monocytogenes evolutionary lineage I strains which possess four exotoxins: a phosphatidylinositol-specific phospholipase C (PlcA), a broad-range phospholipase C (PlcB), Listeriolysin O (LLO) and Listeriolysin S (LLS). The simultaneous contribution of these molecules to virulence has never been explored. Here, the importance of these four exotoxins of an epidemic lineage I L. monocytogenes strain (F2365) in virulence was assessed in chicken embryos infected in the allantoic cavity. We show that LLS does not play a role in virulence while LLO is required to infect and kill chicken embryos both in wild type transcriptional regulator of virulence PrfA (PrfAWT) and constitutively active PrfA (PrfA*) backgrounds. We demonstrate that PlcA, a toxin previously considered as a minor virulence factor, played a major role in virulence in a PrfA* background. Interestingly, GFP transcriptional fusions show that the plcA promoter is less active than the hly promoter in vitro, explaining why the contribution of PlcA to virulence could be observed more importantly in a PrfA* background. Together, our results suggest that PlcA might play a more important role in the infectious lifecycle of L. monocytogenes than previously thought, explaining why all the strains of L. monocytogenes have conserved an intact copy of plcA in their genomes.
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Listeriolysin O-dependent host surfaceome remodeling modulates Listeria monocytogenes invasion
Pathogens and Disease, 2018Co-Authors: Andreas Kuehbacher, Pascale Cossart, Juan J Quereda, Karel Novy, Martin Sachse, Maryse Moya-nilges, Bernd Wollscheid, Javier Pizarro-cerdaAbstract:Listeria monocytogenes is a pathogenic bacterium that invades epithelial cells by activating host signaling cascades, which promote bacterial engulfment within a phagosome. The pore-forming toxin Listeriolysin O (LLO), which is required for bacteria phagosomal escape, has also been associated with the activation of several signaling pathways when secreted by extracellular bacteria, including Ca2+ influx and promotion of L. monocytogenes entry. Quantitative host surfaceome analysis revealed significant quantitative remodeling of a defined set of cell surface glycoproteins upon LLO treatment, including a subset previously identified to play a role in the L. monocytogenes infection process. Our data further shows that the lysosomal-associated membrane proteins LAMP-1 and LAMP-2 are translocated to the cellular surface and those LLO-induced Ca2+ fluxes are required to trigger the surface relocalization of LAMP-1. Finally, we identify late endosomes/lysosomes as the major donor compartments of LAMP-1 upon LLO treatment and by perturbing their function, we suggest that these organelles participate in L. monocytogenes invasion.
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Listeriolysin S: A bacteriocin from epidemic Listeria monocytogenes strains that targets the gut microbiota.
Gut microbes, 2017Co-Authors: Juan J Quereda, Pascale Cossart, Jazmín Meza-torres, Javier Pizarro-cerdaAbstract:Listeria monocytogenes is a Gram-positive food-borne pathogen that in humans may traverse the intestinal, placental and blood/brain barriers, causing gastroenteritis, abortions and meningitis. Crossing of these barriers is dependent on the bacterial ability to enter host cells, and several L. monocytogenes surface and secreted virulence factors are known to facilitate entry and the intracellular lifecycle. The study of L. monocytogenes strains associated to human listeriosis epidemics has revealed the presence of novel virulence factors. One such factor is Listeriolysin S, a thiazole/oxazole modified microcin that displays bactericidal activity and modifies the host microbiota during infection. Our recent results therefore highlight the interaction of L. monocytogenes with gut microbes as a crucial step in epidemic listeriosis. In this article, we will discuss novel implications for this family of toxins in the pathogenesis of diverse medically relevant microorganisms.
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mapping of sumo sites and analysis of sumoylation changes induced by external stimuli
Proceedings of the National Academy of Sciences of the United States of America, 2014Co-Authors: Francis Impens, Lilliana Radoshevich, Pascale Cossart, David RibetAbstract:SUMOylation is an essential ubiquitin-like modification involved in important biological processes in eukaryotic cells. Identification of small ubiquitin-related modifier (SUMO)-conjugated residues in proteins is critical for understanding the role of SUMOylation but remains experimentally challenging. We have set up a powerful and high-throughput method combining quantitative proteomics and peptide immunocapture to map SUMOylation sites and have analyzed changes in SUMOylation in response to stimuli. With this technique we identified 295 SUMO1 and 167 SUMO2 sites on endogenous substrates of human cells. We further used this strategy to characterize changes in SUMOylation induced by Listeriolysin O, a bacterial toxin that impairs the host cell SUMOylation machinery, and identified several classes of host proteins specifically deSUMOylated in response to this toxin. Our approach constitutes an unprecedented tool, broadly applicable to various SUMO-regulated cellular processes in health and disease.
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Listeriolysin O: the Swiss army knife of Listeria
Trends in Microbiology, 2012Co-Authors: Melanie Anne Hamon, David Ribet, Fabrizia Stavru, Pascale CossartAbstract:Listeriolysin O (LLO) is a toxin produced by Listeria monocytogenes, an opportunistic bacterial pathogen responsible for the disease listeriosis. This disease starts with the ingestion of contaminated foods and mainly affects immunocompromised individuals, newborns, and pregnant women. In the laboratory, L. monocytogenes is used as a model organism to study processes such as cell invasion, intracellular survival, and cell-to-cell spreading, as this Gram-positive bacterium has evolved elaborate molecular strategies to subvert host cell functions. LLO is a major virulence factor originally shown to be crucial for bacterial escape from the internalization vacuole after entry into cells. However, recent studies are revisiting the role of LLO during infection and are revealing new insights into the action of LLO, in particular before bacterial entry. These latest findings along with their impact on the infectious process will be discussed.
Daniel A Portnoy - One of the best experts on this subject based on the ideXlab platform.
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Listeriolysin o a phagosome specific cytolysin revisited
Cellular Microbiology, 2019Co-Authors: Brittney N Nguyen, Bret N Peterson, Daniel A PortnoyAbstract:Listeriolysin O (LLO) is an essential determinant of Listeria monocytogenes pathogenesis that mediates the escape of L. monocytogenes from host cell vacuoles, thereby allowing replication in the cytosol without causing appreciable cell death. As a member of the cholesterol-dependent cytolysin (CDC) family of pore-forming toxins, LLO is unique in that it is secreted by a facultative intracellular pathogen, whereas all other CDCs are produced by pathogens that are largely extracellular. Replacement of LLO with other CDCs results in strains that are extremely cytotoxic and 10,000-fold less virulent in mice. LLO has structural and regulatory features that allow it to function intracellularly without causing cell death, most of which map to a unique N-terminal region of LLO referred to as the proline, glutamic acid, serine, threonine (PEST)-like sequence. Yet, while LLO has unique properties required for its intracellular site of action, extracellular LLO, like other CDCs, affects cells in a myriad of ways. Because all CDCs form pores in cholesterol-containing membranes that lead to rapid Ca2+ influx and K+ efflux, they consequently trigger a wide range of host cell responses, including mitogen-activated protein kinase activation, histone modification, and caspase-1 activation. There is no debate that extracellular LLO, like all other CDCs, can stimulate multiple cellular activities, but the primary question we wish to address in this perspective is whether these activities contribute to L. monocytogenes pathogenesis.
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Listeriolysin o a phagosome specific lysin
Microbes and Infection, 2007Co-Authors: Pamela Schnupf, Daniel A PortnoyAbstract:Listeriolysin O (LLO) is a pore-forming toxin of the cholesterol-dependent cytolysin family and a primary virulence factor of the gram-positive, facultative intracellular pathogen Listeria monocytogenes. During the intracellular life cycle of L. monocytogenes, LLO is largely responsible for mediating rupture of the phagosomal membrane, thereby allowing the bacterium access to the host cytosol, its replicative niche. In the host cytosol, LLO activity is controlled at numerous levels to prevent perforation of the plasma membrane and loss of the intracellular environment. In this review, we focus primarily on the role of LLO in phagosomal escape and the multiple regulatory mechanisms that control LLO activity in the host cytosol.
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a specific gene expression program triggered by gram positive bacteria in the cytosol
Proceedings of the National Academy of Sciences of the United States of America, 2004Co-Authors: Ramona L Mccaffrey, Paul Fawcett, Mary X D Oriordan, Edward A Havell, Patrick O Brown, Daniel A PortnoyAbstract:Innate and adaptive immunity depends critically on host recognition of pathogen-associated molecules. Toll-like receptors (TLRs) are key mediators of pathogen surveillance at the cell or phagocytic vacuole surface. However, mechanisms underlying recognition of pathogens in other cellular compartments remain unclear, and responses elicited by cytosolic challenge are poorly characterized. We therefore used mouse cDNA microarrays to investigate gene expression triggered by infection of bone marrow-derived macrophages with cytosol- and vacuole-localized Listeria monocytogenes (Lm), a model cytosolic pathogen. The resulting gene expression program included two basic categories of induced genes: an “early/persistent” cluster consistent with NF-κB-dependent responses downstream of TLRs, and a subsequent “late response” cluster largely composed of IFN-responsive genes (IRGs). The early/persistent cluster was observed upon infection with WT, heat-killed, or mutant Lm lacking Listeriolysin O, the pore-forming hemolysin that promotes escape from phagocytic vacuoles. However, the IRG cluster depended on entry of WT Lm into the cytosol. Infection with Listeriolysin O-expressing, cytosolic Bacillus subtilis (Bs) strikingly recapitulated the expression profile associated with WT Lm, including IRG induction. IRG up-regulation was associated with MyD88-independent induction of IFN-β transcription and activity. Whereas Staphylococcus aureus (Sa) lipoteichoic acid treatment confirmed that many late-response genes could also be stimulated through TLRs, our study identified a cytosol-specific transcriptional program independent of TLR signaling through MyD88. Further characterization of cytosolic surveillance pathway(s) and their points of convergence with TLR- and IFN-dependent pathways will enhance our understanding of the means by which mammals detect and respond to pathogens.
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inducible control of virulence gene expression in listeria monocytogenes temporal requirement of Listeriolysin o during intracellular infection
Journal of Bacteriology, 2002Co-Authors: Christina E Dancz, Daniel A Portnoy, Andrea Haraga, Darren E HigginsAbstract:We have constructed a lac repressor/operator-based system to tightly regulate expression of bacterial genes during intracellular infection by Listeria monocytogenes. An L. monocytogenes strain was constructed in which expression of Listeriolysin O was placed under the inducible control of an isopropyl-β-d-thiogalactopyranoside (IPTG)-dependent promoter. Listeriolysin O (LLO) is a pore-forming cytolysin that mediates lysis of L. monocytogenes-containing phagosomes. Using hemolytic-activity assays and Western blot analysis, we demonstrated dose-dependent IPTG induction of LLO during growth in broth culture. Moreover, intracellular growth of the inducible-LLO (iLLO) strain in the macrophage-like cell line J774 was strictly dependent upon IPTG. We have further shown that iLLO bacteria trapped within primary phagocytic vacuoles can be induced to escape into the cytosol following addition of IPTG to the cell culture medium, thus yielding the ability to control bacterial escape from the phagosome and the initiation of intracellular growth. Using the iLLO strain in plaque-forming assays, we demonstrated an additional requirement for LLO in facilitating cell-to-cell spread in L2 fibroblasts, a nonprofessional phagocytic cell line. Furthermore, the efficiency of cell-to-cell spread of iLLO bacteria in L2 cells was IPTG dose dependent. The potential use of this system for determining the temporal requirements of additional virulence determinants of intracellular pathogenesis is discussed.
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the cell biology of listeria monocytogenes infection the intersection of bacterial pathogenesis and cell mediated immunity
Journal of Cell Biology, 2002Co-Authors: Daniel A Portnoy, Victoria Auerbuch, Ian J GlomskiAbstract:Listeria monocytogenes has emerged as a remarkably tractable pathogen to dissect basic aspects of cell biology, intracellular pathogenesis, and innate and acquired immunity. In order to maintain its intracellular lifestyle, L. monocytogenes has evolved a number of mechanisms to exploit host processes to grow and spread cell to cell without damaging the host cell. The pore-forming protein Listeriolysin O mediates escape from host vacuoles and utilizes multiple fail-safe mechanisms to avoid causing toxicity to infected cells. Once in the cytosol, the L. monocytogenes ActA protein recruits host cell Arp2/3 complexes and enabled/vasodilator-stimulated phosphoprotein family members to mediate efficient actin-based motility, thereby propelling the bacteria into neighboring cells. Alteration in any of these processes dramatically reduces the ability of the bacteria to establish a productive infection in vivo.
Trinad Chakraborty - One of the best experts on this subject based on the ideXlab platform.
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Missing elimination via membrane vesicle shedding contributes to the diminished calcium sensitivity of Listeriolysin O
Nature Publishing Group, 2018Co-Authors: Jana Maurer, Timothy J Mitchell, Sabrina Hupp, Helena Pillich, Trinad Chakraborty, Asparouh I IlievAbstract:Abstract The lytic capacity of cholesterol-dependent cytolysins is enhanced in the extracellular calcium-free environment through a combination of limited membrane repair and diminished membrane toxin removal. For a typical neurotoxin of the group, pneumolysin, this effect has already been observed at reduced (1 mM) calcium conditions, which are pathophysiologically relevant. Here, we tested another neurotoxin of the group, Listeriolysin O from L. monocytogenes, active in the primary vacuole after bacterium phagocytosis in host cells. Reduced calcium did not increase the lytic capacity of Listeriolysin (in contrast to pneumolysin), while calcium-free conditions elevated it 2.5 times compared to 10 times for pneumolysin (at equivalent hemolytic capacities). To clarify these differences, we analyzed membrane vesicle shedding, known to be a calcium-dependent process for toxin removal from eukaryotic cell membranes. Both pneumolysin and Listeriolysin initiated vesicle shedding, which was completely blocked by the lack of extracellular calcium. Lack of calcium, however, elevated the toxin load per a cell only for pneumolysin and not for Listeriolysin. This result indicates that vesicle shedding does not play a role in the membrane removal of Listeriolysin and outlines a major difference between it and other members of the CDC group. Furthermore, it provides new tools for studying membrane vesicle shedding
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sterol and ph interdependence in the binding oligomerization and pore formation of Listeriolysin o
Biochemistry, 2007Co-Authors: Andrej Bavdek, Trinad Chakraborty, Nelson O Gekara, Siegfried Weiss, Ayub Darji, Dragan Priselac, Ion Gutierrez Aguirre, Peter Macek, Jeremy H Lakey, Gregor AnderluhAbstract:Listeriolysin O (LLO) is the most important virulence factor of the intracellular pathogen Listeria monocytogenes. Its main task is to enable escape of bacteria from the phagosomal vacuole into the ...
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induction of suicidal erythrocyte death by Listeriolysin from listeria monocytogenes
Cellular Physiology and Biochemistry, 2007Co-Authors: Michael Foller, Trinad Chakraborty, Ekaterina Shumilina, Rebecca S Lam, Walid Mohamed, Ravi S Kasinathan, Stephan M Huber, Florian LangAbstract:Listeriolysin, the secreted cytolysin of the facultative intracellular bacterium Listeria monocytogenes, is its major virulence factor. Previously, non-lytic concentrations of Listeriolysin were shown to induce Ca2+-permeable nonselective cation channels in human embryonic kidney cells. In erythrocytes, Ca2+ entry is followed by activation of K+ channels resulting in K+-exit as well as by membrane scrambling resulting in phosphatidylserine exposure at the cell surface. Phosphatidylserine-exposing erythrocytes are recognized by macrophages, engulfed, degraded and thus cleared from circulating blood. Phosphatidylserine exposure is a key event of eryptosis, the suicidal death of erythrocytes. The present study utilized patch-clamp technique, Fluo3-fluorescence, and annexin V-binding in FACS analysis to determine the effect of Listeriolysin on cell membrane conductance, cytosolic free Ca2+ concentration, and phosphatidylserine exposure, respectively. Within 30 minutes, exposure of human peripheral blood erythrocytes to low concentrations of Listeriolysin (which were non-hemolytic for the majority of cells) induced a Ca2+-permeable cation conductance in the erythrocyte cell membrane, increased cytosolic Ca2+ concentration, and triggered annexin V-binding. Increase of extracellular K+ concentration blunted, but did not prevent, Listeriolysin-induced annexin V-binding. In conclusion, Listeriolysin triggers suicidal death of erythrocytes, an effect at least partially due to depletion of intracellular K+. Listeriolysin induced suicidal erythrocyte death could well contribute to the pathophysiology of L. monocytogenes infection.
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the cholesterol dependent cytolysin Listeriolysin o aggregates rafts via oligomerization
Cellular Microbiology, 2005Co-Authors: Nelson O Gekara, Trinad Chakraborty, Thomas Jacobs, Siegfried WeissAbstract:The pore-forming toxin Listeriolysin O (LLO) is the main virulence factor of Listeria monocytogenes. LLO is known to act as a pseudo cytokine/chemokine, which induces a broad spectrum of host responses that ultimately influences the outcome of listeriosis. In the present study we demonstrate that LLO is a potent aggregator of lipid rafts. LLO was found to aggregate the raft associated molecules GM1, the GPI-anchored proteins CD14 and CD16 as well as the tyrosine kinase Lyn. Abrogation of the cytolytic activity of LLO by cholesterol pretreatment was found not to interfere with LLO's ability to aggregate rafts or trigger tyrosine phosphorylation in cells. However, a monoclonal antibody that blocks the oligomerization of LLO was found to inhibit rafts' aggregation as well as the induction of tyrosine phosphorylation. This implies that rafts aggregation by LLO which is independent of cytolytic activity, is due to the oligomerization of its membrane bound toxin monomers. Thus, LLO most likely induces signalling through the coaggregation of rafts' associated receptors, kinases and adaptors.
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role of listeria monocytogenes exotoxins Listeriolysin and phosphatidylinositol specific phospholipase c in activation of human neutrophils
Infection and Immunity, 1999Co-Authors: Ulf Sibelius, Trinad Chakraborty, Thomas Jacobs, Siegfried Weiss, Evacathrin Schulz, Frank Rose, Katja Hattar, Werner Seeger, Friedrich GrimmingerAbstract:Polymorphonuclear leukocytes (PMN) are essential for resolution of infections with Listeria monocytogenes. The present study investigated the role of the listerial exotoxins Listeriolysin (LLO) and phosphatidylinositol-specific phospholipase C (PlcA) in human neutrophil activation. Different Listeria strains, mutated in individual virulence genes, as well as purified LLO were used. Coincubation of human neutrophils with wild-type L. monocytogenes provoked PMN activation, occurring independently of phagocytosis events, with concomitant elastase secretion, leukotriene generation, platelet-activating factor (PAF) synthesis, respiratory burst, and enhanced phosphoinositide hydrolysis. Degranulation and leukotriene formation were noted to be solely dependent on LLO expression, as these features were absent when the LLO-defective mutant EGD- and the avirulent strain L. innocua were used. These effects were fully reproduced by a recombinant L. innocua strain expressing LLO (INN+) and by the purified LLO molecule. LLO secretion was also required for PAF synthesis. However, wild-type L. monocytogenes was more potent in eliciting PAF formation than mutants expressing LLO, suggesting the involvement of additional virulence factors. This was even more obvious for phosphoinositide hydrolysis and respiratory burst: these events were provoked not only by INN+ but also by the LLO-defective mutant EGD- and by a recombinant L. innocua strain producing listerial PlcA. We conclude that human neutrophils react to extracellularly provided listerial exotoxins by rapid cell activation. Listeriolysin is centrally involved in triggering degranulation and lipid mediator generation, and further virulence factors such as PlcA apparently contribute to trigger neutrophil phosphoinositide hydrolysis and respiratory burst. In this way, listerial exotoxins may influence the host defense against infections with L. monocytogenes.
Stefan H E Kaufmann - One of the best experts on this subject based on the ideXlab platform.
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from evil to good a cytolysin in vaccine development
Trends in Microbiology, 2001Co-Authors: Guido Dietrich, Jurgen Hess, Stefan H E Kaufmann, Ivaylo Gentschev, Bernhard Knapp, Werner GoebelAbstract:Current vaccination strategies mainly target antigens into the phagosomal, major histocompatibility complex class II antigen-processing pathway and thus lead predominantly to humoral immune responses. The elicitation of cytotoxic T-cell responses instead requires introduction of antigens into the cytosol of professional antigen-presenting cells (APCs). The intracellular bacterium Listeria monocytogenes gains access to the host cell cytosol by means of a cytolysin, Listeriolysin O. Vaccine researchers have successfully employed Listeriolysin in novel vaccination approaches to provide access to the cytosol of professional APCs for purified protein antigens, attenuated bacterial vaccine strains, DNA vaccines and liposome contents.
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effective dna vaccination against listeriosis by prime boost inoculation with the gene gun
Journal of Immunology, 1999Co-Authors: Joachim Fensterle, Leander Grode, Jurgen Hess, Stefan H E KaufmannAbstract:Protective immunity against Listeria monocytogenes strongly depends on CD8+ T lymphocytes, and both IFN-γ secretion and target cell killing are considered relevant to protection. We analyzed whether we could induce a protective type 1 immune response by DNA vaccination with the gene gun using plasmids encoding for two immunodominant listerial Ags, Listeriolysin and p60. To induce a Th1 response, we 1) coprecipitated a plasmid encoding for GM-CSF, 2) employed a prime/boost vaccination schedule with a 45-day interval, and 3) coinjected oligodeoxynucleotides (ODN) containing immunostimulatory CpG motifs. DNA immunization of BALB/c mice with plasmids encoding for Listeriolysin (pChly) and p60 (pCiap) efficiently induced MHC class I-restricted, Ag-specific CD8+ T cells that produced IFN-γ. Coinjection of CpG-ODN significantly increased the frequency of specific IFN-γ-secreting T cells. Although pChly induced specific CD8+ T cells expressing CTL activity, it failed to stimulate CD4+ T cells. Only pCiap induced significant CD4+ T cell and humoral responses, which were predominantly of Th2 type. Vaccination with either plasmid induced protective immunity against listerial challenge, and coinjection of CpG ODN improved vaccine efficacy in some situations. This study demonstrates the feasibility of gene gun administration of plasmid DNA for inducing immunity against an intracellular pathogen for which protection primarily depends on type 1 CD8+ T cells.
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mycobacterium bovis bacille calmette guerin strains secreting Listeriolysin of listeria monocytogenes
Proceedings of the National Academy of Sciences of the United States of America, 1998Co-Authors: Jurgen Hess, Diana Miko, Andre Catic, Vera Lehmensiek, David G Russell, Stefan H E KaufmannAbstract:Recombinant (r) Mycobacterium bovis strains were constructed that secrete biologically active Listeriolysin (Hly) fusion protein of Listeria monocytogenes. The r-BCG strains pAT261:Hly or pMV306:Hly expressed plasmid multicopies or chromosomal single copies of the hly gene, respectively. Human and murine macrophage-like cell lines were infected with r-BCG pAT261:Hly and pMV306:Hly strains. Interestingly, intracellular persistence of both r-BCG strains was reduced in macrophages as compared with the parental BCG strain. By immunogold labeling Hly was detected in membrane structures and within the phagosomal space of macrophages. In addition, Hly was localized within cytoplasmic vacuoles outside the mycobacteria-containing phagosome of host cells infected with r-BCG pAT261:Hly or r-BCG pMV306:Hly. Hly fusions consistently colocalized with a lysosome-associated membrane glycoprotein, suggesting that membrane-attack conformation of Hly was not altered. Although r-BCG pAT261:Hly and r-BCG pMV306:Hly microorganims apparently did not egress into the cytoplasmic compartment of host cells, they both improved major histocompatibility complex class I presentation of cophagocytosed soluble protein as compared with wild-type BCG microbes. These data suggest that Hly secretion endows BCG with an improved capacity to stimulate CD8 T cells. Because CD8 T cells play a major role in protection against tuberculosis such Hly secreting r-BCG constructs are antituberculosis vaccine candidates.
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salmonella strain secreting active Listeriolysin changes its intracellular localization
Infection and Immunity, 1995Co-Authors: Ivaylo Gentschev, Stefan H E Kaufmann, Zeljka Sokolovic, Hansjoachim Mollenkopf, J Hess, Michael Kuhn, G F Krohne, Werner GoebelAbstract:We describe the construction of an attenuated Salmonella dublin aroA strain which secretes via the Escherichia coli hemolysin secretion machinery an active hybrid cytolysin consisting of Listeriolysin from Listeria monocytogenes and the C-terminal secretion signal of E. coli hemolysin. This hemolytic S. dublin strain is partially released into the cytoplasm of the host cell following uptake by J774 macrophage cells, whereas the nonhemolytic control S. dublin aroA strain remains in the phagosome.
Jurgen Hess - One of the best experts on this subject based on the ideXlab platform.
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increased vaccine efficacy against tuberculosis of recombinant mycobacterium bovis bacille calmette guerin mutants that secrete Listeriolysin
Journal of Clinical Investigation, 2005Co-Authors: Leander Grode, Peter Seiler, Sven Baumann, Jurgen Hess, Volker Brinkmann, Ali Nasser Eddine, Peggy Mann, Christian Goosmann, Silke Bandermann, Debbie A SmithAbstract:The tuberculosis vaccine Mycobacterium bovis bacille Calmette-Guerin (BCG) was equipped with the membrane-perforating Listeriolysin (Hly) of Listeria monocytogenes, which was shown to improve protection against Mycobacterium tuberculosis. Following aerosol challenge, the Hly-secreting recombinant BCG (hly+ rBCG) vaccine was shown to protect significantly better against aerosol infection with M. tuberculosis than did the parental BCG strain. The isogenic, urease C–deficient hly+ rBCG (ΔureC hly+ rBCG) vaccine, providing an intraphagosomal pH closer to the acidic pH optimum for Hly activity, exhibited still higher vaccine efficacy than parental BCG. ΔureC hly+ rBCG also induced profound protection against a member of the M. tuberculosis Beijing/W genotype family while parental BCG failed to do so consistently. Hly not only promoted antigen translocation into the cytoplasm but also apoptosis of infected macrophages. We concluded that superior vaccine efficacy of ΔureC hly+ rBCG as compared with parental BCG is primarily based on improved cross-priming, which causes enhanced T cell–mediated immunity.
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from evil to good a cytolysin in vaccine development
Trends in Microbiology, 2001Co-Authors: Guido Dietrich, Jurgen Hess, Stefan H E Kaufmann, Ivaylo Gentschev, Bernhard Knapp, Werner GoebelAbstract:Current vaccination strategies mainly target antigens into the phagosomal, major histocompatibility complex class II antigen-processing pathway and thus lead predominantly to humoral immune responses. The elicitation of cytotoxic T-cell responses instead requires introduction of antigens into the cytosol of professional antigen-presenting cells (APCs). The intracellular bacterium Listeria monocytogenes gains access to the host cell cytosol by means of a cytolysin, Listeriolysin O. Vaccine researchers have successfully employed Listeriolysin in novel vaccination approaches to provide access to the cytosol of professional APCs for purified protein antigens, attenuated bacterial vaccine strains, DNA vaccines and liposome contents.
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effective dna vaccination against listeriosis by prime boost inoculation with the gene gun
Journal of Immunology, 1999Co-Authors: Joachim Fensterle, Leander Grode, Jurgen Hess, Stefan H E KaufmannAbstract:Protective immunity against Listeria monocytogenes strongly depends on CD8+ T lymphocytes, and both IFN-γ secretion and target cell killing are considered relevant to protection. We analyzed whether we could induce a protective type 1 immune response by DNA vaccination with the gene gun using plasmids encoding for two immunodominant listerial Ags, Listeriolysin and p60. To induce a Th1 response, we 1) coprecipitated a plasmid encoding for GM-CSF, 2) employed a prime/boost vaccination schedule with a 45-day interval, and 3) coinjected oligodeoxynucleotides (ODN) containing immunostimulatory CpG motifs. DNA immunization of BALB/c mice with plasmids encoding for Listeriolysin (pChly) and p60 (pCiap) efficiently induced MHC class I-restricted, Ag-specific CD8+ T cells that produced IFN-γ. Coinjection of CpG-ODN significantly increased the frequency of specific IFN-γ-secreting T cells. Although pChly induced specific CD8+ T cells expressing CTL activity, it failed to stimulate CD4+ T cells. Only pCiap induced significant CD4+ T cell and humoral responses, which were predominantly of Th2 type. Vaccination with either plasmid induced protective immunity against listerial challenge, and coinjection of CpG ODN improved vaccine efficacy in some situations. This study demonstrates the feasibility of gene gun administration of plasmid DNA for inducing immunity against an intracellular pathogen for which protection primarily depends on type 1 CD8+ T cells.
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mycobacterium bovis bacille calmette guerin strains secreting Listeriolysin of listeria monocytogenes
Proceedings of the National Academy of Sciences of the United States of America, 1998Co-Authors: Jurgen Hess, Diana Miko, Andre Catic, Vera Lehmensiek, David G Russell, Stefan H E KaufmannAbstract:Recombinant (r) Mycobacterium bovis strains were constructed that secrete biologically active Listeriolysin (Hly) fusion protein of Listeria monocytogenes. The r-BCG strains pAT261:Hly or pMV306:Hly expressed plasmid multicopies or chromosomal single copies of the hly gene, respectively. Human and murine macrophage-like cell lines were infected with r-BCG pAT261:Hly and pMV306:Hly strains. Interestingly, intracellular persistence of both r-BCG strains was reduced in macrophages as compared with the parental BCG strain. By immunogold labeling Hly was detected in membrane structures and within the phagosomal space of macrophages. In addition, Hly was localized within cytoplasmic vacuoles outside the mycobacteria-containing phagosome of host cells infected with r-BCG pAT261:Hly or r-BCG pMV306:Hly. Hly fusions consistently colocalized with a lysosome-associated membrane glycoprotein, suggesting that membrane-attack conformation of Hly was not altered. Although r-BCG pAT261:Hly and r-BCG pMV306:Hly microorganims apparently did not egress into the cytoplasmic compartment of host cells, they both improved major histocompatibility complex class I presentation of cophagocytosed soluble protein as compared with wild-type BCG microbes. These data suggest that Hly secretion endows BCG with an improved capacity to stimulate CD8 T cells. Because CD8 T cells play a major role in protection against tuberculosis such Hly secreting r-BCG constructs are antituberculosis vaccine candidates.