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Ingo B Autenrieth - One of the best experts on this subject based on the ideXlab platform.
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bacterial adhesion and host cell factors leading to effector protein injection by type iii secretion system
International Journal of Medical Microbiology, 2019Co-Authors: Erwin Bohn, Michael Sonnabend, Kristina Klein, Ingo B AutenriethAbstract:Abstract Type III secretion systems (T3SS) play a crucial role for virulence in many Gram-negative bacteria. After tight bacterial contact to host cells, the T3SS injects effector proteins into the host cells, which leads to cell invasion, tissue destruction and/or immune evasion. Over the last decade several attempts were made to characterize the host-cell interactions which precede and determine effector protein injection during Infection. The development of the TEM-β-lactamase reporter was an important breakthrough to achieve this goal. By this means it was demonstrated that during Infection with many Gram-negative pathogens such as Salmonella, Pseudomonas or Yersinia the main targets of T3SS are leukocytes of the myeloid lineage such as neutrophils, macrophages or dendritic cells. This is due to the recruitment of these cells to the site of Infection, but also due to the specific interplay between bacterial and host cells. Comprehensive studies on Yersinia pestis, Yersinia enterocolitica and Yersinia pseudotuberculosis effector translocation show that adhesins such as Invasin (Inv), Yersinia adhesin A (YadA) and attachment and invasion locus (Ail) are critical for effector translocation. Here, mainly the complex interaction of YadA and Ail with various host cell receptor repertoires on leukocytes and the modulatory effects of serum factors direct effector translocation predominantly towards myeloid cells. The current understanding suggests that mostly protein based interactions between bacteria and host determine host cell specific effector translocation during Yersinia Infection. However, for Shigella dysenteriae Infection it was shown that glycan-glycan interactions can also play a critical role for the adhesion preceding effector translocation. In addition, the Shigella Infection model revealed that the activation status of cells is a further criterium directing effector translocation into a distinct cell population. In this review the current understanding of the complex and species-specific interaction between bacteria and host cells leading to type III secretion is discussed.
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Yersinia enterocolitica invasin protein triggers differential production of interleukin 1 interleukin 8 monocyte chemoattractant protein 1 granulocyte macrophage colony stimulating factor and tumor necrosis factor alpha in epithelial cells implications for understanding the early cytokine network in Yersinia Infections
Infection and Immunity, 2000Co-Authors: Daniel Kampik, Ralf Schulte, Ingo B AutenriethAbstract:Yersinia enterocolitica Infection of epithelial cells results in interleukin-8 (IL-8) mRNA expression. Herein we demonstrate that besides IL-8, increased mRNA levels of five other cytokines, IL-1α, IL-1β, monocyte chemoattractant protein 1 (MCP-1), granulocyte-macrophage colony-stimulating factor (GM-CSF), and tumor necrosis factor alpha (TNF-α), can be detected upon Infection of HeLa cells with Yersinia. Yersinia-triggered cytokine production was not affected by blocking phosphatidylinositol-3-phosphate kinase with wortmannin, which inhibited bacterial invasion. Comparable cytokine mRNA responses were triggered by Escherichia coli expressing Yersinia inv, while no response was triggered by an inv-deficient Yersinia mutant. Moreover, cytokine responses were independent from metabolic activity of the bacteria, as killed bacterial cells were sufficient for triggering cytokine responses in HeLa cells. Semiquantitative reverse transcription-PCR analysis was used to assess the kinetics of cytokine mRNA expression in infected HeLa cells. IL-8, IL-1α, IL-1β, MCP-1, GM-CSF, and TNF-α mRNA expression increased within 1 h postInfection, reached a maximum after 3 to 4 h, and then declined to preInfection levels within 3 h. IL-8, MCP-1, and GM-CSF were secreted by HeLa cells, whereas IL-1α and IL-1β were not secreted and thus were found exclusively intracellularly. TNF-α protein could not be detected in cell lysates or supernatants. Stimulation of HeLa cells with IL-1α was followed by increased IL-8 mRNA expression, whereas stimulation with IL-8 did not induce cytokine production. Likewise, MCP-1 and GM-CSF did not induce significant cytokine responses in HeLa cells. Our results implicate that the initial host response to Yersinia Infection might be sustained by IL-8, MCP-1, and GM-CSF produced by epithelial cells.
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interferon consensus sequence binding protein confers resistance against Yersinia enterocolitica
Infection and Immunity, 2000Co-Authors: Joachim Hein, Andreas Sing, Volkhard A J Kempf, Sonja Preger, Joachim Diebold, Nicole Bucheler, Ivan D Horak, Uwe Kramer, Ingo B AutenriethAbstract:Interferon (IFN) consensus sequence binding protein (ICSBP) (19) belongs to the IFN regulatory factor (IRF) family of mammalian transcription factors (see for a review reference 47). Proteins of the IRF family bind to the IFN-stimulated response element (ISRE) and control transcription of genes with ISREs within their promoter regions (57). The IRF family plays an important role in the regulation of both type I (IFN-α/β) and type II (IFN-γ) IFN-inducible genes. ICSBP is exclusively expressed in hematopoietically derived cells and predominantly induced by IFN-γ (25, 55). Analyses of recently generated ICSBP knockout (ICSBP−/−) mice have permitted insights into the in vivo role of ICSBP (34). These mice exhibit a chronic myelogenous leukemia (CML)-like syndrome and display enhanced susceptibility to a variety of intracellular pathogens including Listeria monocytogenes, Leishmania major, and Toxoplasma gondii (22, 28, 34, 64). ICSBP−/− mice fail to develop T helper 1 (Th1)-driven immune responses due to a primary defect in interleukin-12 (IL-12) p40 induction and, as a consequence, IFN-γ-dependent host resistance (28, 34, 64). Furthermore, ICSBP−/− mice show reduced and delayed oxidative burst, whereas nitric oxide (NO) production is normal (22). Th2 immune responses, however, are not affected in these mice. In addition, ICSBP modulates survival of myeloid cells by regulating expression of apoptosis-related genes (26). Yersinia enterocolitica is enteropathogenic for humans and rodents. The bacteria cross the intestinal epithelial barrier by translocating through M cells, spread into the lamina propria, and colonize preferentially the underlying Peyer's patches (2, 4, 14, 29, 30). Virulence plasmid (pYV)-harboring strains are able to migrate from the Peyer's patches to the mesenteric lymph nodes and deeper organs such as the spleen, liver, and lungs, where they multiply extracellularly and lead to the formation of multiple necrotic abscesses (2, 4, 30, 69). In contrast, nonvirulent strains lacking the pYV (pCD1 in Y. pestis) plasmid are contained within granulomas, resulting in a lower rate of Infection before rapid clearance of the bacteria (45, 71, 74). Successful control and elimination of Y. enterocolitica depends on both innate and adaptive immunity. Neutrophils and macrophages are involved in partial restriction of bacterial replication in the early phase of primary Infection in mice (4, 15, 31, 62). Furthermore, despite Y. enterocolitica being an extracellular pathogen, it is well established that T-cell-mediated and IFN-γ-dependent immune mechanisms are essential for resistance (1, 5). Consequently, adoptive transfer of Yersinia-specific CD4+ Th1 cell clones into athymic T-cell-deficient nude mice confers resistance against this pathogen (6). Previous studies have shown that C57BL/6 mice, which produce high levels of IFN-γ, are resistant to Y. enterocolitica, whereas BALB/c mice, which secrete only small quantities of IFN-γ, are susceptible to Yersinia Infection (1). Furthermore, neutralization or genetic deletion of the cytokine tumor necrosis factor alpha (TNF-α), IFN-γ, IL-12, or IL-18 abrogates resistance to Yersinia Infection (3, 7, 10). Based on these results, it is conceivable that antigen-presenting cells such as dendritic cells and macrophages become activated during contact with microbes and start producing IL-12 and IL-18. These cytokines strongly induce the expression of IFN-γ in natural killer (NK) cells and CD4+ Th1 cells. Most likely, IFN-γ produced by these cells synergizes with macrophage-produced TNF-α to activate microbicidal mechanisms such as reactive oxygen intermediates and reactive nitrogen intermediates in macrophages. Although ICSBP has been shown to be essential for immunity to intracellular pathogens, nothing is known about its requirement for immunity to extracellular pathogens, in particular Y. enterocolitica. The aim of this study was to investigate (i) whether ICSBP−/− mice exhibit an altered susceptibility to Yersinia, (ii) which defense mechanisms against Yersinia depend on the coordinate expression of the transcription factor ICSBP, and (iii) whether administration of recombinant cytokines restores impaired immunity in ICSBP−/− mice. The experiments described herein argue for an essential role of ICSBP in resistance against Y. enterocolitica.
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il 18 ifn gamma inducing factor regulates early cytokine production in and promotes resolution of bacterial Infection in mice
Journal of Immunology, 1998Co-Authors: Erwin Bohn, Jurgen Heesemann, Andreas Sing, Robert Zumbihl, Claudia Bielfeldt, Haruki Okamura, Masashi Kurimoto, Ingo B AutenriethAbstract:IL-12-induced IFN-γ production is essential for clearance of Yersinia enterocolitica Infection. Similar to IL-12, the recently described cytokine IL-18 (IFN-γ-inducing factor) is produced by macrophages and induces IFN-γ production in spleen cells. Therefore, we have investigated the role of IL-18 in Yersinia Infection of mice. Heat-killed Yersinia-triggered IL-18-promoted IFN-γ production of splenocytes was predominantly dependent on endogenous IL-12 production, whereas IL-12-promoted IFN-γ production was not IL-18 dependent. IL-18-induced IFN-γ production was to a higher degree dependent on IFN-γR-mediated mechanisms and in synergism with IL-2 resulted in at least fivefold higher IFN-γ levels as compared with the combination of IL-12 plus IL-2. Analysis of the effect of IL-18 on IL-12 production of LPS-stimulated peritoneal macrophages revealed that IL-18 decreased LPS-induced IL-12 production, indicating that IL-18 might be involved in negative regulation of IL-12 production. In vivo studies revealed that Yersinia -resistant C57BL/6 mice expressed fourfold higher IL-18 mRNA levels than did susceptible BALB/c mice. Administration of anti-IL-18 Abs caused a 100- to 1000-fold increase in bacterial counts in the spleen of infected mice but did not change IFN-γ production levels. Taken together, our data demonstrate that IL-18 is involved in regulation of cytokine production during the early phase of bacterial Infections as well as in clearance of Yersinia Infection.
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defense mechanisms in peyer s patches and mesenteric lymph nodes against Yersinia enterocolitica involve integrins and cytokines
Infection and Immunity, 1996Co-Authors: Ingo B Autenrieth, Volkhard A J Kempf, Thomas Sprinz, Sonja Preger, Andastrid SchnellAbstract:Adhesion molecules and cytokines are involved in regulation of cellular host responses in Infection processes. In this study the roles of the integrins Mac-1 and VLA-4, as well as those of the cytokines tumor necrosis factor alpha (TNF-alpha) and gamma interferon (IFN-gamma), in defense mechanisms against Yersinia enterocolitica in Peyer9s patches (PP) and mesenteric lymph nodes (MLN) were investigated by blocking these molecules with antibodies in vivo prior to orogastric Yersinia Infection. Intestinal Yersinia Infection caused abscesses composed of polymorphonuclear (Mac-1+ VLA-4+ Pgp-1+ ICAM-1-) and mononuclear (Mac-1+ VLA-4+ Pgp-1+ ICAM-inhibited phagocytosis of Yersiniae by macrophages, (ii) reduced Yersinia-specific proliferation and IFN-gamma production of T cells from PP and MLN, and (iii) caused increased bacterial growth in PP and MLN followed by profound tissue destruction. Neutralization of TNF-alpha or IFN-gamma had comparable effects, suggesting that cell-mediated host responses including activated macrophages are required for control of Yersiniae in intestinal tissues. The number of Mac-1+ cells in PP and MLN increased after Yersinia Infection, and recruitment of these cells was not blocked by administration of anticytokine or anti-integrin antibodies. While anti-VLA-4, -TNF-alpha, or -IFN-gamma antibody treatment caused an increased dissemination of Yersiniae from PP to the spleen systemic dissemination was reduced by anti-Mac-1 antibodies. The results of this study suggest that the cytokines IFN-gamma and TNF-alpha as well as the integrins Mac-1 and VLA-4 are involved in protective cellular host defense mechanisms in PP and MLN against Y. enterocolitica, the latter probably being involved in both cell-cell and cell-pathogen interactions.
James B Bliska - One of the best experts on this subject based on the ideXlab platform.
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RESEARCH ARTICLE RandomMutagenesis Identifies a C-Terminal Region of YopD Important for Yersinia Type III Secretion Function
2016Co-Authors: Rebecca Solomon, James B Bliska, Weibing Zhang, Grace Mccrann, I. ViboudAbstract:A common virulence mechanism among bacterial pathogens is the use of specialized se-cretion systems that deliver virulence proteins through a translocation channel inserted in the host cell membrane. During Yersinia Infection, the host recognizes the type III secretion system mounting a pro-inflammatory response. However, soon after they are translocated, the effectors efficiently counteract that response. In this study we sought to identify YopD residues responsible for type III secretion system function. Through random mutagenesis, we identified eight Y. pseudotuberculosis yopD mutants with single amino acid changes af-fecting various type III secretion functions. Three severely defective mutants had substitu-tions in residues encompassing a 35 amino acid region (residues 168–203) located between the transmembrane domain and the C-terminal putative coiled-coil region of YopD. These mutations did not affect regulation of the low calcium response or YopB-YopD inter-action but markedly inhibited MAPK and NFκB activation. When some of these mutations were introduced into the native yopD gene, defects in effector translocation and pore forma-tion were also observed. We conclude that this newly identified region is important for YopD translocon function. The role of this domain in vivo remains elusive, as amino acid substitu-tions in that region did not significantly affect virulence of Y. pseudotuberculosis in orogastri-cally-infected mice
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iqgap1 is important for activation of caspase 1 in macrophages and is targeted by Yersinia pestis type iii effector yopm
Mbio, 2014Co-Authors: Lawton K Chung, Naomi H Philip, Igor E Brodsky, Valentina A Schmidt, Antonius Koller, Till Strowig, Richard A Flavell, James B BliskaAbstract:ABSTRACT YopM is a leucine-rich repeat (LRR)-containing effector in several Yersinia species, including Yersinia pestis and Y. pseudotuberculosis. Different Yersinia strains encode distinct YopM isoforms with variable numbers of LRRs but conserved C-terminal tails. A 15-LRR isoform in Y. pseudotuberculosis YPIII was recently shown to bind and inhibit caspase-1 via a YLTD motif in LRR 10, and attenuation of YopM − YPIII was reversed in mice lacking caspase-1, indicating that caspase-1 inhibition is a major virulence function of YopM YPIII . To determine if other YopM proteins inhibit caspase-1, we utilized Y. pseudotuberculosis strains natively expressing a 21-LRR isoform lacking the YLTD motif (YopM 32777 ) or ectopically expressing a Y. pestis 15-LRR version with a functional (YopM KIM ) or inactivated (YopM KIM D 271 A) YLTD motif. Results of mouse and macrophage Infections with these strains showed that YopM 32777 , YopM KIM , and YopM KIM D 271 A inhibit caspase-1 activation, indicating that the YLTD motif is dispensable for this activity. Analysis of YopM KIM deletion variants revealed that LRRs 6 to 15 and the C-terminal tail are required to inhibit caspase-1 activation. YopM 32777 , YopM KIM , and YopM KIM deletion variants were purified, and binding partners in macrophage lysates were identified. Caspase-1 bound to YopM KIM but not YopM 32777 . Additionally, YopM KIM bound IQGAP1 and the use of Iqgap1 −/− macrophages revealed that this scaffolding protein is important for caspase-1 activation upon Infection with YopM − Y. pseudotuberculosis. Thus, while multiple YopM isoforms inhibit caspase-1 activation, their variable LRR domains bind different host proteins to perform this function and the LRRs of YopM KIM target IQGAP1, a novel regulator of caspase-1, in macrophages. IMPORTANCE Activation of caspase-1, mediated by macromolecular complexes termed inflammasomes, is important for innate immune defense against pathogens. Pathogens can, in turn, subvert caspase-1-dependent responses through the action of effector proteins. For example, the Yersinia effector YopM inhibits caspase-1 activation by arresting inflammasome formation. This caspase-1 inhibitory activity has been studied in a specific YopM isoform, and in this case, the protein was shown to act as a pseudosubstrate to bind and inhibit caspase-1. Different Yersinia strains encode distinct YopM isoforms, many of which lack the pseudosubstrate motif. We studied additional isoforms and found that these YopM proteins inhibit caspase-1 activation independently of a pseudosubstrate motif. We also identified IQGAP1 as a novel binding partner of the Yersinia pestis YopM KIM isoform and demonstrated that IQGAP1 is important for caspase-1 activation in macrophages infected with Yersinia . Thus, this study reveals new insights into inflammasome regulation during Yersinia Infection.
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yopj promoted cytotoxicity and systemic colonization are associated with high levels of murine interleukin 18 gamma interferon and neutrophils in a live vaccine model of Yersinia pseudotuberculosis Infection
Infection and Immunity, 2010Co-Authors: Yue Zhang, James B BliskaAbstract:Several Yersinia species have been utilized as live attenuated vaccines to prime protective immunity against Yersiniae and other pathogens. A type III secretion system effector known as YopJ in Y. pseudotuberculosis and Y. pestis and YopP in Y. enterocolitica has been shown to regulate host immune responses to live Yersinia vaccines. YopJ/P kills macrophages and dendritic cells, reduces their production of tumor necrosis factor alpha (TNF-α) and interleukin-12 (IL-12), and promotes systemic colonization in mouse models of intestinal Yersinia Infection. Furthermore, YopP activity decreases antigen presentation by dendritic cells, and a yopP mutant of a live Y. enterocolitica carrier vaccine elicited effective priming of CD8 T cells to a heterologous antigen in mice. These results suggest that YopJ/P activity suppresses both innate and adaptive immune responses to live Yersinia vaccines. Here, a sublethal intragastric mouse Infection model using wild-type and catalytically inactive yopJ mutant strains of Y. pseudotuberculosis was developed to further investigate how YopJ action impacts innate and adaptive immune responses to a live vaccine. Surprisingly, YopJ-promoted cytotoxicity and systemic colonization were associated with significant increases in neutrophils in spleens and the proinflammatory cytokines IL-18 and gamma interferon (IFN-γ) in serum samples of mice vaccinated with Y. pseudotuberculosis. Secretion of IL-18 accompanied YopJ-mediated killing of macrophages infected ex vivo with Y. pseudotuberculosis, suggesting a mechanism by which this effector directly increases proinflammatory cytokine levels in vivo. Mice vaccinated with the wild-type strain or the yopJ mutant produced similar levels of antibodies to Y. pseudotuberculosis antigens and were equally resistant to lethal intravenous challenge with Y. pestis. The findings indicate that a proinflammatory, rather than anti-inflammatory, process accompanies YopJ-promoted cytotoxicity, leading to increased systemic colonization by Y. pseudotuberculosis and potentially enhancing adaptive immunity to a live vaccine.
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aurintricarboxylic acid blocks in vitro and in vivo activity of yoph an essential virulent factor of Yersinia pestis the agent of plague
Journal of Biological Chemistry, 2003Co-Authors: Fubo Liang, James B Bliska, Zhonghui Huang, Seung Yub Lee, Jiao Liang, Maya I Ivanov, Andres Alonso, David S Lawrence, Tomas Mustelin, Zhong Yin ZhangAbstract:Yersinia are causative agents in human diseases ranging from gastrointestinal syndromes to Bubonic Plague. There is increasing risk of misuse of infectious agents, such as Yersinia pestis, as weapons of terror as well as instruments of warfare for mass destruction. YopH is an essential virulence factor whose protein-tyrosine phosphatase (PTP) activity is required for Yersinia pathogenicity. Consequently, there is considerable interest in developing potent and selective YopH inhibitors as novel anti-plague agents. We have screened a library of 720 structurally diverse commercially available carboxylic acids and identified 26 YopH inhibitors with IC50 values below 100 mum. The most potent and specific YopH inhibitor is aurintricarboxylic acid (ATA), which exhibits a Ki value of 5 nm for YopH and displays 6-120-fold selectivity in favor of YopH against a panel of mammalian PTPs. To determine whether ATA can block the activity of YopH in a cellular context, we have examined the effect of ATA on T-cell signaling in human Jurkat cells transfected with YopH. We show that YopH severely decreases the T-cell receptor-induced cellular tyrosine phosphorylation, ERK1/2 activity, and interleukin-2 transcriptional activity. We demonstrate that ATA can effectively block the inhibitory activity of YopH and restore normal T-cell function. These results provide a proof-of-concept for the hypothesis that small molecule inhibitors that selectively target YopH may be therapeutically useful. In addition, it is expected that potent and selective YopH inhibitors, such as ATA, should be useful reagents to delineate YopH's cellular targets in plague and other pathogenic conditions caused by Yersinia Infection.
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role of toll like receptor signaling in the apoptotic response of macrophages to Yersinia Infection
Infection and Immunity, 2003Co-Authors: Yue Zhang, James B BliskaAbstract:Macrophages encode several Toll-like receptors (TLRs) that recognize bacterial components, such as lipoproteins (TLR2) or lipopolysaccharides (TLR4), and activate multiple signaling pathways. Activation of transcription factor NF-κB by TLR2 or TLR4 signaling promotes proinflammatory and cell survival responses. Alternatively, TLR2 or TLR4 signaling can promote apoptosis if the activation of NF-κB is blocked. The gram-negative bacterial pathogen Yersinia pseudotuberculosis secretes into macrophages a protease (YopJ) that inhibits the activation of NF-κB and promotes apoptosis. We show that primary macrophages expressing constitutively active inhibitor κB kinase β (IKKβ) are completely resistant to YopJ-dependent apoptosis, indicating that YopJ inhibits signaling upstream of IKKβ. Apoptosis is reduced two- to threefold in TLR4−/− macrophages infected with Y. pseudotuberculosis, while the apoptotic response of TLR2−/− macrophages to Y. pseudotuberculosis Infection is equivalent to that of wild-type macrophages. Therefore, TLR4 is the primary source of apoptotic signaling in Yersinia-infected macrophages. Our results also show that a small percentage of macrophages can die as a result of an apoptotic process that is YopJ dependent but does not require TLR2 or TLR4 signaling.
Naomi H Philip - One of the best experts on this subject based on the ideXlab platform.
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the card19 locus of murine chromosome 13 regulates terminal cell lysis downstream of caspase activation and gasdermin d cleavage
bioRxiv, 2021Co-Authors: Elisabet Bjanes, Meghan A Wynoskydolfi, Reyna Garcia Sillas, Rina Matsuda, Benjamin Demarco, Timothee Fettrelet, Alexandra Delaney, Daniel Grubaugh, Lopez Emr, Naomi H PhilipAbstract:Cell death plays a critical role in inflammatory responses. During pyroptosis, inflammatory caspases cleave Gasdermin D (GSDMD) to release an N-terminal fragment that generates plasma membrane pores that mediate cell lysis and IL-1 cytokine release. Terminal cell lysis and IL-1β release following caspase activation can be uncoupled in certain cell types or in response to particular stimuli, a state termed hyperactivation. However, the factors and mechanisms that regulate terminal cell lysis downstream of GSDMD cleavage remain poorly understood. In the course of studies to define regulation of pyroptosis during Yersinia Infection, we identified a line of Card19-/- mice whose macrophages were protected from cell death and showed reduced pore formation during apoptosis or pyroptosis, yet had wild-type levels of caspase activation, IL-1 secretion, and GSDMD cleavage. Unexpectedly, CARD19, a mitochondrial CARD-containing protein, was not directly responsible for this, as two independently-generated CRISPR/Cas9 Card19 knockout mice showed no defect in macrophage cell lysis. The original Card19-/- line was generated in a 129SvEvBrd background, and SNP analysis revealed a six megabase region of 129 origin co-segregating with the Card19 locus. Card19 is located on chromosome 13, adjacent to Ninj1, which was recently reported to regulate cell lysis downstream of GSDMD activation. Nonetheless, we could not detect major defects in NINJ1 protein expression or mutations in Ninj1 coding sequence in Card19-/- mice. Mice from the original Card19-/- line exhibited significantly increased susceptibility to Yersinia Infection, demonstrating that cell lysis itself plays a key role in protection against bacterial Infection. Our findings identify a locus on murine chromosome 13 that regulates the ability of macrophages to undergo plasma membrane rupture downstream of gasdermin cleavage, and implicates additional NINJ1-independent factors that control terminal cell lysis.
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cell extrinsic tnf collaborates with trif signaling to promote Yersinia induced apoptosis
Journal of Immunology, 2016Co-Authors: Lance W Peterson, Naomi H Philip, Christopher P Dillon, Douglas R Green, John Bertin, Peter J Gough, Igor E BrodskyAbstract:Innate immune responses that are crucial for control of Infection are often targeted by microbial pathogens. Blockade of NF-κB and MAPK signaling by the Yersinia virulence factor YopJ inhibits cytokine production by innate immune cells but also triggers cell death. This cell death requires RIPK1 kinase activity and caspase-8, which are engaged by TLR4 and the adaptor protein TRIF. Nevertheless, TLR4- and TRIF-deficient cells undergo significant apoptosis, implicating TLR4/TRIF-independent pathways in the death of Yersinia-infected cells. In this article, we report a key role for TNF/TNFR1 in Yersinia-induced cell death of murine macrophages, which occurs despite the blockade of NF-κB and MAPK signaling imposed by Yersinia on infected cells. Intriguingly, direct analysis of YopJ injection revealed a heterogeneous population of injection-high and injection-low cells, and demonstrated that TNF expression came from the injection-low population. Moreover, TNF production by this subpopulation was necessary for maximal apoptosis in the population of highly injected cells, and TNFR-deficient mice displayed enhanced susceptibility to Yersinia Infection. These data demonstrate an important role for collaboration between TNF and pattern recognition receptor signals in promoting maximal apoptosis during bacterial Infection, and demonstrate that heterogeneity in virulence factor injection and cellular responses play an important role in promoting anti-Yersinia immune defense.
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iqgap1 is important for activation of caspase 1 in macrophages and is targeted by Yersinia pestis type iii effector yopm
Mbio, 2014Co-Authors: Lawton K Chung, Naomi H Philip, Igor E Brodsky, Valentina A Schmidt, Antonius Koller, Till Strowig, Richard A Flavell, James B BliskaAbstract:ABSTRACT YopM is a leucine-rich repeat (LRR)-containing effector in several Yersinia species, including Yersinia pestis and Y. pseudotuberculosis. Different Yersinia strains encode distinct YopM isoforms with variable numbers of LRRs but conserved C-terminal tails. A 15-LRR isoform in Y. pseudotuberculosis YPIII was recently shown to bind and inhibit caspase-1 via a YLTD motif in LRR 10, and attenuation of YopM − YPIII was reversed in mice lacking caspase-1, indicating that caspase-1 inhibition is a major virulence function of YopM YPIII . To determine if other YopM proteins inhibit caspase-1, we utilized Y. pseudotuberculosis strains natively expressing a 21-LRR isoform lacking the YLTD motif (YopM 32777 ) or ectopically expressing a Y. pestis 15-LRR version with a functional (YopM KIM ) or inactivated (YopM KIM D 271 A) YLTD motif. Results of mouse and macrophage Infections with these strains showed that YopM 32777 , YopM KIM , and YopM KIM D 271 A inhibit caspase-1 activation, indicating that the YLTD motif is dispensable for this activity. Analysis of YopM KIM deletion variants revealed that LRRs 6 to 15 and the C-terminal tail are required to inhibit caspase-1 activation. YopM 32777 , YopM KIM , and YopM KIM deletion variants were purified, and binding partners in macrophage lysates were identified. Caspase-1 bound to YopM KIM but not YopM 32777 . Additionally, YopM KIM bound IQGAP1 and the use of Iqgap1 −/− macrophages revealed that this scaffolding protein is important for caspase-1 activation upon Infection with YopM − Y. pseudotuberculosis. Thus, while multiple YopM isoforms inhibit caspase-1 activation, their variable LRR domains bind different host proteins to perform this function and the LRRs of YopM KIM target IQGAP1, a novel regulator of caspase-1, in macrophages. IMPORTANCE Activation of caspase-1, mediated by macromolecular complexes termed inflammasomes, is important for innate immune defense against pathogens. Pathogens can, in turn, subvert caspase-1-dependent responses through the action of effector proteins. For example, the Yersinia effector YopM inhibits caspase-1 activation by arresting inflammasome formation. This caspase-1 inhibitory activity has been studied in a specific YopM isoform, and in this case, the protein was shown to act as a pseudosubstrate to bind and inhibit caspase-1. Different Yersinia strains encode distinct YopM isoforms, many of which lack the pseudosubstrate motif. We studied additional isoforms and found that these YopM proteins inhibit caspase-1 activation independently of a pseudosubstrate motif. We also identified IQGAP1 as a novel binding partner of the Yersinia pestis YopM KIM isoform and demonstrated that IQGAP1 is important for caspase-1 activation in macrophages infected with Yersinia . Thus, this study reveals new insights into inflammasome regulation during Yersinia Infection.
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caspase 8 mediates caspase 1 processing and innate immune defense in response to bacterial blockade of nf κb and mapk signaling
Proceedings of the National Academy of Sciences of the United States of America, 2014Co-Authors: Naomi H Philip, Meghan A Wynoskydolfi, Christopher P Dillon, Annelise G Snyder, Patrick Fitzgerald, Erin E Zwack, Louise Fitzgerald, Elizabeth A Mauldin, Alan M Copenhaver, Sunny ShinAbstract:Toll-like receptor signaling and subsequent activation of NF-κB– and MAPK-dependent genes during Infection play an important role in antimicrobial host defense. The YopJ protein of pathogenic Yersinia species inhibits NF-κB and MAPK signaling, resulting in blockade of NF-κB–dependent cytokine production and target cell death. Nevertheless, Yersinia Infection induces inflammatory responses in vivo. Moreover, increasing the extent of YopJ-dependent cytotoxicity induced by Yersinia pestis and Yersinia pseudotuberculosis paradoxically leads to decreased virulence in vivo, suggesting that cell death promotes anti-Yersinia host defense. However, the specific pathways responsible for YopJ-induced cell death and how this cell death mediates immune defense against Yersinia remain poorly defined. YopJ activity induces processing of multiple caspases, including caspase-1, independently of inflammasome components or the adaptor protein ASC. Unexpectedly, caspase-1 activation in response to the activity of YopJ required caspase-8, receptor-interacting serine/threonine kinase 1 (RIPK1), and Fas-associated death domain (FADD), but not RIPK3. Furthermore, whereas RIPK3 deficiency did not affect YopJ-induced cell death or caspase-1 activation, deficiency of both RIPK3 and caspase-8 or FADD completely abrogated Yersinia-induced cell death and caspase-1 activation. Mice lacking RIPK3 and caspase-8 in their hematopoietic compartment showed extreme susceptibility to Yersinia and were deficient in monocyte and neutrophil-derived production of proinflammatory cytokines. Our data demonstrate for the first time to our knowledge that RIPK1, FADD, and caspase-8 are required for YopJ-induced cell death and caspase-1 activation and suggest that caspase-8–mediated cell death overrides blockade of immune signaling by YopJ to promote anti-Yersinia immune defense.
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caspase 8 ripk1 and fadd regulate cell death and caspase 1 activation during Yersinia Infection inm3p 438
Journal of Immunology, 2014Co-Authors: Naomi H Philip, Meghan A Wynoskydolfi, Christopher P Dillon, Annelise G Snyder, Patrick Fitzgerald, Erin E Zwack, Baofeng Hu, Louise Fitzgerald, Elizabeth A Mauldin, Alan M CopenhaverAbstract:Programmed cell death is an evolutionarily conserved response to Infection that can promote host defense or microbial virulence. Pathogens manipulate various immune signaling pathways through the activity of specific virulence factors that access the host cell cytosol. Cell death is a major consequence of Infection with pathogenic Y. pseudotuberculosis and requires the effector YopJ, a potent inhibitor of NF-κB and MAPK signaling. However, the pathways that regulate cell death in response to Yersinia Infection and the precise mechanism by which cell death mediates protective immunity are not well understood. We find a novel requirement for caspase-8, receptor interacting protein 1 (RIPK1) and Fas-associated protein with death domain (FADD) in Yersinia-induced cell death and pro-inflammatory caspase-1 activation. Moreover, mice deficient in caspase-8 were highly susceptible to Yersinia Infection and were unable to sustain innate cytokine production. We hypothesize that activation of these pathways during Yersinia Infection may induce specific pro-inflammatory signals that shape innate and adaptive responses and promote microbial clearance.
Meghan A Wynoskydolfi - One of the best experts on this subject based on the ideXlab platform.
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the card19 locus of murine chromosome 13 regulates terminal cell lysis downstream of caspase activation and gasdermin d cleavage
bioRxiv, 2021Co-Authors: Elisabet Bjanes, Meghan A Wynoskydolfi, Reyna Garcia Sillas, Rina Matsuda, Benjamin Demarco, Timothee Fettrelet, Alexandra Delaney, Daniel Grubaugh, Lopez Emr, Naomi H PhilipAbstract:Cell death plays a critical role in inflammatory responses. During pyroptosis, inflammatory caspases cleave Gasdermin D (GSDMD) to release an N-terminal fragment that generates plasma membrane pores that mediate cell lysis and IL-1 cytokine release. Terminal cell lysis and IL-1β release following caspase activation can be uncoupled in certain cell types or in response to particular stimuli, a state termed hyperactivation. However, the factors and mechanisms that regulate terminal cell lysis downstream of GSDMD cleavage remain poorly understood. In the course of studies to define regulation of pyroptosis during Yersinia Infection, we identified a line of Card19-/- mice whose macrophages were protected from cell death and showed reduced pore formation during apoptosis or pyroptosis, yet had wild-type levels of caspase activation, IL-1 secretion, and GSDMD cleavage. Unexpectedly, CARD19, a mitochondrial CARD-containing protein, was not directly responsible for this, as two independently-generated CRISPR/Cas9 Card19 knockout mice showed no defect in macrophage cell lysis. The original Card19-/- line was generated in a 129SvEvBrd background, and SNP analysis revealed a six megabase region of 129 origin co-segregating with the Card19 locus. Card19 is located on chromosome 13, adjacent to Ninj1, which was recently reported to regulate cell lysis downstream of GSDMD activation. Nonetheless, we could not detect major defects in NINJ1 protein expression or mutations in Ninj1 coding sequence in Card19-/- mice. Mice from the original Card19-/- line exhibited significantly increased susceptibility to Yersinia Infection, demonstrating that cell lysis itself plays a key role in protection against bacterial Infection. Our findings identify a locus on murine chromosome 13 that regulates the ability of macrophages to undergo plasma membrane rupture downstream of gasdermin cleavage, and implicates additional NINJ1-independent factors that control terminal cell lysis.
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serine 25 phosphorylation inhibits ripk1 kinase dependent cell death in models of Infection and inflammation
Nature Communications, 2019Co-Authors: Yves Dondelinger, Dario Priem, Daniel Sorobetea, Piero Giansanti, Tom Delanghe, Meghan A Wynoskydolfi, Diego Rojasrivera, Ria RoelandtAbstract:RIPK1 regulates cell death and inflammation through kinase-dependent and -independent mechanisms. As a scaffold, RIPK1 inhibits caspase-8-dependent apoptosis and RIPK3/MLKL-dependent necroptosis. As a kinase, RIPK1 paradoxically induces these cell death modalities. The molecular switch between RIPK1 pro-survival and pro-death functions remains poorly understood. We identify phosphorylation of RIPK1 on Ser25 by IKKs as a key mechanism directly inhibiting RIPK1 kinase activity and preventing TNF-mediated RIPK1-dependent cell death. Mimicking Ser25 phosphorylation (S > D mutation) protects cells and mice from the cytotoxic effect of TNF in conditions of IKK inhibition. In line with their roles in IKK activation, TNF-induced Ser25 phosphorylation of RIPK1 is defective in TAK1- or SHARPIN-deficient cells and restoring phosphorylation protects these cells from TNF-induced death. Importantly, mimicking Ser25 phosphorylation compromises the in vivo cell death-dependent immune control of Yersinia Infection, a physiological model of TAK1/IKK inhibition, and rescues the cell death-induced multi-organ inflammatory phenotype of the SHARPIN-deficient mice.
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caspase 8 mediates caspase 1 processing and innate immune defense in response to bacterial blockade of nf κb and mapk signaling
Proceedings of the National Academy of Sciences of the United States of America, 2014Co-Authors: Naomi H Philip, Meghan A Wynoskydolfi, Christopher P Dillon, Annelise G Snyder, Patrick Fitzgerald, Erin E Zwack, Louise Fitzgerald, Elizabeth A Mauldin, Alan M Copenhaver, Sunny ShinAbstract:Toll-like receptor signaling and subsequent activation of NF-κB– and MAPK-dependent genes during Infection play an important role in antimicrobial host defense. The YopJ protein of pathogenic Yersinia species inhibits NF-κB and MAPK signaling, resulting in blockade of NF-κB–dependent cytokine production and target cell death. Nevertheless, Yersinia Infection induces inflammatory responses in vivo. Moreover, increasing the extent of YopJ-dependent cytotoxicity induced by Yersinia pestis and Yersinia pseudotuberculosis paradoxically leads to decreased virulence in vivo, suggesting that cell death promotes anti-Yersinia host defense. However, the specific pathways responsible for YopJ-induced cell death and how this cell death mediates immune defense against Yersinia remain poorly defined. YopJ activity induces processing of multiple caspases, including caspase-1, independently of inflammasome components or the adaptor protein ASC. Unexpectedly, caspase-1 activation in response to the activity of YopJ required caspase-8, receptor-interacting serine/threonine kinase 1 (RIPK1), and Fas-associated death domain (FADD), but not RIPK3. Furthermore, whereas RIPK3 deficiency did not affect YopJ-induced cell death or caspase-1 activation, deficiency of both RIPK3 and caspase-8 or FADD completely abrogated Yersinia-induced cell death and caspase-1 activation. Mice lacking RIPK3 and caspase-8 in their hematopoietic compartment showed extreme susceptibility to Yersinia and were deficient in monocyte and neutrophil-derived production of proinflammatory cytokines. Our data demonstrate for the first time to our knowledge that RIPK1, FADD, and caspase-8 are required for YopJ-induced cell death and caspase-1 activation and suggest that caspase-8–mediated cell death overrides blockade of immune signaling by YopJ to promote anti-Yersinia immune defense.
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caspase 8 ripk1 and fadd regulate cell death and caspase 1 activation during Yersinia Infection inm3p 438
Journal of Immunology, 2014Co-Authors: Naomi H Philip, Meghan A Wynoskydolfi, Christopher P Dillon, Annelise G Snyder, Patrick Fitzgerald, Erin E Zwack, Baofeng Hu, Louise Fitzgerald, Elizabeth A Mauldin, Alan M CopenhaverAbstract:Programmed cell death is an evolutionarily conserved response to Infection that can promote host defense or microbial virulence. Pathogens manipulate various immune signaling pathways through the activity of specific virulence factors that access the host cell cytosol. Cell death is a major consequence of Infection with pathogenic Y. pseudotuberculosis and requires the effector YopJ, a potent inhibitor of NF-κB and MAPK signaling. However, the pathways that regulate cell death in response to Yersinia Infection and the precise mechanism by which cell death mediates protective immunity are not well understood. We find a novel requirement for caspase-8, receptor interacting protein 1 (RIPK1) and Fas-associated protein with death domain (FADD) in Yersinia-induced cell death and pro-inflammatory caspase-1 activation. Moreover, mice deficient in caspase-8 were highly susceptible to Yersinia Infection and were unable to sustain innate cytokine production. We hypothesize that activation of these pathways during Yersinia Infection may induce specific pro-inflammatory signals that shape innate and adaptive responses and promote microbial clearance.
Alan M Copenhaver - One of the best experts on this subject based on the ideXlab platform.
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caspase 8 mediates caspase 1 processing and innate immune defense in response to bacterial blockade of nf κb and mapk signaling
Proceedings of the National Academy of Sciences of the United States of America, 2014Co-Authors: Naomi H Philip, Meghan A Wynoskydolfi, Christopher P Dillon, Annelise G Snyder, Patrick Fitzgerald, Erin E Zwack, Louise Fitzgerald, Elizabeth A Mauldin, Alan M Copenhaver, Sunny ShinAbstract:Toll-like receptor signaling and subsequent activation of NF-κB– and MAPK-dependent genes during Infection play an important role in antimicrobial host defense. The YopJ protein of pathogenic Yersinia species inhibits NF-κB and MAPK signaling, resulting in blockade of NF-κB–dependent cytokine production and target cell death. Nevertheless, Yersinia Infection induces inflammatory responses in vivo. Moreover, increasing the extent of YopJ-dependent cytotoxicity induced by Yersinia pestis and Yersinia pseudotuberculosis paradoxically leads to decreased virulence in vivo, suggesting that cell death promotes anti-Yersinia host defense. However, the specific pathways responsible for YopJ-induced cell death and how this cell death mediates immune defense against Yersinia remain poorly defined. YopJ activity induces processing of multiple caspases, including caspase-1, independently of inflammasome components or the adaptor protein ASC. Unexpectedly, caspase-1 activation in response to the activity of YopJ required caspase-8, receptor-interacting serine/threonine kinase 1 (RIPK1), and Fas-associated death domain (FADD), but not RIPK3. Furthermore, whereas RIPK3 deficiency did not affect YopJ-induced cell death or caspase-1 activation, deficiency of both RIPK3 and caspase-8 or FADD completely abrogated Yersinia-induced cell death and caspase-1 activation. Mice lacking RIPK3 and caspase-8 in their hematopoietic compartment showed extreme susceptibility to Yersinia and were deficient in monocyte and neutrophil-derived production of proinflammatory cytokines. Our data demonstrate for the first time to our knowledge that RIPK1, FADD, and caspase-8 are required for YopJ-induced cell death and caspase-1 activation and suggest that caspase-8–mediated cell death overrides blockade of immune signaling by YopJ to promote anti-Yersinia immune defense.
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caspase 8 ripk1 and fadd regulate cell death and caspase 1 activation during Yersinia Infection inm3p 438
Journal of Immunology, 2014Co-Authors: Naomi H Philip, Meghan A Wynoskydolfi, Christopher P Dillon, Annelise G Snyder, Patrick Fitzgerald, Erin E Zwack, Baofeng Hu, Louise Fitzgerald, Elizabeth A Mauldin, Alan M CopenhaverAbstract:Programmed cell death is an evolutionarily conserved response to Infection that can promote host defense or microbial virulence. Pathogens manipulate various immune signaling pathways through the activity of specific virulence factors that access the host cell cytosol. Cell death is a major consequence of Infection with pathogenic Y. pseudotuberculosis and requires the effector YopJ, a potent inhibitor of NF-κB and MAPK signaling. However, the pathways that regulate cell death in response to Yersinia Infection and the precise mechanism by which cell death mediates protective immunity are not well understood. We find a novel requirement for caspase-8, receptor interacting protein 1 (RIPK1) and Fas-associated protein with death domain (FADD) in Yersinia-induced cell death and pro-inflammatory caspase-1 activation. Moreover, mice deficient in caspase-8 were highly susceptible to Yersinia Infection and were unable to sustain innate cytokine production. We hypothesize that activation of these pathways during Yersinia Infection may induce specific pro-inflammatory signals that shape innate and adaptive responses and promote microbial clearance.