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1
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Sheneman KR, Cummins TD, Merchant ML, Hood JL, Uriarte SM, Lawrenz MB. Yersinia pestis Actively Inhibits the Production of Extracellular Vesicles by Human Neutrophils. J Extracell Vesicles 2025; 14:e70074. [PMID: 40240908 PMCID: PMC12003101 DOI: 10.1002/jev2.70074] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/17/2024] [Accepted: 03/23/2025] [Indexed: 04/18/2025] Open
Abstract
Yersinia pestis is the etiologic agent of the plague. A hallmark of plague is subversion of the host immune response by disrupting host signalling pathways required for inflammation. This non-inflammatory environment permits bacterial colonization and has been shown to be essential for disease manifestation. Previous work has shown that Y. pestis inhibits phagocytosis and degranulation by neutrophils. Manipulation of these key vesicular trafficking pathways suggests that Y. pestis influences extracellular vesicle (EV) secretion, cargo selection, trafficking and/or maturation. Our goals were to define the EV population produced by neutrophils in response to Y. pestis and determine how these vesicles might influence inflammation. Towards these goals, EVs were isolated from human neutrophils infected with Y. pestis or a mutant lacking bacterial effector proteins known to manipulate host cell signalling. Mass spectrometry data revealed that cargoes packaged in EVs isolated from mutant infected cells were enriched with antimicrobial and cytotoxic proteins, contents which differed from uninfected and Y. pestis infected cells. Further, EVs produced in response to Y. pestis lacked inflammatory properties observed in those isolated from neutrophils responding to the mutant. Together, these data demonstrate that Y. pestis actively inhibits the production of antimicrobial EVs produced by neutrophils, likely contributing to immune evasion.
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Affiliation(s)
- Katelyn R. Sheneman
- Department of Microbiology and ImmunologyUniversity of LouisvilleLouisvilleKentuckyUSA
| | - Timothy D. Cummins
- Department of Medicine and Proteomics Technology CenterUniversity of LouisvilleLouisvilleKentuckyUSA
| | - Michael L. Merchant
- Department of Pharmacology and ToxicologyUniversity of LouisvilleLouisvilleKentuckyUSA
| | - Joshua L. Hood
- Department of Pharmacology and ToxicologyUniversity of LouisvilleLouisvilleKentuckyUSA
- Hepatobiology and Toxicology COBREUniversity of LouisvilleLouisvilleKentuckyUSA
| | - Silvia M. Uriarte
- Department of Oral Immunology & Infectious DiseaseUniversity of LouisvilleLouisvilleKentuckyUSA
| | - Matthew B. Lawrenz
- Department of Microbiology and ImmunologyUniversity of LouisvilleLouisvilleKentuckyUSA
- Center for Predictive Medicine for Biodefense and Emerging Infectious DiseasesUniversity of LouisvilleLouisvilleKentuckyUSA
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2
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Sheneman KR, Cummins TD, Merchant ML, Hood JL, Uriarte SM, Lawrenz MB. Yersinia pestis actively inhibits the production of extracellular vesicles by human neutrophils. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.12.20.629761. [PMID: 39763979 PMCID: PMC11702605 DOI: 10.1101/2024.12.20.629761] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 01/12/2025]
Abstract
Yersinia pestis is the etiologic agent of the plague. A hallmark of plague is subversion of the host immune response by disrupting host signaling pathways required for inflammation. This non-inflammatory environment permits bacterial colonization and has been shown to be essential for disease manifestation. Previous work has shown that Y. pestis inhibits phagocytosis and degranulation by neutrophils. Manipulation of these key vesicular trafficking pathways suggests that Y. pestis influences EV secretion, cargo selection, trafficking, and/or maturation. Our goal was to define the EV population produced by neutrophils in response to Y. pestis and determine how these vesicles might influence inflammation. Toward these goals, EVs were isolated from human neutrophils infected with Y. pestis or a mutant lacking bacterial effector proteins known to manipulate host cell signaling. Mass spectrometry data revealed that cargoes packaged in EVs isolated from mutant infected cells were enriched with antimicrobials and cytotoxic proteins, contents which differed from uninfected and Y. pestis infected cells. Further, EVs produced in response to Y. pestis lacked inflammatory properties observed in those isolated from neutrophils responding to the mutant. Together, these data demonstrate that Y. pestis actively inhibits the production of antimicrobial EVs produced by neutrophils, likely contributing to immune evasion.
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Affiliation(s)
| | - Timothy D Cummins
- Department of Medicine and Proteomics Technology Center, University of Louisville
| | | | - Joshua L Hood
- Department of Pharmacology and Toxicology, University of Louisville
- Hepatobiology and Toxicology COBRE, University of Louisville
| | - Silvia M Uriarte
- Department of Oral Immunology & Infectious Disease, University of Louisville
| | - Matthew B Lawrenz
- Department of Microbiology and Immunology, University of Louisville
- Center for Predictive Medicine for Biodefense and Emerging Infectious Diseases, University of Louisville
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3
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Brady A, Mora Martinez LC, Hammond B, Whitefoot-Keliin KM, Haribabu B, Uriarte SM, Lawrenz MB. Distinct mechanisms of type 3 secretion system recognition control LTB4 synthesis in neutrophils and macrophages. PLoS Pathog 2024; 20:e1012651. [PMID: 39423229 PMCID: PMC11524448 DOI: 10.1371/journal.ppat.1012651] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/09/2024] [Revised: 10/30/2024] [Accepted: 10/09/2024] [Indexed: 10/21/2024] Open
Abstract
Leukotriene B4 (LTB4) is an inflammatory lipid produced in response to pathogens that is critical for initiating the inflammatory cascade needed to control infection. However, during plague, Yersinia pestis inhibits the timely synthesis of LTB4 and subsequent inflammation. Using bacterial mutants, we previously determined that Y. pestis inhibits LTB4 synthesis via the action of the Yop effector proteins that are directly secreted into host cells through a type 3 secretion system (T3SS). Here, we show that the T3SS is the primary pathogen associated molecular pattern (PAMP) required for production of LTB4 in response to both Yersinia and Salmonella. However, we also unexpectantly discovered that T3SS-mediated LTB4 synthesis by neutrophils and macrophages require the activation of two distinctly different host signaling pathways. We identified that phagocytosis and the NLRP3/CASP1 inflammasome significantly impact LTB4 synthesis by macrophages but not neutrophils. Instead, the SKAP2/PLC signaling pathway is required for T3SS-mediated LTB4 production by neutrophils. Finally, while recognition of the T3SS is required for LTB4 production, we also discovered that a second unrelated PAMP-mediated signal activates the MAP kinase pathway needed for synthesis. Together, these data demonstrate significant differences in the host factors and signaling pathways required by macrophages and neutrophils to quickly produce LTB4 in response to bacteria. Moreover, while macrophages and neutrophils might rely on different signaling pathways for T3SS-dependent LTB4 synthesis, Y. pestis has evolved virulence mechanisms to counteract this response by either leukocyte to inhibit LTB4 synthesis and colonize the host.
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Affiliation(s)
- Amanda Brady
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Leonardo C. Mora Martinez
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Benjamin Hammond
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Kaitlyn M. Whitefoot-Keliin
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Bodduluri Haribabu
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
- Center for Microbiomics, Inflammation and Pathogenicity, Louisville, Kentucky, United States of America
| | - Silvia M. Uriarte
- Deptartment of Oral Immunology & Infectious Diseases, University of Louisville, Louisville, Kentucky, United States of America
- Center for Predictive Medicine for Biodefense and Emerging Infectious Diseases, Louisville, Kentucky, United States of America
| | - Matthew B. Lawrenz
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
- Center for Predictive Medicine for Biodefense and Emerging Infectious Diseases, Louisville, Kentucky, United States of America
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4
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Greene J, Snyder RA, Cotten KL, Huiszoon RC, Chu S, Braza RED, Chapin AA, Stine JM, Bentley WE, Ghodssi R, Davis KM. Yersinia pseudotuberculosis growth arrest during type-III secretion system expression is associated with altered ribosomal protein expression and decreased gentamicin susceptibility. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.09.02.610769. [PMID: 39282321 PMCID: PMC11398311 DOI: 10.1101/2024.09.02.610769] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Indexed: 10/25/2024]
Abstract
It has been long appreciated that expression of the Yersinia type-III secretion system (T3SS) in culture is associated with growth arrest. Here we sought to understand whether this impacts expression of ribosomal protein genes, which were among the most highly abundant transcripts in exponential phase Yersinia pseudotuberculosis based on RNA-seq analysis. To visualize changes in ribosomal protein expression, we generated a fluorescent transcriptional reporter with the promoter upstream of rpsJ/S10 fused to a destabilized gfp variant. We confirmed reporter expression significantly increases in exponential phase and decreases as cells transition to stationary phase. We then utilized a mouse model of systemic Y. pseudotuberculosis infection to compare T3SS and S10 reporter expression during clustered bacterial growth in the spleen, and found that cells expressing high levels of the T3SS had decreased S10 levels, while cells with lower T3SS expression retained higher S10 expression. In bacteriological media, growth inhibition with T3SS induction and a reduction in S10 expression were observed in subsets of cells, while cells with high expression of both T3SS and S10 were also observed. Loss of T3SS genes resulted in rescued growth and heightened S10 expression. To understand if clustered growth impacted bacterial gene expression, we utilized droplet-based microfluidics to encapsulate bacteria in spherical agarose droplets, and also observed growth inhibition with high expression of T3SS and reduced S10 levels that better mirrored phenotypes observed in the mouse spleen. Finally, we show that T3SS expression is sufficient to promote tolerance to the ribosome-targeting antibiotic, gentamicin. Collectively, these data indicate that the growth arrest associated with T3SS induction leads to decreased expression of ribosomal protein genes, and this results in reduced antibiotic susceptibility.
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Affiliation(s)
- Justin Greene
- W. Harry Feinstone Department of Molecular Microbiology and Immunology Johns Hopkins Bloomberg School of Public Health, Baltimore, MD, USA
| | - Rhett A. Snyder
- W. Harry Feinstone Department of Molecular Microbiology and Immunology Johns Hopkins Bloomberg School of Public Health, Baltimore, MD, USA
| | - Katherine L. Cotten
- W. Harry Feinstone Department of Molecular Microbiology and Immunology Johns Hopkins Bloomberg School of Public Health, Baltimore, MD, USA
| | - Ryan C. Huiszoon
- Institute for Systems Research, University of Maryland, College Park, MD, USA
- Fischell Department of Bioengineering, University of Maryland, College Park, MD, USA
- Robert E. Fischell Institute for Biomedical Devices, University of Maryland, College Park, MD, USA
| | - Sangwook Chu
- Institute for Systems Research, University of Maryland, College Park, MD, USA
| | - Rezia Era D. Braza
- W. Harry Feinstone Department of Molecular Microbiology and Immunology Johns Hopkins Bloomberg School of Public Health, Baltimore, MD, USA
| | - Ashley A. Chapin
- Institute for Systems Research, University of Maryland, College Park, MD, USA
- Fischell Department of Bioengineering, University of Maryland, College Park, MD, USA
- Robert E. Fischell Institute for Biomedical Devices, University of Maryland, College Park, MD, USA
| | - Justin M. Stine
- Institute for Systems Research, University of Maryland, College Park, MD, USA
- Department of Electrical and Computer Engineering, University of Maryland, College Park, MD, USA
- Robert E. Fischell Institute for Biomedical Devices, University of Maryland, College Park, MD, USA
| | - William E. Bentley
- Fischell Department of Bioengineering, University of Maryland, College Park, MD, USA
- Robert E. Fischell Institute for Biomedical Devices, University of Maryland, College Park, MD, USA
| | - Reza Ghodssi
- Institute for Systems Research, University of Maryland, College Park, MD, USA
- Department of Electrical and Computer Engineering, University of Maryland, College Park, MD, USA
- Robert E. Fischell Institute for Biomedical Devices, University of Maryland, College Park, MD, USA
| | - Kimberly M. Davis
- W. Harry Feinstone Department of Molecular Microbiology and Immunology Johns Hopkins Bloomberg School of Public Health, Baltimore, MD, USA
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5
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Chen KW, Broz P. Gasdermins as evolutionarily conserved executors of inflammation and cell death. Nat Cell Biol 2024; 26:1394-1406. [PMID: 39187689 DOI: 10.1038/s41556-024-01474-z] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/01/2024] [Accepted: 07/04/2024] [Indexed: 08/28/2024]
Abstract
The gasdermins are a family of pore-forming proteins that have recently emerged as executors of pyroptosis, a lytic form of cell death that is induced by the innate immune system to eradicate infected or malignant cells. Mammalian gasdermins comprise a cytotoxic N-terminal domain, a flexible linker and a C-terminal repressor domain. Proteolytic cleavage in the linker releases the cytotoxic domain, thereby allowing it to form β-barrel membrane pores. Formation of gasdermin pores in the plasma membrane eventually leads to a loss of the electrochemical gradient, cell death and membrane rupture. Here we review recent work that has expanded our understanding of gasdermin biology and function in mammals by revealing their activation mechanism, their regulation and their roles in autoimmunity, host defence and cancer. We further highlight fungal and bacterial gasdermin pore formation pointing to a conserved mechanism of cell death induction.
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Affiliation(s)
- Kaiwen W Chen
- Immunology Translational Research Programme, Department of Microbiology and Immunology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore.
- Immunology Programme, Life Sciences Institute, National University of Singapore, Singapore, Singapore.
| | - Petr Broz
- Department of Immunobiology, University of Lausanne, Lausanne, Switzerland.
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Jetton D, Muendlein HI, Connolly WM, Magri Z, Smirnova I, Batorsky R, Mecsas J, Degterev A, Poltorak A. Non-canonical autophosphorylation of RIPK1 drives timely pyroptosis to control Yersinia infection. Cell Rep 2024; 43:114641. [PMID: 39154339 PMCID: PMC11465231 DOI: 10.1016/j.celrep.2024.114641] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/24/2024] [Revised: 06/17/2024] [Accepted: 07/31/2024] [Indexed: 08/20/2024] Open
Abstract
Caspase-8-dependent pyroptosis has been shown to mediate host protection from Yersinia infection. For this mode of cell death, the kinase activity of receptor-interacting protein kinase 1 (RIPK1) is required, but the autophosphorylation sites required to drive caspase-8 activation have not been determined. Here, we show that non-canonical autophosphorylation of RIPK1 at threonine 169 (T169) is necessary for caspase-8-mediated pyroptosis. Mice with alanine in the T169 position are highly susceptible to Yersinia dissemination. Mechanistically, the delayed formation of a complex containing RIPK1, ZBP1, Fas-associated protein with death domain (FADD), and caspase-8 abrogates caspase-8 maturation in T169A mice and leads to the eventual activation of RIPK3-dependent necroptosis in vivo; however, this is insufficient to protect the host, suggesting that timely pyroptosis during early response is specifically required to control infection. These results position RIPK1 T169 phosphorylation as a driver of pyroptotic cell death critical for host defense.
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Affiliation(s)
- David Jetton
- Graduate Program in Immunology, Tufts Graduate School of Biomedical Sciences, Boston, MA 02111, USA
| | - Hayley I Muendlein
- Department of Immunology, Tufts University School of Medicine, Boston, MA 02111, USA
| | - Wilson M Connolly
- Department of Immunology, Tufts University School of Medicine, Boston, MA 02111, USA
| | - Zoie Magri
- Graduate Program in Immunology, Tufts Graduate School of Biomedical Sciences, Boston, MA 02111, USA
| | - Irina Smirnova
- Department of Immunology, Tufts University School of Medicine, Boston, MA 02111, USA
| | - Rebecca Batorsky
- Data Intensive Studies Center, Tufts University, Medford, MA 02155, USA
| | - Joan Mecsas
- Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, MA 02111, USA
| | - Alexei Degterev
- Department of Cell, Molecular & Developmental Biology, Tufts University School of Medicine, Boston, MA 02111, USA
| | - Alexander Poltorak
- Graduate Program in Immunology, Tufts Graduate School of Biomedical Sciences, Boston, MA 02111, USA; Department of Immunology, Tufts University School of Medicine, Boston, MA 02111, USA.
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7
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Brady A, Mora-Martinez LC, Hammond B, Haribabu B, Uriarte SM, Lawrenz MB. Distinct Mechanisms of Type 3 Secretion System Recognition Control LTB 4 Synthesis in Neutrophils versus Macrophages. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.07.01.601466. [PMID: 39005373 PMCID: PMC11244889 DOI: 10.1101/2024.07.01.601466] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 07/16/2024]
Abstract
Leukotriene B4 (LTB4) is critical for initiating the inflammatory cascade in response to infection. However, Yersinia pestis colonizes the host by inhibiting the timely synthesis of LTB4 and inflammation. Here, we show that the bacterial type 3 secretion system (T3SS) is the primary pathogen associated molecular pattern (PAMP) responsible for LTB4 production by leukocytes in response to Yersinia and Salmonella, but synthesis is inhibited by the Yop effectors during Yersinia interactions. Moreover, we unexpectedly discovered that T3SS-mediated LTB4 synthesis by neutrophils and macrophages require two distinct host signaling pathways. We show that the SKAP2/PLC signaling pathway is essential for LTB4 production by neutrophils but not macrophages. Instead, phagocytosis and the NLRP3/CASP1 inflammasome are needed for LTB4 synthesis by macrophages. Finally, while recognition of the T3SS is required for LTB4 production, we also discovered a second unrelated PAMP-mediated signal independently activates the MAP kinase pathway needed for LTB4 synthesis. Together, these data demonstrate significant differences in the signaling pathways required by macrophages and neutrophils to quickly respond to bacterial infections.
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Affiliation(s)
- Amanda Brady
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Leonardo C. Mora-Martinez
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Benjamin Hammond
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Bodduluri Haribabu
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
- Center for Microbiomics, Inflammation and Pathogenicity, Louisville, Kentucky, United States of America
| | - Silvia M. Uriarte
- Deptartment of Oral Immunology & Infectious Diseases, University of Louisville, Louisville, Kentucky, United States of America
- Center for Predictive Medicine for Biodefense and Emerging Infectious Diseases, Louisville, Kentucky, United States of America
| | - Matthew B. Lawrenz
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
- Center for Predictive Medicine for Biodefense and Emerging Infectious Diseases, Louisville, Kentucky, United States of America
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8
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Brady A, Sheneman KR, Pulsifer AR, Price SL, Garrison TM, Maddipati KR, Bodduluri SR, Pan J, Boyd NL, Zheng JJ, Rai SN, Hellmann J, Haribabu B, Uriarte SM, Lawrenz MB. Type 3 secretion system induced leukotriene B4 synthesis by leukocytes is actively inhibited by Yersinia pestis to evade early immune recognition. PLoS Pathog 2024; 20:e1011280. [PMID: 38271464 PMCID: PMC10846697 DOI: 10.1371/journal.ppat.1011280] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/12/2023] [Revised: 02/06/2024] [Accepted: 01/16/2024] [Indexed: 01/27/2024] Open
Abstract
Subverting the host immune response to inhibit inflammation is a key virulence strategy of Yersinia pestis. The inflammatory cascade is tightly controlled via the sequential action of lipid and protein mediators of inflammation. Because delayed inflammation is essential for Y. pestis to cause lethal infection, defining the Y. pestis mechanisms to manipulate the inflammatory cascade is necessary to understand this pathogen's virulence. While previous studies have established that Y. pestis actively inhibits the expression of host proteins that mediate inflammation, there is currently a gap in our understanding of the inflammatory lipid mediator response during plague. Here we used the murine model to define the kinetics of the synthesis of leukotriene B4 (LTB4), a pro-inflammatory lipid chemoattractant and immune cell activator, within the lungs during pneumonic plague. Furthermore, we demonstrated that exogenous administration of LTB4 prior to infection limited bacterial proliferation, suggesting that the absence of LTB4 synthesis during plague contributes to Y. pestis immune evasion. Using primary leukocytes from mice and humans further revealed that Y. pestis actively inhibits the synthesis of LTB4. Finally, using Y. pestis mutants in the Ysc type 3 secretion system (T3SS) and Yersinia outer protein (Yop) effectors, we demonstrate that leukocytes recognize the T3SS to initiate the rapid synthesis of LTB4. However, several Yop effectors secreted through the T3SS effectively inhibit this host response. Together, these data demonstrate that Y. pestis actively inhibits the synthesis of the inflammatory lipid LTB4 contributing to the delay in the inflammatory cascade required for rapid recruitment of leukocytes to sites of infection.
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Affiliation(s)
- Amanda Brady
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Katelyn R. Sheneman
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Amanda R. Pulsifer
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Sarah L. Price
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Taylor M. Garrison
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Krishna Rao Maddipati
- Department of Pathology, Lipidomics Core Facility, Wayne State University, Detroit, Michigan, United States of America
| | - Sobha R. Bodduluri
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Jianmin Pan
- Biostatistics and Bioinformatics Facility, Brown Cancer Center, University of Louisville, Louisville, Kentucky, United States of America
| | - Nolan L. Boyd
- Center for Cardiometabolic Science, Christina Lee Brown Environment Institute, Division of Environmental Medicine, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Jing-Juan Zheng
- Center for Cardiometabolic Science, Christina Lee Brown Environment Institute, Division of Environmental Medicine, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Shesh N. Rai
- Department of Pharmacology and Toxicology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Jason Hellmann
- Center for Cardiometabolic Science, Christina Lee Brown Environment Institute, Division of Environmental Medicine, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Bodduluri Haribabu
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
| | - Silvia M. Uriarte
- Deptartment of Oral Immunology & Infectious Diseases, University of Louisville, Louisville, Kentucky, United States of America
| | - Matthew B. Lawrenz
- Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, Kentucky, United States of America
- Center for Predictive Medicine for Biodefense and Emerging Infectious Diseases, Louisville, Kentucky, United States of America
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9
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Zhang J, Brodsky IE, Shin S. Yersinia deploys type III-secreted effectors to evade caspase-4 inflammasome activation in human cells. mBio 2023; 14:e0131023. [PMID: 37615436 PMCID: PMC10653943 DOI: 10.1128/mbio.01310-23] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/15/2023] [Accepted: 07/06/2023] [Indexed: 08/25/2023] Open
Abstract
IMPORTANCE Yersinia are responsible for significant disease burden in humans, ranging from recurrent disease outbreaks (yersiniosis) to pandemics (Yersinia pestis plague). Together with rising antibiotic resistance rates, there is a critical need to better understand Yersinia pathogenesis and host immune mechanisms, as this information will aid in developing improved immunomodulatory therapeutics. Inflammasome responses in human cells are less studied relative to murine models of infection, though recent studies have uncovered key differences in inflammasome responses between mice and humans. Here, we dissect human intestinal epithelial cell and macrophage inflammasome responses to Yersinia pseudotuberculosis. Our findings provide insight into species- and cell type-specific differences in inflammasome responses to Yersinia.
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Affiliation(s)
- Jenna Zhang
- Department of Microbiology, University of Pennsylvania, Perelman School of Medicine, Philadelphia, Pennsylvania, USA
| | - Igor E. Brodsky
- Department of Pathobiology, University of Pennsylvania, School of Veterinary Medicine, Philadelphia, Pennsylvania, USA
| | - Sunny Shin
- Department of Microbiology, University of Pennsylvania, Perelman School of Medicine, Philadelphia, Pennsylvania, USA
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10
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Davidson RK, Davis KM. Yersinia pseudotuberculosis: Cultivation, Storage, and Methods for Introducing DNA. ACTA ACUST UNITED AC 2021; 59:e122. [PMID: 33079471 DOI: 10.1002/cpmc.122] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022]
Abstract
Yersinia pseudotuberculosis has been studied for many decades, and research on this microbe has taught us a great deal about host-pathogen interactions, bacterial manipulation of host cells, virulence factors, and the evolution of pathogens. This microbe should not be cultivated at 37°C because this is a trigger that the bacterium uses to sense its presence within a mammalian host and results in expression of genes necessary to colonize a mammalian host. Prolonged growth at this temperature can result in accumulation of mutations that reduce the virulence of the strain, so all protocols need to be modified for growth at room temperature, or 26°C. This article describes protocols for cultivating this microbe and for its long-term storage and its genetic manipulation by transformation and conjugation. © 2020 Wiley Periodicals LLC. Basic Protocol 1: Growth of Y. pseudotuberculosis from a stock Basic Protocol 2: Growth of Y. pseudotuberculosis in liquid medium from a single colony Basic Protocol 3: Freezing Y. pseudotuberculosis in glycerol for long-term storage Basic Protocol 4: Transformation of Y. pseudotuberculosis by electroporation Basic Protocol 5: Tri-parental mating/conjugation.
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Affiliation(s)
- Robert K Davidson
- W. Harry Feinstone Department of Molecular Microbiology and Immunology, Johns Hopkins Bloomberg School of Public Health, Baltimore, Maryland
| | - Kimberly M Davis
- W. Harry Feinstone Department of Molecular Microbiology and Immunology, Johns Hopkins Bloomberg School of Public Health, Baltimore, Maryland
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11
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RIPK1 activates distinct gasdermins in macrophages and neutrophils upon pathogen blockade of innate immune signaling. Proc Natl Acad Sci U S A 2021; 118:2101189118. [PMID: 34260403 DOI: 10.1073/pnas.2101189118] [Citation(s) in RCA: 61] [Impact Index Per Article: 15.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/30/2022] Open
Abstract
Injection of effector proteins to block host innate immune signaling is a common strategy used by many pathogenic organisms to establish an infection. For example, pathogenic Yersinia species inject the acetyltransferase YopJ into target cells to inhibit NF-κB and MAPK signaling. To counteract this, detection of YopJ activity in myeloid cells promotes the assembly of a RIPK1-caspase-8 death-inducing platform that confers antibacterial defense. While recent studies revealed that caspase-8 cleaves the pore-forming protein gasdermin D to trigger pyroptosis in macrophages, whether RIPK1 activates additional substrates downstream of caspase-8 to promote host defense is unclear. Here, we report that the related gasdermin family member gasdermin E (GSDME) is activated upon detection of YopJ activity in a RIPK1 kinase-dependent manner. Specifically, GSDME promotes neutrophil pyroptosis and IL-1β release, which is critical for anti-Yersinia defense. During in vivo infection, IL-1β neutralization increases bacterial burden in wild-type but not Gsdme-deficient mice. Thus, our study establishes GSDME as an important mediator that counteracts pathogen blockade of innate immune signaling.
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12
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Redundant and Cooperative Roles for Yersinia pestis Yop Effectors in the Inhibition of Human Neutrophil Exocytic Responses Revealed by Gain-of-Function Approach. Infect Immun 2020; 88:IAI.00909-19. [PMID: 31871100 DOI: 10.1128/iai.00909-19] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/03/2019] [Accepted: 12/16/2019] [Indexed: 12/13/2022] Open
Abstract
Yersinia pestis causes a rapid, lethal disease referred to as plague. Y. pestis actively inhibits the innate immune system to generate a noninflammatory environment during early stages of infection to promote colonization. The ability of Y. pestis to create this early noninflammatory environment is in part due to the action of seven Yop effector proteins that are directly injected into host cells via a type 3 secretion system (T3SS). While each Yop effector interacts with specific host proteins to inhibit their function, several Yop effectors either target the same host protein or inhibit converging signaling pathways, leading to functional redundancy. Previous work established that Y. pestis uses the T3SS to inhibit neutrophil respiratory burst, phagocytosis, and release of inflammatory cytokines. Here, we show that Y. pestis also inhibits release of granules in a T3SS-dependent manner. Moreover, using a gain-of-function approach, we discovered previously hidden contributions of YpkA and YopJ to inhibition and that cooperative actions by multiple Yop effectors are required to effectively inhibit degranulation. Independent from degranulation, we also show that multiple Yop effectors can inhibit synthesis of leukotriene B4 (LTB4), a potent lipid mediator released by neutrophils early during infection to promote inflammation. Together, inhibition of these two arms of the neutrophil response likely contributes to the noninflammatory environment needed for Y. pestis colonization and proliferation.
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Eichelberger KR, Jones GS, Goldman WE. Inhibition of Neutrophil Primary Granule Release during Yersinia pestis Pulmonary Infection. mBio 2019; 10:e02759-19. [PMID: 31822588 PMCID: PMC6904878 DOI: 10.1128/mbio.02759-19] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/16/2019] [Accepted: 10/21/2019] [Indexed: 12/22/2022] Open
Abstract
Inhalation of Yersinia pestis causes primary pneumonic plague, the most severe manifestation of plague that is characterized by a dramatic neutrophil influx to the lungs. Neutrophils are ineffective during primary pneumonic plague, failing to control Y. pestis growth in the airways. However, the mechanisms by which Y. pestis resists neutrophil killing are incompletely understood. Here, we show that Y. pestis inhibits neutrophil degranulation, an important line of host innate immune defense. We observed that neutrophils from the lungs of mice infected intranasally with Y. pestis fail to release primary granules throughout the course of disease. Using a type III secretion system (T3SS) injection reporter strain, we determined that Y. pestis directly inhibits neutrophil granule release by a T3SS-dependent mechanism. Combinatorial mutant analysis revealed that a Y. pestis strain lacking both effectors YopE and YopH did not inhibit primary granule release and is killed by neutrophils both in vivo and in vitro Similarly, Y. pestis strains injecting only YopE or YopH are able to inhibit the majority of primary granule release from human neutrophils. We determined that YopE and YopH block Rac2 activation and calcium flux, respectively, to inhibit neutrophil primary granule release in isolated human neutrophils. These results demonstrate that Y. pestis coordinates the inhibition of neutrophil primary granule release through the activities of two distinct effectors, and this inhibition promotes Y. pestis survival during primary pneumonic plague.IMPORTANCEYersinia pestis is the causative agent of plague and is one of the deadliest human pathogens. The pneumonic form of Y. pestis infection has played a critical role in the severity of both historical and modern plague outbreaks, yet the host-pathogen interactions that govern the lethality of Yersinia pestis pulmonary infections are incompletely understood. Here, we report that Yersinia pestis inhibits neutrophil degranulation during infection, rendering neutrophils ineffective and allowing unrestricted growth of Y. pestis in the lungs. This coordinated inhibition of granule release not only demonstrates the pathogenic benefit of "silencing" lung neutrophils but also reveals specific host processes and pathways that could be manipulated to reduce the severity of primary pneumonic plague.
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Affiliation(s)
- Kara R Eichelberger
- Department of Microbiology and Immunology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA
| | - Grant S Jones
- Department of Microbiology and Immunology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA
| | - William E Goldman
- Department of Microbiology and Immunology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA
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Li J, Yang Y, Iqbal A, Qadri R, Shi P, Wang Y, Wu Y, Fan H, Wu G. Correlation analysis of cold-related gene expression with physiological and biochemical indicators under cold stress in oil palm. PLoS One 2019; 14:e0225768. [PMID: 31774880 PMCID: PMC6881061 DOI: 10.1371/journal.pone.0225768] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/21/2019] [Accepted: 11/12/2019] [Indexed: 11/19/2022] Open
Abstract
Oil palm (Elaeis guineensis Jacq.) is a representative tropical oil crop that is sensitive to low temperature. Oil palm can experience cold damage when exposed to low temperatures for a long period. During these unfavorable conditions, a series of gene induction/repression and physico-chemical changes occur in oil palm. To better understand the link between these events, we investigated the expression levels of various genes (including COR410, COR413, CBF1, CBF2, CBF3, ICE1-1, ICE1-2, ICE1-4, SIZ1-1, SIZ1-2, ZAT10, ZAT12) and the accumulation of osmolytes (proline, malondialdehyde and sucrose). Likewise, the activity of superoxide dismutase (SOD) in oil palm under cold stress (4°C, 8°C and 12°C) was examined. The results showed a clear link among the expression of CBFs (especially CBF1 and CBF3) and the all genes examined under cold stress (12°C). The expression of CBF1 and CBF2 also exhibited a positive link with the accumulation of sucrose and proline under cold stress in oil palm. At 4°C, the proline content exhibited a very significant correlation with electrolyte leakage in oil palm. The results of this study provide necessary information regarding the mechanism of the response and adaption of oil palm to cold stress. Additionally, they offer clues for the selection or development of cold-tolerant cultivars from the available germplasms of oil palm.
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Affiliation(s)
- Jing Li
- Hainan Key Laboratory of Tropical Oil Crops Biology/Coconut Research Institute, Chinese Academy of Tropical Agricultural Sciences, Wenchang, Hainan, China
| | - Yaodong Yang
- Hainan Key Laboratory of Tropical Oil Crops Biology/Coconut Research Institute, Chinese Academy of Tropical Agricultural Sciences, Wenchang, Hainan, China
- * E-mail:
| | - Amjad Iqbal
- Hainan Key Laboratory of Tropical Oil Crops Biology/Coconut Research Institute, Chinese Academy of Tropical Agricultural Sciences, Wenchang, Hainan, China
| | - Rashad Qadri
- Hainan Key Laboratory of Tropical Oil Crops Biology/Coconut Research Institute, Chinese Academy of Tropical Agricultural Sciences, Wenchang, Hainan, China
| | - Peng Shi
- Hainan Key Laboratory of Tropical Oil Crops Biology/Coconut Research Institute, Chinese Academy of Tropical Agricultural Sciences, Wenchang, Hainan, China
| | - Yong Wang
- Hainan Key Laboratory of Tropical Oil Crops Biology/Coconut Research Institute, Chinese Academy of Tropical Agricultural Sciences, Wenchang, Hainan, China
| | - Yi Wu
- Hainan Key Laboratory of Tropical Oil Crops Biology/Coconut Research Institute, Chinese Academy of Tropical Agricultural Sciences, Wenchang, Hainan, China
| | - Haikuo Fan
- Hainan Key Laboratory of Tropical Oil Crops Biology/Coconut Research Institute, Chinese Academy of Tropical Agricultural Sciences, Wenchang, Hainan, China
| | - Guojiang Wu
- Key Laboratory of Plant Resource Conservation and Sustainable Utilization, South China Botanical Garden, Chinese Academy of Sciences, Guangzhou, China
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15
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Phenotypic Diversification of Microbial Pathogens—Cooperating and Preparing for the Future. J Mol Biol 2019; 431:4645-4655. [DOI: 10.1016/j.jmb.2019.06.024] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/17/2019] [Revised: 06/20/2019] [Accepted: 06/21/2019] [Indexed: 12/22/2022]
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Abstract
The human and animal pathogens Yersinia pestis, which causes bubonic and pneumonic plague, and Yersinia pseudotuberculosis and Yersinia enterocolitica, which cause gastroenteritis, share a type 3 secretion system which injects effector proteins, Yops, into host cells. This system is critical for virulence of all three pathogens in tissue infection. Neutrophils are rapidly recruited to infected sites and all three pathogens frequently interact with and inject Yops into these cells during tissue infection. Host receptors, serum factors, and bacterial adhesins appear to collaborate to promote neutrophil- Yersinia interactions in tissues. The ability of neutrophils to control infection is mixed depending on the stage of infection and points to the efficiency of Yops and other bacterial factors to mitigate bactericidal effects of neutrophils. Yersinia in close proximity to neutrophils has higher levels of expression from yop promoters, and neutrophils in close proximity to Yersinia express higher levels of pro-survival genes than migrating neutrophils. In infected tissues, YopM increases neutrophil survival and YopH targets a SKAP2/SLP-76 signal transduction pathway. Yet the full impact of these and other Yops and other Yersinia factors on neutrophils in infected tissues has yet to be understood.
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Affiliation(s)
- Joan Mecsas
- Department of Molecular Biology and Microbiology, 136 Harrison Ave, Tufts University School of Medicine, Boston, MA, 02111, USA
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17
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Davis KM, Isberg RR. One for All, but Not All for One: Social Behavior during Bacterial Diseases. Trends Microbiol 2018; 27:64-74. [PMID: 30243514 DOI: 10.1016/j.tim.2018.09.001] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/02/2018] [Revised: 08/26/2018] [Accepted: 09/05/2018] [Indexed: 02/07/2023]
Abstract
It has been known for decades that individual cells within pathogenic bacterial populations have reduced antibiotic susceptibility, which is linked to decreased metabolic rates. A similar phenomenon occurs with virulence-associated proteins, as reduced expression is associated with increased fitness of individual cells. Non-producers within the population can benefit from the virulence proteins produced by others in the population without suffering a fitness cost, thus maintaining a genetically uniform population. Cooperative behavior has been reported for Salmonella and Yersinia, consistent with selection of social behavior to retain genes associated with pathogenesis; however, cooperation was unclear within Mycobacterium populations. This review focuses on these recent descriptions of cooperation, discusses the mechanisms driving heterogeneity, and evaluates the evidence that expression of virulence-associated proteins comes at a fitness cost.
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Affiliation(s)
- Kimberly M Davis
- W. Harry Feinstone Department of Molecular Microbiology and Immunology, Johns Hopkins Bloomberg School of Public Health, 615 N. Wolfe St., Baltimore, MD 21205, USA
| | - Ralph R Isberg
- Department of Molecular Biology and Microbiology, Tufts University School of Medicine, 150 Harrison Ave., Boston, MA 02111, USA.
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18
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Westerman TL, Bogomolnaya L, Andrews-Polymenis HL, Sheats MK, Elfenbein JR. The Salmonella type-3 secretion system-1 and flagellar motility influence the neutrophil respiratory burst. PLoS One 2018; 13:e0203698. [PMID: 30204776 PMCID: PMC6133356 DOI: 10.1371/journal.pone.0203698] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/15/2018] [Accepted: 08/24/2018] [Indexed: 11/28/2022] Open
Abstract
Neutrophils are innate immune response cells designed to kill invading microorganisms. One of the mechanisms neutrophils use to kill bacteria is generation of damaging reactive oxygen species (ROS) via the respiratory burst. However, during enteric salmonellosis, neutrophil-derived ROS actually facilitates Salmonella expansion and survival in the gut. This seeming paradox led us to hypothesize that Salmonella may possess mechanisms to influence the neutrophil respiratory burst. In this work, we used an in vitro Salmonella-neutrophil co-culture model to examine the impact of enteric infection relevant virulence factors on the respiratory burst of human neutrophils. We report that neutrophils primed with granulocyte-macrophage colony stimulating factor and suspended in serum containing complement produce a robust respiratory burst when stimulated with viable STm. The magnitude of the respiratory burst increases when STm are grown under conditions to induce the expression of the type-3 secretion system-1. STm mutants lacking the type-3 secretion system-1 induce less neutrophil ROS than the virulent WT. In addition, we demonstrate that flagellar motility is a significant agonist of the neutrophil respiratory burst. Together our data demonstrate that both the type-3 secretion system-1 and flagellar motility, which are established virulence factors in enteric salmonellosis, also appear to directly influence the magnitude of the neutrophil respiratory burst in response to STm in vitro.
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Affiliation(s)
- Trina L. Westerman
- Department of Clinical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC, United States of America
| | - Lydia Bogomolnaya
- Department of Microbial Pathogenesis and Immunology, College of Medicine, Texas A&M University, Bryan, TX, United States of America
- Institute of Fundamental Medicine and Biology, Kazan Federal University, Kazan, Russia
| | - Helene L. Andrews-Polymenis
- Department of Microbial Pathogenesis and Immunology, College of Medicine, Texas A&M University, Bryan, TX, United States of America
| | - M. Katherine Sheats
- Department of Clinical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC, United States of America
| | - Johanna R. Elfenbein
- Department of Clinical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC, United States of America
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Kang E, Crouse A, Chevallier L, Pontier SM, Alzahrani A, Silué N, Campbell-Valois FX, Montagutelli X, Gruenheid S, Malo D. Enterobacteria and host resistance to infection. Mamm Genome 2018; 29:558-576. [PMID: 29785663 DOI: 10.1007/s00335-018-9749-4] [Citation(s) in RCA: 25] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/16/2018] [Accepted: 05/14/2018] [Indexed: 02/06/2023]
Abstract
Enterobacteriaceae are a large family of Gram-negative, non-spore-forming bacteria. Although many species exist as part of the natural flora of animals including humans, some members are associated with both intestinal and extraintestinal diseases. In this review, we focus on members of this family that have important roles in human disease: Salmonella, Escherichia, Shigella, and Yersinia, providing a brief overview of the disease caused by these bacteria, highlighting the contribution of animal models to our understanding of their pathogenesis and of host genetic determinants involved in susceptibility or resistance to infection.
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Affiliation(s)
- Eugene Kang
- Department of Microbiology and Immunology, McGill University, Montreal, QC, Canada
- McGill Research Center on Complex Traits, McGill University, Montreal, QC, Canada
| | - Alanna Crouse
- McGill Research Center on Complex Traits, McGill University, Montreal, QC, Canada
- Department of Human Genetics, McGill University, Montreal, QC, Canada
| | - Lucie Chevallier
- U955 - IMRB, Team 10 - Biology of the neuromuscular system, Inserm, École Nationale Vétérinaire d'Alfort, UPEC, Maisons-Alfort, France
- Mouse Genetics Laboratory, Department of Genomes and Genetics, Institut Pasteur, Paris, France
| | - Stéphanie M Pontier
- Department of Chemistry and Biomolecular Sciences, Centre for Chemical and Synthetic Biology, University of Ottawa, Ottawa, ON, Canada
| | - Ashwag Alzahrani
- Department of Chemistry and Biomolecular Sciences, Centre for Chemical and Synthetic Biology, University of Ottawa, Ottawa, ON, Canada
| | - Navoun Silué
- Department of Chemistry and Biomolecular Sciences, Centre for Chemical and Synthetic Biology, University of Ottawa, Ottawa, ON, Canada
| | - François-Xavier Campbell-Valois
- Department of Chemistry and Biomolecular Sciences, Centre for Chemical and Synthetic Biology, University of Ottawa, Ottawa, ON, Canada
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
| | - Xavier Montagutelli
- U955 - IMRB, Team 10 - Biology of the neuromuscular system, Inserm, École Nationale Vétérinaire d'Alfort, UPEC, Maisons-Alfort, France
| | - Samantha Gruenheid
- Department of Microbiology and Immunology, McGill University, Montreal, QC, Canada
- McGill Research Center on Complex Traits, McGill University, Montreal, QC, Canada
| | - Danielle Malo
- McGill Research Center on Complex Traits, McGill University, Montreal, QC, Canada.
- Department of Human Genetics, McGill University, Montreal, QC, Canada.
- Department of Medicine, McGill University, Montreal, QC, Canada.
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20
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Carneiro MBH, Roma EH, Ranson AJ, Doria NA, Debrabant A, Sacks DL, Vieira LQ, Peters NC. NOX2-Derived Reactive Oxygen Species Control Inflammation during Leishmania amazonensis Infection by Mediating Infection-Induced Neutrophil Apoptosis. THE JOURNAL OF IMMUNOLOGY 2017; 200:196-208. [PMID: 29158417 DOI: 10.4049/jimmunol.1700899] [Citation(s) in RCA: 36] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 06/20/2017] [Accepted: 10/23/2017] [Indexed: 12/25/2022]
Abstract
Reactive oxygen species (ROS) produced by NADPH phagocyte oxidase isoform (NOX2) are critical for the elimination of intracellular pathogens in many infections. Despite their importance, the role of ROS following infection with the eukaryotic pathogen Leishmania has not been fully elucidated. We addressed the role of ROS in C57BL/6 mice following intradermal infection with Leishmania amazonensis. Despite equivalent parasite loads compared with wild-type (WT) mice, mice deficient in ROS production by NOX2 due to the absence of the gp91 subunit (gp91phox-/-) had significantly more severe pathology in the later stages of infection. Pathology in gp91phox-/- mice was not associated with alterations in CD4+ T cell-mediated immunity but was preceded by enhanced neutrophil accumulation at the dermal infection site. Ex vivo analysis of infected versus uninfected neutrophils revealed a deficiency in infection-driven apoptosis in gp91phox-/- mice versus WT mice. gp91phox-/- mice presented with higher percentages of healthy or necrotic neutrophils but lower percentages of apoptotic neutrophils at early and chronic time points. In vitro infection of gp91phox-/- versus WT neutrophils also revealed reduced apoptosis and CD95 expression but increased necrosis in infected cells at 10 h postinfection. Provision of exogenous ROS in the form of H2O2 reversed the necrotic phenotype and restored CD95 expression on infected gp91phox-/- neutrophils. Although ROS production is typically viewed as a proinflammatory event, our observations identify the importance of ROS in mediating appropriate neutrophil apoptosis and the importance of apoptosis in inflammation and pathology during chronic infection.
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Affiliation(s)
- Matheus B H Carneiro
- Snyder Institute for Chronic Diseases, Department of Microbiology, Immunology and Infectious Diseases, Cumming School of Medicine, University of Calgary, Calgary, Alberta T2N 4Z6, Canada.,Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais 31270-901, Brazil
| | - Eric H Roma
- Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais 31270-901, Brazil.,Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20852; and
| | - Adam J Ranson
- Snyder Institute for Chronic Diseases, Department of Microbiology, Immunology and Infectious Diseases, Cumming School of Medicine, University of Calgary, Calgary, Alberta T2N 4Z6, Canada
| | - Nicole A Doria
- Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20852; and
| | - Alain Debrabant
- Division of Emerging and Transfusion Transmitted Diseases, Office of Blood Research and Review, Center for Biologics Evaluation and Research, U.S. Food and Drug Administration, Bethesda, MD 20993
| | - David L Sacks
- Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20852; and
| | - Leda Q Vieira
- Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais 31270-901, Brazil
| | - Nathan C Peters
- Snyder Institute for Chronic Diseases, Department of Microbiology, Immunology and Infectious Diseases, Cumming School of Medicine, University of Calgary, Calgary, Alberta T2N 4Z6, Canada;
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Herrera Estrada L, Wu H, Ling K, Zhang G, Sumagin R, Parkos CA, Jones RM, Champion JA, Neish AS. Bioengineering Bacterially Derived Immunomodulants: A Therapeutic Approach to Inflammatory Bowel Disease. ACS NANO 2017; 11:9650-9662. [PMID: 28872828 PMCID: PMC7653663 DOI: 10.1021/acsnano.7b03239] [Citation(s) in RCA: 24] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/10/2023]
Abstract
Bacterial enteric pathogens have evolved efficient mechanisms to suppress mammalian inflammatory and immunoregulatory pathways. By exploiting the evolutionary relationship between the gut and pathogenic bacteria, we have developed a potential mucosal therapeutic. Our findings suggest that engineered preparations of the Salmonella acetyltransferase, AvrA, suppress acute inflammatory responses such as those observed in inflammatory bowel disease (IBD). We created 125 nm diameter cross-linked protein nanoparticles directly from AvrA and carrier protein to deliver AvrA in the absence of Salmonella. AvrA nanoparticles are internalized in vitro and in vivo into barrier epithelial and lamina propria monocytic cells. AvrA nanoparticles inhibit inflammatory signaling and confer cytoprotection in vitro, and in murine colitis models, we observe decreased clinical and histological indices of inflammation. Thus, we have combined naturally evolved immunomodulatory proteins with modern bioengineering to produce AvrA nanoparticles, a potential treatment for IBD.
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Affiliation(s)
- Lina Herrera Estrada
- School of Chemical & Biomolecular Engineering, Georgia Institute of Technology, 950 Atlantic Drive NW, Atlanta, Georgia 30332, United States
| | - Huixia Wu
- Department of Pathology, Emory University School of Medicine, Whitehead Bldg., 615 Michael Street, Atlanta, Georgia 30322, United States
| | - Kevin Ling
- School of Chemical & Biomolecular Engineering, Georgia Institute of Technology, 950 Atlantic Drive NW, Atlanta, Georgia 30332, United States
| | - Guikai Zhang
- Department of Pathology, Emory University School of Medicine, Whitehead Bldg., 615 Michael Street, Atlanta, Georgia 30322, United States
| | - Ronen Sumagin
- Department of Pathology, Emory University School of Medicine, Whitehead Bldg., 615 Michael Street, Atlanta, Georgia 30322, United States
| | - Charles A. Parkos
- Department of Pathology, Emory University School of Medicine, Whitehead Bldg., 615 Michael Street, Atlanta, Georgia 30322, United States
| | - Rheinallt M. Jones
- Department of Pathology, Emory University School of Medicine, Whitehead Bldg., 615 Michael Street, Atlanta, Georgia 30322, United States
| | - Julie A. Champion
- School of Chemical & Biomolecular Engineering, Georgia Institute of Technology, 950 Atlantic Drive NW, Atlanta, Georgia 30332, United States
- Corresponding Authors: Phone: 404-894-2874. . Phone: 404-727-8545.
| | - Andrew S. Neish
- Department of Pathology, Emory University School of Medicine, Whitehead Bldg., 615 Michael Street, Atlanta, Georgia 30322, United States
- Corresponding Authors: Phone: 404-894-2874. . Phone: 404-727-8545.
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22
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Grabowski B, Schmidt MA, Rüter C. Immunomodulatory Yersinia outer proteins (Yops)-useful tools for bacteria and humans alike. Virulence 2017; 8:1124-1147. [PMID: 28296562 DOI: 10.1080/21505594.2017.1303588] [Citation(s) in RCA: 26] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
Human-pathogenic Yersinia produce plasmid-encoded Yersinia outer proteins (Yops), which are necessary to down-regulate anti-bacterial responses that constrict bacterial survival in the host. These Yops are effectively translocated directly from the bacterial into the target cell cytosol by the type III secretion system (T3SS). Cell-penetrating peptides (CPPs) in contrast are characterized by their ability to autonomously cross cell membranes and to transport cargo - independent of additional translocation systems. The recent discovery of bacterial cell-penetrating effector proteins (CPEs) - with the prototype being the T3SS effector protein YopM - established a new class of autonomously translocating immunomodulatory proteins. CPEs represent a vast source of potential self-delivering, anti-inflammatory therapeutics. In this review, we give an update on the characteristic features of the plasmid-encoded Yops and, based on recent findings, propose the further development of these proteins for potential therapeutic applications as natural or artificial cell-penetrating forms of Yops might be of value as bacteria-derived biologics.
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Affiliation(s)
- Benjamin Grabowski
- a Institute of Infectiology - Centre for Molecular Biology of Inflammation (ZMBE), University of Münster , Münster , Germany
| | - M Alexander Schmidt
- a Institute of Infectiology - Centre for Molecular Biology of Inflammation (ZMBE), University of Münster , Münster , Germany
| | - Christian Rüter
- a Institute of Infectiology - Centre for Molecular Biology of Inflammation (ZMBE), University of Münster , Münster , Germany
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23
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Delgado-Rizo V, Martínez-Guzmán MA, Iñiguez-Gutierrez L, García-Orozco A, Alvarado-Navarro A, Fafutis-Morris M. Neutrophil Extracellular Traps and Its Implications in Inflammation: An Overview. Front Immunol 2017; 8:81. [PMID: 28220120 PMCID: PMC5292617 DOI: 10.3389/fimmu.2017.00081] [Citation(s) in RCA: 462] [Impact Index Per Article: 57.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/14/2016] [Accepted: 01/17/2017] [Indexed: 12/14/2022] Open
Abstract
In addition to physical barriers, neutrophils are considered a part of the first line of immune defense. They can be found in the bloodstream, with a lifespan of 6–8 h, and in tissue, where they can last up to 7 days. The mechanisms that neutrophils utilize for host defense are phagocytosis, degranulation, cytokine production, and, the most recently described, neutrophil extracellular trap (NET) production. NETs are DNA structures released due to chromatin decondensation and spreading, and they thus occupy three to five times the volume of condensed chromatin. Several proteins adhere to NETs, including histones and over 30 components of primary and secondary granules, among them components with bactericidal activity such as elastase, myeloperoxidase, cathepsin G, lactoferrin, pentraxin 3, gelatinase, proteinase 3, LL37, peptidoglycan-binding proteins, and others with bactericidal activity able to destroy virulence factors. Three models for NETosis are known to date. (a) Suicidal NETosis, with a duration of 2–4 h, is the best described model. (b) In vital NETosis with nuclear DNA release, neutrophils release NETs without exhibiting loss of nuclear or plasma membrane within 5–60 min, and it is independent of reactive oxygen species (ROS) and the Raf/MERK/ERK pathway. (c) The final type is vital NETosis with release of mitochondrial DNA that is dependent on ROS and produced after stimuli with GM-CSF and lipopolysaccharide. Recent research has revealed neutrophils as more sophisticated immune cells that are able to precisely regulate their granular enzymes release by ion fluxes and can release immunomodulatory cytokines and chemokines that interact with various components of the immune system. Therefore, they can play a key role in autoimmunity and in autoinflammatory and metabolic diseases. In this review, we intend to show the two roles played by neutrophils: as a first line of defense against microorganisms and as a contributor to the pathogenesis of various illnesses, such as autoimmune, autoinflammatory, and metabolic diseases.
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Shi G, Su M, Liang J, Duan R, Gu W, Xiao Y, Zhang Z, Qiu H, Zhang Z, Li Y, Zhang X, Ling Y, Song L, Chen M, Zhao Y, Wu J, Jing H, Xiao J, Wang X. Complete genome sequence and comparative genome analysis of a new special Yersinia enterocolitica. Arch Microbiol 2016; 198:673-87. [PMID: 27129539 DOI: 10.1007/s00203-016-1229-1] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/17/2015] [Revised: 04/15/2016] [Accepted: 04/25/2016] [Indexed: 10/21/2022]
Abstract
Yersinia enterocolitica is the most diverse species among the Yersinia genera and shows more polymorphism, especially for the non-pathogenic strains. Individual non-pathogenic Y. enterocolitica strains are wrongly identified because of atypical phenotypes. In this study, we isolated an unusual Y. enterocolitica strain LC20 from Rattus norvegicus. The strain did not utilize urea and could not be classified as the biotype. API 20E identified Escherichia coli; however, it grew well at 25 °C, but E. coli grew well at 37 °C. We analyzed the genome of LC20 and found the whole chromosome of LC20 was collinear with Y. enterocolitica 8081, and the urease gene did not exist on the genome which is consistent with the result of API 20E. Also, the 16 S and 23 SrRNA gene of LC20 lay on a branch of Y. enterocolitica. Furthermore, the core-based and pan-based phylogenetic trees showed that LC20 was classified into the Y. enterocolitica cluster. Two plasmids (80 and 50 k) from LC20 shared low genetic homology with pYV from the Yersinia genus, one was an ancestral Yersinia plasmid and the other was novel encoding a number of transposases. Some pathogenic and non-pathogenic Y. enterocolitica-specific genes coexisted in LC20. Thus, although it could not be classified into any Y. enterocolitica biotype due to its special biochemical metabolism, we concluded the LC20 was a Y. enterocolitica strain because its genome was similar to other Y. enterocolitica and it might be a strain with many mutations and combinations emerging in the processes of its evolution.
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Affiliation(s)
- Guoxiang Shi
- Zhejiang Provincial Centre for Disease Control and Prevention, Hangzhou, People's Republic of China
| | - Mingming Su
- CAS Key Laboratory of Genome Sciences and Information, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, People's Republic of China.,University of Chinese Academy of Sciences, Beijing, People's Republic of China
| | - Junrong Liang
- National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, State Key Laboratory of Infectious Disease Prevention and Control, 155 Changbai Road Changping District, Beiing, 102206, People's Republic of China.,Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Beijing, People's Republic of China
| | - Ran Duan
- National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, State Key Laboratory of Infectious Disease Prevention and Control, 155 Changbai Road Changping District, Beiing, 102206, People's Republic of China.,Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Beijing, People's Republic of China
| | - Wenpeng Gu
- Yunnan Provincial Centre for Disease Control and Prevention, Kunming, People's Republic of China
| | - Yuchun Xiao
- National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, State Key Laboratory of Infectious Disease Prevention and Control, 155 Changbai Road Changping District, Beiing, 102206, People's Republic of China.,Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Beijing, People's Republic of China
| | - Zhewen Zhang
- CAS Key Laboratory of Genome Sciences and Information, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, People's Republic of China
| | - Haiyan Qiu
- National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, State Key Laboratory of Infectious Disease Prevention and Control, 155 Changbai Road Changping District, Beiing, 102206, People's Republic of China.,Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Beijing, People's Republic of China
| | - Zheng Zhang
- Zhejiang Provincial Centre for Disease Control and Prevention, Hangzhou, People's Republic of China
| | - Yi Li
- Wenzhou Municipal Centre for Disease Control and Prevention, Wenzhou, People's Republic of China
| | - Xiaohe Zhang
- Wenzhou Municipal Centre for Disease Control and Prevention, Wenzhou, People's Republic of China
| | - Yunchao Ling
- CAS Key Laboratory of Genome Sciences and Information, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, People's Republic of China.,University of Chinese Academy of Sciences, Beijing, People's Republic of China
| | - Lai Song
- CAS Key Laboratory of Genome Sciences and Information, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, People's Republic of China.,University of Chinese Academy of Sciences, Beijing, People's Republic of China
| | - Meili Chen
- CAS Key Laboratory of Genome Sciences and Information, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, People's Republic of China.,University of Chinese Academy of Sciences, Beijing, People's Republic of China
| | - Yongbing Zhao
- CAS Key Laboratory of Genome Sciences and Information, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, People's Republic of China.,University of Chinese Academy of Sciences, Beijing, People's Republic of China
| | - Jiayan Wu
- CAS Key Laboratory of Genome Sciences and Information, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, People's Republic of China
| | - Huaiqi Jing
- National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, State Key Laboratory of Infectious Disease Prevention and Control, 155 Changbai Road Changping District, Beiing, 102206, People's Republic of China.,Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Beijing, People's Republic of China
| | - Jingfa Xiao
- CAS Key Laboratory of Genome Sciences and Information, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, People's Republic of China
| | - Xin Wang
- National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, State Key Laboratory of Infectious Disease Prevention and Control, 155 Changbai Road Changping District, Beiing, 102206, People's Republic of China. .,Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Beijing, People's Republic of China.
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Martínez-Chavarría LC. Yersinia pestis-Host Immune Cells Interactions at Early Events During Bubonic Plague Infection. CURRENT TROPICAL MEDICINE REPORTS 2016. [DOI: 10.1007/s40475-016-0071-5] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/22/2022]
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26
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Ye Q, Wu Q, Hu H, Zhang J, Huang H. Prevalence and characterization of Yersinia enterocolitica isolated from retail foods in China. Food Control 2016. [DOI: 10.1016/j.foodcont.2015.09.016] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/27/2022]
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27
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Pha K, Navarro L. Yersinia type III effectors perturb host innate immune responses. World J Biol Chem 2016; 7:1-13. [PMID: 26981193 PMCID: PMC4768113 DOI: 10.4331/wjbc.v7.i1.1] [Citation(s) in RCA: 32] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/25/2015] [Revised: 09/02/2015] [Accepted: 11/04/2015] [Indexed: 02/05/2023] Open
Abstract
The innate immune system is the first line of defense against invading pathogens. Innate immune cells recognize molecular patterns from the pathogen and mount a response to resolve the infection. The production of proinflammatory cytokines and reactive oxygen species, phagocytosis, and induced programmed cell death are processes initiated by innate immune cells in order to combat invading pathogens. However, pathogens have evolved various virulence mechanisms to subvert these responses. One strategy utilized by Gram-negative bacterial pathogens is the deployment of a complex machine termed the type III secretion system (T3SS). The T3SS is composed of a syringe-like needle structure and the effector proteins that are injected directly into a target host cell to disrupt a cellular response. The three human pathogenic Yersinia spp. (Y. pestis, Y. enterocolitica, and Y. pseudotuberculosis) are Gram-negative bacteria that share in common a 70 kb virulence plasmid which encodes the T3SS. Translocation of the Yersinia effector proteins (YopE, YopH, YopT, YopM, YpkA/YopO, and YopP/J) into the target host cell results in disruption of the actin cytoskeleton to inhibit phagocytosis, downregulation of proinflammatory cytokine/chemokine production, and induction of cellular apoptosis of the target cell. Over the past 25 years, studies on the Yersinia effector proteins have unveiled tremendous knowledge of how the effectors enhance Yersinia virulence. Recently, the long awaited crystal structure of YpkA has been solved providing further insights into the activation of the YpkA kinase domain. Multisite autophosphorylation by YpkA to activate its kinase domain was also shown and postulated to serve as a mechanism to bypass regulation by host phosphatases. In addition, novel Yersinia effector protein targets, such as caspase-1, and signaling pathways including activation of the inflammasome were identified. In this review, we summarize the recent discoveries made on Yersinia effector proteins and their contribution to Yersinia pathogenesis.
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28
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Herrera Estrada L, Padmore TJ, Champion JA. Bacterial Effector Nanoparticles as Breast Cancer Therapeutics. Mol Pharm 2016; 13:710-9. [DOI: 10.1021/acs.molpharmaceut.5b00377] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022]
Affiliation(s)
- Lina Herrera Estrada
- Department of Chemical & Biomolecular Engineering, Georgia Institute of Technology, 950 Atlantic Drive NW, Atlanta, Georgia 30332, United States
| | - Trudy J. Padmore
- Department of Chemical & Biomolecular Engineering, Georgia Institute of Technology, 950 Atlantic Drive NW, Atlanta, Georgia 30332, United States
| | - Julie A. Champion
- Department of Chemical & Biomolecular Engineering, Georgia Institute of Technology, 950 Atlantic Drive NW, Atlanta, Georgia 30332, United States
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Abstract
During pneumonic plague, the bacterium Yersinia pestis elicits the development of inflammatory lung lesions that continue to expand throughout infection. This lesion development and persistence are poorly understood. Here, we examine spatially distinct regions of lung lesions using laser capture microdissection and transcriptome sequencing (RNA-seq) analysis to identify transcriptional differences between lesion microenvironments. We show that cellular pathways involved in leukocyte migration and apoptosis are downregulated in the center of lung lesions compared to the periphery. Probing for the bacterial factor(s) important for the alteration in neutrophil survival, we show both in vitro and in vivo that Y. pestis increases neutrophil survival in a manner that is dependent on the type III secretion system effector YopM. This research explores the complexity of spatially distinct host-microbe interactions and emphasizes the importance of cell relevance in assays in order to fully understand Y. pestis virulence. Yersinia pestis is a high-priority pathogen and continues to cause outbreaks worldwide. The ability of Y. pestis to be transmitted via respiratory droplets and its history of weaponization has led to its classification as a select agent most likely to be used as a biological weapon. Unrestricted bacterial growth during the initial preinflammatory phase primes patients to be infectious once disease symptoms begin in the proinflammatory phase, and the rapid disease progression can lead to death before Y. pestis infection can be diagnosed and treated. Using in vivo analyses and focusing on relevant cell types during pneumonic plague infection, we can identify host pathways that may be manipulated to extend the treatment window for pneumonic plague patients.
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30
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Spinner JL, Hasenkrug AM, Shannon JG, Kobayashi SD, Hinnebusch BJ. Role of the Yersinia YopJ protein in suppressing interleukin-8 secretion by human polymorphonuclear leukocytes. Microbes Infect 2015; 18:21-9. [PMID: 26361732 DOI: 10.1016/j.micinf.2015.08.015] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/24/2015] [Revised: 08/20/2015] [Accepted: 08/31/2015] [Indexed: 12/19/2022]
Abstract
Polymorphonuclear leukocytes, in addition to their direct bactericidal activities, produce cytokines involved in the activation and regulation of the innate and adaptive immune response to infection. In this study we evaluated the cytokine response of human PMNs following incubation with the pathogenic Yersinia species. Yersinia pestis strains with the pCD1 virulence plasmid, which encodes cytotoxic Yop proteins that are translocated into host cells, stimulated little or no cytokine production compared to pCD1-negative strains. In particular, PMNs incubated with pCD1-negative Y. pestis secreted 1000-fold higher levels of interleukin-8 (IL-8 or CXCL8), a proinflammatory chemokine important for PMN recruitment and activation. Deletion of yopE, -H, -T, -M or ypkA had no effect on pCD1-dependent inhibition, whereas deletion of yopJ resulted in significantly increased IL-8 production. Like Y. pestis, the enteropathogenic Yersinia species inhibited IL-8 secretion by PMNs, and strains lacking the virulence plasmid induced high levels of IL-8. Our results show that virulence plasmid-encoded effector Yops, particularly YopJ, prevent IL-8 secretion by human PMNs. Suppression of the chemotactic IL-8 response by Y. pestis may contribute to the delayed PMN recruitment to the infected lymph node that typifies bubonic plague.
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Affiliation(s)
- Justin L Spinner
- Laboratory of Zoonotic Pathogens, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 905 S. 4th St., Hamilton, Montana 59840, USA
| | - Aaron M Hasenkrug
- Laboratory of Zoonotic Pathogens, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 905 S. 4th St., Hamilton, Montana 59840, USA
| | - Jeffrey G Shannon
- Laboratory of Zoonotic Pathogens, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 905 S. 4th St., Hamilton, Montana 59840, USA
| | - Scott D Kobayashi
- Laboratory of Human Bacterial Pathogenesis, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 905 S. 4th St., Hamilton, Montana 59840, USA
| | - B Joseph Hinnebusch
- Laboratory of Zoonotic Pathogens, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 905 S. 4th St., Hamilton, Montana 59840, USA.
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31
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Community behavior and spatial regulation within a bacterial microcolony in deep tissue sites serves to protect against host attack. Cell Host Microbe 2014; 17:21-31. [PMID: 25500192 DOI: 10.1016/j.chom.2014.11.008] [Citation(s) in RCA: 99] [Impact Index Per Article: 9.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/23/2014] [Revised: 09/09/2014] [Accepted: 10/27/2014] [Indexed: 12/20/2022]
Abstract
Bacterial pathogens express virulence-specific transcriptional programs that allow tissue colonization. Although phenotypic variation has been noted in the context of antibiotic exposure, no direct evidence exists for heterogeneity in virulence-specific transcriptional programs within tissues. In a mouse model of Yersinia pseudotuberculosis infection, we show that at least three subpopulations of bacteria develop within a single tissue site in response to distinct host signals. Bacteria growing on the exterior of spleen microcolonies responded to soluble signals and induced the nitric oxide (NO)-detoxifying gene, hmp. Hmp effectively eliminated NO diffusion and protected the interior bacterial population from exposure to NO-derived inducing signals. A third subpopulation, constituting the most peripherally localized bacteria, directly contacted neutrophils and transcriptionally upregulated a virulence factor. These studies demonstrate that growth within tissues results in transcriptional specialization within a single focus of microbial replication, facilitating directed pathogen counterattack against the host response.
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32
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Ye Z, Gorman AA, Uittenbogaard AM, Myers-Morales T, Kaplan AM, Cohen DA, Straley SC. Caspase-3 mediates the pathogenic effect of Yersinia pestis YopM in liver of C57BL/6 mice and contributes to YopM's function in spleen. PLoS One 2014; 9:e110956. [PMID: 25372388 PMCID: PMC4220956 DOI: 10.1371/journal.pone.0110956] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/07/2013] [Accepted: 09/26/2014] [Indexed: 12/15/2022] Open
Abstract
The virulence protein YopM of the plague bacterium Yersinia pestis has different dominant effects in liver and spleen. Previous studies focused on spleen, where YopM inhibits accumulation of inflammatory dendritic cells. In the present study we focused on liver, where PMN function may be directly undermined by YopM without changes in inflammatory cell numbers in the initial days of infection, and foci of inflammation are easily identified. Mice were infected with parent and ΔyopM-1 Y. pestis KIM5, and effects of YopM were assessed by immunohistochemistry and determinations of bacterial viable numbers in organs. The bacteria were found associated with myeloid cells in foci of inflammation and in liver sinusoids. A new in-vivo phenotype of YopM was revealed: death of inflammatory cells, evidenced by TUNEL staining beginning at d 1 of infection. Based on distributions of Ly6G+, F4/80+, and iNOS+ cells within foci, the cells that were killed could have included both PMNs and macrophages. By 2 d post-infection, YopM had no effect on distribution of these cells, but by 3 d cellular decomposition had outstripped acute inflammation in foci due to parent Y. pestis, while foci due to the ΔyopM-1 strain still contained many inflammatory cells. The destruction depended on the presence of both PMNs in the mice and YopM in the bacteria. In mice that lacked the apoptosis mediator caspase-3 the infection dynamics were novel: the parent Y. pestis was limited in growth comparably to the ΔyopM-1 strain in liver, and in spleen a partial growth limitation for parent Y. pestis was seen. This result identified caspase-3 as a co-factor or effector in YopM's action and supports the hypothesis that in liver YopM's main pathogenic effect is mediated by caspase-3 to cause apoptosis of PMNs.
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Affiliation(s)
- Zhan Ye
- Department of Microbiology, Immunology, and Molecular Genetics, University of Kentucky, Lexington, KY, United States of America
| | - Amanda A. Gorman
- Department of Microbiology, Immunology, and Molecular Genetics, University of Kentucky, Lexington, KY, United States of America
| | - Annette M. Uittenbogaard
- Department of Microbiology, Immunology, and Molecular Genetics, University of Kentucky, Lexington, KY, United States of America
| | - Tanya Myers-Morales
- Department of Microbiology, Immunology, and Molecular Genetics, University of Kentucky, Lexington, KY, United States of America
| | - Alan M. Kaplan
- Department of Microbiology, Immunology, and Molecular Genetics, University of Kentucky, Lexington, KY, United States of America
| | - Donald A. Cohen
- Department of Microbiology, Immunology, and Molecular Genetics, University of Kentucky, Lexington, KY, United States of America
| | - Susan C. Straley
- Department of Microbiology, Immunology, and Molecular Genetics, University of Kentucky, Lexington, KY, United States of America
- * E-mail:
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33
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Effect of thermolabile toxin from Yersinia pseudotuberculosis on functions of innate immunity cells. Bull Exp Biol Med 2014; 157:483-7. [PMID: 25110089 DOI: 10.1007/s10517-014-2597-7] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/08/2013] [Indexed: 10/24/2022]
Abstract
The thermolabile toxin of Yersinia pseudotuberculosis produces a selective dose-dependent stimulating effect on functional activity of innate immunity cells. Prolonged apoptosis-inducing action of the toxin was associated with activation of enzymes of the oxygen-dependent system (LDH and myeloperoxidase) at the early terms of observation (up to 3 h). In turn, increased number of macrophages with apoptotic changes was noted at the early stages of contact with the thermolabile toxin (5 h), and its further growth was observed against the background of activation of mitochondrial enzymes and production of NO metabolites.
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34
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Caulfield AJ, Lathem WW. Disruption of fas-fas ligand signaling, apoptosis, and innate immunity by bacterial pathogens. PLoS Pathog 2014; 10:e1004252. [PMID: 25101900 PMCID: PMC4125287 DOI: 10.1371/journal.ppat.1004252] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/19/2022] Open
Affiliation(s)
- Adam J. Caulfield
- Department of Microbiology-Immunology, Northwestern University Feinberg School of Medicine, Chicago, Illinois, United States of America
| | - Wyndham W. Lathem
- Department of Microbiology-Immunology, Northwestern University Feinberg School of Medicine, Chicago, Illinois, United States of America
- * E-mail:
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35
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Wang X, Liang J, Xi J, Yang J, Wang M, Tian K, Li J, Qiu H, Xiao Y, Duan R, Yang H, Li K, Cui Z, Qi M, Jing H. Canis lupus familiaris involved in the transmission of pathogenic Yersinia spp. in China. Vet Microbiol 2014; 172:339-44. [DOI: 10.1016/j.vetmic.2014.04.015] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/31/2013] [Revised: 03/28/2014] [Accepted: 04/18/2014] [Indexed: 11/15/2022]
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36
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Liang J, Bi Z, Shi G, Xiao Y, Qiu H, Kou Z, Hu B, Jing H, Wang X. Two novel ail-positive biotype 1A strains of Yersinia enterocolitica isolated in China with unequal adhesion and invasion properties. INFECTION GENETICS AND EVOLUTION 2014; 27:83-8. [PMID: 25038297 DOI: 10.1016/j.meegid.2014.07.009] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 03/19/2014] [Revised: 05/20/2014] [Accepted: 07/07/2014] [Indexed: 10/25/2022]
Abstract
Yersinia enterocolitica is an enteric pathogen having six biotypes: 1A, 1B, 2, 3, 4, and 5. Different bioserotypes have been associated with varying pathogenicity, and the strains of biotype 1A lack the virulence-associated pYV-bearing genes and were once considered to be avirulent. However, there is growing epidemiological, clinical, and experimental evidence to suggest some biotype 1A isolates are virulent and can cause gastrointestinal disease. Here, we describe two biotype 1A strains discovered from 3807 isolates that carry the ail (attachment and invasion locus) gene. The two strains showed unique PFGE patterns compared to all other isolates in the Chinese Y. enterocolitica isolate PFGE database. Strain SDWL-003 isolated from a sheep shared ail sequence identical to A1 pattern, and the foxA (ferrioxamine receptor) sequence was identical to the pathogenic F5 pattern, besides, the PFGE patterns of SDWL-003 was also cluster to pathogenic branch; however it does not attach to or invade Hep-2 cells. The ail sequence of strain 2006RAT isolated from a Microtus fortis showed several mutations compared to other published genomes, and therefore formed an entirely new pathogenic pattern. Though it clustered to non-pathogenic block with foxA sequence polymorphism analysis or PFGE assay, the strain 2006RAT showed adhesion properties. The data here bring new insights into the molecular genetics of Y. enterocolitica biotype 1A, show some isolates of 1A biotype gaining potential pathogenicity using the function of the virulence gene - ail, and indicate the lateral gene transfer of ail virulence genes proceeded between pathogenic and nonpathogenic Y. enterocolitica.
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Affiliation(s)
- Junrong Liang
- National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, State Key Laboratory for Infectious Disease Prevention and Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, 102206 Beijing, China
| | - Zhenqiang Bi
- Shandong Provincial Centre for Disease Control and Prevention, 650022 Jinan, China
| | - Guoxiang Shi
- Zhejiang Provincial Centre for Disease Control and Prevention, 450016 Hangzhou, China
| | - Yuchun Xiao
- National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, State Key Laboratory for Infectious Disease Prevention and Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, 102206 Beijing, China
| | - Haiyan Qiu
- National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, State Key Laboratory for Infectious Disease Prevention and Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, 102206 Beijing, China
| | - Zengqiang Kou
- Shandong Provincial Centre for Disease Control and Prevention, 650022 Jinan, China
| | - Bin Hu
- Shandong Provincial Centre for Disease Control and Prevention, 650022 Jinan, China
| | - Huaiqi Jing
- National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, State Key Laboratory for Infectious Disease Prevention and Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, 102206 Beijing, China.
| | - Xin Wang
- National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, State Key Laboratory for Infectious Disease Prevention and Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, 102206 Beijing, China.
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37
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Robinet P, Baychelier F, Fontaine T, Picard C, Debré P, Vieillard V, Latgé JP, Elbim C. A polysaccharide virulence factor of a human fungal pathogen induces neutrophil apoptosis via NK cells. THE JOURNAL OF IMMUNOLOGY 2014; 192:5332-42. [PMID: 24790151 DOI: 10.4049/jimmunol.1303180] [Citation(s) in RCA: 58] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/08/2023]
Abstract
Aspergillus fumigatus is an opportunistic human fungal pathogen that sheds galactosaminogalactan (GG) into the environment. Polymorphonuclear neutrophils (PMNs) and NK cells are both part of the first line of defense against pathogens. We recently reported that GG induces PMN apoptosis. In this study, we show that PMN apoptosis occurs via a new NK cell-dependent mechanism. Reactive oxygen species, induced by the presence of GG, play an indispensable role in this apoptotic effect by increasing MHC class I chain-related molecule A expression at the PMN surface. This increased expression enables interaction between MHC class I chain-related molecule A and NKG2D, leading to NK cell activation, which in turn generates a Fas-dependent apoptosis-promoting signal in PMNs. Taken together, our results demonstrate that the crosstalk between PMNs and NK cells is essential to GG-induced PMN apoptosis. NK cells might thus play a role in the induction of PMN apoptosis in situations such as unexplained neutropenia or autoimmune diseases.
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Affiliation(s)
- Pauline Robinet
- Université Pierre et Marie Curie Paris 06, Unité Mixte de Recherche-S CR7, Sorbonne Université, F-75005 Paris, France; INSERM, Centre d'Immunologie et des Maladies Infectieuses, Unité Mixte de Recherche-S CR7, INSERM U1135, F-75013 Paris, France
| | - Florence Baychelier
- INSERM, Centre d'Immunologie et des Maladies Infectieuses, Unité Mixte de Recherche-S CR7, INSERM U1135, F-75013 Paris, France
| | | | - Capucine Picard
- Centre D'étude des Déficits Immunitaires, Assistance Publique-Hôpitaux de Paris, L'hôpital Necker - Enfants Malades, 75743 Paris, France; Paris Descartes University, Sorbonne Paris Cité, 75006 Paris, France; Laboratory of Human Genetics of Infectious Diseases, Necker Branch, INSERM U980, Necker Medical School, 75015 Paris, France; and
| | - Patrice Debré
- Université Pierre et Marie Curie Paris 06, Unité Mixte de Recherche-S CR7, Sorbonne Université, F-75005 Paris, France; INSERM, Centre d'Immunologie et des Maladies Infectieuses, Unité Mixte de Recherche-S CR7, INSERM U1135, F-75013 Paris, France; Département d'Immunologie, Assistance Publique - Hôpitaux de Paris, Hôpital Pitié-Salpêtrière, 75013 Paris, France
| | - Vincent Vieillard
- INSERM, Centre d'Immunologie et des Maladies Infectieuses, Unité Mixte de Recherche-S CR7, INSERM U1135, F-75013 Paris, France
| | - Jean-Paul Latgé
- Institut Pasteur, Unité des Aspergillus, 75015 Paris, France
| | - Carole Elbim
- Université Pierre et Marie Curie Paris 06, Unité Mixte de Recherche-S CR7, Sorbonne Université, F-75005 Paris, France; INSERM, Centre d'Immunologie et des Maladies Infectieuses, Unité Mixte de Recherche-S CR7, INSERM U1135, F-75013 Paris, France;
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38
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Spinner JL, Winfree S, Starr T, Shannon JG, Nair V, Steele-Mortimer O, Hinnebusch BJ. Yersinia pestis survival and replication within human neutrophil phagosomes and uptake of infected neutrophils by macrophages. J Leukoc Biol 2013; 95:389-98. [PMID: 24227798 DOI: 10.1189/jlb.1112551] [Citation(s) in RCA: 45] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/24/2022] Open
Abstract
Yersinia pestis, the bacterial agent of plague, is transmitted by fleas. The bite of an infected flea deposits Y. pestis into the dermis and triggers recruitment of innate immune cells, including phagocytic PMNs. Y. pestis can subvert this PMN response and survive at the flea-bite site, disseminate, and persist in the host. Although its genome encodes a number of antiphagocytic virulence factors, phagocytosis of Y. pestis by PMNs has been observed. This study tests the hypotheses that Y. pestis, grown at the ambient temperature of the flea vector (21°C), where the major antiphagocytic virulence factors are not produced, can survive and replicate within human PMNs and can use PMNs as a route to infect macrophages subsequently. We show that Y. pestis is localized within PMN phagosomes, predominately as individual bacteria, and that intracellular bacteria can survive and replicate. Within 12 h of infection, ~70% of infected PMNs had PS on their surface and were plausibly competent for efferocytosis. With the use of live cell confocal imaging, we show that autologous HMDMs recognize and internalize infected PMNs and that Y. pestis survives and replicates within these HMDMs following efferocytosis. Addition of HMDMs to infected PMNs resulted in decreased secretion of inflammatory cytokines (compared with HMDMs incubated directly with pCD1(-) Y. pestis) and increased secretion of the anti-inflammatory cytokine IL-1ra. Thus, Y. pestis can survive and replicate within PMNs, and infected PMNs may be a route for noninflammatory infection of macrophages.
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Affiliation(s)
- Justin L Spinner
- 1.Rocky Mountain Laboratories, NIAID, NIH, 903 South 4th St., Hamilton, MT 59840, USA.
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Abstract
The majority of human Yersinia pestis infections result from introduction of bacteria into the skin by the bite of an infected flea. Once in the dermis, Y. pestis can evade the host’s innate immune response and subsequently disseminate to the draining lymph node (dLN). There, the pathogen replicates to large numbers, causing the pathognomonic bubo of bubonic plague. In this study, several cytometric and microscopic techniques were used to characterize the early host response to intradermal (i.d.) Y. pestis infection. Mice were infected i.d. with fully virulent or attenuated strains of dsRed-expressing Y. pestis, and tissues were analyzed by flow cytometry. By 4 h postinfection, there were large numbers of neutrophils in the infected dermis and the majority of cell-associated bacteria were associated with neutrophils. We observed a significant effect of the virulence plasmid (pCD1) on bacterial survival and neutrophil activation in the dermis. Intravital microscopy of i.d. Y. pestis infection revealed dynamic interactions between recruited neutrophils and bacteria. In contrast, very few bacteria interacted with dendritic cells (DCs), indicating that this cell type may not play a major role early in Y. pestis infection. Experiments using neutrophil depletion and a CCR7 knockout mouse suggest that dissemination of Y. pestis from the dermis to the dLN is not dependent on neutrophils or DCs. Taken together, the results of this study show a very rapid, robust neutrophil response to Y. pestis in the dermis and that the virulence plasmid pCD1 is important for the evasion of this response. Yersinia pestis remains a public health concern today because of sporadic plague outbreaks that occur throughout the world and the potential for its illegitimate use as a bioterrorism weapon. Since bubonic plague pathogenesis is initiated by the introduction of Y. pestis into the skin, we sought to characterize the response of the host’s innate immune cells to bacteria early after intradermal infection. We found that neutrophils, innate immune cells that engulf and destroy microbes, are rapidly recruited to the injection site, irrespective of strain virulence, indicating that Y. pestis is unable to subvert neutrophil recruitment to the site of infection. However, we saw a decreased activation of neutrophils that were associated with Y. pestis strains harboring the pCD1 plasmid, which is essential for virulence. These findings indicate a role for pCD1-encoded factors in suppressing the activation/stimulation of these cells in vivo.
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Nörenberg D, Wieser A, Magistro G, Hoffmann C, Meyer C, Messerer M, Schubert S. Molecular analysis of a novel Toll/interleukin-1 receptor (TIR)-domain containing virulence protein of Y. pseudotuberculosis among Far East scarlet-like fever serotype I strains. Int J Med Microbiol 2013; 303:583-94. [PMID: 24018301 DOI: 10.1016/j.ijmm.2013.08.002] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2013] [Revised: 07/29/2013] [Accepted: 08/07/2013] [Indexed: 01/05/2023] Open
Abstract
Pathogenicity of Yersinia pseudotuberculosis is determined by an arsenal of virulence factors. Particularly, the Yersinia outer proteins (Yops) and the Type III secretion system (T3SS) encoded on the pYV virulence plasmid are required for Yersinia pathogenicity. A specific group of Y. pseudotuberculosis, responsible for the clinical syndrome described as Far East scarlet-like fever (FESLF), is known to have an altered virulence gene cluster. Far East strains cause unique clinical symptoms for which the pYV virulence plasmid plays apparently a rather secondary role. Here, we characterize a previously unknown protein of Y. pseudotuberculosis serotype I strains (TcpYI) which can be found particularly among the FESLF strain group. The TcpYI protein shares considerable sequence homology to members of the Toll/IL-1 receptor family. Bacterial TIR domain containing proteins (Tcps) interact with the innate immune system by TIR-TIR interactions and subvert host defenses via individual, multifaceted mechanisms. In terms of virulence, it appears that the TcpYI protein of Y. pseudotuberculosis displays its own virulence phenotype compared to the previously characterized bacterial Tcps. Our results clearly demonstrate that TcpYI increases the intracellular survival of the respective strains in vitro. Furthermore, we show here that the intracellular survival benefit of the wild-type strain correlates with an increase in tcpYI gene expression inside murine macrophages. In support of this, we found that TcpYI enhances the survival inside the spleens of mice in a mouse model of peritonitis. Our results may point toward involvement of the TcpYI protein in inhibition of phagocytosis, particularly in distinct Y. pseudotuberculosis strains of the FESLF strain group where the pYV virulence plasmid is absent.
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Affiliation(s)
- Dominik Nörenberg
- Max von Pettenkofer-Institut für Hygiene und Medizinische Mikrobiologie, Marchioninistr. 17, D-81377 München, Germany
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Vagima Y, Levy Y, Gur D, Tidhar A, Aftalion M, Abramovich H, Zahavy E, Zauberman A, Flashner Y, Shafferman A, Mamroud E. Early sensing of Yersinia pestis airway infection by bone marrow cells. Front Cell Infect Microbiol 2012. [PMID: 23189271 PMCID: PMC3505838 DOI: 10.3389/fcimb.2012.00143] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/05/2023] Open
Abstract
Bacterial infection of the lungs triggers a swift innate immune response that involves the production of cytokines and chemokines that promote recruitment of immune cells from the bone marrow (BM) into the infected tissue and limit the ability of the pathogen to replicate. Recent in vivo studies of pneumonic plague in animal models indicate that the pulmonary pro-inflammatory response to airway infection with Yersinia pestis is substantially delayed in comparison to other pathogens. Consequently, uncontrolled proliferation of the pathogen in the lungs is observed, followed by dissemination to internal organs and death. While the lack of an adequate early immune response in the lung is well described, the response of BM-derived cells is poorly understood. In this study, we show that intranasal (i.n.) infection of mice with a fully virulent Y. pestis strain is sensed early by the BM compartment, resulting in a reduction in CXCR4 levels on BM neutrophils and their subsequent release into the blood 12 hours (h) post infection. In addition, increased levels of BM-derived hematopoietic stem and progenitor cells (HSPC) were detected in the blood early after infection. Mobilization of both immature and mature cells was accompanied by the reduction of BM SDF-1 (CXCL-12) levels and the reciprocal elevation of SDF-1 in the blood 24 h post infection. RT-PCR analysis of RNA collected from total BM cells revealed an early induction of myeloid-associated genes, suggesting a prompt commitment to myeloid lineage differentiation. These findings indicate that lung infection by Y. pestis is sensed by BM cells early after infection, although bacterial colonization of the BM occurs at late disease stages, and point on a potential cross-talk between the lung and the BM at early stages of pneumonic plague.
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Affiliation(s)
- Yaron Vagima
- Department of Biochemistry and Molecular Genetics, Israel Institute for Biological Research Ness Ziona, Israel
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Role of YopK in Yersinia pseudotuberculosis resistance against polymorphonuclear leukocyte defense. Infect Immun 2012; 81:11-22. [PMID: 23090955 DOI: 10.1128/iai.00650-12] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022] Open
Abstract
The enteropathogen Yersinia pseudotuberculosis can survive in the harsh environment of lymphoid compartments that abounds in immune cells. This capacity is dependent on the plasmid-encoded Yersinia outer proteins (Yops) that are delivered into the host cell via a mechanism involving the Yersinia type III secretion system. We show that the virulence protein YopK has a role in the mechanism by which Y. pseudotuberculosis avoids the polymorphonuclear leukocyte or neutrophil (PMN) defense. A yopK mutant, which is attenuated in the mouse infection model, where it fails to cause systemic infection, was found to colonize Peyer's patches and mesenteric lymph nodes more rapidly than the wild-type strain. Further, in mice lacking PMNs, the yopK mutant caused full disease with systemic spread and typical symptoms. Analyses of effects on PMNs revealed that both the wild-type strain and the yopK mutant inhibited internalization and reactive oxygen species production, as well as neutrophil extracellular trap formation by PMNs. However, the wild-type strain effectively avoided induction of PMN death, whereas the mutant caused a necrosis-like PMN death. Taken together, our results indicate that YopK is required for the ability of Yersinia to resist the PMN defense, which is critical for the virulence of the pathogen. We suggest a mechanism whereby YopK functions to prevent unintended Yop delivery and thereby PMN disruption, resulting in necrosis-like cell death, which would enhance the inflammatory response favoring the host.
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Neutrophil extracellular traps exhibit antibacterial activity against burkholderia pseudomallei and are influenced by bacterial and host factors. Infect Immun 2012; 80:3921-9. [PMID: 22927051 DOI: 10.1128/iai.00806-12] [Citation(s) in RCA: 64] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022] Open
Abstract
Burkholderia pseudomallei is the causative pathogen of melioidosis, of which a major predisposing factor is diabetes mellitus. Polymorphonuclear neutrophils (PMNs) kill microbes extracellularly by the release of neutrophil extracellular traps (NETs). PMNs play a key role in the control of melioidosis, but the involvement of NETs in killing of B. pseudomallei remains obscure. Here, we showed that bactericidal NETs were released from human PMNs in response to B. pseudomallei in a dose- and time-dependent manner. B. pseudomallei-induced NET formation required NADPH oxidase activation but not phosphatidylinositol-3 kinase, mitogen-activated protein kinases, or Src family kinase signaling pathways. B. pseudomallei mutants defective in the virulence-associated Bsa type III protein secretion system (T3SS) or capsular polysaccharide I (CPS-I) induced elevated levels of NETs. NET induction by such mutants was associated with increased bacterial killing, phagocytosis, and oxidative burst by PMNs. Taken together the data imply that T3SS and the capsule may play a role in evading the induction of NETs. Importantly, PMNs from diabetic subjects released NETs at a lower level than PMNs from healthy subjects. Modulation of NET formation may therefore be associated with the pathogenesis and control of melioidosis.
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Ponnusamy D, Clinkenbeard KD. Yersinia pestis intracellular parasitism of macrophages from hosts exhibiting high and low severity of plague. PLoS One 2012; 7:e42211. [PMID: 22848745 PMCID: PMC3407133 DOI: 10.1371/journal.pone.0042211] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/22/2012] [Accepted: 07/03/2012] [Indexed: 12/24/2022] Open
Abstract
BACKGROUND Yersinia pestis causes severe disease in natural rodent hosts, but mild to inapparent disease in certain rodent predators such as dogs. Y. pestis initiates infection in susceptible hosts by parasitizing and multiplying intracellularly in local macrophages prior to systemic dissemination. Thus, we hypothesize that Y. pestis disease severity may depend on the degree to which intracellular Y. pestis overcomes the initial host macrophage imposed stress. METHODOLOGY/PRINCIPAL FINDINGS To test this hypothesis, the progression of in vitro infection by Y. pestis KIM62053.1+ of mouse splenic and RAW264.7 tissue culture macrophages and dog peripheral blood-derived and DH82 tissue culture macrophages was studied using microscopy and various parameters of infection. The study showed that during the early stage of infection, intracellular Y. pestis assumed filamentous cellular morphology with multiple copies of the genome per bacterium in both mouse and dog macrophages. Later, in mouse macrophages, the infection elicited spacious vacuolar extension of Yersinia containing vacuoles (YCV), and the filamentous Y. pestis reverted to coccobacillary morphology with genomic equivalents approximately equaling colony forming units. In contrast, Y. pestis infected dog macrophages did not show noticeable extension of YCV, and intracellular Y. pestis retained the filamentous cellular morphology for the entire experiment in DH82 cells or were killed by blood-derived macrophages. In addition, during the later stage of infection, Y. pestis infected mouse macrophages exhibited cell lysis whereas dog macrophages did not. CONCLUSION/SIGNIFICANCE Overall, these results support our hypothesis that Y. pestis in mouse macrophages can overcome the initial intracellular stress necessary for subsequent systemic infection. However, in dogs, failure of Y. pestis to overcome macrophage imposed stress may result in mild or in apparent disease in dogs.
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Affiliation(s)
- Duraisamy Ponnusamy
- Department of Veterinary Pathobiology, Center for Veterinary Health Sciences, Oklahoma State University, Stillwater, Oklahoma, United States of America
| | - Kenneth D. Clinkenbeard
- Department of Veterinary Pathobiology, Center for Veterinary Health Sciences, Oklahoma State University, Stillwater, Oklahoma, United States of America
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Zheng Y, Lilo S, Mena P, Bliska JB. YopJ-induced caspase-1 activation in Yersinia-infected macrophages: independent of apoptosis, linked to necrosis, dispensable for innate host defense. PLoS One 2012; 7:e36019. [PMID: 22563435 PMCID: PMC3338577 DOI: 10.1371/journal.pone.0036019] [Citation(s) in RCA: 29] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2012] [Accepted: 03/27/2012] [Indexed: 12/16/2022] Open
Abstract
Yersinia outer protein J (YopJ) is a type III secretion system (T3SS) effector of pathogenic Yersinia (Yersinia pestis, Yersinia enterocolitica and Yersinia pseudotuberculosis) that is secreted into host cells. YopJ inhibits survival response pathways in macrophages, causing cell death. Allelic variation of YopJ is responsible for differential cytotoxicity in Yersinia strains. YopJ isoforms in Y. enterocolitica O:8 (YopP) and Y. pestis KIM (YopJKIM) strains have high cytotoxic activity. In addition, YopJKIM-induced macrophage death is associated with caspase-1 activation and interleukin-1β (IL-1β secretion. Here, the mechanism of YopJKIM-induced cell death, caspase-1 activation, and IL-1β secretion in primary murine macrophages was examined. Caspase-3/7 activity was low and the caspase-3 substrate poly (ADP-ribose) polymerase (PARP) was not cleaved in Y. pestis KIM5-infected macrophages. In addition, cytotoxicity and IL-1β secretion were not reduced in the presence of a caspase-8 inhibitor, or in B-cell lymphoma 2 (Bcl-2)-associated X protein (Bax)/Bcl-2 homologous antagonist/killer (Bak) knockout macrophages, showing that YopJKIM-mediated cell death and caspase-1 activation occur independent of mitochondrial-directed apoptosis. KIM5-infected macrophages released high mobility group protein B1 (HMGB1), a marker of necrosis, and microscopic analysis revealed that necrotic cells contained active caspase-1, indicating that caspase-1 activation is associated with necrosis. Inhibitor studies showed that receptor interacting protein 1 (RIP1) kinase and reactive oxygen species (ROS) were not required for cytotoxicity or IL-β release in KIM5-infected macrophages. IL-1β secretion was reduced in the presence of cathepsin B inhibitors, suggesting that activation of caspase-1 requires cathepsin B activity. Ectopically-expressed YopP caused higher cytotoxicity and secretion of IL-1β in Y. pseudotuberculosis-infected macrophages than YopJKIM. Wild-type and congenic caspase 1 knockout C57BL/6 mice were equally susceptible to lethal infection with Y. pseudotuberculosis ectopically expressing YopP. These data suggest that YopJ-induced caspase-1 activation in Yersinia-infected macrophages is a downstream consequence of necrotic cell death and is dispensable for innate host resistance to a strain with enhanced cytotoxicity.
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Affiliation(s)
- Ying Zheng
- Department of Molecular Genetics and Microbiology, and Center for Infectious Diseases, Stony Brook University, Stony Brook, New York, United States of America
| | - Sarit Lilo
- Department of Molecular Genetics and Microbiology, and Center for Infectious Diseases, Stony Brook University, Stony Brook, New York, United States of America
| | - Patricio Mena
- Department of Molecular Genetics and Microbiology, and Center for Infectious Diseases, Stony Brook University, Stony Brook, New York, United States of America
| | - James B. Bliska
- Department of Molecular Genetics and Microbiology, and Center for Infectious Diseases, Stony Brook University, Stony Brook, New York, United States of America
- * E-mail:
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Eisele NA, Anderson DM. Host Defense and the Airway Epithelium: Frontline Responses That Protect against Bacterial Invasion and Pneumonia. J Pathog 2011; 2011:249802. [PMID: 22567325 PMCID: PMC3335569 DOI: 10.4061/2011/249802] [Citation(s) in RCA: 53] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/16/2011] [Revised: 07/18/2011] [Accepted: 07/21/2011] [Indexed: 01/31/2023] Open
Abstract
Airway epithelial cells are the first line of defense against invading microbes, and they protect themselves through the production of carbohydrate and protein matrices concentrated with antimicrobial products. In addition, they act as sentinels, expressing pattern recognition receptors that become activated upon sensing bacterial products and stimulate downstream recruitment and activation of immune cells which clear invading microbes. Bacterial pathogens that successfully colonize the lungs must resist these mechanisms or inhibit their production, penetrate the epithelial barrier, and be prepared to resist a barrage of inflammation. Despite the enormous task at hand, relatively few virulence factors coordinate the battle with the epithelium while simultaneously providing resistance to inflammatory cells and causing injury to the lung. Here we review mechanisms whereby airway epithelial cells recognize pathogens and activate a program of antibacterial pathways to prevent colonization of the lung, along with a few examples of how bacteria disrupt these responses to cause pneumonia.
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Affiliation(s)
- Nicholas A. Eisele
- Department of Veterinary Pathobiology, University of Missouri, Columbia, MO 65211, USA
- Department of Molecular Microbiology and Immunology, University of Missouri, Columbia, MO 65211, USA
- The Laboratory for Infectious Disease Research, University of Missouri, Columbia, MO 65211, USA
| | - Deborah M. Anderson
- Department of Veterinary Pathobiology, University of Missouri, Columbia, MO 65211, USA
- The Laboratory for Infectious Disease Research, University of Missouri, Columbia, MO 65211, USA
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Resistance to Yersinia pestis infection decreases with age in B10.T(6R) mice. Infect Immun 2011; 79:4438-46. [PMID: 21859850 DOI: 10.1128/iai.05267-11] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/14/2023] Open
Abstract
We demonstrate that 2-month-old female B10.T(6R) mice are highly resistant to systemic infection with the KIM5 strain of Yersinia pestis and that B10.T(6R) mice become susceptible to Y. pestis infection by the age of 5 months. In this study, young (2-month-old) and middle-aged (5- to 12-month-old) B10.T(6R) mice were infected with equal CFU counts of Y. pestis. The 50% lethal dose (LD(50)) for young B10.T(6R) mice was ∼1.4 × 10(4) CFU, while middle-aged B10.T(6R) mice exhibited an LD(50) of ∼60 CFU. Elevated bacterial burdens were found in the spleens of middle-aged mice at 24 and 60 h and in the livers at 60 h postinfection. Immune cell infiltration was greater in the livers of resistant young mice than in those of middle-aged mice and mice of the susceptible C57BL/6N strain. Unlike susceptible mice, young B10.T(6R) mice did not develop necrotic lesions throughout the liver. Instead, livers from young B10.T(6R) mice contained granuloma-like structures. Immunohistochemical staining of liver sections from these mice at 60 h postinfection revealed that the majority of immune cells present in these structures were neutrophils. These findings suggest that resistance to plague in B10.T(6R) mice correlates with early formation of neutrophilic lesions in the liver.
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Laws TR, Davey MS, Green C, Cooper IAM, Titball RW, Lukaszewski RA. Yersinia pseudotuberculosis is resistant to killing by human neutrophils. Microbes Infect 2011; 13:607-11. [PMID: 21354325 DOI: 10.1016/j.micinf.2011.02.004] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/21/2010] [Revised: 01/05/2011] [Accepted: 02/09/2011] [Indexed: 11/19/2022]
Abstract
The interaction between human neutrophils and the Gram negative gastrointestinal pathogen Yersinia pseudotuberculosis was investigated in vitro. Despite the wealth of data describing how Yersinia can affect the function of neutrophils, there are no published studies describing if neutrophil cells can affect the viability of Y. pseudotuberculosis. The wild-type IP32953 strain of Y. pseudotuberculosis was found to be resistant to killing by human neutrophils. Confocal examination and flow-cytometric analysis of this interaction revealed that bacteria were taken up.
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Affiliation(s)
- Thomas R Laws
- Defence Science and Technology Laboratory, Salisbury, Wiltshire SP4 OJQ, UK.
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Spooner R, Yilmaz Ö. The role of reactive-oxygen-species in microbial persistence and inflammation. Int J Mol Sci 2011; 12:334-52. [PMID: 21339989 PMCID: PMC3039955 DOI: 10.3390/ijms12010334] [Citation(s) in RCA: 164] [Impact Index Per Article: 11.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/04/2010] [Revised: 12/19/2010] [Accepted: 01/05/2011] [Indexed: 02/08/2023] Open
Abstract
The mechanisms of chronic infections caused by opportunistic pathogens are of keen interest to both researchers and health professionals globally. Typically, chronic infectious disease can be characterized by an elevation in immune response, a process that can often lead to further destruction. Reactive-Oxygen-Species (ROS) have been strongly implicated in the aforementioned detrimental response by host that results in self-damage. Unlike excessive ROS production resulting in robust cellular death typically induced by acute infection or inflammation, lower levels of ROS produced by host cells are increasingly recognized to play a critical physiological role for regulating a variety of homeostatic cellular functions including growth, apoptosis, immune response, and microbial colonization. Sources of cellular ROS stimulation can include "danger-signal-molecules" such as extracellular ATP (eATP) released by stressed, infected, or dying cells. Particularly, eATP-P2X(7) receptor mediated ROS production has been lately found to be a key modulator for controlling chronic infection and inflammation. There is growing evidence that persistent microbes can alter host cell ROS production and modulate eATP-induced ROS for maintaining long-term carriage. Though these processes have yet to be fully understood, exploring potential positive traits of these "injurious" molecules could illuminate how opportunistic pathogens maintain persistence through physiological regulation of ROS signaling.
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Affiliation(s)
- Ralee Spooner
- Department of Periodontology, University of Florida, Gainesville, FL 32610, USA; E-Mail:
| | - Özlem Yilmaz
- Department of Periodontology, University of Florida, Gainesville, FL 32610, USA; E-Mail:
- Emerging Pathogens Institute, University of Florida, Gainesville, FL 32610, USA
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