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Tapscott T, Kim JS, Crawford MA, Fitzsimmons L, Liu L, Jones-Carson J, Vázquez-Torres A. Guanosine tetraphosphate relieves the negative regulation of Salmonella pathogenicity island-2 gene transcription exerted by the AT-rich ssrA discriminator region. Sci Rep 2018; 8:9465. [PMID: 29930310 PMCID: PMC6013443 DOI: 10.1038/s41598-018-27780-9] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/01/2017] [Accepted: 06/01/2018] [Indexed: 01/09/2023] Open
Abstract
The repressive activity of ancestral histone-like proteins helps integrate transcription of foreign genes with discrepant AT content into existing regulatory networks. Our investigations indicate that the AT-rich discriminator region located between the −10 promoter element and the transcription start site of the regulatory gene ssrA plays a distinct role in the balanced expression of the Salmonella pathogenicity island-2 (SPI2) type III secretion system. The RNA polymerase-binding protein DksA activates the ssrAB regulon post-transcriptionally, whereas the alarmone guanosine tetraphosphate (ppGpp) relieves the negative regulation imposed by the AT-rich ssrA discriminator region. An increase in the GC-content of the ssrA discriminator region enhances ssrAB transcription and SsrB translation, thus activating the expression of downstream SPI2 genes. A Salmonella strain expressing a GC-rich ssrA discriminator region is attenuated in mice and grows poorly intracellularly. The combined actions of ppGpp and DksA on SPI2 expression enable Salmonella to grow intracellularly, and cause disease in a murine model of infection. Collectively, these findings indicate that (p)ppGpp relieves the negative regulation associated with the AT-rich discriminator region in the promoter of the horizontally-acquired ssrA gene, whereas DksA activates ssrB gene expression post-transcriptionally. The combined effects of (p)ppGpp and DksA on the ssrAB locus facilitate a balanced SPI2 virulence gene transcription that is essential for Salmonella pathogenesis.
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Affiliation(s)
- Timothy Tapscott
- Molecular Biology Program, University of Colorado School of Medicine, Aurora, CO, USA
| | - Ju-Sim Kim
- Department of Immunology and Microbiology, University of Colorado School of Medicine, Aurora, CO, USA
| | - Matthew A Crawford
- Department of Immunology and Microbiology, University of Colorado School of Medicine, Aurora, CO, USA
| | - Liam Fitzsimmons
- Department of Immunology and Microbiology, University of Colorado School of Medicine, Aurora, CO, USA
| | - Lin Liu
- Department of Immunology and Microbiology, University of Colorado School of Medicine, Aurora, CO, USA
| | - Jessica Jones-Carson
- Department of Immunology and Microbiology, University of Colorado School of Medicine, Aurora, CO, USA.,Division of Infectious Diseases, University of Colorado School of Medicine, Aurora, CO, USA
| | - Andrés Vázquez-Torres
- Molecular Biology Program, University of Colorado School of Medicine, Aurora, CO, USA. .,Department of Immunology and Microbiology, University of Colorado School of Medicine, Aurora, CO, USA. .,Veterans Affairs Eastern Colorado Health Care System, Denver, CO, USA.
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Brenz Y, Ohnezeit D, Winther-Larsen HC, Hagedorn M. Nramp1 and NrampB Contribute to Resistance against Francisella in Dictyostelium. Front Cell Infect Microbiol 2017; 7:282. [PMID: 28680861 PMCID: PMC5478718 DOI: 10.3389/fcimb.2017.00282] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/20/2017] [Accepted: 06/09/2017] [Indexed: 12/16/2022] Open
Abstract
The Francisella genus comprises highly pathogenic bacteria that can cause fatal disease in their vertebrate and invertebrate hosts including humans. In general, Francisella growth depends on iron availability, hence, iron homeostasis must be tightly regulated during Francisella infection. We used the system of the professional phagocyte Dictyostelium and the fish pathogen F. noatunensis subsp. noatunensis (F.n.n.) to investigate the role of the host cell iron transporters Nramp (natural resistance associated macrophage proteins) during Francisella infection. Like its mammalian ortholog, Dictyostelium Nramp1 transports iron from the phagosome into the cytosol, whereas the paralog NrampB is located on the contractile vacuole and controls, together with Nramp1, the cellular iron homeostasis. In Dictyostelium, Nramp1 localized to the F.n.n.-phagosome but disappeared from the compartment dependent on the presence of IglC, an established Francisella virulence factor. In the absence of Nramp transporters the bacteria translocated more efficiently from the phagosome into the host cell cytosol, its replicative niche. Increased escape rates coincided with increased proteolytic activity in bead-containing phagosomes indicating a role of the Nramp transporters for phagosomal maturation. In the nramp mutants, a higher bacterial load was observed in the replicative phase compared to wild-type host cells. Upon bacterial access to the cytosol of wt cells, mRNA levels of bacterial iron uptake factors were transiently upregulated. Decreased iron levels in the nramp mutants were compensated by a prolonged upregulation of the iron scavenging system. These results show that Nramps contribute to host cell immunity against Francisella infection by influencing the translocation efficiency from the phagosome to the cytosol but not by restricting access to nutritional iron in the cytosol.
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Affiliation(s)
- Yannick Brenz
- Department of Parasitology, Bernhard Nocht Institute for Tropical MedicineHamburg, Germany
| | - Denise Ohnezeit
- Institute for Medical Microbiology, Hygiene and Virology, University Medical Center Hamburg-EppendorfHamburg, Germany
| | - Hanne C Winther-Larsen
- Centre for Integrative Microbial Evolution and Department of Pharmaceutical Biosciences, University of OsloOslo, Norway
| | - Monica Hagedorn
- Department of Life Sciences and Chemistry, Jacobs UniversityBremen, Germany
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3
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Huang L, Luo Y, Sun X, Ju H, Tian J, Yu BY. An artemisinin-mediated ROS evolving and dual protease light-up nanocapsule for real-time imaging of lysosomal tumor cell death. Biosens Bioelectron 2017; 92:724-732. [DOI: 10.1016/j.bios.2016.10.004] [Citation(s) in RCA: 24] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/12/2016] [Revised: 09/18/2016] [Accepted: 10/03/2016] [Indexed: 01/15/2023]
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Muhsinin M, Ulupi N, Gunawan A, Wibawan IWT, Sumantri C. Association of NRAMP1 Polymorphisms with Immune Traits in
Indonesian Native Chickens. ACTA ACUST UNITED AC 2016. [DOI: 10.3923/ijps.2016.401.406] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/15/2022]
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Iwaki T, Fujita Y, Tanaka N, Giga-Hama Y, Takegawa K. Mitochondrial ABC Transporter Atm1p Is Required for Protection against Oxidative Stress and Vacuolar Functions inSchizosaccharomyces pombe. Biosci Biotechnol Biochem 2014; 69:2109-16. [PMID: 16306692 DOI: 10.1271/bbb.69.2109] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/12/2023]
Abstract
A potential correlation between mitochondrial and vacuolar functions is known to exit in yeast. Fission yeast atm1(+), SPAC15A10.01, encodes a putative half-type ABC transporter with an N-terminal mitochondrial-targeting signal. In an attempt to evaluate the possible involvement of mitochondrion in vacuole function, a functional analysis of atm1(+) was performed by gene disruption. Growth of the atm1 mutant was inhibited in the presence of oxidizing agents, and S. cerevisiae Atm1p was found to complement this growth defect. atm1Delta cells exhibited defects in fluid-phase endocytosis and vacuolar fusion under hypotonic stress. GFP-tagged Atm1p was observed to be localized in the mitochondria. These data strongly suggest that fission yeast Atm1p was not only involved in protection against oxidative stress, but also played a role in vacuolar functions.
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Affiliation(s)
- Tomoko Iwaki
- Department of Life Sciences, Faculty of Agriculture, Kagawa University, Japan
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Wang L, Wang D, Li F. Insight into the structures of the second and fifth transmembrane domains of Slc11a1 in membrane mimics. J Pept Sci 2014; 20:165-72. [DOI: 10.1002/psc.2593] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/09/2013] [Revised: 09/29/2013] [Accepted: 10/30/2013] [Indexed: 01/03/2023]
Affiliation(s)
- Li Wang
- State Key Laboratory of Supramolecular Structure and Materials; Jilin University; Changchun 130012 China
| | - Dan Wang
- State Key Laboratory of Supramolecular Structure and Materials; Jilin University; Changchun 130012 China
| | - Fei Li
- State Key Laboratory of Supramolecular Structure and Materials; Jilin University; Changchun 130012 China
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Upregulation of the host SLC11A1 gene by Clostridium difficile toxin B facilitates glucosylation of Rho GTPases and enhances toxin lethality. Infect Immun 2013; 81:2724-32. [PMID: 23690404 DOI: 10.1128/iai.01177-12] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022] Open
Abstract
Pseudomembranous enterocolitis associated with Clostridium difficile infection is an important cause of morbidity and mortality in patients being treated with antibiotics. Two closely related large protein toxins produced by C. difficile, TcdA and TcdB, which act identically but at different efficiencies to glucosylate low-molecular-weight Rho GTPases, underlie the microbe's pathogenicity. Using antisense RNA encoded by a library of human expressed sequence tags (ESTs), we randomly inactivated host chromosomal genes in HeLa cells and isolated clones that survived exposure to ordinarily lethal doses of TcdB. This phenotypic screening and subsequent analysis identified solute carrier family 11 member 1 (SLC11A1; formerly NRAMP1), a divalent cation transporter crucial to host defense against certain microbes, as an enhancer of TcdB lethality. Whereas SLC11A1 normally is poorly expressed in human cells of nonmyeloid lineage, TcdB increased SLC11A1 mRNA abundance in such cells through the actions of the RNA-binding protein HuR. We show that short hairpin RNA (shRNA) directed against SLC11A1 reduced TcdB glucosylation of small Rho GTPases and, consequently, toxin lethality. Consistent with the previously known role of SLC11A1 in cation transport, these effects were enhanced by elevation of Mn(2+) in media; conversely, they were decreased by treatment with a chelator of divalent cations. Our findings reveal an unsuspected role for SLC11A1 in determining C. difficile pathogenicity, demonstrate the novel ability of a bacterial toxin to increase its cytotoxicity, establish a mechanistic basis for these effects, and suggest a therapeutic approach to mitigate cell killing by C. difficile toxins A and B.
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He XM, Fang MX, Zhang ZT, Hu YS, Jia XZ, He DL, Liang SD, Nie QH, Zhang XQ. Characterization of chicken natural resistance-associated macrophage protein encoding genes (Nramp1 and Nramp2) and association with salmonellosis resistance. GENETICS AND MOLECULAR RESEARCH 2013; 12:618-30. [PMID: 23408449 DOI: 10.4238/2013.january.30.5] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/03/2022]
Abstract
Natural resistance-associated macrophage protein 1 and 2 encoding genes (Nramp1 and Nramp2) are related to many diseases. We cloned the cDNA of chicken Nramp1 and Nramp2 genes, characterized their expression and polymorphisms, and investigated the association of some SNPs with resistance to salmonellosis. The Nramp1 cDNA was 1746 bp long and the Nramp2 cDNA was 1938 bp long. These cDNAs are similar to previously reported cDNAs, varying by two and one amino acids, respectively. The chicken Nramp1 gene expressed predominantly in liver, thymus and spleen in both females and males. The Nramp2 gene expressed in almost all tissues, but predominantly in breast muscle, leg muscle, cerebrum, cerebellum, lung, kidney, and heart in both females and males. We identified 45 SNPs and 2 indels in the chicken Nramp1 gene; three of 13 SNPs in the exons were missense mutations (Arg223Gln, Ala273Glu and Arg497Gln). Association analysis indicated that A24101991G is significantly associated with chicken salmonellosis resistance. These results will be useful for functional investigation of chicken Nramp1 and Nramp2 genes.
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Affiliation(s)
- X M He
- Department of Animal Genetics, Breeding and Reproduction, College of Animal Science, South China Agricultural University, Guangzhou, Guangdong, China
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Subcellular localization of iron and heme metabolism related proteins at early stages of erythrophagocytosis. PLoS One 2012; 7:e42199. [PMID: 22860081 PMCID: PMC3408460 DOI: 10.1371/journal.pone.0042199] [Citation(s) in RCA: 49] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/27/2012] [Accepted: 07/02/2012] [Indexed: 12/26/2022] Open
Abstract
Background Senescent red blood cells (RBC) are recognized, phagocytosed and cleared by tissue macrophages. During this erythrophagocytosis (EP), RBC are engulfed and processed in special compartments called erythrophagosomes. We previously described that following EP, heme is rapidly degraded through the catabolic activity of heme oxygenase (HO). Extracted heme iron is then either exported or stored by macrophages. However, the cellular localization of the early steps of heme processing and iron extraction during EP remains to be clearly defined. Methodology/Principal Findings We took advantage of our previously described cellular model of EP, using bone marrow-derived macrophages (BMDM). The subcellular localization of both inducible and constitutive isoforms of HO (HO-1 and HO-2), of the divalent metal transporters (Nramp1, Nramp2/DMT1, Fpn), and of the recently identified heme transporter HRG-1, was followed by fluorescence and electron microscopy during the earliest steps of EP. We also looked at some ER [calnexin, glucose-6-phosphatase (G6Pase) activity] and lysosomes (Lamp1) markers during EP. In both quiescent and LPS-activated BMDM, Nramp1 and Lamp1 were shown to be strong markers of the erythrophagolysosomal membrane. HRG-1 was also recruited to the erythrophagosome. Furthermore, we observed calnexin labeling and G6Pase activity at the erythrophagosomal membrane, indicating the contribution of ER in this phagocytosis model. In contrast, Nramp2/DMT1, Fpn, HO-1 and HO-2 were not detected at the membrane of erythrophagosomes. Conclusions/Significance Our study highlights the subcellular localization of various heme- and iron-related proteins during early steps of EP, thereby suggesting a model for heme catabolism occurring outside the phagosome, with heme likely being transported into the cytosol through HRG1. The precise function of Nramp1 at the phagosomal membrane in this model remains to be determined.
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Bozzaro S, Eichinger L. The professional phagocyte Dictyostelium discoideum as a model host for bacterial pathogens. Curr Drug Targets 2011; 12:942-54. [PMID: 21366522 PMCID: PMC3267156 DOI: 10.2174/138945011795677782] [Citation(s) in RCA: 82] [Impact Index Per Article: 5.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/21/2010] [Accepted: 10/26/2010] [Indexed: 01/24/2023]
Abstract
The use of simple hosts such as Dictyostelium discoideum in the study of host pathogen interactions offers a number of advantages and has steadily increased in recent years. Infection-specific genes can often only be studied in a very limited way in man and even in the mouse model their analysis is usually expensive, time consuming and technically challenging or sometimes even impossible. In contrast, their functional analysis in D. discoideum and other simple model organisms is often easier, faster and cheaper. Because host-pathogen interactions necessarily involve two organisms, it is desirable to be able to genetically manipulate both the pathogen and its host. Particularly suited are those hosts, like D. discoideum, whose genome sequence is known and annotated and for which excellent genetic and cell biological tools are available in order to dissect the complex crosstalk between host and pathogen. The review focusses on host-pathogen interactions of D. discoideum with Legionella pneumophila, mycobacteria, and Salmonella typhimurium which replicate intracellularly.
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Affiliation(s)
- Salvatore Bozzaro
- Department of Clinical and Biological Sciences, University of Turin, Ospedale S. Luigi, 10043 Orbassano, Italy.
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Abstract
Iron is almost ubiquitous in living organisms due to the utility of its redox chemistry. It is also dangerous as it can catalyse the formation of reactive free radicals - a classical double-edged sword. In this review, we examine the uptake and usage of iron by trypanosomatids and discuss how modulation of host iron metabolism plays an important role in the protective response. Trypanosomatids require iron for crucial processes including DNA replication, antioxidant defence, mitochondrial respiration, synthesis of the modified base J and, in African trypanosomes, the alternative oxidase. The source of iron varies between species. Bloodstream-form African trypanosomes acquire iron from their host by uptake of transferrin, and Leishmania amazonensis expresses a ZIP family cation transporter in the plasma membrane. In other trypanosomatids, iron uptake has been poorly characterized. Iron-withholding responses by the host can be a major determinant of disease outcome. Their role in trypanosomatid infections is becoming apparent. For example, the cytosolic sequestration properties of NRAMP1, confer resistance against leishmaniasis. Conversely, cytoplasmic sequestration of iron may be favourable rather than detrimental to Trypanosoma cruzi. The central role of iron in both parasite metabolism and the host response is attracting interest as a possible point of therapeutic intervention.
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Czachorowski M, Lam-Yuk-Tseung S, Cellier M, Gros P. Transmembrane topology of the mammalian Slc11a2 iron transporter. Biochemistry 2009; 48:8422-34. [PMID: 19621945 PMCID: PMC2736113 DOI: 10.1021/bi900606y] [Citation(s) in RCA: 39] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Abstract
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The mammalian Slc11a1 and Slc11a2 proteins define a large family of secondary metal transporters. Slc11a1 and Slc11a2 function as pH-dependent divalent cation transporters that play a critical role in host defenses against infections and in Fe2+ homeostasis, respectively. The position and polarity of individual transmembrane domains (TMD) of Slc11a2 were studied by an epitope tagging method based on the insertion of small antigenic hemagglutinin A (HA) peptides (YPYDVPDYAS) in predicted intra- or extracellular loops of the protein. The tagged proteins were expressed in transfected LLC-PK1 kidney cells and tested for transport activity, and the polarity of inserted tags with respect to the plasma membrane was determined by immunofluorescence in intact and permeabilized cells. HA epitope tags were inserted at positions 1, 98, 131, 175, 201, 243, 284, 344, 403, 432, 468, 504, and 561. Insertions at positions 98, 131, 175, 403, and 432 abrogated metal transport by Slc11a2, while insertions at positions 1, 201, 243, 284, 344, 468, 504, and 561 resulted in functional proteins. Topology mapping in functional HA-tagged Slc11a2 proteins indicated that the N-terminus (1), as well as loops delineated by TMD4−5 (201), TMD6−7 (284), and TMD10−11 (468), and C-terminus (561) are intracellular, while loops separating TMD5−6 (243), TMD7−8 (344), and TMD11−12 (504) are extracellular. These results are compatible with a topology of 12 transmembrane domains, with intracellular amino and carboxy termini. Structural models constructed by homology threading support this 12TMD topology and show 2-fold structural symmetry in the arrangement of membrane helices for TM1−5 and TM6−10 (conserved Slc11 hydrophobic core).
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Affiliation(s)
- Maciej Czachorowski
- Department of Biochemistry, McGill University, Montreal, Quebec, Canada H3G-0B1
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Nairz M, Fritsche G, Crouch MLV, Barton HC, Fang FC, Weiss G. Slc11a1 limits intracellular growth of Salmonella enterica sv. Typhimurium by promoting macrophage immune effector functions and impairing bacterial iron acquisition. Cell Microbiol 2009; 11:1365-81. [PMID: 19500110 DOI: 10.1111/j.1462-5822.2009.01337.x] [Citation(s) in RCA: 81] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022]
Abstract
The natural resistance-associated macrophage protein 1, Slc11a1, is a phagolysosomal transporter for protons and divalent ions including iron that confers host protection against diverse intracellular pathogens including Salmonella. We investigated and compared the regulation of iron homeostasis and immune function in RAW264.7 murine phagocytes stably transfected with non-functional Slc11a1 and functional Slc11a1 controls in response to an infection with Salmonella enterica serovar Typhimurium. We report that macrophages lacking functional Slc11a1 displayed an increased expression of transferrin receptor 1, resulting in enhanced acquisition of transferrin-bound iron. In contrast, cellular iron release mediated via ferroportin 1 was significantly lower in Salmonella-infected Slc11a1-negative macrophages in comparison with phagocytes bearing Slc11a1. Lack of Slc11a1 led to intracellular persistence of S. enterica serovar Typhimurium within macrophages, which was paralleled by a reduced formation of nitric oxide, tumour necrosis factor-alpha and interleukin-6 in Slc11a1-negative macrophages following Salmonella infection, whereas interleukin-10 production was increased. Moreover, Slc11a1-negative phagocytes exhibited higher cellular iron content, resulting in increased iron acquisition by intracellular Salmonella. Our observations indicate a bifunctional role for Slc11a1 within phagocytes. Slc11a restricts iron availability, which first augments pro-inflammatory macrophage effector functions and second concomitantly limits microbial iron access.
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Affiliation(s)
- Manfred Nairz
- Department of Internal Medicine I, Clinical Immunology and Infectious Diseases, Innsbruck Medical University, Innsbruck, Austria
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Vidal SM, Malo D, Marquis JF, Gros P. Forward genetic dissection of immunity to infection in the mouse. Annu Rev Immunol 2008; 26:81-132. [PMID: 17953509 DOI: 10.1146/annurev.immunol.26.021607.090304] [Citation(s) in RCA: 50] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
Forward genetics is an experimental approach in which gene mapping and positional cloning are used to elucidate the molecular mechanisms underlying phenotypic differences between two individuals for a given trait. This strategy has been highly successful for the study of inbred mouse strains that show differences in innate susceptibility to bacterial, parasitic, fungal, and viral infections. Over the past 20 years, these studies have led to the identification of a number of cell populations and critical biochemical pathways and proteins that are essential for the early detection of and response to invading pathogens. Strikingly, the macrophage is the point of convergence for many of these genetic studies. This has led to the identification of diverse pathways involved in extracellular and intracellular pathogen recognition, modification of the properties and content of phagosomes, transcriptional response, and signal transduction for activation of adaptive immune mechanisms. In models of viral infections, elegant genetic studies highlighted the pivotal role of natural killer cells in the detection and destruction of infected cells.
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Affiliation(s)
- S M Vidal
- Department of Microbiology and Immunology, McGill University, Montreal, Quebec, Canada H3G 1Y6
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Marquis JF, Gros P. Genetic analysis of resistance to infections in mice: A/J meets C57BL/6J. Curr Top Microbiol Immunol 2008; 321:27-57. [PMID: 18727486 DOI: 10.1007/978-3-540-75203-5_2] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/30/2022]
Abstract
Susceptibility to infectious diseases has long been known to have a genetic component in human populations. This genetic effect is often complex and difficult to study as it is further modified by environmental factors including the disease-causing pathogen itself. The laboratory mouse has proved a useful alternative to implement a genetic approach to study host defenses against infections. Our laboratory has used genetic analysis and positional cloning to characterize single and multi-gene effects regulating inter-strain differences in the susceptibility of A/J and C57BL/6J mice to infection with several bacterial and parasitic pathogens. This has led to the identification of several proteins including Nrampl (Slc11a1), Birc1e, Icsbp, C5a, and others that play critical roles in the antimicrobial defenses of macrophages against intracellular pathogens. The use of AcB/BcA recombinant congenic strains has further facilitated the characterization of single gene effects in complex traits such as susceptibility to malaria. The genetic identification of erythrocyte pyruvate kinase (Pklr) and myeloid pantetheinase enzymes (Vnn1/3) as key regulators of blood-stage parasitemia has suggested that cellular redox potential may be a key biochemical determinant of Plasmodium parasite replication. Expanding these types of studies to additional inbred strains and to emerging stocks of mutagenized mice will undoubtedly continue to unravel the molecular basis of host defense against infections.
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Affiliation(s)
- J-F Marquis
- Department of Biochemistry, McGill University, McIntyre Medical Building, Montreal, QC H3G 1Y6, Canada
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Phagocytosis and host-pathogen interactions in Dictyostelium with a look at macrophages. INTERNATIONAL REVIEW OF CELL AND MOLECULAR BIOLOGY 2008; 271:253-300. [PMID: 19081545 DOI: 10.1016/s1937-6448(08)01206-9] [Citation(s) in RCA: 65] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/23/2022]
Abstract
Research into phagocytosis and host-pathogen interactions in the lower eukaryote Dictyostelium discoideum has flourished in recent years. This chapter presents a glimpse of where this research stands, with emphasis on the cell biology of the phagocytic process and on the wealth of molecular genetic data that have been gathered. The basic mechanistic machinery and most of the underlying genes appear to be evolutionarily conserved, reflecting the fact that phagocytosis arose as an efficient way to ingest food in single protozoan cells devoid of a rigid cell wall. In spite of some differences, the signal transduction pathways regulating phagosome biogenesis are also emerging as ultimately similar between Dictyostelium and macrophages. Both cell types are hosts for many pathogenic invasive bacteria, which exploit phagocytosis to grow intracellularly. We present an overwiew, based on the analysis of mutants, on how Dictyostelium contributes as a genetic model system to decipher the complexity of host-pathogen interactions.
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Hautefort I, Thompson A, Eriksson-Ygberg S, Parker ML, Lucchini S, Danino V, Bongaerts RJM, Ahmad N, Rhen M, Hinton JCD. During infection of epithelial cells Salmonella enterica serovar Typhimurium undergoes a time-dependent transcriptional adaptation that results in simultaneous expression of three type 3 secretion systems. Cell Microbiol 2007; 10:958-84. [PMID: 18031307 PMCID: PMC2343689 DOI: 10.1111/j.1462-5822.2007.01099.x] [Citation(s) in RCA: 189] [Impact Index Per Article: 10.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/10/2023]
Abstract
The biogenesis of the Salmonella-containing vacuole within mammalian cells has been intensively studied over recent years. However, the ability of Salmonella to sense and adapt to the intracellular environment of different types of host cells has received much less attention. To address this issue, we report the transcriptome of Salmonella enterica serovar Typhimurium SL1344 within epithelial cells and show comparisons with Salmonella gene expression inside macrophages. We report that S. Typhimurium expresses a characteristic intracellular transcriptomic signature in response to the environments it encounters within different cell types. The signature involves the upregulation of the mgtBC, pstACS and iro genes for magnesium, phosphate and iron uptake, and Salmonella pathogenicity island 2 (SPI2). Surprisingly, in addition to SPI2, the invasion-associated SPI1 pathogenicity island and the genes involved in flagellar biosynthesis were expressed inside epithelial cells at later stages of the infection, while they were constantly downregulated in macrophage-like cells. To our knowledge, this is the first report of the simultaneous transcription of all three Type Three Secretion Systems (T3SS) within an intracellular Salmonella population. We discovered that S. Typhimurium strain SL1344 was strongly cytotoxic to epithelial cells after 6 h of infection and hypothesize that the time-dependent changes in Salmonella gene expression within epithelial cells reflects the bacterial response to host cells that have been injured by the infection process.
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Affiliation(s)
- I Hautefort
- Molecular Microbiology Group, Institute of Food Research, Norwich NR4 7UA, UK.
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Reduced in vitro functional activity of human NRAMP1 (SLC11A1) allele that predisposes to increased risk of pediatric tuberculosis disease. Genes Immun 2007; 8:691-8. [DOI: 10.1038/sj.gene.6364435] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/08/2022]
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Abstract
Progress in the characterization of genes involved in the control of iron homeostasis in humans and in mice has improved the definition of iron overload and of the cells affected by it. The cell involved in iron overload with the greatest effect on immunity is the macrophage. Intriguing evidence has emerged, however, in the last 12 years indicating that parenchymal iron overload is linked to genes classically associated with the immune system. This review offers an update of the genes and proteins relevant to iron metabolism expressed in cells of the innate immune system, and addresses the question of how this system is affected in clinical situations of iron overload. The relationship between iron and the major cells of adaptive immunity, the T lymphocytes, will also be reviewed. Most studies addressing this last question in humans were performed in the clinical model of Hereditary Hemochromatosis. Data will also be reviewed demonstrating how the disruption of molecules essentially involved in adaptive immune responses result in the spontaneous development of iron overload and how they act as modifiers of iron overload.
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Affiliation(s)
- Graça Porto
- Institute of Molecular and Cellular Biology, Rua do Campo Alegre, Porto 8234150, Portugal.
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20
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Niño-Moreno P, Portales-Pérez D, Hernández-Castro B, Portales-Cervantes L, Flores-Meraz V, Baranda L, Gómez-Gómez A, Acuña-Alonzo V, Granados J, González-Amaro R. P2X7 and NRAMP1/SLC11 A1 gene polymorphisms in Mexican mestizo patients with pulmonary tuberculosis. Clin Exp Immunol 2007; 148:469-77. [PMID: 17493019 PMCID: PMC1941940 DOI: 10.1111/j.1365-2249.2007.03359.x] [Citation(s) in RCA: 60] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022] Open
Abstract
Tuberculosis remains one of the most important infectious diseases worldwide. Several studies have suggested that genetic factors may affect susceptibility to tuberculosis, but the specific genes involved have not yet been fully characterized. NRAMP1/SLC11 A1 and P2X(7) genes have been linked to increased risk for tuberculosis in some African and Asiatic populations. To explore the potential role of these genes in the susceptibility to pulmonary tuberculosis in a Mexican mestizo population, we evaluated the association of D543N and 3'-UTR polymorphisms in NRAMP1/SLC11 A1 and - 762 and A1513C polymorphisms in P2X(7) genes with the risk for tuberculosis. Polymerase chain reaction (PCR) amplification of genomic DNA followed by restriction fragment length polymorphism analysis, and allelic-specific PCR was employed. We found no significant differences in allelic frequency in NRAMP1/SLC11 A1 gene polymorphisms in 94 patients with tuberculosis compared to 100 healthy contacts. Similarly, no significant association of the P2X(7)-762 gene polymorphism with tuberculosis was detected. In contrast, the P2X(7) A1513C polymorphism was associated significantly with tuberculosis (P = 0.02, odds ratio = 5.28, 95% CI, 0.99-37.69), an association that had not been reported previously. However, when the function of P2X(7) was assessed by an L-selectin loss assay, we did not find significant differences in patients compared to healthy contacts or between PPD(+) and PPD(-) control individuals. This study further supports the complex role of P2X(7) gene in host regulation of Mycobacterium tuberculosis infection, and demonstrates that different associations of gene polymorphisms and tuberculosis are found in distinct racial populations.
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Affiliation(s)
- P Niño-Moreno
- Dpto de Inmunología, Facultad de Medicina, Universidad Autónoma de San Luis Potosí, San Luis Potosí, México
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21
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Courville P, Chaloupka R, Cellier MFM. Recent progress in structure-function analyses of Nramp proton-dependent metal-ion transporters. Biochem Cell Biol 2007; 84:960-78. [PMID: 17215883 DOI: 10.1139/o06-193] [Citation(s) in RCA: 93] [Impact Index Per Article: 5.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022] Open
Abstract
The natural resistance-associated macrophage protein (Nramp) homologs form a family of proton-coupled transporters that facilitate the cellular absorption of divalent metal ions (Me2+, including Mn2+, Fe2+, Co2+, and Cd2+). The Nramp, or solute carrier 11 (SLC11), family is conserved in eukaryotes and bacteria. Humans and rodents express 2 parologous genes that are associated with iron disorders and immune diseases. The NRAMP1 (SLC11A1) protein is specific to professional phagocytes and extrudes Me2+ from the phagosome to defend against ingested microbes; polymorphisms in the NRAMP1 gene are associated with various immune diseases. Several isoforms of NRAMP2 (SLC11A2, DMT1, DCT1) are expressed ubiquitously in recycling endosomes or specifically at the apical membrane of epithelial cells in intestine and kidneys, and can contribute to iron overload, whereas mutations impairing NRAMP2 function cause a form of congenital microcytic hypochromic anemia. Structure-function studies, using various experimental models, and mutagenesis approaches have begun to reveal the overall transmembrane organization of Nramp, some of the transmembrane segments (TMS) that are functionally important, and an unusual mechanism coupling Me2+ and proton H+ transport. The approaches used include functional complementation of yeast knockout strains, electrophysiology analyses in Xenopus oocytes, and transport assays that use mammalian and bacterial cells and direct and indirect measurements of SLC11 transporter properties. These complementary studies enabled the identification of TMS1 and 6 as crucial structural segments for Me2+ and H+ symport, and will help develop a deeper understanding of the Nramp transport mechanism and its contribution to Me2+ homeostasis in human health and diseases.
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Affiliation(s)
- P Courville
- Institut National de la Recherche Scientifique, INRS-Institut Armand-Frappier, 531, Bd. des prairies, Laval, QC H7V 1B7, Canada
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22
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Marquis JF, Gros P. Intracellular Leishmania: your iron or mine? Trends Microbiol 2007; 15:93-5. [PMID: 17257847 DOI: 10.1016/j.tim.2007.01.001] [Citation(s) in RCA: 26] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/15/2006] [Revised: 01/04/2007] [Accepted: 01/17/2007] [Indexed: 11/29/2022]
Abstract
Iron is a co-factor for several essential enzymes and biochemical pathways, including those required for replication of pathogens such as Leishmania in macrophages. Iron acquisition is emerging as a key battleground in which the iron import systems of microbes are pitted against the iron withdrawal and sequestration systems of macrophages, with both competing for iron at the interface of host-pathogen interaction. The recent characterization of a ferrous iron transport system (LIT1) in Leishmania amazonensis that is induced intracellularly and is required for survival in macrophages and for virulence in vivo provides an elegant example of the adaptation of protozoa to the iron-poor phagosomal environment.
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Affiliation(s)
- Jean-François Marquis
- Centre for the Study of Host Resistance, Department of Biochemistry, McGill University, Montréal, Québec, H3G 1Y6, Canada
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23
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Abstract
Phagosomes are fascinating subcellular structures. After all, there are only a few compartments that are born before our very eyes and whose development we can follow in a light microscope until their contents disintegrate and are completely absorbed. Yet, some phagosomes are taken advantage of by pathogenic microorganisms, which change their fate. Research into phagosome biogenesis has flourished in recent years - the purpose of this review is to give a glimpse of where this research stands, with emphasis on the cell biology of macrophage phagosomes, on new model organisms for the study of phagosome biogenesis and on intracellular pathogens and their interference with normal phagosome function.
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Affiliation(s)
- Albert Haas
- Cell Biology Institute, University of Bonn, Ulrich-Haberland-Str. 61a, 53121 Bonn, Germany.
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24
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Fortin A, Abel L, Casanova JL, Gros P. Host genetics of mycobacterial diseases in mice and men: forward genetic studies of BCG-osis and tuberculosis. Annu Rev Genomics Hum Genet 2007; 8:163-92. [PMID: 17492906 DOI: 10.1146/annurev.genom.8.080706.092315] [Citation(s) in RCA: 123] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
In humans, genetic factors have long been suspected to contribute to the onset and outcome of tuberculosis. Such effects are difficult to identify owing to their complex inheritance, and to the confounding impact of environmental factors, notably pathogen-associated virulence determinants. Recently, forward genetic approaches in mouse models and in human populations have been used to elucidate a molecular basis for predisposition to mycobacterial diseases. The genetic dissection of host predisposition to infection with Mycobacterium bovis BCG and M. tuberculosis will help to define the key molecules involved in host antituberculous immunity and should provide new insights into this important infectious disease.
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Affiliation(s)
- A Fortin
- Emerillon Therapeutics, Montréal, Canada
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25
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Lam-Yuk-Tseung S, Picard V, Gros P. Identification of a Tyrosine-based Motif (YGSI) in the Amino Terminus of Nramp1 (Slc11a1) That Is Important for Lysosomal Targeting. J Biol Chem 2006. [DOI: 10.1016/s0021-9258(19)84081-4] [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] Open
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26
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Lam-Yuk-Tseung S, Picard V, Gros P. Identification of a tyrosine-based motif (YGSI) in the amino terminus of Nramp1 (Slc11a1) that is important for lysosomal targeting. J Biol Chem 2006; 281:31677-88. [PMID: 16905747 DOI: 10.1074/jbc.m601828200] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/04/2023] Open
Abstract
In macrophages, Nramp1 (Slc11a1) is expressed in lysosomes and restricts replication of intracellular pathogens by removing divalent metals (Mn2+ and Fe2+) from the phagolysosome. Nramp2 (DMT1, Slc11a2) is expressed both at the duodenal brush border where it mediates uptake of dietary iron and ubiquitously at the plasma membrane/recycling endosomes of many cell types where it transports transferrin-associated iron across the endosomal membrane. In Nramp2, a carboxyl-terminal cytoplasmic motif ((555)YLLNT(559)) is critical for internalization and recycling of the transporter from the plasma membrane. Here we studied the subcellular trafficking properties of Nramp1 and investigated the cis-acting sequences responsible for targeting to lysosomes. For this, we constructed and studied Nramp1/Nramp2 chimeric proteins where homologous domains of each protein were exchanged. Chimeras exchanging the amino-(upstream TM1) and carboxyl-terminal (downstream TM12) cytoplasmic segments of both transporters were stably expressed in porcine LLC-PK1 kidney cells and were studied with respect to expression, maturation, stability, cell surface targeting, transport activity, and subcellular localization. An Nramp2 isoform II chimera bearing the amino terminus of Nramp1 was not expressed at the cell surface but was targeted to lysosomes. This lysosomal targeting was abolished by single alanine substitutions at Tyr15 and Ile18 of a (15)YGSI(18) motif present in the amino terminus of Nramp1. These results identify YGSI as a tyrosine-based sorting signal responsible for lysosomal targeting of Nramp1.
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Affiliation(s)
- Steven Lam-Yuk-Tseung
- Department of Biochemistry, McGill Cancer Center and Center for Host Resistance, McGill University, Montreal, Quebec H3G 1Y6, Canada
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27
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Caron J, Larivière L, Nacache M, Tam M, Stevenson MM, McKerly C, Gros P, Malo D. Influence of Slc11a1 on the outcome of Salmonella enterica serovar Enteritidis infection in mice is associated with Th polarization. Infect Immun 2006; 74:2787-802. [PMID: 16622216 PMCID: PMC1459719 DOI: 10.1128/iai.74.5.2787-2802.2006] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022] Open
Abstract
Genetic analyses identified Ses1 as a significant quantitative trait locus influencing the carrier state of 129S6 mice following a sublethal challenge with Salmonella enterica serovar Enteritidis. Previous studies have determined that Slc11a1 was an excellent candidate gene for Ses1. Kinetics of infection in 129S6 mice and Slc11a1-deficient (129S6-Slc11a1(tm1Mcg)) mice demonstrated that the wild-type allele of Slc11a1 contributed to the S. enterica serovar Enteritidis carrier state as early as 7 days postinfection. Gene expression profiling demonstrated that 129S6 mice had a significant up-regulation of proinflammatory genes associated with macrophage activation at day 10 postinfection, followed by a gradual increase in immunoglobulin transcripts, whereas 129S6-Slc11a1(tm1Mcg) mice had higher levels of immunoglobulins earlier in the infection. Quantitative reverse transcription-PCR revealed an increase in Th1 cytokine (Ifng and Il12) and Th1-specific transcription factor Tbx21 expression during infection in both the 129S6 and 129S6-Slc11a1(tm1Mcg) strains. However, the expression of Gata3, a transcription factor involved in Th2 polarization, Cd28, and Il4 was markedly increased in Slc11a1-deficient mice during infection, suggesting a predominant Th2 phenotype in 129S6-Slc11a1(tm1Mcg) animals following S. enterica serovar Enteritidis infection. A strong immunoglobulin G2a response, reflecting Th1 activity, was observed only in 129S6 mice. All together, these results are consistent with an impact of Slc11a1 on Th cell differentiation during chronic S. enterica serovar Enteritidis infection. The presence of a Th2 bias in Slc11a1-deficient mice is associated with improved bacterial clearance.
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Affiliation(s)
- Judith Caron
- Department of Human Genetics, McGill University, Montreal, QC, Canada H3G 1A4
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28
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Forbes J, Lam-Yuk-Tseung S, Gros P. Modulation of Iron Availability at the Host-Pathogen Interface in Phagocytic Cells. EcoSal Plus 2006; 2. [PMID: 26443573 DOI: 10.1128/ecosalplus.8.8.10] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/07/2005] [Indexed: 06/05/2023]
Abstract
This review summarizes recent data on iron metabolism in macrophages, with a special emphasis on possible bacteriostatic and bactericidal consequences for intracellular pathogens. It includes the role of biological chelators and transporters in normal macrophage physiology and antimicrobial defense. Iron is an essential metal cofactor for many biochemical pathways in mammals. However, excess iron promotes the formation of cytotoxic oxygen derivatives so that systemic iron levels must be tightly regulated. The mechanism of iron recycling by macrophages including iron efflux from erythrocyte-containing phagosomes, iron release from macrophages, and entry into the transferrin (Tf) cycle remain poorly understood. Ferroportin expression in the liver, spleen, and bone marrow cells appears to be restricted to macrophages. Mutant mice bearing a conditional deletion of the ferroportin gene in macrophages show retention of iron by hepatic Kupffer cells and splenic macrophages. Hepcidin is induced by lipopolysaccharide (LPS) in mouse spleens and splenic macrophage in vitro and appears to mediate the LPS-induced down-regulation of ferroportin in the intestine and in splenic macrophages, suggesting that inflammatory agents may regulate iron metabolism through modulation of ferroportin expression. The host transporter Nramp1 may compete directly with bacterial divalent-metal transport systems for the acquisition of divalent metals within the phagosomal space. The ultimate outcome of these competing interactions influences the ability of pathogens to survive and replicate intracellularly. This seems particularly relevant to the Salmonella, Leishmania, and Mycobacterium spp., in which inactivating mutations in Nramp1 abrogate the natural resistance of macrophages to these pathogens.
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Intracellular Voyeurism: Examining the Modulation of Host Cell Activities bySalmonella enterica Serovar Typhimurium. EcoSal Plus 2005; 1. [PMID: 26443522 DOI: 10.1128/ecosalplus.2.2.2] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022]
Abstract
Salmonella spp. can infect host cells by gaining entry through phagocytosis or by inducing host cell membrane ruffling that facilitates bacterial uptake. With its wide host range, Salmonella enterica serovar Typhimurium has proven to be an important model organism for studying intracellular bacterial pathogenesis. Upon entry into host cells, serovar Typhimurium typically resides within a membrane-bound compartment termed the Salmonella-containing vacuole (SCV). From the SCV, serovar Typhimurium can inject several effector proteins that subvert many normal host cell systems, including endocytic trafficking, cytoskeletal rearrangements, lipid signaling and distribution, and innate and adaptive host defenses. The study of these intracellular events has been made possible through the use of various imaging techniques, ranging from classic methods of transmission electron microscopy to advanced livecell fluorescence confocal microscopy. In addition, DNA microarrays have now been used to provide a "snapshot" of global gene expression in serovar Typhimurium residing within the infected host cell. This review describes key aspects of Salmonella-induced subversion of host cell activities, providing examples of imaging that have been used to elucidate these events. Serovar Typhimurium engages specific host cell machinery from initial contact with the host cell to replication within the SCV. This continuous interaction with the host cell has likely contributed to the extensive arsenal that serovar Typhimurium now possesses, including two type III secretion systems, a range of ammunition in the form of TTSS effectors, and a complex genetic regulatory network that coordinates the expression of hundreds of virulence factors.
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30
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Caron J, Loredo-Osti JC, Morgan K, Malo D. Mapping of interactions and mouse congenic strains identified novel epistatic QTLs controlling the persistence of Salmonella Enteritidis in mice. Genes Immun 2005; 6:500-8. [PMID: 15973461 DOI: 10.1038/sj.gene.6364234] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/29/2022]
Abstract
The host response to infection in humans is multifactorial and involves the complex interaction between two genomes (the host and the pathogen) and the environment. Using an experimental mouse model of chronic infection, we have previously identified the individual effect of three significant and one suggestive quantitative trait loci (QTLs) (Ses1, Ses2, Ses3 and Ses1.1) on Salmonella Enteritidis persistence in target organs of 129S6/SvEvTac mice. Congenic strain construction was performed by transferring each of these QTLs from C57BL/6J onto the 129S6/SvEvTac background, and phenotypic analysis confirmed that Ses1 and Ses1.1 contribute to bacterial clearance. Additional QTLs regulating Salmonella carriage in 129S6/SvEvTac mice were identified using a two-locus epistasis QTL linkage mapping approach conducted separately in females and males. The epistatic model for females included the individual effect of Ses3 and two significant interactions (Ses1-D7Mit267 and Ses1-DXMit48) accounting for 47% of the total phenotypic variance. The model for males included the individual effect of Ses1.1, three interactions (Ses1-D9Mit218, D2Mit197-D4Mit2 and D3Mit256-D13Mit36) and explained 47% of the phenotypic variance. Our results suggest that the oligogenic nature of Salmonella persistence and epistasis are important constituents of the genetic architecture of the host response to chronic Salmonella infection.
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Affiliation(s)
- J Caron
- Department of Human Genetics, McGill University, Montréal, Québec, Canada
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31
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Poon AH, Laprise C, Lemire M, Hudson TJ, Schurr E. NRAMP1 is not associated with asthma, atopy, and serum immunoglobulin E levels in the French Canadian population. Genes Immun 2005; 6:519-27. [PMID: 15988535 DOI: 10.1038/sj.gene.6364238] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
Reduced infection by mycobacteria, including Mycobacterium tuberculosis, may be partly responsible for increased prevalence of allergic and autoimmune diseases in developed countries. In a murine model of innate resistance to mycobacteria, the Nramp1 gene has been shown to affect asthma susceptibility. From this observation, it was proposed that human NRAMP1 may be a modulator of asthma risk in human populations. To experimentally test the candidacy of NRAMP1 in asthma susceptibility, we characterized five genetic variants of NRAMP1 (5'CAn, 274C>T, 469+14G>C, D543N, and 1729+del4) in an asthma family-based cohort from northeastern Quebec. We did not observe any significant association between NRAMP1 variants (either allele or haplotype specific) with asthma, atopy, or serum immunoglobulin E levels. These results demonstrate that, in spite of direct involvement of Nramp1 in a murine asthma model, in human populations NRAMP1 is not likely to be a major contributor to the genetic etiology of asthma and asthma-related phenotypes.
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Affiliation(s)
- A H Poon
- McGill Centre for the Study of Host Resistance, Research Institute of the McGill University Health Centre, Montreal, QC, Canada
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32
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Fortier A, Min-Oo G, Forbes J, Lam-Yuk-Tseung S, Gros P. Single gene effects in mouse models of host: pathogen interactions. J Leukoc Biol 2005; 77:868-77. [PMID: 15653750 DOI: 10.1189/jlb.1004616] [Citation(s) in RCA: 50] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/24/2022] Open
Abstract
Inbred mouse strains have been known for many years to vary in their degree of susceptibility to different types of infectious diseases. The genetic basis of these interstrain differences is sometimes simple but often complex. In a few cases, positional cloning has been used successfully to identify single gene effects. The natural resistance-associated macrophage protein 1 (Nramp1) gene (Slc11a1) codes for a metal transporter active at the phagosomal membrane of macrophages, and Nramp1 mutations cause susceptibility to Mycobacterium, Salmonella, and Leishmania. Furthermore, recent advances in gene transfer technologies in transgenic mice have enabled the functional dissection of gene effects mapping to complex, repeated parts of the genome, such as the Lgn1 locus, causing susceptibility to Legionella pneumophila in macrophages. Finally, complex traits such as the genetically determined susceptibility to malaria can sometimes be broken down into multiple single gene effects. One such example is the case of pyruvate kinase, where a loss-of-function mutation was recently shown by our group to be protective against blood-stage infection with Plasmodium chabaudi. In all three cases reviewed, the characterization of the noted gene effect(s) has shed considerable light on the pathophysiology of the infection, including host response mechanisms.
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Affiliation(s)
- Anne Fortier
- Department of Biochemistry, McGill University, Montreal, QC, Canada
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Abstract
The use of iron as a cofactor in basic metabolic pathways is essential to both pathogenic microorganisms and their hosts. It is also a pivotal component of the innate immune response through its role in the generation of toxic oxygen and nitrogen intermediates. During evolution, the shared requirement of micro- and macroorganisms for this important nutrient has shaped the pathogen-host relationship. Here, we discuss how pathogens compete with the host for iron, and also how the host uses iron to counteract this threat.
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Affiliation(s)
- Ulrich E Schaible
- Max-Planck-Institute for Infection Biology, Department of Immunology, Schumannstrasse 21-22, D-10117, Berlin, Germany
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34
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White JK, Mastroeni P, Popoff JF, Evans CAW, Blackwell JM. Slc11a1-mediated resistance to Salmonella enterica serovar Typhimurium and Leishmania donovani infections does not require functional inducible nitric oxide synthase or phagocyte oxidase activity. J Leukoc Biol 2004; 77:311-20. [PMID: 15601666 DOI: 10.1189/jlb.0904546] [Citation(s) in RCA: 42] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/01/2023] Open
Abstract
Solute carrier family 11a member 1 (Slc11a1; formerly natural resistance-associated macrophage protein 1) encodes a late endosomal/lysosomal protein/divalent cation transporter, which regulates iron homeostasis in macrophages. During macrophage activation, Slc11a1 exerts pleiotropic effects on gene regulation and function, including generation of nitric oxide (NO) via inducible NO synthase (iNOS; encoded by Nos2A) and of reactive oxygen intermediates (ROI) via the phagocyte oxidase complex. As NO and ROI have potent antimicrobial activity in macrophages, it was assumed that their activities would contribute to Slc11a1-regulated innate resistance to Salmonella enterica serovar Typhimurium and Leishmania donovani. By intercrossing mice with gene disruptions at Nos2A and Cybb (encoding gp91phox, the heavy chain subunit of cytochrome b-245 and an essential component of phagocyte NADPH oxidase) onto equivalent Slc11a1 wild-type and mutant genetic backgrounds, we demonstrate that neither iNOS nor gp91phox activity is required for Slc11a1-mediated innate resistance to either infection. Functional gp91phox and iNOS are required to control S. enterica serovar Typhimurium in non-Slc11a1-regulated phases of infection. For L. donovani, an organ-specific requirement for iNOS to clear parasites from the spleen was observed at 50 days post-infection, but neither iNOS nor gp91phox influenced late-phase infection in the liver. This contrasted with Leishmania major infection, which caused rapid lesion growth and death in iNOS knockout mice and some exacerbation of disease with gp91phox deficiency. This highlights the adaptive differences in tissue and cellular tropisms between L. donovani and L. major and the different genes and mechanisms that regulate visceral versus cutaneous forms of the disease.
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Affiliation(s)
- Jacqueline K White
- Wellcome Trust/MRC Building, University of Cambridge School of Clinical Medicine, Addenbrookes Hospital, Hills Road, Cambridge CB2 2XY, UK
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35
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Wicker LS, Chamberlain G, Hunter K, Rainbow D, Howlett S, Tiffen P, Clark J, Gonzalez-Munoz A, Cumiskey AM, Rosa RL, Howson JM, Smink LJ, Kingsnorth A, Lyons PA, Gregory S, Rogers J, Todd JA, Peterson LB. Fine Mapping, Gene Content, Comparative Sequencing, and Expression Analyses Support Ctla4 and Nramp1 as Candidates for Idd5.1 and Idd5.2 in the Nonobese Diabetic Mouse. THE JOURNAL OF IMMUNOLOGY 2004; 173:164-73. [PMID: 15210771 DOI: 10.4049/jimmunol.173.1.164] [Citation(s) in RCA: 91] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/10/2023]
Abstract
At least two loci that determine susceptibility to type 1 diabetes in the NOD mouse have been mapped to chromosome 1, Idd5.1 (insulin-dependent diabetes 5.1) and Idd5.2. In this study, using a series of novel NOD.B10 congenic strains, Idd5.1 has been defined to a 2.1-Mb region containing only four genes, Ctla4, Icos, Als2cr19, and Nrp2 (neuropilin-2), thereby excluding a major candidate gene, Cd28. Genomic sequence comparison of the two functional candidate genes, Ctla4 and Icos, from the B6 (resistant at Idd5.1) and the NOD (susceptible at Idd5.1) strains revealed 62 single nucleotide polymorphisms (SNPs), only two of which were in coding regions. One of these coding SNPs, base 77 of Ctla4 exon 2, is a synonymous SNP and has been correlated previously with type 1 diabetes susceptibility and differential expression of a CTLA-4 isoform. Additional expression studies in this work support the hypothesis that this SNP in exon 2 is the genetic variation causing the biological effects of Idd5.1. Analysis of additional congenic strains has also localized Idd5.2 to a small region (1.52 Mb) of chromosome 1, but in contrast to the Idd5.1 interval, Idd5.2 contains at least 45 genes. Notably, the Idd5.2 region still includes the functionally polymorphic Nramp1 gene. Future experiments to test the identity of Idd5.1 and Idd5.2 as Ctla4 and Nramp1, respectively, can now be justified using approaches to specifically alter or mimic the candidate causative SNPs.
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Affiliation(s)
- Linda S Wicker
- Juvenile Diabetes Research Foundation/Wellcome Trust Diabetes and Inflammation Laboratory, Department of Medical Genetics, Cambridge Institute for Medical Research, University of Cambridge, Cambridge CB2 2XY, UK.
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36
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37
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Abstract
The Gram-negative pathogen Salmonella enterica can survive and replicate within a variety of mammalian cells. Regardless of the cell type, internalized bacteria survive and replicate within the Salmonella-containing vacuole, the biogenesis of which is dependent on bacterially encoded virulence factors. In particular, Type III secretion systems translocate bacterial effector proteins into the eukaryotic cell where they can specifically interact with a variety of targets. Salmonella has two distinct Type III secretion systems that are believed to have completely different functions. The SPI2 system is induced intracellularly and is required for intracellular survival in macrophages; it plays no role in invasion but is categorized as being required for Salmonella-containing vacuole biogenesis. In contrast, the SPI1 Type III secretion system is induced extracellularly and is essential for invasion of nonphagocytic cells. Its role in post-invasion processes has not been well studied. Recent studies indicate that Salmonella-containing vacuole biogenesis may be more dependent on SPI1 than previously believed. Other non-SPI2 virulence factors and the host cell itself may play critical roles in determining the intracellular environment of this facultative intracellular pathogen. In this review we discuss the recent advances in determining the mechanisms by which Salmonella regulate Salmonella-containing vacuole biogenesis and the implications of these findings.
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Affiliation(s)
- Leigh A Knodler
- Host-Parasite Interactions Section, Laboratory of Intracellular Parasites, National Institutes of Allergy and Infectious Diseases/NIH, Rocky Mountain Laboratories, Hamilton, MT 59840, USA
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