Publications
2014 |
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Articles de journaux |
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| 23. | Majzoub, Karim; Hafirassou, Mohamed Lamine ; Meignin, Carine ; Goto, Akira ; Marzi, Stefano ; Fedorova, Antonina ; Verdier, Yann ; Vinh, Joëlle ; Hoffmann, Jules A; Martin, Franck ; Baumert, Thomas F; Schuster, Catherine ; Imler, Jean-Luc RACK1 controls IRES-mediated translation of viruses Article de journal Cell, 159 (5), p. 1086–1095, 2014, ISSN: 1097-4172. Résumé | Liens | BibTeX | Étiquettes: Cell Line, Cell Surface, Cytoplasmic and Nuclear, Dicistroviridae, GTP-Binding Proteins, Hepacivirus, Hepatocytes, Humans, Insect Viruses, Models, Molecular, Neoplasm Proteins, Peptide Initiation Factors, Protein Biosynthesis, Receptors, Regulatory Sequences, Ribonucleic Acid, Tumor, Virus Replication @article{majzoub_rack1_2014, title = {RACK1 controls IRES-mediated translation of viruses}, author = { Karim Majzoub and Mohamed Lamine Hafirassou and Carine Meignin and Akira Goto and Stefano Marzi and Antonina Fedorova and Yann Verdier and Joëlle Vinh and Jules A. Hoffmann and Franck Martin and Thomas F. Baumert and Catherine Schuster and Jean-Luc Imler}, doi = {10.1016/j.cell.2014.10.041}, issn = {1097-4172}, year = {2014}, date = {2014-11-01}, journal = {Cell}, volume = {159}, number = {5}, pages = {1086--1095}, abstract = {Fighting viral infections is hampered by the scarcity of viral targets and their variability, resulting in development of resistance. Viruses depend on cellular molecules-which are attractive alternative targets-for their life cycle, provided that they are dispensable for normal cell functions. Using the model organism Drosophila melanogaster, we identify the ribosomal protein RACK1 as a cellular factor required for infection by internal ribosome entry site (IRES)-containing viruses. We further show that RACK1 is an essential determinant for hepatitis C virus translation and infection, indicating that its function is conserved for distantly related human and fly viruses. Inhibition of RACK1 does not affect Drosophila or human cell viability and proliferation, and RACK1-silenced adult flies are viable, indicating that this protein is not essential for general translation. Our findings demonstrate a specific function for RACK1 in selective mRNA translation and uncover a target for the development of broad antiviral intervention.}, keywords = {Cell Line, Cell Surface, Cytoplasmic and Nuclear, Dicistroviridae, GTP-Binding Proteins, Hepacivirus, Hepatocytes, Humans, Insect Viruses, Models, Molecular, Neoplasm Proteins, Peptide Initiation Factors, Protein Biosynthesis, Receptors, Regulatory Sequences, Ribonucleic Acid, Tumor, Virus Replication}, pubstate = {published}, tppubtype = {article} } Fighting viral infections is hampered by the scarcity of viral targets and their variability, resulting in development of resistance. Viruses depend on cellular molecules-which are attractive alternative targets-for their life cycle, provided that they are dispensable for normal cell functions. Using the model organism Drosophila melanogaster, we identify the ribosomal protein RACK1 as a cellular factor required for infection by internal ribosome entry site (IRES)-containing viruses. We further show that RACK1 is an essential determinant for hepatitis C virus translation and infection, indicating that its function is conserved for distantly related human and fly viruses. Inhibition of RACK1 does not affect Drosophila or human cell viability and proliferation, and RACK1-silenced adult flies are viable, indicating that this protein is not essential for general translation. Our findings demonstrate a specific function for RACK1 in selective mRNA translation and uncover a target for the development of broad antiviral intervention. |
2013 |
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Articles de journaux |
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| 22. | Fukuyama, Hidehiro; Verdier, Yann; Guan, Yongsheng; Makino-Okamura, Chieko; Shilova, Victoria; Liu, Xi; Maksoud, Elie; Matsubayashi, Jun; Haddad, Iman; Spirohn, Kerstin; Ono, Kenichiro; Hetru, Charles; Rossier, Jean; Ideker, Trey; Boutros, Michael; Vinh, Joëlle; Hoffmann, Jules A Landscape of protein-protein interactions in Drosophila immune deficiency signaling during bacterial challenge Article de journal Proc. Natl. Acad. Sci. U.S.A., 110 (26), p. 10717–10722, 2013, ISSN: 1091-6490. Résumé | Liens | BibTeX | Étiquettes: Amino Acid, Chromatin Assembly and Disassembly, Escherichia coli, functional proteomics, Genes, Genetically Modified, Histone Acetyltransferases, Host-Pathogen Interactions, Humans, IMD interactome, Insect, Models, Molecular, Protein Interaction Maps, Sequence Homology, Signal Transduction, small ubiquitin-like modifier @article{fukuyama_landscape_2013, title = {Landscape of protein-protein interactions in Drosophila immune deficiency signaling during bacterial challenge}, author = { Hidehiro Fukuyama and Yann Verdier and Yongsheng Guan and Chieko Makino-Okamura and Victoria Shilova and Xi Liu and Elie Maksoud and Jun Matsubayashi and Iman Haddad and Kerstin Spirohn and Kenichiro Ono and Charles Hetru and Jean Rossier and Trey Ideker and Michael Boutros and Joëlle Vinh and Jules A. Hoffmann}, doi = {10.1073/pnas.1304380110}, issn = {1091-6490}, year = {2013}, date = {2013-06-01}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, volume = {110}, number = {26}, pages = {10717--10722}, abstract = {The Drosophila defense against pathogens largely relies on the activation of two signaling pathways: immune deficiency (IMD) and Toll. The IMD pathway is triggered mainly by Gram-negative bacteria, whereas the Toll pathway responds predominantly to Gram-positive bacteria and fungi. The activation of these pathways leads to the rapid induction of numerous NF-κB-induced immune response genes, including antimicrobial peptide genes. The IMD pathway shows significant similarities with the TNF receptor pathway. Recent evidence indicates that the IMD pathway is also activated in response to various noninfectious stimuli (i.e., inflammatory-like reactions). To gain a better understanding of the molecular machinery underlying the pleiotropic functions of this pathway, we first performed a comprehensive proteomics analysis to identify the proteins interacting with the 11 canonical members of the pathway initially identified by genetic studies. We identified 369 interacting proteins (corresponding to 291 genes) in heat-killed Escherichia coli-stimulated Drosophila S2 cells, 92% of which have human orthologs. A comparative analysis of gene ontology from fly or human gene annotation databases points to four significant common categories: (i) the NuA4, nucleosome acetyltransferase of H4, histone acetyltransferase complex, (ii) the switching defective/sucrose nonfermenting-type chromatin remodeling complex, (iii) transcription coactivator activity, and (iv) translation factor activity. Here we demonstrate that sumoylation of the IκB kinase homolog immune response-deficient 5 plays an important role in the induction of antimicrobial peptide genes through a highly conserved sumoylation consensus site during bacterial challenge. Taken together, the proteomics data presented here provide a unique avenue for a comparative functional analysis of proteins involved in innate immune reactions in flies and mammals.}, keywords = {Amino Acid, Chromatin Assembly and Disassembly, Escherichia coli, functional proteomics, Genes, Genetically Modified, Histone Acetyltransferases, Host-Pathogen Interactions, Humans, IMD interactome, Insect, Models, Molecular, Protein Interaction Maps, Sequence Homology, Signal Transduction, small ubiquitin-like modifier}, pubstate = {published}, tppubtype = {article} } The Drosophila defense against pathogens largely relies on the activation of two signaling pathways: immune deficiency (IMD) and Toll. The IMD pathway is triggered mainly by Gram-negative bacteria, whereas the Toll pathway responds predominantly to Gram-positive bacteria and fungi. The activation of these pathways leads to the rapid induction of numerous NF-κB-induced immune response genes, including antimicrobial peptide genes. The IMD pathway shows significant similarities with the TNF receptor pathway. Recent evidence indicates that the IMD pathway is also activated in response to various noninfectious stimuli (i.e., inflammatory-like reactions). To gain a better understanding of the molecular machinery underlying the pleiotropic functions of this pathway, we first performed a comprehensive proteomics analysis to identify the proteins interacting with the 11 canonical members of the pathway initially identified by genetic studies. We identified 369 interacting proteins (corresponding to 291 genes) in heat-killed Escherichia coli-stimulated Drosophila S2 cells, 92% of which have human orthologs. A comparative analysis of gene ontology from fly or human gene annotation databases points to four significant common categories: (i) the NuA4, nucleosome acetyltransferase of H4, histone acetyltransferase complex, (ii) the switching defective/sucrose nonfermenting-type chromatin remodeling complex, (iii) transcription coactivator activity, and (iv) translation factor activity. Here we demonstrate that sumoylation of the IκB kinase homolog immune response-deficient 5 plays an important role in the induction of antimicrobial peptide genes through a highly conserved sumoylation consensus site during bacterial challenge. Taken together, the proteomics data presented here provide a unique avenue for a comparative functional analysis of proteins involved in innate immune reactions in flies and mammals. |
| 21. | Baron, Olga Lucia; van West, Pieter; Industri, Benoit; Ponchet, Michel; Dubreuil, Géraldine; Gourbal, Benjamin; Reichhart, Jean-Marc; Coustau, Christine Parental transfer of the antimicrobial protein LBP/BPI protects Biomphalaria glabrata eggs against oomycete infections Article de journal PLoS Pathog., 9 (12), p. e1003792, 2013, ISSN: 1553-7374. Résumé | Liens | BibTeX | Étiquettes: Acute-Phase Proteins, Antimicrobial Cationic Peptides, Biomphalaria, Blood Proteins, Carrier Proteins, Cell Membrane, Cell Membrane Permeability, Cloning, Escherichia coli, Female, Immunity, Infection, Maternally-Acquired, Membrane Glycoproteins, Microbial Sensitivity Tests, Molecular, Oomycetes, Recombinant Proteins, Zygote @article{baron_parental_2013, title = {Parental transfer of the antimicrobial protein LBP/BPI protects Biomphalaria glabrata eggs against oomycete infections}, author = { Olga Lucia Baron and Pieter van West and Benoit Industri and Michel Ponchet and Géraldine Dubreuil and Benjamin Gourbal and Jean-Marc Reichhart and Christine Coustau}, doi = {10.1371/journal.ppat.1003792}, issn = {1553-7374}, year = {2013}, date = {2013-01-01}, journal = {PLoS Pathog.}, volume = {9}, number = {12}, pages = {e1003792}, abstract = {Vertebrate females transfer antibodies via the placenta, colostrum and milk or via the egg yolk to protect their immunologically immature offspring against pathogens. This evolutionarily important transfer of immunity is poorly documented in invertebrates and basic questions remain regarding the nature and extent of parental protection of offspring. In this study, we show that a lipopolysaccharide binding protein/bactericidal permeability increasing protein family member from the invertebrate Biomphalaria glabrata (BgLBP/BPI1) is massively loaded into the eggs of this freshwater snail. Native and recombinant proteins displayed conserved LPS-binding, antibacterial and membrane permeabilizing activities. A broad screening of various pathogens revealed a previously unknown biocidal activity of the protein against pathogenic water molds (oomycetes), which is conserved in human BPI. RNAi-dependent silencing of LBP/BPI in the parent snails resulted in a significant reduction of reproductive success and extensive death of eggs through oomycete infections. This work provides the first functional evidence that a LBP/BPI is involved in the parental immune protection of invertebrate offspring and reveals a novel and conserved biocidal activity for LBP/BPI family members.}, keywords = {Acute-Phase Proteins, Antimicrobial Cationic Peptides, Biomphalaria, Blood Proteins, Carrier Proteins, Cell Membrane, Cell Membrane Permeability, Cloning, Escherichia coli, Female, Immunity, Infection, Maternally-Acquired, Membrane Glycoproteins, Microbial Sensitivity Tests, Molecular, Oomycetes, Recombinant Proteins, Zygote}, pubstate = {published}, tppubtype = {article} } Vertebrate females transfer antibodies via the placenta, colostrum and milk or via the egg yolk to protect their immunologically immature offspring against pathogens. This evolutionarily important transfer of immunity is poorly documented in invertebrates and basic questions remain regarding the nature and extent of parental protection of offspring. In this study, we show that a lipopolysaccharide binding protein/bactericidal permeability increasing protein family member from the invertebrate Biomphalaria glabrata (BgLBP/BPI1) is massively loaded into the eggs of this freshwater snail. Native and recombinant proteins displayed conserved LPS-binding, antibacterial and membrane permeabilizing activities. A broad screening of various pathogens revealed a previously unknown biocidal activity of the protein against pathogenic water molds (oomycetes), which is conserved in human BPI. RNAi-dependent silencing of LBP/BPI in the parent snails resulted in a significant reduction of reproductive success and extensive death of eggs through oomycete infections. This work provides the first functional evidence that a LBP/BPI is involved in the parental immune protection of invertebrate offspring and reveals a novel and conserved biocidal activity for LBP/BPI family members. |
2011 |
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Articles de journaux |
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| 20. | Aoun, Richard Bou; Hetru, Charles; Troxler, Laurent; Doucet, Daniel; Ferrandon, Dominique; Matt, Nicolas Analysis of thioester-containing proteins during the innate immune response of Drosophila melanogaster Article de journal J Innate Immun, 3 (1), p. 52–64, 2011, ISSN: 1662-8128. Résumé | Liens | BibTeX | Étiquettes: bioinformatic, DNA, Evolution, Gene Expression Regulation, Hemocytes, Immunity, In Situ Hybridization, Innate, Molecular, Mutation, Phylogeny, Sequence Analysis @article{bou_aoun_analysis_2011, title = {Analysis of thioester-containing proteins during the innate immune response of Drosophila melanogaster}, author = { Richard Bou Aoun and Charles Hetru and Laurent Troxler and Daniel Doucet and Dominique Ferrandon and Nicolas Matt}, doi = {10.1159/000321554}, issn = {1662-8128}, year = {2011}, date = {2011-01-01}, journal = {J Innate Immun}, volume = {3}, number = {1}, pages = {52--64}, abstract = {Thioester-containing proteins (TEPs) are conserved proteins among insects that are thought to be involved in innate immunity. In Drosophila, the Tep family is composed of 6 genes named Tep1-Tep6. In this study, we investigated the phylogeny, expression pattern and roles of these genes in the host defense of Drosophila. Protostomian Tep genes are clustered in 3 distinct branches, 1 of which is specific to mosquitoes. Most D. melanogaster Tep genes are expressed in hemocytes, can be induced in the fat body, and are expressed in specific regions of the hypodermis. This expression pattern is consistent with a role in innate immunity. However, we find that TEP1, TEP2, and TEP4 are not strictly required in the body cavity to fight several bacterial and fungal infections. One possibility is that Drosophila TEPs act redundantly or that their absence can be compensated by other components of the immune response. TEPs may thus provide a subtle selective advantage during evolution. Alternatively, they may be required in host defense against specific as yet unidentified natural pathogens of Drosophila.}, keywords = {bioinformatic, DNA, Evolution, Gene Expression Regulation, Hemocytes, Immunity, In Situ Hybridization, Innate, Molecular, Mutation, Phylogeny, Sequence Analysis}, pubstate = {published}, tppubtype = {article} } Thioester-containing proteins (TEPs) are conserved proteins among insects that are thought to be involved in innate immunity. In Drosophila, the Tep family is composed of 6 genes named Tep1-Tep6. In this study, we investigated the phylogeny, expression pattern and roles of these genes in the host defense of Drosophila. Protostomian Tep genes are clustered in 3 distinct branches, 1 of which is specific to mosquitoes. Most D. melanogaster Tep genes are expressed in hemocytes, can be induced in the fat body, and are expressed in specific regions of the hypodermis. This expression pattern is consistent with a role in innate immunity. However, we find that TEP1, TEP2, and TEP4 are not strictly required in the body cavity to fight several bacterial and fungal infections. One possibility is that Drosophila TEPs act redundantly or that their absence can be compensated by other components of the immune response. TEPs may thus provide a subtle selective advantage during evolution. Alternatively, they may be required in host defense against specific as yet unidentified natural pathogens of Drosophila. |
2009 |
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Articles de journaux |
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| 19. | Kemp, Cordula; Imler, Jean-Luc Antiviral immunity in drosophila Article de journal Current Opinion in Immunology, 21 (1), p. 3–9, 2009, ISSN: 1879-0372. Résumé | Liens | BibTeX | Étiquettes: Argonaute Proteins, Caspases, DEAD-box RNA Helicases, Evolution, Gene Expression Regulation, Host-Pathogen Interactions, Membrane Proteins, Molecular, Nuclear Proteins, Ribonuclease III, RNA, RNA Helicases, RNA Interference, RNA Virus Infections, RNA Viruses, RNA-Induced Silencing Complex, Viral, Virulence @article{kemp_antiviral_2009, title = {Antiviral immunity in drosophila}, author = { Cordula Kemp and Jean-Luc Imler}, doi = {10.1016/j.coi.2009.01.007}, issn = {1879-0372}, year = {2009}, date = {2009-02-01}, journal = {Current Opinion in Immunology}, volume = {21}, number = {1}, pages = {3--9}, abstract = {Genetic analysis of the drosophila antiviral response indicates that RNA interference plays a major role. This contrasts with the situation in mammals, where interferon-induced responses mediate innate antiviral host-defense. An inducible response also contributes to antiviral immunity in drosophila, and similarities in the sensing and signaling of viral infection are becoming apparent between drosophila and mammals. In particular, DExD/H box helicases appear to play a crucial role in the cytosolic detection of viral RNAs in flies and mammals.}, keywords = {Argonaute Proteins, Caspases, DEAD-box RNA Helicases, Evolution, Gene Expression Regulation, Host-Pathogen Interactions, Membrane Proteins, Molecular, Nuclear Proteins, Ribonuclease III, RNA, RNA Helicases, RNA Interference, RNA Virus Infections, RNA Viruses, RNA-Induced Silencing Complex, Viral, Virulence}, pubstate = {published}, tppubtype = {article} } Genetic analysis of the drosophila antiviral response indicates that RNA interference plays a major role. This contrasts with the situation in mammals, where interferon-induced responses mediate innate antiviral host-defense. An inducible response also contributes to antiviral immunity in drosophila, and similarities in the sensing and signaling of viral infection are becoming apparent between drosophila and mammals. In particular, DExD/H box helicases appear to play a crucial role in the cytosolic detection of viral RNAs in flies and mammals. |
| 18. | Garrett, Matthew; Fullaondo, Ane; Troxler, Laurent; Micklem, Gos; Gubb, David Identification and analysis of serpin-family genes by homology and synteny across the 12 sequenced Drosophilid genomes Article de journal BMC Genomics, 10 , p. 489, 2009, ISSN: 1471-2164. Résumé | Liens | BibTeX | Étiquettes: bioinformatic, Comparative Genomic Hybridization, Conserved Sequence, DNA, Drosophilidae, Evolution, Genome, Insect, Molecular, Multigene Family, Sequence Alignment, Sequence Analysis, Serpins, Synteny @article{garrett_identification_2009, title = {Identification and analysis of serpin-family genes by homology and synteny across the 12 sequenced Drosophilid genomes}, author = { Matthew Garrett and Ane Fullaondo and Laurent Troxler and Gos Micklem and David Gubb}, doi = {10.1186/1471-2164-10-489}, issn = {1471-2164}, year = {2009}, date = {2009-01-01}, journal = {BMC Genomics}, volume = {10}, pages = {489}, abstract = {BACKGROUND: The Drosophila melanogaster genome contains 29 serpin genes, 12 as single transcripts and 17 within 6 gene clusters. Many of these serpins have a conserved "hinge" motif characteristic of active proteinase inhibitors. However, a substantial proportion (42%) lacks this motif and represents non-inhibitory serpin-fold proteins of unknown function. Currently, it is not known whether orthologous, inhibitory serpin genes retain the same target proteinase specificity within the Drosophilid lineage, nor whether they give rise to non-inhibitory serpin-fold proteins or other, more diverged, proteins. RESULTS: We collated 188 orthologues to the D. melanogaster serpins from the other 11 Drosophilid genomes and used synteny to find further family members, raising the total to 226, or 71% of the number of orthologues expected assuming complete conservation across all 12 Drosophilid species. In general the sequence constraints on the serpin-fold itself are loose. The critical Reactive Centre Loop (RCL) sequence, including the target proteinase cleavage site, is strongly conserved in inhibitory serpins, although there are 3 exceptional sets of orthologues in which the evolutionary constraints are looser. Conversely, the RCL of non-inhibitory serpin orthologues is less conserved, with 3 exceptions that presumably bind to conserved partner molecules. We derive a consensus hinge motif, for Drosophilid inhibitory serpins, which differs somewhat from that of the vertebrate consensus. Three gene clusters appear to have originated in the melanogaster subgroup, Spn28D, Spn77B and Spn88E, each containing one inhibitory serpin orthologue that is present in all Drosophilids. In addition, the Spn100A transcript appears to represent a novel serpin-derived fold. CONCLUSION: In general, inhibitory serpins rarely change their range of proteinase targets, except by a duplication/divergence mechanism. Non-inhibitory serpins appear to derive from inhibitory serpins, but not the reverse. The conservation of different family members varied widely across the 12 sequenced Drosophilid genomes. An approach considering synteny as well as homology was important to find the largest set of orthologues.}, keywords = {bioinformatic, Comparative Genomic Hybridization, Conserved Sequence, DNA, Drosophilidae, Evolution, Genome, Insect, Molecular, Multigene Family, Sequence Alignment, Sequence Analysis, Serpins, Synteny}, pubstate = {published}, tppubtype = {article} } BACKGROUND: The Drosophila melanogaster genome contains 29 serpin genes, 12 as single transcripts and 17 within 6 gene clusters. Many of these serpins have a conserved "hinge" motif characteristic of active proteinase inhibitors. However, a substantial proportion (42%) lacks this motif and represents non-inhibitory serpin-fold proteins of unknown function. Currently, it is not known whether orthologous, inhibitory serpin genes retain the same target proteinase specificity within the Drosophilid lineage, nor whether they give rise to non-inhibitory serpin-fold proteins or other, more diverged, proteins. RESULTS: We collated 188 orthologues to the D. melanogaster serpins from the other 11 Drosophilid genomes and used synteny to find further family members, raising the total to 226, or 71% of the number of orthologues expected assuming complete conservation across all 12 Drosophilid species. In general the sequence constraints on the serpin-fold itself are loose. The critical Reactive Centre Loop (RCL) sequence, including the target proteinase cleavage site, is strongly conserved in inhibitory serpins, although there are 3 exceptional sets of orthologues in which the evolutionary constraints are looser. Conversely, the RCL of non-inhibitory serpin orthologues is less conserved, with 3 exceptions that presumably bind to conserved partner molecules. We derive a consensus hinge motif, for Drosophilid inhibitory serpins, which differs somewhat from that of the vertebrate consensus. Three gene clusters appear to have originated in the melanogaster subgroup, Spn28D, Spn77B and Spn88E, each containing one inhibitory serpin orthologue that is present in all Drosophilids. In addition, the Spn100A transcript appears to represent a novel serpin-derived fold. CONCLUSION: In general, inhibitory serpins rarely change their range of proteinase targets, except by a duplication/divergence mechanism. Non-inhibitory serpins appear to derive from inhibitory serpins, but not the reverse. The conservation of different family members varied widely across the 12 sequenced Drosophilid genomes. An approach considering synteny as well as homology was important to find the largest set of orthologues. |
2007 |
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Articles de journaux |
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| 17. | Croker, Ben; Crozat, Karine; Berger, Michael; Xia, Yu; Sovath, Sosathya; Schaffer, Lana; Eleftherianos, Ioannis; Imler, Jean-Luc; Beutler, Bruce ATP-sensitive potassium channels mediate survival during infection in mammals and insects Article de journal Nature Genetics, 39 (12), p. 1453–1460, 2007, ISSN: 1546-1718. Résumé | Liens | BibTeX | Étiquettes: ATP-Binding Cassette Transporters, Cloning, Coronary Vessels, Crosses, Ethylnitrosourea, Genetic, Homozygote, Infection, Inwardly Rectifying, KATP Channels, Lipopolysaccharides, Mice, Molecular, Mutagenesis, Potassium Channels, Sulfonylurea Receptors @article{croker_atp-sensitive_2007, title = {ATP-sensitive potassium channels mediate survival during infection in mammals and insects}, author = { Ben Croker and Karine Crozat and Michael Berger and Yu Xia and Sosathya Sovath and Lana Schaffer and Ioannis Eleftherianos and Jean-Luc Imler and Bruce Beutler}, doi = {10.1038/ng.2007.25}, issn = {1546-1718}, year = {2007}, date = {2007-01-01}, journal = {Nature Genetics}, volume = {39}, number = {12}, pages = {1453--1460}, abstract = {Specific homeostatic mechanisms confer stability in innate immune responses, preventing injury or death from infection. Here we identify, from a screen of N-ethyl-N-nitrosourea-mutagenized mice, a mutation causing both profound susceptibility to infection by mouse cytomegalovirus and approximately 20,000-fold sensitization to lipopolysaccharide (LPS), poly(I.C) and immunostimulatory (CpG) DNA. The LPS hypersensitivity phenotype is not suppressed by mutations in Myd88, Trif, Tnf, Tnfrsf1a, Ifnb, Ifng or Stat1, genes contributing to LPS responses, and results from an abnormality extrinsic to hematopoietic cells. The phenotype is due to a null allele of Kcnj8, encoding Kir6.1, a protein that combines with SUR2 to form an ATP-sensitive potassium channel (K(ATP)) expressed in coronary artery smooth muscle and endothelial cells. In Drosophila melanogaster, suppression of dSUR by RNA interference similarly causes hypersensitivity to infection by flock house virus. Thus, K(ATP) evolved to serve a homeostatic function during infection, and in mammals it prevents coronary artery vasoconstriction induced by cytokines dependent on TLR and/or MDA5 immunoreceptors.}, keywords = {ATP-Binding Cassette Transporters, Cloning, Coronary Vessels, Crosses, Ethylnitrosourea, Genetic, Homozygote, Infection, Inwardly Rectifying, KATP Channels, Lipopolysaccharides, Mice, Molecular, Mutagenesis, Potassium Channels, Sulfonylurea Receptors}, pubstate = {published}, tppubtype = {article} } Specific homeostatic mechanisms confer stability in innate immune responses, preventing injury or death from infection. Here we identify, from a screen of N-ethyl-N-nitrosourea-mutagenized mice, a mutation causing both profound susceptibility to infection by mouse cytomegalovirus and approximately 20,000-fold sensitization to lipopolysaccharide (LPS), poly(I.C) and immunostimulatory (CpG) DNA. The LPS hypersensitivity phenotype is not suppressed by mutations in Myd88, Trif, Tnf, Tnfrsf1a, Ifnb, Ifng or Stat1, genes contributing to LPS responses, and results from an abnormality extrinsic to hematopoietic cells. The phenotype is due to a null allele of Kcnj8, encoding Kir6.1, a protein that combines with SUR2 to form an ATP-sensitive potassium channel (K(ATP)) expressed in coronary artery smooth muscle and endothelial cells. In Drosophila melanogaster, suppression of dSUR by RNA interference similarly causes hypersensitivity to infection by flock house virus. Thus, K(ATP) evolved to serve a homeostatic function during infection, and in mammals it prevents coronary artery vasoconstriction induced by cytokines dependent on TLR and/or MDA5 immunoreceptors. |
2004 |
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Articles de journaux |
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| 16. | Blandin, Stephanie A; Shiao, Shin-Hong; Moita, Luis F; Janse, Chris J; Waters, Andrew P; Kafatos, Fotis C; Levashina, Elena A Complement-like protein TEP1 is a determinant of vectorial capacity in the malaria vector Anopheles gambiae Article de journal Cell, 116 (5), p. 661–670, 2004, ISSN: 0092-8674. Résumé | BibTeX | Étiquettes: Anopheles, Female, Genetic, Humans, Insect Proteins, Insect Vectors, Malaria, Models, Molecular, Plasmodium berghei, Polymorphism, Protein Structure, RNA, Sequence Alignment, Tertiary @article{blandin_complement-like_2004, title = {Complement-like protein TEP1 is a determinant of vectorial capacity in the malaria vector Anopheles gambiae}, author = { Stephanie A. Blandin and Shin-Hong Shiao and Luis F. Moita and Chris J. Janse and Andrew P. Waters and Fotis C. Kafatos and Elena A. Levashina}, issn = {0092-8674}, year = {2004}, date = {2004-01-01}, journal = {Cell}, volume = {116}, number = {5}, pages = {661--670}, abstract = {Anopheles mosquitoes are major vectors of human malaria in Africa. Large variation exists in the ability of mosquitoes to serve as vectors and to transmit malaria parasites, but the molecular mechanisms that determine vectorial capacity remain poorly understood. We report that the hemocyte-specific complement-like protein TEP1 from the mosquito Anopheles gambiae binds to and mediates killing of midgut stages of the rodent malaria parasite Plasmodium berghei. The dsRNA knockdown of TEP1 in adults completely abolishes melanotic refractoriness in a genetically selected refractory strain. Moreover, in susceptible mosquitoes this knockdown increases the number of developing parasites. Our results suggest that the TEP1-dependent parasite killing is followed by a TEP1-independent clearance of dead parasites by lysis and/or melanization. Further elucidation of the molecular mechanisms of TEP1-mediated parasite killing will be of great importance for our understanding of the principles of vectorial capacity in insects.}, keywords = {Anopheles, Female, Genetic, Humans, Insect Proteins, Insect Vectors, Malaria, Models, Molecular, Plasmodium berghei, Polymorphism, Protein Structure, RNA, Sequence Alignment, Tertiary}, pubstate = {published}, tppubtype = {article} } Anopheles mosquitoes are major vectors of human malaria in Africa. Large variation exists in the ability of mosquitoes to serve as vectors and to transmit malaria parasites, but the molecular mechanisms that determine vectorial capacity remain poorly understood. We report that the hemocyte-specific complement-like protein TEP1 from the mosquito Anopheles gambiae binds to and mediates killing of midgut stages of the rodent malaria parasite Plasmodium berghei. The dsRNA knockdown of TEP1 in adults completely abolishes melanotic refractoriness in a genetically selected refractory strain. Moreover, in susceptible mosquitoes this knockdown increases the number of developing parasites. Our results suggest that the TEP1-dependent parasite killing is followed by a TEP1-independent clearance of dead parasites by lysis and/or melanization. Further elucidation of the molecular mechanisms of TEP1-mediated parasite killing will be of great importance for our understanding of the principles of vectorial capacity in insects. |
2003 |
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| 15. | Luna, C; Hoa, N T; Zhang, J; Kanzok, S M; Brown, S E; Imler, Jean-Luc; Knudson, D L; Zheng, L Characterization of three Toll-like genes from mosquito Aedes aegypti Article de journal Insect Molecular Biology, 12 (1), p. 67–74, 2003, ISSN: 0962-1075. Résumé | BibTeX | Étiquettes: Aedes, Base Sequence, Cell Surface, Chimera, Cloning, Developmental, Female, Gene Expression Regulation, Genetic, Insect Proteins, Male, Messenger, Models, Molecular, Mutagenesis, Promoter Regions, Receptors, Reverse Transcriptase Polymerase Chain Reaction, RNA, Sequence Alignment, Signal Transduction, Site-Directed, Transfection @article{luna_characterization_2003, title = {Characterization of three Toll-like genes from mosquito Aedes aegypti}, author = { C. Luna and N. T. Hoa and J. Zhang and S. M. Kanzok and S. E. Brown and Jean-Luc Imler and D. L. Knudson and L. Zheng}, issn = {0962-1075}, year = {2003}, date = {2003-02-01}, journal = {Insect Molecular Biology}, volume = {12}, number = {1}, pages = {67--74}, abstract = {Three Toll-related genes (AeToll1A, AeToll1B and AeToll5) were cloned and characterized from the yellow fever vector mosquito, Aedes aegypti. All three genes exhibited high levels of amino acid sequence similarity with Drosophila melanogaster (Dm)Toll1 and DmTehao (Toll5). AeToll1A and AeToll1B are 1124 and 1076 amino acid residues long, respectively. Both contain a carboxyl extension downstream of the Toll/interleukin-1 receptor (TIR) domain. AeToll5 is 1007 residues long and, like DmTehao, lacks the carboxyl terminal extension. Expression of these three genes was examined throughout development and after immune challenge. Both AeToll1A and AeToll5, like their Drosophila counterparts, activate transcription of drosomycin promoter in both Aedes and Drosophila cell lines. Deletion of the carboxyl extension of AeToll1A did not result in a further elevated level of the antifungal response. The intracellular signalling process appears to be species specific based on two observations. (1) DmToll is completely inactive in an Aedes cell line, suggesting a higher specificity requirement for DmToll in the intracellular signalling process. (2) Only one of three amino acid residues essential for DmToll function is required for AeToll1A function.}, keywords = {Aedes, Base Sequence, Cell Surface, Chimera, Cloning, Developmental, Female, Gene Expression Regulation, Genetic, Insect Proteins, Male, Messenger, Models, Molecular, Mutagenesis, Promoter Regions, Receptors, Reverse Transcriptase Polymerase Chain Reaction, RNA, Sequence Alignment, Signal Transduction, Site-Directed, Transfection}, pubstate = {published}, tppubtype = {article} } Three Toll-related genes (AeToll1A, AeToll1B and AeToll5) were cloned and characterized from the yellow fever vector mosquito, Aedes aegypti. All three genes exhibited high levels of amino acid sequence similarity with Drosophila melanogaster (Dm)Toll1 and DmTehao (Toll5). AeToll1A and AeToll1B are 1124 and 1076 amino acid residues long, respectively. Both contain a carboxyl extension downstream of the Toll/interleukin-1 receptor (TIR) domain. AeToll5 is 1007 residues long and, like DmTehao, lacks the carboxyl terminal extension. Expression of these three genes was examined throughout development and after immune challenge. Both AeToll1A and AeToll5, like their Drosophila counterparts, activate transcription of drosomycin promoter in both Aedes and Drosophila cell lines. Deletion of the carboxyl extension of AeToll1A did not result in a further elevated level of the antifungal response. The intracellular signalling process appears to be species specific based on two observations. (1) DmToll is completely inactive in an Aedes cell line, suggesting a higher specificity requirement for DmToll in the intracellular signalling process. (2) Only one of three amino acid residues essential for DmToll function is required for AeToll1A function. |
2001 |
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| 14. | Levashina, Elena A; Moita, L F; Blandin, Stéphanie A; Vriend, G; Lagueux, Marie; Kafatos, F C Conserved role of a complement-like protein in phagocytosis revealed by dsRNA knockout in cultured cells of the mosquito, Anopheles gambiae Article de journal Cell, 104 (5), p. 709–718, 2001, ISSN: 0092-8674. Résumé | BibTeX | Étiquettes: alpha-Macroglobulins, Anopheles, Cells, Cloning, Complement C3, Cultured, DNA Fragmentation, Double-Stranded, Female, Genetic, Gram-Negative Bacteria, Hemocytes, Insect Proteins, Molecular, Nucleic Acid Denaturation, Phagocytosis, Protein Structure, RNA, Tertiary, Transcription @article{levashina_conserved_2001, title = {Conserved role of a complement-like protein in phagocytosis revealed by dsRNA knockout in cultured cells of the mosquito, Anopheles gambiae}, author = { Elena A. Levashina and L. F. Moita and Stéphanie A. Blandin and G. Vriend and Marie Lagueux and F. C. Kafatos}, issn = {0092-8674}, year = {2001}, date = {2001-01-01}, journal = {Cell}, volume = {104}, number = {5}, pages = {709--718}, abstract = {We characterize a novel hemocyte-specific acute phase glycoprotein from the malaria vector, Anopheles gambiae. It shows substantial structural and functional similarities, including the highly conserved thioester motif, to both a central component of mammalian complement system, factor C3, and to a pan-protease inhibitor, alpha2-macroglobulin. Most importantly, this protein serves as a complement-like opsonin and promotes phagocytosis of some Gram-negative bacteria in a mosquito hemocyte-like cell line. Chemical inactivation by methylamine and depletion by double-stranded RNA knockout demonstrate that this function is dependent on the internal thioester bond. This evidence of a complement-like function in a protostome animal adds substantially to the accumulating evidence of a common ancestry of immune defenses in insects and vertebrates.}, keywords = {alpha-Macroglobulins, Anopheles, Cells, Cloning, Complement C3, Cultured, DNA Fragmentation, Double-Stranded, Female, Genetic, Gram-Negative Bacteria, Hemocytes, Insect Proteins, Molecular, Nucleic Acid Denaturation, Phagocytosis, Protein Structure, RNA, Tertiary, Transcription}, pubstate = {published}, tppubtype = {article} } We characterize a novel hemocyte-specific acute phase glycoprotein from the malaria vector, Anopheles gambiae. It shows substantial structural and functional similarities, including the highly conserved thioester motif, to both a central component of mammalian complement system, factor C3, and to a pan-protease inhibitor, alpha2-macroglobulin. Most importantly, this protein serves as a complement-like opsonin and promotes phagocytosis of some Gram-negative bacteria in a mosquito hemocyte-like cell line. Chemical inactivation by methylamine and depletion by double-stranded RNA knockout demonstrate that this function is dependent on the internal thioester bond. This evidence of a complement-like function in a protostome animal adds substantially to the accumulating evidence of a common ancestry of immune defenses in insects and vertebrates. |
1999 |
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| 13. | Lowenberger, C A; Smartt, C T; Bulet, Philippe; Ferdig, M T; Severson, D W; Hoffmann, Jules A; Christensen, B M Insect immunity: molecular cloning, expression, and characterization of cDNAs and genomic DNA encoding three isoforms of insect defensin in Aedes aegypti Article de journal Insect Mol. Biol., 8 (1), p. 107–118, 1999, ISSN: 0962-1075. Résumé | BibTeX | Étiquettes: Aedes, Amino Acid, Base Sequence, Blotting, Chromosome Mapping, Cloning, Complementary, Defensins, DNA, Gene Expression, Hemolymph, Molecular, Northern, Protein Isoforms, Proteins, Sequence Homology @article{lowenberger_insect_1999, title = {Insect immunity: molecular cloning, expression, and characterization of cDNAs and genomic DNA encoding three isoforms of insect defensin in Aedes aegypti}, author = { C. A. Lowenberger and C. T. Smartt and Philippe Bulet and M. T. Ferdig and D. W. Severson and Jules A. Hoffmann and B. M. Christensen}, issn = {0962-1075}, year = {1999}, date = {1999-02-01}, journal = {Insect Mol. Biol.}, volume = {8}, number = {1}, pages = {107--118}, abstract = {Aedes aegypti were immune activated by injection with bacteria, and the expression of insect defensins was measured over time. Northern analyses indicated that defensin transcriptional activity continued for at least 21 days after bacterial injection, and up to 10 days after saline inoculation. Mature defensin levels in the haemolymph reached approximately 45 microM at 24 h post inoculation. cDNAs encoding the preprodefensins of three previously described mature Ae. aegypti defensins were amplified by PCR, cloned and sequenced. Genomic clones were amplified using primers designed against the cDNA sequence. Sequence comparison indicates that there is significant inter- and intra-isoform variability in the signal peptide and prodefensin sequences of defensin genes. Preprodefensin sequences of isoforms A and B are very similar, consisting of a signal peptide region of twenty amino acids, a prodefensin region of thirty-eight amino acids and a forty amino acid mature peptide domain. The sequence encoding isoform C is significantly different, comprising a signal peptide region of twenty-three amino acids, a prodefensin region of thirty-six amino acids, and the mature protein domain of forty amino acids. Analysis of the genomic clones of each isoform revealed one intron spatially conserved in the prodefensin region of all sequences. The intron in isoforms A and B is 64 nt long, and except for a 4 nt substitution in one clone, these intron sequences are identical. The intron in isoform C is 76 nt long and does not share significant identity with the intron sequences of isoforms A or B. The defensin gene mapped to chromosome 3, between two known loci, blt and LF168.}, keywords = {Aedes, Amino Acid, Base Sequence, Blotting, Chromosome Mapping, Cloning, Complementary, Defensins, DNA, Gene Expression, Hemolymph, Molecular, Northern, Protein Isoforms, Proteins, Sequence Homology}, pubstate = {published}, tppubtype = {article} } Aedes aegypti were immune activated by injection with bacteria, and the expression of insect defensins was measured over time. Northern analyses indicated that defensin transcriptional activity continued for at least 21 days after bacterial injection, and up to 10 days after saline inoculation. Mature defensin levels in the haemolymph reached approximately 45 microM at 24 h post inoculation. cDNAs encoding the preprodefensins of three previously described mature Ae. aegypti defensins were amplified by PCR, cloned and sequenced. Genomic clones were amplified using primers designed against the cDNA sequence. Sequence comparison indicates that there is significant inter- and intra-isoform variability in the signal peptide and prodefensin sequences of defensin genes. Preprodefensin sequences of isoforms A and B are very similar, consisting of a signal peptide region of twenty amino acids, a prodefensin region of thirty-eight amino acids and a forty amino acid mature peptide domain. The sequence encoding isoform C is significantly different, comprising a signal peptide region of twenty-three amino acids, a prodefensin region of thirty-six amino acids, and the mature protein domain of forty amino acids. Analysis of the genomic clones of each isoform revealed one intron spatially conserved in the prodefensin region of all sequences. The intron in isoforms A and B is 64 nt long, and except for a 4 nt substitution in one clone, these intron sequences are identical. The intron in isoform C is 76 nt long and does not share significant identity with the intron sequences of isoforms A or B. The defensin gene mapped to chromosome 3, between two known loci, blt and LF168. |
1998 |
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Articles de journaux |
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| 12. | Uttenweiler-Joseph, S; Moniatte, M; Lagueux, Marie; Dorsselaer, Van A; Hoffmann, Jules A; Bulet, Philippe Differential display of peptides induced during the immune response of Drosophila: a matrix-assisted laser desorption ionization time-of-flight mass spectrometry study Article de journal Proc. Natl. Acad. Sci. U.S.A., 95 (19), p. 11342–11347, 1998, ISSN: 0027-8424. Résumé | BibTeX | Étiquettes: Bacteria, Chromatography, Cloning, Hemolymph, High Pressure Liquid, Immunity, Insect Proteins, Mass, Matrix-Assisted Laser Desorption-Ionization, Messenger, Molecular, Peptides, Protein Precursors, RNA, Sequence Analysis, Spectrometry, Time Factors @article{uttenweiler-joseph_differential_1998, title = {Differential display of peptides induced during the immune response of Drosophila: a matrix-assisted laser desorption ionization time-of-flight mass spectrometry study}, author = { S. Uttenweiler-Joseph and M. Moniatte and Marie Lagueux and A. Van Dorsselaer and Jules A. Hoffmann and Philippe Bulet}, issn = {0027-8424}, year = {1998}, date = {1998-09-01}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, volume = {95}, number = {19}, pages = {11342--11347}, abstract = {We have developed an approach based on a differential mass spectrometric analysis to detect molecules induced during the immune response of Drosophila, regardless of their biological activities. For this, we have applied directly matrix-assisted laser desorption/ionization MS to hemolymph samples from individual flies before and after an immune challenge. This method provided precise information on the molecular masses of immune-induced molecules and allowed the detection, in the molecular range of 1.5-11 kDa, of 24 Drosophila immune-induced molecules (DIMs). These molecules are all peptides, and four correspond to already characterized antimicrobial peptides. We have further analyzed the induction of the various peptides by immune challenge in wild-type flies and in mutants with a compromised antimicrobial response. We also describe a methodology combining matrix-assisted laser desorption ionization time-of-flight MS, HPLC, and Edman degradation, which yielded the peptide sequence of three of the DIMs. Finally, molecular cloning and Northern blot analyses revealed that one of the DIMs is produced as a prepropeptide and is inducible on a bacterial challenge.}, keywords = {Bacteria, Chromatography, Cloning, Hemolymph, High Pressure Liquid, Immunity, Insect Proteins, Mass, Matrix-Assisted Laser Desorption-Ionization, Messenger, Molecular, Peptides, Protein Precursors, RNA, Sequence Analysis, Spectrometry, Time Factors}, pubstate = {published}, tppubtype = {article} } We have developed an approach based on a differential mass spectrometric analysis to detect molecules induced during the immune response of Drosophila, regardless of their biological activities. For this, we have applied directly matrix-assisted laser desorption/ionization MS to hemolymph samples from individual flies before and after an immune challenge. This method provided precise information on the molecular masses of immune-induced molecules and allowed the detection, in the molecular range of 1.5-11 kDa, of 24 Drosophila immune-induced molecules (DIMs). These molecules are all peptides, and four correspond to already characterized antimicrobial peptides. We have further analyzed the induction of the various peptides by immune challenge in wild-type flies and in mutants with a compromised antimicrobial response. We also describe a methodology combining matrix-assisted laser desorption ionization time-of-flight MS, HPLC, and Edman degradation, which yielded the peptide sequence of three of the DIMs. Finally, molecular cloning and Northern blot analyses revealed that one of the DIMs is produced as a prepropeptide and is inducible on a bacterial challenge. |
| 11. | Levashina, Elena A; Ohresser, S; Lemaitre, Bruno; Imler, Jean-Luc Two distinct pathways can control expression of the gene encoding the Drosophila antimicrobial peptide metchnikowin Article de journal Journal of Molecular Biology, 278 (3), p. 515–527, 1998, ISSN: 0022-2836. Résumé | Liens | BibTeX | Étiquettes: Anti-Infective Agents, Antimicrobial Cationic Peptides, Base Sequence, Cloning, Gene Expression Regulation, Genes, Genetic, Genetically Modified, Glycopeptides, Insect, Insect Proteins, Larva, Molecular, Mutation, Peptides, Promoter Regions, Recombinant Fusion Proteins, Reporter, Restriction Mapping, Transcription @article{levashina_two_1998, title = {Two distinct pathways can control expression of the gene encoding the Drosophila antimicrobial peptide metchnikowin}, author = { Elena A. Levashina and S. Ohresser and Bruno Lemaitre and Jean-Luc Imler}, doi = {10.1006/jmbi.1998.1705}, issn = {0022-2836}, year = {1998}, date = {1998-01-01}, journal = {Journal of Molecular Biology}, volume = {278}, number = {3}, pages = {515--527}, abstract = {Metchnikowin is a recently discovered proline-rich peptide from Drosophila with antibacterial and antifungal properties. Like most other antimicrobial peptides from insects, its expression is immune-inducible. Here we present evidence that induction of metchnikowin gene expression can be mediated either by the TOLL pathway or by the imd gene product. We show that the gene remains inducible in Toll-deficient mutants, in which the antifungal response is blocked, as well as in imd mutants, which fail to mount an antibacterial response. However, in Toll-deficient;imd double mutants, metchnikowin gene expression can no longer be detected after immune challenge. Our results suggest that expression of this peptide with dual activity can be triggered by signals generated by either bacterial or fungal infection. Cloning of the metchnikowin gene revealed the presence in the 5' flanking region of several putative cis-regulatory motifs characterized in the promoters of insect immune genes: namely, Rel sites, GATA motifs, interferon consensus response elements and NF-IL6 response elements. Establishment of transgenic fly lines in which the GFP reporter gene was placed under the control of 1.5 kb of metchnikowin gene upstream sequences indicates that this fragment is able to confer full immune inducibility and tissue specificity of expression on the transgene.}, keywords = {Anti-Infective Agents, Antimicrobial Cationic Peptides, Base Sequence, Cloning, Gene Expression Regulation, Genes, Genetic, Genetically Modified, Glycopeptides, Insect, Insect Proteins, Larva, Molecular, Mutation, Peptides, Promoter Regions, Recombinant Fusion Proteins, Reporter, Restriction Mapping, Transcription}, pubstate = {published}, tppubtype = {article} } Metchnikowin is a recently discovered proline-rich peptide from Drosophila with antibacterial and antifungal properties. Like most other antimicrobial peptides from insects, its expression is immune-inducible. Here we present evidence that induction of metchnikowin gene expression can be mediated either by the TOLL pathway or by the imd gene product. We show that the gene remains inducible in Toll-deficient mutants, in which the antifungal response is blocked, as well as in imd mutants, which fail to mount an antibacterial response. However, in Toll-deficient;imd double mutants, metchnikowin gene expression can no longer be detected after immune challenge. Our results suggest that expression of this peptide with dual activity can be triggered by signals generated by either bacterial or fungal infection. Cloning of the metchnikowin gene revealed the presence in the 5' flanking region of several putative cis-regulatory motifs characterized in the promoters of insect immune genes: namely, Rel sites, GATA motifs, interferon consensus response elements and NF-IL6 response elements. Establishment of transgenic fly lines in which the GFP reporter gene was placed under the control of 1.5 kb of metchnikowin gene upstream sequences indicates that this fragment is able to confer full immune inducibility and tissue specificity of expression on the transgene. |
1996 |
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Articles de journaux |
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| 10. | Richman, A M; Bulet, Philippe; Hetru, Charles; Barillas-Mury, Carolina; Hoffmann, Jules A; Kafalos, Fotis C Inducible immune factors of the vector mosquito Anopheles gambiae: biochemical purification of a defensin antibacterial peptide and molecular cloning of preprodefensin cDNA Article de journal Insect Mol. Biol., 5 (3), p. 203–210, 1996, ISSN: 0962-1075. Résumé | BibTeX | Étiquettes: Amino Acid, Anopheles, Base Sequence, Blood Bactericidal Activity, Blood Proteins, Cloning, Complementary, Defensins, DNA, Escherichia coli, Female, Gene Expression, Genes, Insect, Insect Vectors, Larva, Micrococcus luteus, Molecular, Sequence Homology @article{richman_inducible_1996, title = {Inducible immune factors of the vector mosquito Anopheles gambiae: biochemical purification of a defensin antibacterial peptide and molecular cloning of preprodefensin cDNA}, author = { A. M. Richman and Philippe Bulet and Charles Hetru and Carolina Barillas-Mury and Jules A. Hoffmann and Fotis C. Kafalos}, issn = {0962-1075}, year = {1996}, date = {1996-08-01}, journal = {Insect Mol. Biol.}, volume = {5}, number = {3}, pages = {203--210}, abstract = {Larvae of the mosquito vector of human malaria, Anopheles gambiae, were inoculated with bacteria and extracts were biochemically fractionated by reverse-phase HPLC. Multiple induced polypeptides and antibacterial activities were observed following bacterial infection, including a member of the insect defensin family of antibacterial proteins. A cDNA encoding An. gambiae preprodefensin was isolated using PCR primers based on phylogenetically conserved sequences. The mature peptide is highly conserved, but the signal and propeptide segments are not, relative to corresponding defensin sequences of other insects. Defensin expression is induced in response to bacterial infection, in both adult and larval stages. In contrast, pupae express defensin mRNA constitutively. Defensin expression may prove a valuable molecular marker to monitor the An. gambiae host response to infection by parasitic protozoa of medical importance.}, keywords = {Amino Acid, Anopheles, Base Sequence, Blood Bactericidal Activity, Blood Proteins, Cloning, Complementary, Defensins, DNA, Escherichia coli, Female, Gene Expression, Genes, Insect, Insect Vectors, Larva, Micrococcus luteus, Molecular, Sequence Homology}, pubstate = {published}, tppubtype = {article} } Larvae of the mosquito vector of human malaria, Anopheles gambiae, were inoculated with bacteria and extracts were biochemically fractionated by reverse-phase HPLC. Multiple induced polypeptides and antibacterial activities were observed following bacterial infection, including a member of the insect defensin family of antibacterial proteins. A cDNA encoding An. gambiae preprodefensin was isolated using PCR primers based on phylogenetically conserved sequences. The mature peptide is highly conserved, but the signal and propeptide segments are not, relative to corresponding defensin sequences of other insects. Defensin expression is induced in response to bacterial infection, in both adult and larval stages. In contrast, pupae express defensin mRNA constitutively. Defensin expression may prove a valuable molecular marker to monitor the An. gambiae host response to infection by parasitic protozoa of medical importance. |
1995 |
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| 9. | Levashina, Elena A; Ohresser, S; Bulet, Philippe ; Reichhart, Jean-Marc ; Hetru, Charles ; Hoffmann, Jules A Metchnikowin, a novel immune-inducible proline-rich peptide from Drosophila with antibacterial and antifungal properties Article de journal Eur. J. Biochem., 233 (2), p. 694–700, 1995, ISSN: 0014-2956. Résumé | BibTeX | Étiquettes: Anti-Bacterial Agents, Antifungal Agents, Antimicrobial Cationic Peptides, Bacteria, Base Sequence, Cells, Chromosome Mapping, Cloning, Cultured, Genetic, Molecular, Peptides, Proline, Transcription @article{levashina_metchnikowin_1995, title = {Metchnikowin, a novel immune-inducible proline-rich peptide from Drosophila with antibacterial and antifungal properties}, author = { Elena A. Levashina and S. Ohresser and Philippe Bulet and Jean-Marc Reichhart and Charles Hetru and Jules A. Hoffmann}, issn = {0014-2956}, year = {1995}, date = {1995-10-01}, journal = {Eur. J. Biochem.}, volume = {233}, number = {2}, pages = {694--700}, abstract = {One of the characteristics of the host defense of higher insects is the rapid and transient synthesis of a variety of potent antimicrobial peptides. To date, several distinct inducible antimicrobial peptides or peptide families have been totally or partially characterized. We present here the isolation and characterization of a novel 26-residue proline-rich immune-inducible peptide from Drosophila, which exhibits both antibacterial (Gram-positive) and antifungal activities. Peptide sequencing and cDNA cloning indicate the presense of two isoforms in our Drosophila Oregon strain, which differ by one residue (His compared to Arg) as a consequence of a single nucleotide change. The gene, which maps in position 52A1-2 on the right arm of the second chromosome, is expressed in the fat body after immune challenge. The novel peptide, which we propose to name metchnikowin, is a member of a family of proline-rich peptides, and we discuss the possible evolutionary relationships within this family.}, keywords = {Anti-Bacterial Agents, Antifungal Agents, Antimicrobial Cationic Peptides, Bacteria, Base Sequence, Cells, Chromosome Mapping, Cloning, Cultured, Genetic, Molecular, Peptides, Proline, Transcription}, pubstate = {published}, tppubtype = {article} } One of the characteristics of the host defense of higher insects is the rapid and transient synthesis of a variety of potent antimicrobial peptides. To date, several distinct inducible antimicrobial peptides or peptide families have been totally or partially characterized. We present here the isolation and characterization of a novel 26-residue proline-rich immune-inducible peptide from Drosophila, which exhibits both antibacterial (Gram-positive) and antifungal activities. Peptide sequencing and cDNA cloning indicate the presense of two isoforms in our Drosophila Oregon strain, which differ by one residue (His compared to Arg) as a consequence of a single nucleotide change. The gene, which maps in position 52A1-2 on the right arm of the second chromosome, is expressed in the fat body after immune challenge. The novel peptide, which we propose to name metchnikowin, is a member of a family of proline-rich peptides, and we discuss the possible evolutionary relationships within this family. |
| 8. | Cornet, B; Bonmatin, J M; Hetru, Charles; Hoffmann, Jules A; Ptak, M; Vovelle, F Refined three-dimensional solution structure of insect defensin A Article de journal Structure, 3 (5), p. 435–448, 1995, ISSN: 0969-2126. Résumé | BibTeX | Étiquettes: Amino Acid, Bacteriolysis, Chemistry, Defensins, Diptera, Gram-Positive Bacteria, Hydrogen Bonding, Insect Hormones, Magnetic Resonance Spectroscopy, Models, Molecular, Physical, Physicochemical Phenomena, Protein Conformation, Recombinant Proteins, Sequence Homology, Solutions, Structure-Activity Relationship @article{cornet_refined_1995, title = {Refined three-dimensional solution structure of insect defensin A}, author = { B. Cornet and J. M. Bonmatin and Charles Hetru and Jules A. Hoffmann and M. Ptak and F. Vovelle}, issn = {0969-2126}, year = {1995}, date = {1995-05-01}, journal = {Structure}, volume = {3}, number = {5}, pages = {435--448}, abstract = {BACKGROUND: Insect defensin A is a basic 4 kDa protein secreted by Phormia terranovae larvae in response to bacterial challenges or injuries. Previous biological tests suggest that the bacterial cytoplasmic membrane is the target of defensin A. The structural study of this protein is the first step towards establishing a structure-activity relationship and forms the basis for understanding its antibiotic activity at the molecular level. RESULTS: We describe a refined model of the three-dimensional structure of defensin A derived from an extensive analysis of 786 inter-proton nuclear Overhauser effects. The backbone fold involves an N-terminal loop and an alpha-helical fragment followed by an antiparallel beta-structure. The helix and the beta-structure are connected by two of the three disulphide bridges present in defensin A, forming a so-called 'cysteine-stabilized alpha beta' (CS alpha beta) motif. The N-terminal loop, which is locally well defined, can occupy different positions with respect to the other moieties of the molecule. CONCLUSIONS: The CS alpha beta motif, which forms the core of the defensin A structure, appears to be a common organization for several families of small proteins with toxic properties. The distribution of amino acid side chains in the protein structure creates several hydrophobic or hydrophilic patches. This leads us to propose that the initial step in the action of positively charged defensin A molecules with cytoplasmic membranes may involve interactions with acidic phospholipids.}, keywords = {Amino Acid, Bacteriolysis, Chemistry, Defensins, Diptera, Gram-Positive Bacteria, Hydrogen Bonding, Insect Hormones, Magnetic Resonance Spectroscopy, Models, Molecular, Physical, Physicochemical Phenomena, Protein Conformation, Recombinant Proteins, Sequence Homology, Solutions, Structure-Activity Relationship}, pubstate = {published}, tppubtype = {article} } BACKGROUND: Insect defensin A is a basic 4 kDa protein secreted by Phormia terranovae larvae in response to bacterial challenges or injuries. Previous biological tests suggest that the bacterial cytoplasmic membrane is the target of defensin A. The structural study of this protein is the first step towards establishing a structure-activity relationship and forms the basis for understanding its antibiotic activity at the molecular level. RESULTS: We describe a refined model of the three-dimensional structure of defensin A derived from an extensive analysis of 786 inter-proton nuclear Overhauser effects. The backbone fold involves an N-terminal loop and an alpha-helical fragment followed by an antiparallel beta-structure. The helix and the beta-structure are connected by two of the three disulphide bridges present in defensin A, forming a so-called 'cysteine-stabilized alpha beta' (CS alpha beta) motif. The N-terminal loop, which is locally well defined, can occupy different positions with respect to the other moieties of the molecule. CONCLUSIONS: The CS alpha beta motif, which forms the core of the defensin A structure, appears to be a common organization for several families of small proteins with toxic properties. The distribution of amino acid side chains in the protein structure creates several hydrophobic or hydrophilic patches. This leads us to propose that the initial step in the action of positively charged defensin A molecules with cytoplasmic membranes may involve interactions with acidic phospholipids. |
1994 |
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| 7. | Fehlbaum, P; Bulet, Philippe; Michaut, L; Lagueux, Marie; Broekaert, W F; Hetru, Charles; Hoffmann, Jules A Insect immunity. Septic injury of Drosophila induces the synthesis of a potent antifungal peptide with sequence homology to plant antifungal peptides Article de journal J. Biol. Chem., 269 (52), p. 33159–33163, 1994, ISSN: 0021-9258. Résumé | BibTeX | Étiquettes: Amino Acid, Antifungal Agents, Base Sequence, Cloning, Complementary, DNA, Insect Proteins, Male, Messenger, Microbial Sensitivity Tests, Molecular, Peptide Biosynthesis, Peptides, Plants, Protein Biosynthesis, Protein Precursors, Proteins, RNA, Sequence Homology @article{fehlbaum_insect_1994, title = {Insect immunity. Septic injury of Drosophila induces the synthesis of a potent antifungal peptide with sequence homology to plant antifungal peptides}, author = {P. Fehlbaum and Philippe Bulet and L. Michaut and Marie Lagueux and W. F. Broekaert and Charles Hetru and Jules A. Hoffmann}, issn = {0021-9258}, year = {1994}, date = {1994-12-01}, journal = {J. Biol. Chem.}, volume = {269}, number = {52}, pages = {33159--33163}, abstract = {In response to a septic injury (pricking with a bacteria-soaked needle) larvae and adults of Drosophila produce considerable amounts of a 44-residue peptide containing 8 cysteines engaged in intramolecular disulfide bridges. The peptide is synthesized in the fat body, a functional homologue of the mammalian liver, and secreted into the blood of the insect. It exhibits potent antifungal activity but is inactive against bacteria. This novel inducible peptide, which we propose to name drosomycin, shows a significant homology with a family of 5-kDa cysteine-rich plant antifungal peptides recently isolated from seeds of Brassicaceae. This finding underlines that plants and insects can rely on similar molecules in their innate host defense.}, keywords = {Amino Acid, Antifungal Agents, Base Sequence, Cloning, Complementary, DNA, Insect Proteins, Male, Messenger, Microbial Sensitivity Tests, Molecular, Peptide Biosynthesis, Peptides, Plants, Protein Biosynthesis, Protein Precursors, Proteins, RNA, Sequence Homology}, pubstate = {published}, tppubtype = {article} } In response to a septic injury (pricking with a bacteria-soaked needle) larvae and adults of Drosophila produce considerable amounts of a 44-residue peptide containing 8 cysteines engaged in intramolecular disulfide bridges. The peptide is synthesized in the fat body, a functional homologue of the mammalian liver, and secreted into the blood of the insect. It exhibits potent antifungal activity but is inactive against bacteria. This novel inducible peptide, which we propose to name drosomycin, shows a significant homology with a family of 5-kDa cysteine-rich plant antifungal peptides recently isolated from seeds of Brassicaceae. This finding underlines that plants and insects can rely on similar molecules in their innate host defense. |
| 6. | Dimarcq, Jean-Luc; Hoffmann, Danièle ; Meister, Marie ; Bulet, Philippe ; Lanot, R; Reichhart, Jean-Marc ; Hoffmann, Jules A Characterization and transcriptional profiles of a Drosophila gene encoding an insect defensin. A study in insect immunity Article de journal Eur. J. Biochem., 221 (1), p. 201–209, 1994, ISSN: 0014-2956. Résumé | BibTeX | Étiquettes: Base Sequence, Blood Proteins, Chromosome Mapping, Cloning, Complementary, Defensins, DNA, Gene Expression, Genetic, Gram-Positive Bacteria, Larva, Molecular, Molecular Structure, Nucleic Acid, Protein Precursors, Regulatory Sequences, Transcription @article{dimarcq_characterization_1994, title = {Characterization and transcriptional profiles of a Drosophila gene encoding an insect defensin. A study in insect immunity}, author = { Jean-Luc Dimarcq and Danièle Hoffmann and Marie Meister and Philippe Bulet and R. Lanot and Jean-Marc Reichhart and Jules A. Hoffmann}, issn = {0014-2956}, year = {1994}, date = {1994-04-01}, journal = {Eur. J. Biochem.}, volume = {221}, number = {1}, pages = {201--209}, abstract = {Insect defensins are a family of 4-kDa, cationic, inducible antibacterial peptides which bear six cysteine residues engaged in three intramolecular disulfide bridges. They owe their name to certain sequence similarities with defensins from mammalian neutrophiles and macrophages. We report the characterization of a novel defensin isoform from Drosophila and the cloning of the gene encoding a preprodefensin. The gene, which is intronless and present in a single copy/haploid genome, maps at position 46CD on the right arm of the second chromosome. The analysis of the upstream region of the gene reveals the presence of multiple putative cis-regulatory sequences similar to mammalian regulatory motifs of acute-phase-response genes. Transcriptional profiles indicate that the Drosophila defensin gene is induced by bacterial challenge with acute-phase kinetics. It is also expressed in the absence of immune challenge during metamorphosis. These and other data on the Drosophila defensin gene lead us to suggest that insect and mammalian defensins have evolved independently.}, keywords = {Base Sequence, Blood Proteins, Chromosome Mapping, Cloning, Complementary, Defensins, DNA, Gene Expression, Genetic, Gram-Positive Bacteria, Larva, Molecular, Molecular Structure, Nucleic Acid, Protein Precursors, Regulatory Sequences, Transcription}, pubstate = {published}, tppubtype = {article} } Insect defensins are a family of 4-kDa, cationic, inducible antibacterial peptides which bear six cysteine residues engaged in three intramolecular disulfide bridges. They owe their name to certain sequence similarities with defensins from mammalian neutrophiles and macrophages. We report the characterization of a novel defensin isoform from Drosophila and the cloning of the gene encoding a preprodefensin. The gene, which is intronless and present in a single copy/haploid genome, maps at position 46CD on the right arm of the second chromosome. The analysis of the upstream region of the gene reveals the presence of multiple putative cis-regulatory sequences similar to mammalian regulatory motifs of acute-phase-response genes. Transcriptional profiles indicate that the Drosophila defensin gene is induced by bacterial challenge with acute-phase kinetics. It is also expressed in the absence of immune challenge during metamorphosis. These and other data on the Drosophila defensin gene lead us to suggest that insect and mammalian defensins have evolved independently. |
1993 |
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Articles de journaux |
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| 5. | Bulet, Philippe; Dimarcq, Jean-Luc; Hetru, Charles; Lagueux, Marie; Charlet, Maurice; Hegy, G; Dorsselaer, Alan Van; Hoffmann, Jules A A novel inducible antibacterial peptide of Drosophila carries an O-glycosylated substitution Article de journal J. Biol. Chem., 268 (20), p. 14893–14897, 1993, ISSN: 0021-9258. Résumé | BibTeX | Étiquettes: Anti-Bacterial Agents, Base Sequence, Carbohydrates, Cloning, DNA, Escherichia coli, Gas Chromatography-Mass Spectrometry, Glycopeptides, Glycosylation, Molecular @article{bulet_novel_1993, title = {A novel inducible antibacterial peptide of Drosophila carries an O-glycosylated substitution}, author = {Philippe Bulet and Jean-Luc Dimarcq and Charles Hetru and Marie Lagueux and Maurice Charlet and G. Hegy and Alan Van Dorsselaer and Jules A. Hoffmann}, issn = {0021-9258}, year = {1993}, date = {1993-07-01}, journal = {J. Biol. Chem.}, volume = {268}, number = {20}, pages = {14893--14897}, abstract = {One of the facets of the host defense of higher insects is the rapid and transient synthesis, following bacterial challenge or trauma, of a battery of potent antibacterial peptides (Steiner, H., Hultmark, D., Engström, A., Bennich, H., and Boman, H. G. (1981) Nature 292, 246-248). The best characterized of these peptides are the cecropins (ibid.), 4-kDa peptides devoid of cysteines, and the insect defensins (Hoffmann, J. A., and Hetru, C. (1992) Immunol. Today 13, 411-415), 4-kDa peptides with three intramolecular disulfide bridges. Several other inducible antibacterial peptides have been characterized only at the level of their amino acid sequences (Hoffmann, J. A., Dimarcq, J. L., and Bulet, P. (1992) Médecine & Sciences 8, 432-439). We report here the isolation of a novel 19-residue proline-rich inducible antibacterial peptide from Drosophila. In contrast to all previous reports on antibacterial peptides, this molecule carries a substitution as evidenced by molecular mass determinations; our data show that this reflects the O-glycosylation of a Thr residue by an N-acetylgalactosamine plus a galactose. A synthetic nonsubstituted peptide of identical amino acid sequence has an activity several times lower (5-10) than the native compound. Our data suggest that this substitution represents a post-translational modification essential for the full biological activity of this novel peptide.}, keywords = {Anti-Bacterial Agents, Base Sequence, Carbohydrates, Cloning, DNA, Escherichia coli, Gas Chromatography-Mass Spectrometry, Glycopeptides, Glycosylation, Molecular}, pubstate = {published}, tppubtype = {article} } One of the facets of the host defense of higher insects is the rapid and transient synthesis, following bacterial challenge or trauma, of a battery of potent antibacterial peptides (Steiner, H., Hultmark, D., Engström, A., Bennich, H., and Boman, H. G. (1981) Nature 292, 246-248). The best characterized of these peptides are the cecropins (ibid.), 4-kDa peptides devoid of cysteines, and the insect defensins (Hoffmann, J. A., and Hetru, C. (1992) Immunol. Today 13, 411-415), 4-kDa peptides with three intramolecular disulfide bridges. Several other inducible antibacterial peptides have been characterized only at the level of their amino acid sequences (Hoffmann, J. A., Dimarcq, J. L., and Bulet, P. (1992) Médecine & Sciences 8, 432-439). We report here the isolation of a novel 19-residue proline-rich inducible antibacterial peptide from Drosophila. In contrast to all previous reports on antibacterial peptides, this molecule carries a substitution as evidenced by molecular mass determinations; our data show that this reflects the O-glycosylation of a Thr residue by an N-acetylgalactosamine plus a galactose. A synthetic nonsubstituted peptide of identical amino acid sequence has an activity several times lower (5-10) than the native compound. Our data suggest that this substitution represents a post-translational modification essential for the full biological activity of this novel peptide. |
| 4. | Kappler, Christine; Meister, Marie ; Lagueux, Marie ; Gateff, E; Hoffmann, Jules A; Reichhart, Jean-Marc Insect immunity. Two 17 bp repeats nesting a kappa B-related sequence confer inducibility to the diptericin gene and bind a polypeptide in bacteria-challenged Drosophila Article de journal EMBO J., 12 (4), p. 1561–1568, 1993, ISSN: 0261-4189. Résumé | BibTeX | Étiquettes: Anti-Bacterial Agents, Base Sequence, Cloning, Gene Expression Regulation, Genes, Genetic, Genetically Modified, Insect, Insect Hormones, Insect Proteins, Lipopolysaccharides, Messenger, Molecular, NF-kappa B, Nucleic Acid, Oligodeoxyribonucleotides, Promoter Regions, Regulatory Sequences, RNA, Transfection @article{kappler_insect_1993, title = {Insect immunity. Two 17 bp repeats nesting a kappa B-related sequence confer inducibility to the diptericin gene and bind a polypeptide in bacteria-challenged Drosophila}, author = { Christine Kappler and Marie Meister and Marie Lagueux and E. Gateff and Jules A. Hoffmann and Jean-Marc Reichhart}, issn = {0261-4189}, year = {1993}, date = {1993-04-01}, journal = {EMBO J.}, volume = {12}, number = {4}, pages = {1561--1568}, abstract = {The Drosophila diptericin gene codes for a 9 kDa antibacterial peptide and is rapidly and transiently expressed in larvae and adults after bacterial challenge. It is also induced in a tumorous Drosophila blood cell line by the addition of lipopolysaccharide (LPS). The promoter of this gene contains two 17 bp repeats located closely upstream of the TATA-box and harbouring a decameric kappa B-related sequence. This study reports that the replacement of the two 17 bp repeats by random sequences abolishes bacteria inducibility in transgenic fly lines. In transfected tumorous blood cells, the replacement of both or either of the 17 bp motifs reduces dramatically LPS inducibility, whereas multiple copies significantly increase the level of transcriptional activation by LPS challenge. A specific DNA-protein binding activity is evidenced in cytoplasmic and nuclear extracts of induced blood cells and fat body. It is absent in controls. It is proposed that induction of the diptericin gene mediated by the two 17 bp repeats occurs via a mechanism similar to that of mammalian NF-kappa B.}, keywords = {Anti-Bacterial Agents, Base Sequence, Cloning, Gene Expression Regulation, Genes, Genetic, Genetically Modified, Insect, Insect Hormones, Insect Proteins, Lipopolysaccharides, Messenger, Molecular, NF-kappa B, Nucleic Acid, Oligodeoxyribonucleotides, Promoter Regions, Regulatory Sequences, RNA, Transfection}, pubstate = {published}, tppubtype = {article} } The Drosophila diptericin gene codes for a 9 kDa antibacterial peptide and is rapidly and transiently expressed in larvae and adults after bacterial challenge. It is also induced in a tumorous Drosophila blood cell line by the addition of lipopolysaccharide (LPS). The promoter of this gene contains two 17 bp repeats located closely upstream of the TATA-box and harbouring a decameric kappa B-related sequence. This study reports that the replacement of the two 17 bp repeats by random sequences abolishes bacteria inducibility in transgenic fly lines. In transfected tumorous blood cells, the replacement of both or either of the 17 bp motifs reduces dramatically LPS inducibility, whereas multiple copies significantly increase the level of transcriptional activation by LPS challenge. A specific DNA-protein binding activity is evidenced in cytoplasmic and nuclear extracts of induced blood cells and fat body. It is absent in controls. It is proposed that induction of the diptericin gene mediated by the two 17 bp repeats occurs via a mechanism similar to that of mammalian NF-kappa B. |
1992 |
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Articles de journaux |
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| 3. | Bonmatin, J M; Bonnat, J L; Gallet, X; Vovelle, F; Ptak, M; Reichhart, Jean-Marc ; Hoffmann, Jules A; Keppi, E; Legrain, M; Achstetter, T Two-dimensional 1H NMR study of recombinant insect defensin A in water: resonance assignments, secondary structure and global folding Article de journal J. Biomol. NMR, 2 (3), p. 235–256, 1992, ISSN: 0925-2738. Résumé | BibTeX | Étiquettes: Defensins, Hydrogen, Insect Hormones, Insects, Magnetic Resonance Spectroscopy, Models, Molecular, Protein Conformation, Recombinant Proteins, Saccharomyces cerevisiae, Thermodynamics @article{bonmatin_two-dimensional_1992, title = {Two-dimensional 1H NMR study of recombinant insect defensin A in water: resonance assignments, secondary structure and global folding}, author = { J. M. Bonmatin and J. L. Bonnat and X. Gallet and F. Vovelle and M. Ptak and Jean-Marc Reichhart and Jules A. Hoffmann and E. Keppi and M. Legrain and T. Achstetter}, issn = {0925-2738}, year = {1992}, date = {1992-01-01}, journal = {J. Biomol. NMR}, volume = {2}, number = {3}, pages = {235--256}, abstract = {A 500 MHz 2D 1H NMR study of recombinant insect defensin A is reported. This defense protein of 40 residues contains 3 disulfide bridges, is positively charged and exhibits antibacterial properties. 2D NMR maps of recombinant defensin A were fully assigned and secondary structure elements were localized. The set of NOE connectivities, 3JNH-alpha H coupling constants as well as 1H/2H exchange rates and delta delta/delta T temperature coefficients of NH protons strongly support the existence of an alpha-helix (residues 14-24) and of an antiparallel beta-sheet (residues 27-40). Models of the backbone folding were generated by using the DISMAN program and energy refined by using the AMBER program. This was done on the basis of: (i) 133 selected NOEs, (ii) 21 dihedral restraints from 3JNH-alpha H coupling constants, (iii) 12 hydrogen bonds mostly deduced from 1H/2H exchange rates or temperature coefficients, in addition to 9 initial disulfide bridge covalent constraints. The two secondary structure elements and the two bends connecting them involve approximately 70% of the total number of residues, which impose some stability in the C-terminal part of the molecule. The remaining N-terminal fragment forms a less well defined loop. This spatial organization, in which a beta-sheet is linked to an alpha-helix by two disulfide bridges and to a large loop by a third disulfide bridge, is rather similar to that found in scorpion charybdotoxin and seems to be partly present in several invertebrate toxins.}, keywords = {Defensins, Hydrogen, Insect Hormones, Insects, Magnetic Resonance Spectroscopy, Models, Molecular, Protein Conformation, Recombinant Proteins, Saccharomyces cerevisiae, Thermodynamics}, pubstate = {published}, tppubtype = {article} } A 500 MHz 2D 1H NMR study of recombinant insect defensin A is reported. This defense protein of 40 residues contains 3 disulfide bridges, is positively charged and exhibits antibacterial properties. 2D NMR maps of recombinant defensin A were fully assigned and secondary structure elements were localized. The set of NOE connectivities, 3JNH-alpha H coupling constants as well as 1H/2H exchange rates and delta delta/delta T temperature coefficients of NH protons strongly support the existence of an alpha-helix (residues 14-24) and of an antiparallel beta-sheet (residues 27-40). Models of the backbone folding were generated by using the DISMAN program and energy refined by using the AMBER program. This was done on the basis of: (i) 133 selected NOEs, (ii) 21 dihedral restraints from 3JNH-alpha H coupling constants, (iii) 12 hydrogen bonds mostly deduced from 1H/2H exchange rates or temperature coefficients, in addition to 9 initial disulfide bridge covalent constraints. The two secondary structure elements and the two bends connecting them involve approximately 70% of the total number of residues, which impose some stability in the C-terminal part of the molecule. The remaining N-terminal fragment forms a less well defined loop. This spatial organization, in which a beta-sheet is linked to an alpha-helix by two disulfide bridges and to a large loop by a third disulfide bridge, is rather similar to that found in scorpion charybdotoxin and seems to be partly present in several invertebrate toxins. |
1990 |
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Articles de journaux |
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| 2. | Dimarcq, Jean-Luc; Zachary, Daniel ; Hoffmann, Jules A; Hoffmann, Danièle ; Reichhart, Jean-Marc Insect immunity: expression of the two major inducible antibacterial peptides, defensin and diptericin, in Phormia terranovae Article de journal EMBO J., 9 (8), p. 2507–2515, 1990, ISSN: 0261-4189. Résumé | BibTeX | Étiquettes: Anti-Bacterial Agents, Base Sequence, Blood Proteins, Cloning, Defensins, Diptera, Gene Expression, Insect Hormones, Insect Proteins, Larva, Molecular, Nucleic Acid Hybridization, Oligonucleotide Probes, Protein Conformation @article{dimarcq_insect_1990, title = {Insect immunity: expression of the two major inducible antibacterial peptides, defensin and diptericin, in Phormia terranovae}, author = { Jean-Luc Dimarcq and Daniel Zachary and Jules A. Hoffmann and Danièle Hoffmann and Jean-Marc Reichhart}, issn = {0261-4189}, year = {1990}, date = {1990-08-01}, journal = {EMBO J.}, volume = {9}, number = {8}, pages = {2507--2515}, abstract = {Injections of low doses of bacteria into larvae of Phormia terranovae induce the appearance of potent bactericidal peptides in the blood, among which predominate the anti-Gram positive insect defensins and the anti-Gram negative diptericins. Insect defensins show significant homologies to mammalian (including human) microbicidal peptides present in polymorphonuclear leukocytes and macrophages. We report the molecular cloning of cDNAs and primer extension studies which indicate that insect defensin is produced as a prepro-peptide yielding mature defensin A (40 residues) after cleavage of a putative signal peptide (23 residues) and a prosequence (34 residues). Previous studies have established that diptericin (82 residues) is matured from a pre-peptide by cleavage of a putative signal peptide (19 residues) and C-terminal amidation. Using oligonucleotide probes complementary to the sequences of the mRNAs for defensin and diptericin, we show by in situ hybridization that both antibacterial peptides are concomitantly synthesized by the same cells: thrombocytoids, a specialized blood cell type, and adipocytes. Transcriptional studies based on hybridization of RNAs to cDNAs of defensin and diptericin indicate that the transcription of both genes is induced regardless of the nature of the stimulus (injection of Gram positive or Gram negative bacteria, lipopolysaccharides). Even a sterile injury applied to axenically raised larvae is efficient in inducing the transcription of both genes suggesting that the local disruption of the integument aspecifically initiates a signalling mechanism which the thrombocytoids and the adipocytes are able to interpret. The transcription of immune genes is relatively short lived and a second challenge yields a response similar to that of the first stimulus, indicating that the experimental insects do not keep a 'memory' of their first injection.}, keywords = {Anti-Bacterial Agents, Base Sequence, Blood Proteins, Cloning, Defensins, Diptera, Gene Expression, Insect Hormones, Insect Proteins, Larva, Molecular, Nucleic Acid Hybridization, Oligonucleotide Probes, Protein Conformation}, pubstate = {published}, tppubtype = {article} } Injections of low doses of bacteria into larvae of Phormia terranovae induce the appearance of potent bactericidal peptides in the blood, among which predominate the anti-Gram positive insect defensins and the anti-Gram negative diptericins. Insect defensins show significant homologies to mammalian (including human) microbicidal peptides present in polymorphonuclear leukocytes and macrophages. We report the molecular cloning of cDNAs and primer extension studies which indicate that insect defensin is produced as a prepro-peptide yielding mature defensin A (40 residues) after cleavage of a putative signal peptide (23 residues) and a prosequence (34 residues). Previous studies have established that diptericin (82 residues) is matured from a pre-peptide by cleavage of a putative signal peptide (19 residues) and C-terminal amidation. Using oligonucleotide probes complementary to the sequences of the mRNAs for defensin and diptericin, we show by in situ hybridization that both antibacterial peptides are concomitantly synthesized by the same cells: thrombocytoids, a specialized blood cell type, and adipocytes. Transcriptional studies based on hybridization of RNAs to cDNAs of defensin and diptericin indicate that the transcription of both genes is induced regardless of the nature of the stimulus (injection of Gram positive or Gram negative bacteria, lipopolysaccharides). Even a sterile injury applied to axenically raised larvae is efficient in inducing the transcription of both genes suggesting that the local disruption of the integument aspecifically initiates a signalling mechanism which the thrombocytoids and the adipocytes are able to interpret. The transcription of immune genes is relatively short lived and a second challenge yields a response similar to that of the first stimulus, indicating that the experimental insects do not keep a 'memory' of their first injection. |
| 1. | Wicker, C; Reichhart, Jean-Marc ; Hoffmann, Danièle ; Hultmark, D; Samakovlis, C; Hoffmann, Jules A Insect immunity. Characterization of a Drosophila cDNA encoding a novel member of the diptericin family of immune peptides Article de journal J. Biol. Chem., 265 (36), p. 22493–22498, 1990, ISSN: 0021-9258. Résumé | BibTeX | Étiquettes: Anti-Bacterial Agents, Base Sequence, Cloning, Diptera, DNA, Escherichia coli, Insect Hormones, Insect Proteins, Molecular, Multigene Family, Nucleic Acid, Oligonucleotide Probes, Sequence Homology @article{wicker_insect_1990, title = {Insect immunity. Characterization of a Drosophila cDNA encoding a novel member of the diptericin family of immune peptides}, author = { C. Wicker and Jean-Marc Reichhart and Danièle Hoffmann and D. Hultmark and C. Samakovlis and Jules A. Hoffmann}, issn = {0021-9258}, year = {1990}, date = {1990-01-01}, journal = {J. Biol. Chem.}, volume = {265}, number = {36}, pages = {22493--22498}, abstract = {Drosophila shows an immune response when challenged by injection of low doses of bacteria. To date, the molecules involved in this immune reaction have remained elusive, with the exception of cecropins (4-kDa antibacterial peptides initially isolated from the moth Hyalophora cecropia) for which three closely related genes have been characterized recently. We report the molecular cloning and sequencing of a cDNA from a library of immune Drosophila which encodes a novel member of the family of diptericins (9-kDa antibacterial peptides initially isolated from the fly Phormia terranovae). Transcripts for the Drosophila diptericin are detected 2 h after injection of bacteria. They are apparently derived from a single gene mapping at position 56 A on the right arm of the second chromosome. We discuss the existence of a distant relationship between the diptericins and two other groups of anti-bacterial insect proteins, the attacins, and the sarcotoxins II.}, keywords = {Anti-Bacterial Agents, Base Sequence, Cloning, Diptera, DNA, Escherichia coli, Insect Hormones, Insect Proteins, Molecular, Multigene Family, Nucleic Acid, Oligonucleotide Probes, Sequence Homology}, pubstate = {published}, tppubtype = {article} } Drosophila shows an immune response when challenged by injection of low doses of bacteria. To date, the molecules involved in this immune reaction have remained elusive, with the exception of cecropins (4-kDa antibacterial peptides initially isolated from the moth Hyalophora cecropia) for which three closely related genes have been characterized recently. We report the molecular cloning and sequencing of a cDNA from a library of immune Drosophila which encodes a novel member of the family of diptericins (9-kDa antibacterial peptides initially isolated from the fly Phormia terranovae). Transcripts for the Drosophila diptericin are detected 2 h after injection of bacteria. They are apparently derived from a single gene mapping at position 56 A on the right arm of the second chromosome. We discuss the existence of a distant relationship between the diptericins and two other groups of anti-bacterial insect proteins, the attacins, and the sarcotoxins II. |
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