Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 19 de 19
Filtrar
1.
PLoS Pathog ; 9(1): e1003098, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23326228

RESUMO

Upon infection, B-lymphocytes expressing antibodies specific for the intruding pathogen develop clonal responses triggered by pathogen recognition via the B-cell receptor. The constant region of antibodies produced by such responding clones dictates their functional properties. In teleost fish, the clonal structure of B-cell responses and the respective contribution of the three isotypes IgM, IgD and IgT remain unknown. The expression of IgM and IgT are mutually exclusive, leading to the existence of two B-cell subsets expressing either both IgM and IgD or only IgT. Here, we undertook a comprehensive analysis of the variable heavy chain (VH) domain repertoires of the IgM, IgD and IgT in spleen of homozygous isogenic rainbow trout (Onchorhynchus mykiss) before, and after challenge with a rhabdovirus, the Viral Hemorrhagic Septicemia Virus (VHSV), using CDR3-length spectratyping and pyrosequencing of immunoglobulin (Ig) transcripts. In healthy fish, we observed distinct repertoires for IgM, IgD and IgT, respectively, with a few amplified µ and τ junctions, suggesting the presence of IgM- and IgT-secreting cells in the spleen. In infected animals, we detected complex and highly diverse IgM responses involving all VH subgroups, and dominated by a few large public and private clones. A lower number of robust clonal responses involving only a few VH were detected for the mucosal IgT, indicating that both IgM(+) and IgT(+) spleen B cells responded to systemic infection but at different degrees. In contrast, the IgD response to the infection was faint. Although fish IgD and IgT present different structural features and evolutionary origin compared to mammalian IgD and IgA, respectively, their implication in the B-cell response evokes these mouse and human counterparts. Thus, it appears that the general properties of antibody responses were already in place in common ancestors of fish and mammals, and were globally conserved during evolution with possible functional convergences.


Assuntos
Células Clonais/metabolismo , Imunoglobulina M/metabolismo , Imunoglobulinas/metabolismo , Novirhabdovirus/imunologia , Oncorhynchus mykiss/imunologia , Baço/imunologia , Animais , Subpopulações de Linfócitos B , Células Clonais/citologia , Células Clonais/imunologia , Evolução Molecular , Doenças dos Peixes/imunologia , Proteínas de Peixes , Humanos , Imunoglobulina D/genética , Imunoglobulina D/metabolismo , Imunoglobulina M/genética , Imunoglobulinas/genética , Imuno-Histoquímica , Camundongos , Mucosa/imunologia , Mucosa/metabolismo , Análise de Sequência de DNA , Especificidade da Espécie , Baço/citologia , Baço/metabolismo , Coloração e Rotulagem
2.
Nat Biotechnol ; 25(7): 763-9, 2007 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-17592475

RESUMO

We report here the complete genome sequence of the virulent strain JIP02/86 (ATCC 49511) of Flavobacterium psychrophilum, a widely distributed pathogen of wild and cultured salmonid fish. The genome consists of a 2,861,988-base pair (bp) circular chromosome with 2,432 predicted protein-coding genes. Among these predicted proteins, stress response mediators, gliding motility proteins, adhesins and many putative secreted proteases are probably involved in colonization, invasion and destruction of the host tissues. The genome sequence provides the basis for explaining the relationships of the pathogen to the host and opens new perspectives for the development of more efficient disease control strategies. It also allows for a better understanding of the physiology and evolution of a significant representative of the family Flavobacteriaceae, whose members are associated with an interesting diversity of lifestyles and habitats.


Assuntos
Biotecnologia/métodos , Peixes/microbiologia , Flavobacterium/metabolismo , Genoma Bacteriano , Animais , Biofilmes , Adesão Celular , Membrana Celular/metabolismo , Infecções por Flavobacteriaceae/metabolismo , Genoma , Modelos Biológicos , Fases de Leitura Aberta , Parasitos
3.
BMC Biol ; 7: 7, 2009 Feb 05.
Artigo em Inglês | MEDLINE | ID: mdl-19196451

RESUMO

BACKGROUND: In mammals, the members of the tripartite motif (TRIM) protein family are involved in various cellular processes including innate immunity against viral infection. Viruses exert strong selective pressures on the defense system. Accordingly, antiviral TRIMs have diversified highly through gene expansion, positive selection and alternative splicing. Characterizing immune TRIMs in other vertebrates may enlighten their complex evolution. RESULTS: We describe here a large new subfamily of TRIMs in teleosts, called finTRIMs, identified in rainbow trout as virus-induced transcripts. FinTRIMs are formed of nearly identical RING/B-box regions and C-termini of variable length; the long variants include a B30.2 domain. The zebrafish genome harbors a striking diversity of finTRIMs, with 84 genes distributed in clusters on different chromosomes. A phylogenetic analysis revealed different subsets suggesting lineage-specific diversification events. Accordingly, the number of fintrim genes varies greatly among fish species. Conserved syntenies were observed only for the oldest fintrims. The closest mammalian relatives are trim16 and trim25, but they are not true orthologs. The B30.2 domain of zebrafish finTRIMs evolved under strong positive selection. The positions under positive selection are remarkably congruent in finTRIMs and in mammalian antiviral TRIM5alpha, concentrated within a viral recognition motif in mammals. The B30.2 domains most closely related to finTRIM are found among NOD-like receptors (NLR), indicating that the evolution of TRIMs and NLRs was intertwined by exon shuffling. CONCLUSION: The diversity, evolution, and features of finTRIMs suggest an important role in fish innate immunity; this would make them the first TRIMs involved in immunity identified outside mammals.


Assuntos
Motivos de Aminoácidos/genética , Duplicação Gênica , Variação Genética/genética , Família Multigênica , Seleção Genética , Sequência de Aminoácidos , Animais , Células COS , Chlorocebus aethiops , Evolução Molecular , Dados de Sequência Molecular , Oncorhynchus mykiss , Filogenia , Domínios RING Finger/genética , Peixe-Zebra
4.
Mol Immunol ; 46(3): 457-72, 2009 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19081138

RESUMO

In mammals, T cell activation requires specific recognition of the peptide-MHC complex by the TcR and co-stimulatory signals. Important co-stimulatory receptors expressed by T cells are the molecules of the CD28 family, that regulate T cell activation, proliferation and tolerance. These receptors recognize B7s and B7-homologous (B7H) molecules that are typically expressed by the antigen presenting cells. In teleost fish, typical T cell responses have been described and the TcR, MHC and CD28/CTLA4 genes have been characterized. In contrast, the members of the B7 gene family have only been described in mammals and birds and have yet to be addressed in lower vertebrates. To learn more about the evolution of components guiding T cell activation in vertebrates, we performed a systematic genomic survey for the B7 co-stimulatory and co-inhibitory IgSF receptors in lower vertebrates with an emphasis on teleost fish. Our search identified fish sequences that are orthologous to B7, B7-H1/B7-DC, B7-H3 and B7-H4 as defined by sequence identity, phylogeny and combinations of short or long-range syntenic relationships. However, we were unable to identify clear orthologs for B7-H2 (CD275, ICOS ligand) in bony fish, which correlates with our prior inability to find ICOS in fish. Interestingly, our results indicate that teleost fish possess a single B7.1/B7.2 (CD80/86) molecule that likely interacts with CD28/CTLA4 as the ligand-binding regions seem to be conserved in both partners. Overall, our analyses implies that gene duplication (and loss) have shaped a molecular repertoire of B7-like molecules that was recruited for the refinement of T cell activation during the evolution of the vertebrates.


Assuntos
Antígenos CD28/genética , Antígenos CD28/imunologia , Evolução Molecular , Sequência de Aminoácidos , Animais , Antígeno B7-1/química , Antígenos CD28/química , Sequência Conservada , Peixes/imunologia , Ligação Genética , Humanos , Modelos Imunológicos , Dados de Sequência Molecular , Filogenia , Proteína 2 Ligante de Morte Celular Programada 1 , Alinhamento de Sequência , Análise de Sequência de Proteína , Sintenia
5.
Mol Immunol ; 45(9): 2437-45, 2008 May.
Artigo em Inglês | MEDLINE | ID: mdl-18279958

RESUMO

In vertebrates, the world of antigenic motifs is matched to large populations of lymphocytes through specific recognition of an epitope by a given receptor unique to a lymphocyte clone. The concept of immune repertoire was proposed to describe this diversity of lymphocyte receptors - Ig and TCR - required by the network of interactions. The immune repertoires became useful tools to describe lymphocyte and receptor populations through the development of the immune system and in pathological situations. Recently, the development of mass technologies made possible a comprehensive survey of immune repertoires at the genome, transcript and protein levels, and some of these techniques have been already adapted to TCR and Ig repertoire analyses. Such approaches generate very big datasets, which necessitates complex and multi-parametric annotations in dedicated databases. They also require new analysis methods, leading to the integration of structure and dynamics of the immune repertoires, at different time scales (immune response, development of the individual, evolution of the species). Such methods may be extended to the analysis of new classes of adaptive-like receptors, which were recently discovered in different invertebrates and in agnathans. Ultimately, they may allow a parallel monitoring of pathogen and immune repertoires addressing the reciprocal influences that decide for the host survival or death. In this review, we first study the characteristics of Ig and TCR repertoires, and we examine several systematic approaches developed for the analysis of these "classical" immune repertoires at different levels. We then consider examples of the recent developments of modeling and statistical analysis, and we discuss their relevance and their importance for the study of the immune diversity. An extended view of immune repertoires is proposed, integrating the diversity of other receptors involved in immune recognition. Also, we discuss how repertoire studies could link pathogen variation and immune diversity to reveal regulatory patterns and rules driving their co-diversification race.


Assuntos
Subpopulações de Linfócitos B/imunologia , Rearranjo Gênico do Linfócito T , Receptores de Antígenos de Linfócitos T , Receptores Imunológicos , Subpopulações de Linfócitos T/imunologia , Receptores Toll-Like , Animais , Genômica/métodos , Humanos , Imunoglobulinas , Receptores de Antígenos de Linfócitos T/genética , Receptores Imunológicos/genética , Receptores Toll-Like/genética
6.
Dev Comp Immunol ; 31(3): 255-71, 2007.
Artigo em Inglês | MEDLINE | ID: mdl-16928399

RESUMO

CD28 family of costimulatory receptors is comprised of molecules with a single V-type extracellular Ig domain, a transmembrane and an intracytoplasmic region with signaling motifs. CD28 and cytotoxic T lymphocyte antigen-4 (CTLA4) homologs have been recently identified in rainbow trout. Other sequences similar to mammalian CD28 family members have now been identified using teleost, Xenopus and chicken databases. CD28- and CTLA4 homologs were found in all vertebrate classes whereas inducible costimulatory signal (ICOS) was restricted to tetrapods, and programmed cell death-1 (PD1) was limited to mammals and chicken. Multiple B and T Lymphocyte Attenuator (BTLA) sequences were found in teleosts, but not in Xenopus or in avian genomes. The intron/exon structure of btlas was different from that of cd28 and other members of the family. The Ig domain encoded in all the btla genes has features of the C-type structure, which suggests that BTLA does not belong to the CD28 family. The genomic localization of these genes in vertebrate genomes supports the split between the BTLA and CD28 families.


Assuntos
Antígenos CD/genética , Antígenos de Diferenciação/genética , Antígenos CD28/genética , Peixes/genética , Filogenia , Receptores Imunológicos/genética , Motivos de Aminoácidos/genética , Sequência de Aminoácidos , Animais , Antígeno CTLA-4 , Galinhas/genética , Galinhas/imunologia , Bases de Dados de Proteínas , Peixes/imunologia , Dados de Sequência Molecular , Alinhamento de Sequência , Linfócitos T/imunologia , Xenopus/genética , Xenopus/imunologia
7.
Antiviral Res ; 66(2-3): 147-52, 2005 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-15885817

RESUMO

Innate, non-specific resistance mechanisms are important to pathogens, particularly for delaying virus replication at the onset of infection. Innate immunity constitutes the first line of defense in vertebrates and is the only one in invertebrates. Little is known about possible antiviral substances in invertebrates. The present work concerns a study of antiviral substances in hemolymph from adult Crassostrea gigas oysters. Despite the detection of cytotoxicity in fresh filtered hemolymph for both mammalian (CC50: 750 microg/ml) and fish cells (CC50: > 2000 microg/ml for EPC cells and 345 microg/ml for RTG-2 cells), an antiviral substance was detected. Fresh filtered hemolymph was capable of inhibiting the replication of herpes simplex virus type 1 in vitro at an EC50 of 425 microg/ml (total proteins) and the replication of infectious pancreatic necrosis virus in EPC and RTG-2 cells at 217 and 156 microg/ml (total proteins), respectively.


Assuntos
Antivirais/farmacologia , Vírus de DNA/efeitos dos fármacos , Hemolinfa/química , Ostreidae/química , Vírus de RNA/efeitos dos fármacos , Animais , Antivirais/isolamento & purificação , Chlorocebus aethiops , Vírus de DNA/crescimento & desenvolvimento , Hemolinfa/enzimologia , Testes de Sensibilidade Microbiana , Ostreidae/metabolismo , Vírus de RNA/crescimento & desenvolvimento , Células Vero
8.
Dev Comp Immunol ; 28(7-8): 793-801, 2004 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-15043947

RESUMO

A rainbow trout interferon (IFN) reporter system has been established by selection of a stable cell line, RTG-P1, transfected with a plasmid expressing the firefly luciferase gene under the control of the promoter for the IFN-induced gene Mx1. After 148 passages, the luciferase expression was still highly induced by polyinosinic:polycytidylic acid (poly I:C) in RTG-P1 cells. Different IFN inducers (dsRNA, viral hemorrhagic septicaemia virus or conditioned medium containing rainbow trout antiviral activity) were able to stimulate the IFN-reporter system in RTG-P1, showing that this cell line can be used to study the activation of the IFN pathway in various contexts. Pyrrolidine dithiocarbamate (PDTC), an NF-kappaB inhibitor, significantly blocked poly I:C induced luciferase accumulation in RTG-P1 at intermediate doses (1-10 microM), suggesting that Mx1 induction through the IFN signalling pathway is NF-kappaB-dependent in fish. This inhibition was not observed for doses of 50 microM or higher. The RTG-P1 reporter system constitutes an interesting tool to study the induction and regulation of IFN signalling in teleost fish.


Assuntos
Interferons/metabolismo , Oncorhynchus mykiss/metabolismo , Regiões Promotoras Genéticas/genética , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Antivirais/farmacologia , Células Cultivadas , Genes Reporter/genética , Interferons/genética , Novirhabdovirus/imunologia , Oncorhynchus mykiss/genética , Oncorhynchus mykiss/virologia , Poli I-C/farmacologia , Pirrolidinas/farmacologia , Tiocarbamatos/farmacologia , Quinase Induzida por NF-kappaB
9.
PLoS One ; 7(4): e33935, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22514610

RESUMO

Genetic factors of resistance and predisposition to viral diseases explain a significant part of the clinical variability observed within host populations. Predisposition to viral diseases has been associated to MHC haplotypes and T cell immunity, but a growing repertoire of innate/intrinsic factors are implicated in the genetic determinism of the host susceptibility to viruses. In a long-term study of the genetics of host resistance to fish rhabdoviruses, we produced a collection of double-haploid rainbow trout clones showing a wide range of susceptibility to Viral Hemorrhagic Septicemia Virus (VHSV) waterborne infection. The susceptibility of fibroblastic cell lines derived from these clonal fish was fully consistent with the susceptibility of the parental fish clones. The mechanisms determining the host resistance therefore did not associate with specific host immunity, but rather with innate or intrinsic factors. One cell line was resistant to rhabdovirus infection due to the combination of an early interferon IFN induction--that was not observed in the susceptible cells--and of yet unknown factors that hamper the first steps of the viral cycle. The implication of IFN was well consistent with the wide range of resistance of this genetic background to VSHV and IHNV, to the birnavirus IPNV and the orthomyxovirus ISAV. Another cell line was even more refractory to the VHSV infection through different antiviral mechanisms. This collection of clonal fish and isogenic cell lines provides an interesting model to analyze the relative contribution of antiviral pathways to the resistance to different viruses.


Assuntos
Doenças dos Peixes/genética , Doenças dos Peixes/virologia , Peixes/genética , Peixes/virologia , Infecções por Rhabdoviridae/genética , Animais , Linhagem Celular , Suscetibilidade a Doenças , Doenças dos Peixes/metabolismo , Peixes/metabolismo , Interferons/metabolismo
10.
Dev Comp Immunol ; 35(12): 1204-14, 2011 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-21414349

RESUMO

In fish as in mammals, virus infections induce changes in the expression of many host genes. Studies conducted during the last fifteen years revealed a major contribution of the interferon system in fish antiviral response. This review describes the screening methods applied to compare the impact of virus infections on the transcriptome in different fish species. These approaches identified a "core" set of genes that are strongly induced in most viral infections. The "core" interferon-induced genes (ISGs) are generally conserved in vertebrates, some of them inhibiting a wide range of viruses in mammals. A selection of ISGs -PKR, vig-1/viperin, Mx, ISG15 and finTRIMs - is further analyzed here to illustrate the diversity and complexity of the mechanisms involved in establishing an antiviral state. Most of the ISG-based pathways remain to be directly determined in fish. Fish ISGs are often duplicated and the functional specialization of multigenic families will be of particular interest for future studies.


Assuntos
Doenças dos Peixes/imunologia , Proteínas de Peixes/imunologia , Peixes/virologia , Imunidade Inata , Interferons/imunologia , Viroses/imunologia , Vírus/imunologia , Animais , Doenças dos Peixes/virologia , Proteínas de Peixes/genética , Proteínas de Peixes/metabolismo , Peixes/genética , Peixes/imunologia , GTP Fosfo-Hidrolases/genética , GTP Fosfo-Hidrolases/imunologia , GTP Fosfo-Hidrolases/metabolismo , Regulação da Expressão Gênica/imunologia , Proteínas Quinases/genética , Proteínas Quinases/imunologia , Proteínas Quinases/metabolismo , Transdução de Sinais/imunologia , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/imunologia , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinas/genética , Ubiquitinas/imunologia , Ubiquitinas/metabolismo , Viroses/virologia
11.
PLoS One ; 6(7): e22022, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21789205

RESUMO

Tripartite motif proteins (TRIM) constitute a large family of proteins containing a RING-Bbox-Coiled Coil motif followed by different C-terminal domains. Involved in ubiquitination, TRIM proteins participate in many cellular processes including antiviral immunity. The TRIM family is ancient and has been greatly diversified in vertebrates and especially in fish. We analyzed the complete sets of trim genes of the large zebrafish genome and of the compact pufferfish genome. Both contain three large multigene subsets--adding the hsl5/trim35-like genes (hltr) to the ftr and the btr that we previously described--all containing a B30.2 domain that evolved under positive selection. These subsets are conserved among teleosts. By contrast, most human trim genes of the other classes have only one or two orthologues in fish. Loss or gain of C-terminal exons generated proteins with different domain organizations; either by the deletion of the ancestral domain or, remarkably, by the acquisition of a new C-terminal domain. Our survey of fish trim genes in fish identifies subsets with different evolutionary dynamics. trims encoding RBCC-B30.2 proteins show the same evolutionary trends in fish and tetrapods: they evolve fast, often under positive selection, and they duplicate to create multigenic families. We could identify new combinations of domains, which epitomize how new trim classes appear by domain insertion or exon shuffling. Notably, we found that a cyclophilin-A domain replaces the B30.2 domain of a zebrafish fintrim gene, as reported in the macaque and owl monkey antiretroviral TRIM5α. Finally, trim genes encoding RBCC-B30.2 proteins are preferentially located in the vicinity of MHC or MHC gene paralogues, which suggests that such trim genes may have been part of the ancestral MHC.


Assuntos
Evolução Molecular , Proteínas de Peixes/genética , Tetraodontiformes/genética , Peixe-Zebra/genética , Sequência de Aminoácidos , Animais , Sequência Conservada/genética , Proteínas de Peixes/química , Proteínas de Peixes/metabolismo , Duplicação Gênica , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Genoma/genética , Humanos , Complexo Principal de Histocompatibilidade/genética , Dados de Sequência Molecular , Família Multigênica/genética , Filogenia , Estrutura Terciária de Proteína , Seleção Genética , Homologia de Sequência de Aminoácidos , Sintenia/genética
12.
Fish Shellfish Immunol ; 22(5): 510-9, 2007 May.
Artigo em Inglês | MEDLINE | ID: mdl-17085058

RESUMO

Inbred lines differentially susceptible to diseases are a powerful tool to get insights into the mechanisms of genetic resistance to pathogens. In fish, chromosome manipulation techniques allow a quick production of such homozygous lines. Using gynogenesis, we produced nine homozygous clones of rainbow trout from a domestic population (INRA Sy strain). We examined the variability between clones for resistance to two rhabdoviruses, the viral haemorrhagic septicaemia virus (VHSV) and the infectious haematopoietic necrosis virus (IHNV). Intraperitoneal injections and waterborne infections were performed in parallel for both viruses. No survival was recorded after intraperitoneal injection of VHSV or IHNV, indicating that fish from all clones were fully susceptible to both viruses by this route of infection. In contrast, the different clones showed a wide range of survival frequency after waterborne infection. The resistance levels to VHSV ranged from 0 to 99% and resistance was not abrogated when resistant and sensitive animals were mixed and subjected to waterborne infection. VHSV was recovered from 10% of resistant fish after waterborne infection, confirming that virus replication was possible in this context but effective only in a low proportion of the population. The different clones also exhibited a wide range of survival (0-68%) after a waterborne infection with IHNV. Although VHSV-resistant clones were not fully resistant to IHNV, the susceptibility to IHNV and VHSV tended to be correlated, suggesting that non-specific mechanisms common to both viruses were involved.


Assuntos
Doenças dos Peixes/imunologia , Vírus da Necrose Hematopoética Infecciosa/patogenicidade , Novirhabdovirus/patogenicidade , Oncorhynchus mykiss/imunologia , Infecções por Rhabdoviridae/veterinária , Animais , Peso Corporal , Clonagem de Organismos/veterinária , Feminino , Doenças dos Peixes/genética , Doenças dos Peixes/virologia , Variação Genética , Imunidade Inata/genética , Vírus da Necrose Hematopoética Infecciosa/imunologia , Masculino , Novirhabdovirus/imunologia , Oncorhynchus mykiss/genética , Oncorhynchus mykiss/virologia , Infecções por Rhabdoviridae/genética , Infecções por Rhabdoviridae/imunologia , Baço/virologia , Análise de Sobrevida , Água
13.
J Immunol ; 178(7): 4385-94, 2007 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-17371995

RESUMO

The recent description of virus-induced fish IFNs has raised questions about the evolution of this complex antiviral system. Identification of the receptor of the zebrafish virus-induced IFN (zIFN) was sought to help resolve these questions. We set up an experimental system to study the zIFN system in the course of a viral infection of zebrafish embryos. In this setting, zIFN was induced by viral infection, and we identified zIFN-dependent induced transcripts. Embryos quickly died from the infection, but zIFN overexpression increased their survival. We took advantage of this experimental system to perform in vivo loss and gain of function analysis of candidate receptors of the class II helical receptor family and identified zCRFB1 and zCRFB5 as the two subunits of the zebrafish IFN receptor. Based on the organization of the zIFN gene and the protein structure of the identified receptor components, the virus-induced fish IFNs appear as orthologs of mammalian IFN-lambda, specifying type III IFN as the ancestral antiviral system of vertebrates.


Assuntos
Interferons/metabolismo , Subunidades Proteicas/fisiologia , Receptores de Interferon/fisiologia , Proteínas de Peixe-Zebra/fisiologia , Peixe-Zebra/imunologia , Processamento Alternativo , Sequência de Aminoácidos , Animais , Embrião não Mamífero/química , Embrião não Mamífero/imunologia , Embrião não Mamífero/virologia , Interferons/genética , Dados de Sequência Molecular , Regiões Promotoras Genéticas , Subunidades Proteicas/análise , Subunidades Proteicas/genética , Receptores de Interferon/análise , Receptores de Interferon/genética , Transcrição Gênica , Peixe-Zebra/genética , Peixe-Zebra/virologia , Proteínas de Peixe-Zebra/análise , Proteínas de Peixe-Zebra/genética
14.
J Immunol ; 176(7): 4191-200, 2006 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-16547256

RESUMO

T cell activation requires both specific recognition of the peptide-MHC complex by the TCR and additional signals delivered by costimulatory receptors. We have identified rainbow trout sequences similar to CD28 (rbtCD28) and CTLA4 (rbtCTLA4). rbtCD28 and rbtCTLA4 are composed of an extracellular Ig-superfamily V domain, a transmembrane region, and a cytoplasmic tail. The presence of a conserved ligand binding site within the V domain of both molecules suggests that these receptors likely recognize the fish homologues of the B7 family. The mRNA expression pattern of rbtCD28 and rbtCTLA4 in naive trout is reminiscent to that reported in humans and mice, because rbtCTLA4 expression within trout leukocytes was quickly up-regulated following PHA stimulation and virus infection. The cytoplasmic tail of rbtCD28 possesses a typical motif that is conserved in mammalian costimulatory receptors for signaling purposes. A chimeric receptor made of the extracellular domain of human CD28 fused to the cytoplasmic tail of rbtCD28 promoted TCR-induced IL-2 production in a human T cell line, indicating that rbtCD28 is indeed a positive costimulator. The cytoplasmic tail of rbtCTLA4 lacked obvious signaling motifs and accordingly failed to signal when fused to the huCD28 extracellular domain. Interestingly, rbtCTLA4 and rbtCD28 are not positioned on the same chromosome and thus do not belong to a unique costimulatory cluster as in mammals. Finally, our results raise questions about the origin and evolution of positive and negative costimulation in vertebrate immune systems.


Assuntos
Antígenos de Diferenciação/metabolismo , Antígenos CD28/metabolismo , Oncorhynchus mykiss/metabolismo , Sequência de Aminoácidos , Animais , Antígenos CD , Antígenos de Diferenciação/química , Antígenos de Diferenciação/genética , Antígenos de Diferenciação/imunologia , Sequência de Bases , Sítios de Ligação , Antígenos CD28/química , Antígenos CD28/genética , Antígenos CD28/imunologia , Antígeno CTLA-4 , Cromossomos/genética , Biologia Computacional , Sequência Conservada , Citoplasma/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Expressão Gênica , Humanos , Interleucina-2/biossíntese , Tecido Linfoide/metabolismo , Dados de Sequência Molecular , Oncorhynchus mykiss/genética , Oncorhynchus mykiss/imunologia , Especificidade de Órgãos , Fosforilação , RNA Mensageiro/genética , Alinhamento de Sequência , Tirosina/metabolismo
15.
J Immunol ; 176(7): 3942-9, 2006 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-16547228

RESUMO

Gut-associated lymphocytes were described in fish, but their involvement in immune responses is still unknown. In rainbow trout, intraepithelial lymphocytes (IELs) are scattered between gut epithelial cells, but neither Peyer's patches nor mesenteric lymph nodes were identified. Rainbow trout IELs contain mainly T cells, because they expressed transcripts of T cell marker homologs of CD8, CD4, CD28, CD3epsilon, TCRzeta, TCRgamma, and TCRbeta and lacked IgM. However, trout IELs did not show specific homing to the gut mucosa, which in mammals defines IELs as a distinctive mucosal population. A detailed analysis of the TCRbeta repertoire of rainbow trout IELs was performed in both naive and virus-infected animals. TCRbeta transcripts of rainbow trout IELs were highly diverse and polyclonal in adult naive individuals, in sharp contrast with the restricted diversity of IEL oligoclonal repertoires described in birds and mammals. Significant modifications of the trout IEL TCRbeta repertoire were observed after a systemic infection with a fish rhabdovirus and were especially marked for Vbeta4-bearing receptors as previously reported for spleen cells. Thus, we could not find any specific properties of the trout IEL TCRbeta repertoire compared with the spleen and pronephros TCRbeta repertoire, which questions the reality of a distinct IEL compartment in teleosts. Our findings suggest that a highly diversified alphabeta TauCR repertoire is maintained in fish IELs in the absence of Peyer's patches and mesenteric lymph nodes, whereas the restricted diversity of mouse alphabeta IELs is attributed to multiple cycles of activation and recirculation, allowing a progressive narrowing of the repertoire.


Assuntos
Células Epiteliais/imunologia , Intestinos/imunologia , Oncorhynchus mykiss/imunologia , Linfócitos T/citologia , Linfócitos T/imunologia , Animais , Biomarcadores , Movimento Celular/imunologia , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Mucosa Intestinal/metabolismo , Intestinos/citologia , Intestinos/virologia , Novirhabdovirus/imunologia , Oncorhynchus mykiss/metabolismo , Oncorhynchus mykiss/virologia , Fenótipo , Receptores de Antígenos de Linfócitos B/imunologia , Receptores de Antígenos de Linfócitos B/metabolismo , Infecções por Rhabdoviridae/imunologia , Infecções por Rhabdoviridae/metabolismo , Infecções por Rhabdoviridae/virologia , Linfócitos T/metabolismo
16.
J Immunol ; 169(11): 6244-52, 2002 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-12444130

RESUMO

VDJ rearrangement at the teleost TCRbeta locus leads to a highly diverse repertoire of junctions for each VbetaJbeta combination. From a rainbow trout 5' RACE library of TCRbeta transcripts, 47 clones encompassing a full Vbeta-Dbeta-Jbeta-Cbeta sequence were selected and analyzed. A similarity analysis of the sequences evidenced 10 Vbeta families, of which 6 were not previously described. Immunoscope and sequence analysis of the Vbeta-Dbeta-Jbeta junctions of the new families confirmed that they create a polyclonal and diverse repertoire. Multiple alignments showed that rainbow trout Vbetas possess most of the conserved residues typical of Vbeta segments. However, this study revealed a high complementarity-determining region 2 (CDR2) and CDR1 length diversity among rainbow trout Vbeta families, suggesting that the spatial orientation of the TCR could fluctuate in the TCR/peptide/MHC complex, depending on the Vbeta expressed. Among the new Vbeta families, Vbeta6 displayed the strongest deviance from typical hypervariable CDR1 and CDR2 loops, with an unusually short CDR2. Moreover, the Vbeta6 sequence is overall divergent from typical Vbeta sequence, raising the question of its functional relevance. Immunoscope experiments identified a Vbeta6-Jbeta3 junction, which was amplified during the response against viral hemorrhagic septicemia virus, a fish rhabdovirus. Vbeta6 seems therefore to be expressed functionally in a selected TCR. However, the shorter Vbeta6 transcripts produced through an alternative splicing lack the C', C", D, and E strands of the Vbeta domain and are probably nonfunctional.


Assuntos
Regiões Determinantes de Complementaridade , Genes Codificadores da Cadeia beta de Receptores de Linfócitos T , Oncorhynchus mykiss/genética , Oncorhynchus mykiss/imunologia , Processamento Alternativo , Sequência de Aminoácidos , Animais , Sequência de Bases , DNA Complementar/genética , Doenças dos Peixes/genética , Doenças dos Peixes/imunologia , Rearranjo Gênico da Cadeia beta dos Receptores de Antígenos dos Linfócitos T , Variação Genética , Dados de Sequência Molecular , Família Multigênica , Novirhabdovirus/imunologia , Oncorhynchus mykiss/virologia , Receptores de Antígenos de Linfócitos T alfa-beta/química , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Infecções por Rhabdoviridae/genética , Infecções por Rhabdoviridae/imunologia , Infecções por Rhabdoviridae/veterinária , Homologia de Sequência de Aminoácidos
17.
J Gen Virol ; 85(Pt 10): 3099-3108, 2004 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-15448373

RESUMO

T-cell responses to viruses are still poorly investigated in lower vertebrates. In rainbow trout, a specific clonal expansion of T cells in response to infection with viral haemorrhagic septicaemia virus (VHSV) was recently identified. Expanded T-cell clones expressed a unique 8 aa Vbeta4-Jbeta1 junction (SSGDSYSE) in different individuals, reminiscent of a typical public response. To get further insight into the nature of this response the modifications of the T-cell repertoire following immunization with plasmid expressing the VHSV external glycoprotein (G), which is the only protein involved in protective immunity, was analysed. After G-based DNA immunization, CDR3-length spectratypes were skewed for several Vbeta-Jbeta combinations, including Vbeta4-Jbeta1. In Vbeta4-Jbeta1, biases consisted of 6 and 8 aa junctions that were detected from day 52, and were still present 3 months after DNA immunization. Sequence analysis of the Vbeta4-Jbeta1 junctions showed that the 8 aa junction (SSGDSYSE) was clearly expanded, indicating that viral G protein was probably the target of the anti-VHSV public response. Additional 6 and 8 aa Vbeta4-Jbeta1 junctions were also expanded in G-DNA-vaccinated fish, showing that significant clonotypic diversity was selected in response to the plasmid-delivered G protein. This higher clonotypic diversity may be related to the demonstrated higher efficiency of G-based DNA vaccines over whole virus immunization. The use of infectious hematopietic necrosis virus (IHNV) recombinant viruses, expressing the VHSV G protein, further substantiated the VHSV G-protein specificity of the 8 aa Vbeta4-Jbeta1 response and designated the 6 aa Vbeta4-Jbeta1 response as potentially directed to a T-cell epitope common to VHSV and IHNV.


Assuntos
Novirhabdovirus/imunologia , Oncorhynchus mykiss/virologia , Linfócitos T/imunologia , Vacinas de DNA/imunologia , Proteínas do Envelope Viral/imunologia , Vacinas Virais/imunologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Dados de Sequência Molecular , Oncorhynchus mykiss/imunologia , Vacinação , Vacinas Sintéticas/imunologia
18.
J Virol ; 76(16): 8040-9, 2002 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-12134009

RESUMO

Virus infections induce changes in the expression of host cell genes. A global knowledge of these modifications should help to better understand the virus/host cell interactions. To obtain a more comprehensive view of the rainbow trout response to a viral infection, we used the subtractive suppressive hybridization methodology in the viral hemorrhagic septicemia model of infection. We infected rainbow trout leukocytes with viral hemorrhagic septicemia virus (VHSV), and total RNA from infected and mock-infected cells was compared at 40 h postinfection. Twenty-four virus-induced genes were ultimately retrieved from the subtracted cDNA library, and their differential expression was further confirmed by semiquantitative reverse transcription-PCR and Northern blot analysis. Among these sequences, three were already described as VHSV-induced genes. Eight sequences with known homologs were extended to full-length cDNA using 5' and 3' rapid amplification of cDNA ends, and they were subsequently divided into three functional subsets. Four genes were homologous to mammalian interferon responsive genes, three were similar to chemo-attractant molecules (CXC chemokine, galectin), and two had nucleic acid binding domains. All of the virus-induced genes were also induced by rainbow trout interferon, indicating that the interferon pathway is the predominant component of the anti-VHSV response. They were also expressed in vivo in experimentally infected fish, indicating their biological relevance in natural infection.


Assuntos
Doenças dos Peixes/genética , Oncorhynchus mykiss/genética , Infecções por Rhabdoviridae/veterinária , Sequência de Aminoácidos , Animais , Sequência de Bases , DNA Complementar/genética , Etiquetas de Sequências Expressas , Doenças dos Peixes/imunologia , Expressão Gênica , Técnicas In Vitro , Interferons/biossíntese , Interferons/genética , Leucócitos/metabolismo , Dados de Sequência Molecular , Oncorhynchus mykiss/imunologia , Oncorhynchus mykiss/virologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Rhabdoviridae/patogenicidade , Infecções por Rhabdoviridae/genética , Infecções por Rhabdoviridae/imunologia , Homologia de Sequência de Aminoácidos , Homologia de Sequência do Ácido Nucleico
19.
J Gen Virol ; 81(Pt 11): 2675-2682, 2000 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-11038379

RESUMO

The homologous genes vig1 and cig5 were identified by differential display PCR as virus-induced genes in rainbow trout and humans, respectively. These genes are significantly related to sequences required for the biosynthesis of metal cofactors, but their function remains unknown. In this study, it is shown that the mouse homologue of vig1/cig5 was induced by vesicular stomatitis virus (VSV) and pseudorabies virus (PrV) in mouse spleen cells. Among a collection of cell lines from dendritic, myeloid, lymphoid or fibroblast lineages, only the dendritic cell line, D2SC1, showed expression of mvig after virus infection. This dendritic restriction was confirmed by our finding that mvig was also induced by both VSV and PrV in CD11c(++) spleen cells, separated by magnetic purification or derived from bone marrow precursor cells. Similar to the fish rhabdovirus viral haemorrhagic septicaemia virus in trout cells, VSV directly induced mvig in the dendritic cell line D2SC1, but the PrV-mediated induction required the integrity of the interferon pathway. This result indicates that mvig is interferon-inducible like its fish and human homologues. Furthermore, mvig was also induced by LPS in bone marrow-derived cells. Thus, mvig expression seems to correlate with an activated state of dendritic cells subjected to different pathogen-associated stimuli.


Assuntos
Células Dendríticas/virologia , Regulação Viral da Expressão Gênica/fisiologia , Herpesvirus Suídeo 1/fisiologia , Proteínas , Pseudorraiva/virologia , Infecções por Rhabdoviridae/virologia , Vírus da Estomatite Vesicular Indiana/fisiologia , Proteínas Virais/fisiologia , Sequência de Aminoácidos , Animais , Células Cultivadas , Células Dendríticas/fisiologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Dados de Sequência Molecular , Transdução de Sinais/fisiologia
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA