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J Virol ; 96(18): e0098622, 2022 09 28.
Artigo em Inglês | MEDLINE | ID: mdl-36069548


All lentiviruses encode the accessory protein Rev, whose main biological function is to mediate the nuclear export of unspliced and incompletely spliced viral transcripts by binding to a viral cis-acting element (termed the Rev-responsive element, RRE) within the env-encoding region. Equine infectious anemia virus (EIAV) is a member of the lentivirus genus in the Retroviridae family and is considered an important model for the study of lentivirus pathogenesis. Here, we identified a novel transcript from the EIAV genome that encoded a viral protein, named Mat, with an unknown function. The transcript mat was fully spliced and comprised parts of the coding regions of MA and TM. Interestingly, the expression of Mat depended on Rev and the chromosome region maintenance 1 (CRM1) pathway. Rev could specifically bind to Mat mRNA to promote its nuclear export. We further identified that the first exon of Mat mRNA, which was located within the Gag-encoding region, acted as an unreported RRE. Altogether, we identified a novel fully spliced transcript mat with an unusual RRE, which interacted with Rev for nuclear export through the CRM1 pathway. These findings updated the EIAV genome structure, highlighted the diversification of posttranscriptional regulation patterns in EIAV, and may help to expand the understanding of gene transcription and expression of lentivirus. IMPORTANCE In lentiviruses, the nuclear export of viral transcripts is an important step in controlling viral gene expression. Generally, the unspliced and incompletely spliced transcripts are exported via the CRM1-dependent export pathway in a process mediated by the viral Rev protein by binding to the Rev-responsive element (RRE) located within the Env-coding region. However, the completely spliced transcripts are exported via an endogenous cellular pathway, which was Rev independent. Here, we identified a novel fully spliced transcript from EIAV and demonstrated that it encoded a viral protein, termed Mat. Interestingly, we determined that the expression of Mat depended on Rev and identified that the first exon of Mat mRNA could specifically bind to Rev and be exported to the cytoplasm, which suggested that the first exon of Mat mRNA was a second RRE of EIAV. These findings provided important insights into the Rev-dependent nuclear export of completely spliced transcripts in lentiviruses.

Produtos do Gene rev , Vírus da Anemia Infecciosa Equina , Lentivirus Equinos , Animais , Produtos do Gene rev/genética , Cavalos , Vírus da Anemia Infecciosa Equina/metabolismo , Splicing de RNA , RNA Mensageiro/genética , RNA Viral/genética
Autophagy ; 17(10): 2800-2817, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-33172327


The innate immune restriction factor SAMHD1 can inhibit diverse viruses in myeloid cells. Mechanistically, SAMHD1 inhibits lentiviral replication including HIV-1 by depleting the nucleotide pool to interfere with their reverse transcription. Equine infectious anemia virus (EIAV) is an ancient lentivirus that preferentially attacks macrophages. However, the mechanism by which EIAV successfully establishes infection in macrophages with functional SAMHD1 remains unclear. Here, we demonstrate that while equine SAMDH1 can limit EIAV replication in equine macrophages at the reverse transcription stage, the antiviral effect is counteracted by the well-known transcriptional regulator Rev, which downregulates equine SAMHD1 through the lysosomal pathway. Remarkably, Rev hijacks BECN1 (beclin 1) and PIK3C3 to mediate SAMHD1 degradation in a canonical macroautophagy/autophagy-independent pathway. Our study illustrates that equine lentiviral Rev possesses important functions in evading cellular innate immunity in addition to its RNA regulatory function, and may provide new insights into the co-evolutionary arms race between SAMHD1 and lentiviruses.Abbreviations:3-MA: 3-methyladenine; AA: amino acid; ACTB: actin beta; AD: activation domain; ATG: autophagy related; Baf A1: bafilomycin A1; BD: binding domain; BECN1: beclin 1; BH3: BCL2-homology-3 domain; BiFC: bimolecular fluorescence complementation; CCD: coiled-coil domain; class III PtdIns3K: class III phosphatidylinositol 3-kinase; CQ: chloroquine; Co-IP: co-immunoprecipitation; dNTPase: dGTP-stimulated deoxynucleoside triphosphate triphosphohydrolase; ECD: evolutionarily conserved domain; EIAV: equine infectious anemia virus; eMDMs: equine monocyte-derived macrophages; GFP: green fluorescent protein; HD: histidine-aspartic; HIV-1: human immunodeficiency virus-1; hpi: hours post infection; hpt: hours post transfection; KO: knockout; LAMP2: lysosomal associated membrane protein 2; LMB: leptomycin B; PMA: phorbol 12-myristate 13-acetate; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; ND: unknown non-essential domain; NES: nuclear export signal; NLS: localization signal; NS: statistically non-significant; PIK3C3: phosphatidylinositol 3-kinase catalytic subunit type 3; RBD: RNA binding domain; RT: reverse transcriptase; siRNAs: small interfering RNAs; SAMHD1: SAM and HD domain containing deoxynucleoside triphosphate triphosphohydrolase 1; SIV: simian immunodeficiency virus; VN: C-terminal residues of Venus 174 to 238; VC: N-terminal residues 2 to 173 of Venus.

Autofagia , Lentivirus Equinos , Animais , Proteína Beclina-1/metabolismo , Cavalos , Lisossomos/metabolismo , Proteína 1 com Domínio SAM e Domínio HD/genética
Rev. Inst. Nac. Hig ; 46(1/2): 64-74, dic. 2015. ilus
Artigo em Espanhol | LILACS, LIVECS | ID: lil-798274


La Anemia Infecciosa Equina (en ingles Equine Infectious Anemia EIA), es una enfermedad viral que afecta a los équidos a nivel mundial. El agente causal, pertenece al género Lentivirus, de la familia Retroviridae, subfamilia Orthoretrovirinae. La enfermedad se caracteriza por episodios febriles recurrentes, trombocitopenia, anemia, pérdida de peso y edema de las partes bajas del cuerpo; si no se produce la muerte en el curso de los ataques clínicos agudos, se produce una fase crónica y la enfermedad tiende a convertirse en latente. La AIE debe notificarse ante la OIE: Organización Mundial de Sanidad Animal. En 2015 la OIE mediante Código Sanitario para los Animales Terrestres se estableció a la Anemia Infecciosa Equina en la lista única de enfermedades e infecciones de los équidos. Los veterinarios que detecten un caso de anemia infecciosa equina deben seguir las pautas nacionales y/o locales para la notificación y las pruebas de diagnóstico correspondientes. Este artículo describe los aspectos más relevantes de la Anemia Infecciosa Equina.

Equine Infectious Anemia (EIA) is a viral disease that affects horses worldwide. The causative agent belongs to the genus Lentivirus, the family Retroviridae, subfamily Orthoretrovirinae. The disease is characterized by recurrent episodes of fever, thrombocytopenia, anemia, weight loss and edema of the lower parts of the body; if death does not occur in the course of the acute clinical attacks, a chronic stage occurs and the disease tends to become dormant. The IEA must be reported to the OIE: World Organization for Animal Health. In 2015 by Health Code OIE Terrestrial Animal it was established Equine Infectious Anemia in the single list of diseases and infections of horses. Veterinarians detected a case of equine infectious anemia should follow national and / or local to the notification and guidelines appropriate diagnostic tests. This article describes the most relevant aspects of the Equine Infectious Anemia.

Animais , Masculino , Feminino , Lentivirus Equinos , Vírus da Anemia Infecciosa Equina , Infecções por Retroviridae/virologia , Saúde Pública , Doenças Transmissíveis/veterinária
Protein Sci ; 24(5): 633-42, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25559821


Equine lentivirus receptor 1 (ELR1) has been identified as a functional cellular receptor for equine infectious anemia virus (EIAV). Herein, recombinant ELR1 and EIAV surface glycoprotein gp90 were respectively expressed in Drosophila melanogaster S2 cells, and purified to homogeneity by Ni-NTA affinity chromatography and gel filtration chromatography. Gel filtration chromatography and analytical ultracentrifugation (AUC) analyses indicated that both ELR1 and gp90 existed as individual monomers in solution and formed a complex with a stoichiometry of 1:1 when mixed. The structure of ELR1 was first determined with the molecular replacement method, which belongs to the space group P42 21 2 with one molecule in an asymmetric unit. It contains eight antiparallel ß-sheets, of which four are in cysteine rich domain 1 (CRD1) and two are in CRD2 and CRD3, respectively. Alignment of ELR1 with HVEM and CD134 indicated that Tyr61, Leu70, and Gly72 in CRD1 of ELR1 are important residues for binding to gp90. Isothermal titration calorimetry (ITC) experiments further confirmed that Leu70 and Gly72 are the critical residues.

Lentivirus Equinos/química , Glicoproteínas de Membrana/química , Estrutura Secundária de Proteína , Receptores Virais/química , Proteínas Recombinantes/química , Animais , Drosophila melanogaster , Anemia Infecciosa Equina/genética , Anemia Infecciosa Equina/virologia , Cavalos/virologia , Vírus da Anemia Infecciosa Equina/patogenicidade , Glicoproteínas de Membrana/biossíntese , Glicoproteínas de Membrana/genética , Complexos Multiproteicos/química , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética
BMC Cell Biol ; 11: 73, 2010 Sep 23.
Artigo em Inglês | MEDLINE | ID: mdl-20863390


BACKGROUND: Adipose-Derived Stromal Cells have been shown to have multiple lineage differentiation properties and to be suitable for tissues regeneration in many degenerative processes. Their use has been proposed for the therapy of joint diseases and tendon injuries in the horse. In the present report the genetic manipulation of Equine Adipose-Derived Stromal Cells has been investigated. RESULTS: Equine Adipose-Derived Stromal Cells were successfully virally transduced as well as transiently and stably transfected with appropriate parameters, without detrimental effect on their differentiation properties. Moreover, green fluorescent protein alone, fused to neo gene, or co-expressed as bi-cistronic reporter constructs, driven by viral and house-keeping gene promoters, were tested. The better expressed cassette was employed to stably transfect Adipose-Derived Stromal Cells for cell therapy purposes. Stably transfected Equine Adipose-Derived Stromal Cells with a heterologous secreted viral antigen were able to immunize horses upon injection into the lateral wall of the neck. CONCLUSION: This study provides the methods to successfully transgenize Adipose-Derived Stromal Cells both by lentiviral vector and by transfection using optimized constructs with suitable promoters and reporter genes. In conclusion these findings provide a working platform for the delivery of potentially therapeutic proteins to the site of cells injection via transgenized Equine Adipose-Derived Stromal Cells.

Artropatias/terapia , Infecções por Lentivirus/terapia , Células Estromais/transplante , Transdução Genética/métodos , Transfecção/métodos , Tecido Adiposo/citologia , Animais , Antígenos Virais/genética , Antígenos Virais/imunologia , Antígenos Virais/metabolismo , Diferenciação Celular/genética , Clonagem Molecular , Estudos de Viabilidade , Terapia Genética , Regeneração Tecidual Guiada , Cavalos , Imunidade/genética , Imunização , Artropatias/patologia , Infecções por Lentivirus/genética , Infecções por Lentivirus/imunologia , Lentivirus Equinos , Transplante de Células-Tronco , Células Estromais/imunologia , Células Estromais/metabolismo , Células Estromais/patologia
Biosalud ; 8(1): 10-12, ene.-dic. 2009.
Artigo em Espanhol | LILACS | ID: lil-555153


He leído, en el trabajo titulado “Degradación de lípidos de la dieta por los equinos, ventajas y desventajas del tubo digestivo”, la anotación que refiere: “En animales de estómago simple laabsorción de lípidos ocurre en el intestino delgado, pero en los equinos es diferente debido actividades de microorganismos en el ciego”. La anotación no es muy transcendente para el aspecto fundamental del trabajo, pero hace una anotación quizás errónea al respecto de la tradicional concepción de la anatomía comparada animal, pues indica que hay animales de estómago simple y por consecuencia de extracción, y que hay otros con estómago compuesto.

Lentivirus Equinos , Lipídeos , Estômago
Proc Natl Acad Sci U S A ; 104(38): 15105-10, 2007 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-17846425


Lentiviral envelope antigenic variation and associated immune evasion are believed to present major obstacles to effective vaccine development. Although this perception is widely assumed by the scientific community, there is, to date, no rigorous experimental data assessing the effect of increasing levels of lentiviral Env variation on vaccine efficacy. It is our working hypothesis that Env is, in fact, a primary determinant of vaccine effectiveness. We previously reported that a successful experimental attenuated equine infectious anemia virus vaccine, derived by mutation of the viral S2 accessory gene, provided 100% protection from disease after virulent virus challenge. Here, we sought to comprehensively test our hypothesis by challenging vaccinated animals with proviral strains of defined, increasing Env variation, using variant envelope SU genes that arose naturally during experimental infection of ponies with equine infectious anemia virus. The reference attenuated vaccine combined with these variant Env challenge strains facilitated evaluation of the protection conferred by ancestral immunogens, because the Env of the attenuated vaccine is a direct ancestor to the variant proviral strain Envs. The results demonstrated that ancestral Env proteins did not impart broad levels of protection against challenge. Furthermore, the results displayed a significant inverse linear correlation of Env divergence and protection from disease. This study demonstrates potential obstacles to the use of single isolate ancestral Env immunogens. Finally, these findings reveal that relatively minor Env variation can pose a substantial challenge to lentiviral vaccine immunity, even when attenuated vaccines are used that, to date, achieve the highest levels of vaccine protection.

Variação Antigênica , Produtos do Gene env/imunologia , Vacinas Virais/imunologia , Animais , Anticorpos Antivirais/imunologia , Anemia Infecciosa Equina/imunologia , Anemia Infecciosa Equina/prevenção & controle , Feminino , Cavalos , Vírus da Anemia Infecciosa Equina/imunologia , Lentivirus Equinos/patogenicidade , Masculino , Fatores de Tempo , Proteínas Virais/imunologia , Vacinas Virais/administração & dosagem , Virulência
Rev. med. vet. (Bogota) ; (7): 27-46, mayo 2004. tab, graf
Artigo em Espanhol | LILACS | ID: lil-560462


Con el fin de determinar los parámetros fisiológicos del hematocrito y hemoglobina y ácido-base en una población de equinos atletas de salto alto en Bogotá, se realizaron pruebas en la Escuela de equitación del Ejercito Nacional, CESPO y Club los Arrayanes, localizadas al norte de la ciudad de Bogotá a una altura sobre el nivel del mar de 2.600 metros, con una temperatura promedio de 13 grados centígrados. Se conto con 40 equinos hembras y machos, los cuales se encontraron entre los 6 y los 12 años de edad. La selección de los atletas se realizo de acuerdo con la categoría en la que participaran y a la normalidad encontrada al examen físico. En los tiempos de muestreo se determinaron las variables sanguíneas en reposo y después del ejercicio, teniendo en cuenta los siguientes parámetros: Hemoglobina, hematocrito, pH de la sangre venosa, presión de gas carbónico venoso, bicarbonato en sangre venosa. Estos ejemplares seleccionados se hicieron saltar 15 obstáculos a la mano. El calentamiento previo a la ejecución de los saltos, consistió en el galopeo libre por 3 minutos y las realización de los 15 obstáculos ya mencionados. Durante esta investigación se pudo observar que el ejercicio de salto no es suficiente extenuante para producir cambios drásticos en la sangre y pH comparados con los que se presentan en los caballos de carreras o de enduro, sin embargo se observa una diferencia dentro de los 3 grupos de rendimiento: alto, medio y bajo...

Animais , Anemia Falciforme , Hematócrito , Concentração de Íons de Hidrogênio , Lentivirus Equinos , Prática Profissional
Immunogenetics ; 55(7): 508-14, 2003 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-12942208


Three moderately to broadly recognized equine infectious anemia virus (EIAV) peptides that contained helper T-lymphocyte (Th) 1 epitopes were previously identified. Although lipopeptide immunization was only weakly immunostimulatory in a preliminary study, as measured by T-lymphocyte proliferation responses, it was of interest to define additional broadly recognized Th1 epitopes to include in future immunization trials. Using broadly cross-reactive and conserved Th epitopes known in the related human immunodeficiency virus-1 (HIV-1) and binding motifs defined in human leukocyte antigen DR molecules as guides, this work identified three new peptides containing Th1 epitopes recognized by 60-75% of EIAV infected horses. The observed similarity across species of major histocompatibility complex (MHC) class II binding motifs and the conservation of Th peptides between related viruses should allow easier targeting of Th epitope regions in less well characterized pathogens and/or in species whose MHC class II molecules are poorly defined.

Proteínas de Ligação a DNA/imunologia , HIV-1/imunologia , Antígenos HLA-DR/imunologia , Lentivirus Equinos/imunologia , Peptídeos/imunologia , Fatores de Transcrição/imunologia , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos , Cavalos/imunologia , Humanos