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1.
BMC Plant Biol ; 14: 210, 2014 Aug 03.
Artículo en Inglés | MEDLINE | ID: mdl-25084837

RESUMEN

BACKGROUND: Plant viral infections disturb defense regulatory networks during tissue invasion. Emerging evidence demonstrates that a significant proportion of these alterations are mediated by hormone imbalances. Although the DELLA proteins have been reported to be central players in hormone cross-talk, their role in the modulation of hormone signaling during virus infections remains unknown. RESULTS: This work revealed that TMV-Cg coat protein (CgCP) suppresses the salicylic acid (SA) signaling pathway without altering defense hormone SA or jasmonic acid (JA) levels in Arabidopsis thaliana. Furthermore, it was observed that the expression of CgCP reduces plant growth and delays the timing of floral transition. Quantitative RT-qPCR analysis of DELLA target genes showed that CgCP alters relative expression of several target genes, indicating that the DELLA proteins mediate transcriptional changes produced by CgCP expression. Analyses by fluorescence confocal microscopy showed that CgCP stabilizes DELLA proteins accumulation in the presence of gibberellic acid (GA) and that the DELLA proteins are also stabilized during TMV-Cg virus infections. Moreover, DELLA proteins negatively modulated defense transcript profiles during TMV-Cg infection. As a result, TMV-Cg accumulation was significantly reduced in the quadruple-DELLA mutant Arabidopsis plants compared to wild type plants. CONCLUSIONS: Taken together, these results demonstrate that CgCP negatively regulates the salicylic acid-mediated defense pathway by stabilizing the DELLA proteins during Arabidopsis thaliana viral infection, suggesting that CgCP alters the stability of DELLAs as a mechanism of negative modulation of antiviral defense responses.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/inmunología , Proteínas de la Cápside/fisiología , Regulación de la Expresión Génica de las Plantas , Interacciones Huésped-Patógeno , Arabidopsis/metabolismo , Arabidopsis/virología , Ciclopentanos/metabolismo , Oxilipinas/metabolismo , Desarrollo de la Planta , Plantas Modificadas Genéticamente , Ácido Salicílico/metabolismo , Tobamovirus
2.
Trends Mol Med ; 20(9): 519-28, 2014 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-25164066

RESUMEN

Therapies that selectively target cancer cells for death have been the center of intense research recently. One potential therapy may involve apoptin proteins, which are able to induce apoptosis in cancer cells leaving normal cells unharmed. Apoptin was originally discovered in the Chicken anemia virus (CAV); however, human gyroviruses (HGyV) have recently been found that also harbor apoptin-like proteins. Although the cancer cell specific activity of these apoptins appears to be well conserved, the precise functions and mechanisms of action are yet to be fully elucidated. Strategies for both delivering apoptin to treat tumors and disseminating the protein inside the tumor body are now being developed, and have shown promise in preclinical animal studies.


Asunto(s)
Antineoplásicos/farmacología , Proteínas de la Cápside/farmacología , Sistemas de Liberación de Medicamentos/métodos , Animales , Antineoplásicos/administración & dosificación , Apoptosis/efectos de los fármacos , Proteínas de la Cápside/fisiología , Muerte Celular/efectos de los fármacos , Virus de la Anemia del Pollo/química , Gyrovirus/química , Humanos , Proteínas Virales/aislamiento & purificación , Proteínas Virales/farmacología
3.
J Virol ; 84(19): 10004-15, 2010 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-20668087

RESUMEN

Venezuelan equine encephalitis virus (VEEV) is a significant human and animal pathogen. The highlight of VEEV replication in vitro, in cells of vertebrate origin, is the rapid development of cytopathic effect (CPE), which is strongly dependent upon the expression of viral capsid protein. Besides being an integral part of virions, the latter protein is capable of (i) binding both the nuclear import and nuclear export receptors, (ii) accumulating in the nuclear pore complexes, (iii) inhibiting nucleocytoplasmic trafficking, and (iv) inhibiting transcription of cellular ribosomal and messenger RNAs. Using our knowledge of the mechanism of VEEV capsid protein function in these processes, we designed VEEV variants containing combinations of mutations in the capsid-coding sequences. These mutations made VEEV dramatically less cytopathic but had no effect on infectious virus production. In cell lines that have defects in type I interferon (IFN) signaling, the capsid mutants demonstrated very efficient persistent replication. In other cells, which have no defects in IFN production or signaling, the same mutants were capable of inducing a long-term antiviral state, downregulating virus replication to an almost undetectable level. However, ultimately, these cells also developed a persistent infection, characterized by continuous virus replication and beta IFN (IFN-beta) release. The results of this study demonstrate that the long-term cellular antiviral state is determined by the synergistic effects of type I IFN signaling and the antiviral reaction induced by replicating viral RNA and/or the expression of VEEV-specific proteins. The designed mutants represent an important model for studying the mechanisms of cell interference with VEEV replication and development of persistent infection.


Asunto(s)
Proteínas de la Cápside/genética , Virus de la Encefalitis Equina Venezolana/genética , Virus de la Encefalitis Equina Venezolana/patogenicidad , Encefalomielitis Equina Venezolana/virología , Enfermedad Aguda , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Proteínas de la Cápside/fisiología , Células Cultivadas , Cricetinae , Efecto Citopatogénico Viral/genética , Efecto Citopatogénico Viral/fisiología , ADN Viral/genética , Virus de la Encefalitis Equina Venezolana/inmunología , Virus de la Encefalitis Equina Venezolana/fisiología , Encefalomielitis Equina Venezolana/inmunología , Genes Virales , Enfermedades de los Caballos/inmunología , Enfermedades de los Caballos/virología , Caballos , Humanos , Interferón Tipo I/inmunología , Ratones , Datos de Secuencia Molecular , Mutación , Células 3T3 NIH , Homología de Secuencia de Aminoácido , Transducción de Señal/inmunología , Virus Sindbis/genética , Virus Sindbis/patogenicidad , Virus Sindbis/fisiología , Replicación Viral
4.
J Virol ; 82(3): 1496-504, 2008 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-18032499

RESUMEN

Early during the infection process, rotavirus causes the shutoff of cell protein synthesis, with the nonstructural viral protein NSP3 playing a vital role in the phenomenon. In this work, we have found that the translation initiation factor 2alpha (eIF2alpha) in infected cells becomes phosphorylated early after virus infection and remains in this state throughout the virus replication cycle, leading to a further inhibition of cell protein synthesis. Under these restrictive conditions, however, the viral proteins and some cellular proteins are efficiently translated. The phosphorylation of eIF2alpha was shown to depend on the synthesis of three viral proteins, VP2, NSP2, and NSP5, since in cells in which the expression of any of these three proteins was knocked down by RNA interference, the translation factor was not phosphorylated. The modification of this factor is, however, not needed for the replication of the virus, since mutant cells that produce a nonphosphorylatable eIF2alpha sustained virus replication as efficiently as wild-type cells. In uninfected cells, the phosphorylation of eIF2alpha induces the formation of stress granules, aggregates of stalled translation complexes that prevent the translation of mRNAs. In rotavirus-infected cells, even though eIF2alpha is phosphorylated these granules are not formed, suggesting that the virus prevents the assembly of these structures to allow the translation of its mRNAs. Under these conditions, some of the cellular proteins that form part of these structures were found to change their intracellular localization, with some of them having dramatic changes, like the poly(A) binding protein, which relocates from the cytoplasm to the nucleus in infected cells, a relocation that depends on the viral protein NSP3.


Asunto(s)
Gránulos Citoplasmáticos/metabolismo , Factor 2 Eucariótico de Iniciación/metabolismo , Rotavirus/fisiología , Animales , Proteínas de la Cápside/fisiología , Línea Celular , Núcleo Celular/química , Células Cultivadas , Silenciador del Gen , Macaca mulatta , Ratones , Fosforilación , Proteínas de Unión a Poli(A)/metabolismo , Proteínas de Unión al ARN/fisiología , Proteínas no Estructurales Virales/fisiología
5.
Curr Top Microbiol Immunol ; 309: 39-66, 2006.
Artículo en Inglés | MEDLINE | ID: mdl-16909896

RESUMEN

Rotaviruses, the leading cause of severe dehydrating diarrhea in infants and young children worldwide, are non-enveloped viruses formed by three concentric layers of protein that enclose a genome of double-stranded RNA. These viruses have a specific cell tropism in vivo, infecting primarily the mature enterocytes of the villi of the small intestine. It has been found that rotavirus cell entry is a complex multistep process, in which different domains of the rotavirus surface proteins interact sequentially with different cell surface molecules, which act as attachment and entry receptors. These recently described molecules include integrins (alpha2beta1, alphavbeta3, and alphaxbeta2) and a heat shock protein (hsc70), and have been found to be associated with cell membrane lipid microdomains. The requirement for several cell molecules, which might need to be present and organized in a precise fashion, could explain the cell and tissue tropism of these viruses. This review focuses on recent data describing the interactions between the virus and its receptors, the role of lipid microdomains in rotavirus infection, and the possible mechanism of rotavirus cell entry.


Asunto(s)
Rotavirus/fisiología , Animales , Proteínas de la Cápside/química , Proteínas de la Cápside/fisiología , Polaridad Celular , Humanos , Integrinas/fisiología , Microdominios de Membrana/fisiología , Modelos Biológicos , Ácido N-Acetilneuramínico/metabolismo , Receptores Virales/fisiología , Tropismo , Proteínas no Estructurales Virales/química , Proteínas no Estructurales Virales/fisiología
6.
J Virol ; 79(16): 10369-75, 2005 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-16051829

RESUMEN

Infection by some rotavirus strains requires the presence of sialic acid on the cell surface, its infectivity being reduced in cells treated with neuraminidase. A neuraminidase treatment-resistant mutant was isolated from the porcine rotavirus strain OSU. In reassortant strains, the neuraminidase-resistant phenotype segregated with the gene coding for VP4. The mutant retained its capacity to bind to sialic acid. The VP4 sequence of the mutant differed from that of the parental OSU strain in an Asp-to-Asn substitution at position 100. Neutralization escape mutants selected from an OSU neuraminidase-sensitive clone by monoclonal antibodies that failed to recognize the neuraminidase-resistant mutant strain carried the same mutation at position 100 and were also neuraminidase resistant. Neuraminidase sensitivity was restored when the mutation at position 100 was compensated for by a second mutation (Gln to Arg) at position 125. Molecular mechanics simulations suggest that the neuraminidase-resistant phenotype associated with mutation of OSU residue 100 from Asp to Asn reflects the conformational changes of the sialic acid cleft that accompany sialic acid binding.


Asunto(s)
Neuraminidasa/farmacología , Rotavirus/genética , Porcinos/virología , Animales , Anticuerpos Monoclonales/inmunología , Proteínas de la Cápside/química , Proteínas de la Cápside/fisiología , Línea Celular , Ácido N-Acetilneuramínico/metabolismo , Conformación Proteica , Proteínas de Unión al ARN/química , Proteínas de Unión al ARN/metabolismo , Rotavirus/química , Proteínas no Estructurales Virales/química , Proteínas no Estructurales Virales/metabolismo
7.
Virology ; 333(1): 54-65, 2005 Mar 01.
Artículo en Inglés | MEDLINE | ID: mdl-15708592

RESUMEN

Rotavirus infection of cultured cells induces a progressive increase in plasma membrane permeability to Ca2+. The viral product responsible for this effect is not known. We have used tunicamycin and brefeldin A to prevent glycosylation and membrane traffic and study the involvement of viral glycoproteins, NSP4 and/or VP7, in rotavirus-infected HT29 and MA104 cells. In infected cells, we observed an increase of plasma membrane Ca2+ permeability and a progressive depletion of agonist-releasable ER pools measured with fura 2 and an enhancement of total Ca2+ content measured as 45Ca2+ uptake. Tunicamycin inhibited the increase in membrane Ca2+ permeability, induced a depletion of agonist-releasable and 45Ca2+-sequestered pools. Brefeldin A inhibited the increase of Ca2+ permeability and the increase in 45Ca2+ uptake induced by infection. We propose that the glycosylated viral product NSP4 (and/or VP7) travels to the plasma membrane to form a Ca2+ channel and hence elevate Ca2+ permeability.


Asunto(s)
Brefeldino A/farmacología , Calcio/metabolismo , Permeabilidad de la Membrana Celular/efectos de los fármacos , Permeabilidad de la Membrana Celular/fisiología , Rotavirus/fisiología , Tunicamicina/farmacología , Animales , Antígenos Virales/fisiología , Proteínas de la Cápside/fisiología , Línea Celular , ARN Polimerasas Dirigidas por ADN/fisiología , Glicosilación/efectos de los fármacos , Procesamiento Proteico-Postraduccional/efectos de los fármacos , Transporte de Proteínas/efectos de los fármacos , Proteínas no Estructurales Virales/fisiología
8.
J Virol ; 77(13): 7254-60, 2003 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-12805424

RESUMEN

Rotavirus infection seems to be a multistep process in which the viruses are required to interact with several cell surface molecules to enter the cell. The virus spike protein VP4, which is cleaved by trypsin into two subunits, VP5 and VP8, is involved in some of these interactions. We have previously shown that the neuraminidase-sensitive rotavirus strain RRV initially attaches to a sialic acid-containing cell molecule through the VP8 subunit of VP4 and subsequently interacts with integrin alpha2beta1 through VP5. After these initial contacts, the virus interacts with at least two additional proteins located at the cell surface, the integrin alphavbeta3 and the heat shock cognate protein Hsc70. In this work, we have shown that rotavirus RRV and its neuraminidase-resistant variant nar3 interact with Hsc70 through a VP5 domain located between amino acids 642 and 658 of the protein. This conclusion is based on the observation that a recombinant protein comprising the 300 carboxy-terminal amino acids of VP5 binds specifically to Hsc70 and a synthetic peptide containing amino acids 642 to 658 competes with the binding of the RRV and nar3 viruses to the heat shock protein. The VP5 peptide also competed with the binding to Hsc70 of the recombinant VP5 protein, and an antibody to Hsc70 reduced the binding of the recombinant protein to the surface of MA104 cells. The fact that the synthetic peptide blocks the infectivity of rotaviruses RRV and nar3 but not their binding to cells indicates that the interaction of VP5 with Hsc70 most probably occurs at a postattachment step during the virus entry process.


Asunto(s)
Antígenos Virales , Proteínas de la Cápside/fisiología , Proteínas HSP70 de Choque Térmico/fisiología , Rotavirus/fisiología , Secuencia de Aminoácidos , Proteínas de la Cápside/química , Línea Celular , Ensayo de Inmunoadsorción Enzimática , Proteínas del Choque Térmico HSC70 , Datos de Secuencia Molecular , Homología de Secuencia de Aminoácido
9.
J Virol ; 77(3): 1757-63, 2003 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-12525609

RESUMEN

Rotavirus is a nonenveloped virus with a three-layered capsid. The inner layer, made of VP2, encloses the genomic RNA and two minor proteins, VP1 and VP3, with which it forms the viral core. Core assembly is coupled with RNA viral replication and takes place in definite cellular structures termed viroplasms. Replication and encapsidation mechanisms are still not fully understood, and little information is available about the intermolecular interactions that may exist among the viroplasmic proteins. NSP2 and NSP5 are two nonstructural viroplasmic proteins that have been shown to interact with each other. They have also been found to be associated with precore replication intermediates that are precursors of the viral core. In this study, we show that NSP5 interacts with VP2 in infected cells. This interaction was demonstrated with recombinant proteins expressed from baculovirus recombinants or in bacterial systems. NSP5-VP2 interaction also affects the stability of VP6 bound to VP2 assemblies. The data presented showed evidence, for the first time, of an interaction between VP2 and a nonstructural rotavirus protein. Published data and the interaction demonstrated here suggest a possible role for NSP5 as an adapter between NSP2 and the replication complex VP2-VP1-VP3 in core assembly and RNA encapsidation, modulating the role of NSP2 as a molecular motor involved in the packaging of viral mRNA.


Asunto(s)
Antígenos Virales , Proteínas de la Cápside/fisiología , Rotavirus/fisiología , Proteínas Virales/fisiología , Ensamble de Virus , Proteínas de la Cápside/análisis , Proteínas de la Cápside/química , Pruebas de Precipitina , Proteínas Virales/análisis
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