The Experts below are selected from a list of 162 Experts worldwide ranked by ideXlab platform
García, María Laura - One of the best experts on this subject based on the ideXlab platform.
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The psorosis disease of citrus: a pale light at the end of the tunnel
2020Co-Authors: Moreno Pedro, Guerri José, García, María LauraAbstract:First reported in 1896, psorosis was the first citrus disease proven to be graft transmissible and also the first for which eradication and budwood certification programs were launched to prevent its economic damage. For many years psorosis etiology remained elusive, and only in 1986 was the disease associated with the presence of virus-like particles in infected plants. However, in the last 2 decades a virus with unusual morphology (Citrus psorosis virus, CPsV) was characterized and closely associated with psorosis disease as previously defined by field symptoms and by biological indexing in sensitive indicator plants. With a tripartite, negative-sense, RNA genome and a ~48 kDa coat protein, CPsV, the presumed causal agent of psorosis, is the type member of the genus Ophiovirus, within the new family Ophioviridae. Availability of the complete genomic sequence of 2 CPsV isolates and partial sequences of many others has enabled i) setting up rapid and sensitive RNA-based detection methods, ii) testing different citrus and relatives for resistance to CPsV, iii) identification of the 2 components (psorosis A and psorosis B) traditionally associated with non-scaled and scaled bark inoculum, respectively, from psorosis-infected plants and study their interactions, iv) analysis of genetic variation and evolutionary forces shaping the CPsV populations, v) preliminary studies on the interactions between virus and host factors, and vi) development of transgenic citrus plants expressing variable degrees of resistance to CPsV. In summary, 120 years after the first report on psorosis we start seeing a pale light at the end of the tunnel.Instituto de Biotecnologia y Biologia Molecula
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Bioinformatic and mutational analysis of Ophiovirus movement proteins, belonging to the 30K superfamily
2019Co-Authors: Borniego, María Belén, Karlin David, Peña, Eduardo José, Robles Luna Gabriel, García, María LauraAbstract:Ophioviridae is a family of segmented, negative-sense, single-stranded RNA plant viruses. We showed that their cell-to-cell movement protein (MP) is an isolated member of the 30K MP superfamily with a unique structural organization. All 30K MPs share a core domain that contains a nearly-invariant signature aspartate. We examined its role in the MP of Citrus psorosis virus (CPsV) and Mirafiori lettuce big-vein virus (MiLBVV). Alanine substitution of this aspartate prevented plasmodesmata accumulation of MPMiLBVV, while MPCPsV was not affected. The capacity of Ophiovirus MPs to increase the plasmodesmata size exclusion limit and non-cell autonomous protein feature was abolished in both mutants. To investigate the role of the signature aspartate in cell-to-cell movement, we constructed a new movement-deficient Tobacco mosaic virus vector used for trans-complementation assays. We showed that both Ophiovirus MP mutants lack the cell-to-cell movement capacity, confirming that this signature aspartate is essential for viral cell-to-cell movement.Instituto de Biotecnologia y Biologia Molecula
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Ophioviruses CPsV and MiLBVV movement protein is encoded in RNA 2 and interacts with the coat protein
2019Co-Authors: Robles Luna Gabriel, Borniego, María Belén, Peña, Eduardo José, Heinlein Manfred, García, María LauraAbstract:Citrus psorosis virus (CPsV) and Mirafiori lettuce big-vein virus (MiLBVV), members of the Ophioviridae family, have segmented negative-sense single-stranded RNA genomes. To date no reports have described how Ophioviruses spread within host plants and/or the proteins involved in this process. Here we show that the 54K protein of CPsV is encoded by RNA 2 and describe its subcellular distribution. Upon transient expression in Nicotiana benthamiana epidermal cells the 54K protein, and also its 54K counterpart protein of MiLBVV, localize to plasmodesmata and enhance GFP cell-to-cell diffusion between cells. Both proteins, but not the coat proteins (CP) of the respective viruses, functionally trans-complement cell-to-cell movement-defective Potato virus X (PVX) and Tobacco mosaic virus (TMV) mutants. The 54K and 54K proteins interact with the virus-specific CP in the cytoplasm, suggesting a potential role of CP in Ophiovirus movement. This is the first study characterizing the movement proteins (MP) of Ophioviruses.Facultad de Ciencias ExactasInstituto de Biotecnologia y Biologia Molecula
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Identification of Mirafiori lettuce big-vein virus and Lettuce big-vein associated virus infecting Lactuca sativa with symptoms of lettuce big-vein disease in Argentina
2019Co-Authors: Barcala Tabarrozzi, Andrés E., Robles Luna Gabriel, Peña E. J., Dal Bó Elena, Reyes C. A., García, María LauraAbstract:Lettuce big-vein disease (BVD) affects all major lettuce-producing areas of the world. The causal agent is Mirafiori lettuce big-vein virus (MLBVV), an Ophiovirus transmitted by the soil-borne fungus Olpidium brassicae (Lot et al., 2002). MLBVV has been detected in many different areas of the world but never in Argentina. La Plata has about 700 ha of lettuce with a production of about 13 000 tonnes, and with about 70% of the total production from Buenos Aires Province. BVD has been detected in different areas in the north and west of the La Plata horticultural green belt. Many of the plants with BVD symptoms had leaf distortions of moderate severity, which affected their commercial value.Facultad de Ciencias ExactasFacultad de Ciencias Agrarias y Forestale
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Genetic variation of populations of Citrus psorosis virus
2019Co-Authors: Martín Susan, García, María Laura, Moreno Pedro, Troisi Antonella, Rubio Luis, Legarreta Gonzalo, Grau Oscar, Allioto Daniela, Guerri JoséAbstract:Citrus psorosis virus (CPsV), the type species of genus Ophiovirus, has a segmented, negative-stranded RNA genome. We examined the population structure and genetic variation of CPsV in three coding regions located in RNAs 1, 2 and 3, analysing 22 isolates from Argentina, California, Florida, Italy and Spain. Most isolates contained a predominant sequence and some minor variants. Estimations of the genetic diversity and phylogenetic clustering of isolates disclosed two populations, one comprising isolates from Spain, Italy, Florida and California and the other including the Argentinean isolates. Isolate CPV-4 (from Texas) included for comparison was distant from both groups, suggesting that it belongs to a third group. The low ratio between non-synonymous and synonymous nucleotide substitutions indicated strong selection for amino acid sequence conservation, particularly in the coat protein gene. Incongruent phylogenetic relationships in different genomic regions suggested that exchange of genomic segments may have contributed to CPsV evolution.Instituto de Biotecnologia y Biologia Molecula
Robles Luna Gabriel - One of the best experts on this subject based on the ideXlab platform.
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Bioinformatic and mutational analysis of Ophiovirus movement proteins, belonging to the 30K superfamily
2019Co-Authors: Borniego, María Belén, Karlin David, Peña, Eduardo José, Robles Luna Gabriel, García, María LauraAbstract:Ophioviridae is a family of segmented, negative-sense, single-stranded RNA plant viruses. We showed that their cell-to-cell movement protein (MP) is an isolated member of the 30K MP superfamily with a unique structural organization. All 30K MPs share a core domain that contains a nearly-invariant signature aspartate. We examined its role in the MP of Citrus psorosis virus (CPsV) and Mirafiori lettuce big-vein virus (MiLBVV). Alanine substitution of this aspartate prevented plasmodesmata accumulation of MPMiLBVV, while MPCPsV was not affected. The capacity of Ophiovirus MPs to increase the plasmodesmata size exclusion limit and non-cell autonomous protein feature was abolished in both mutants. To investigate the role of the signature aspartate in cell-to-cell movement, we constructed a new movement-deficient Tobacco mosaic virus vector used for trans-complementation assays. We showed that both Ophiovirus MP mutants lack the cell-to-cell movement capacity, confirming that this signature aspartate is essential for viral cell-to-cell movement.Instituto de Biotecnologia y Biologia Molecula
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Identification of Mirafiori lettuce big-vein virus and Lettuce big-vein associated virus infecting Lactuca sativa with symptoms of lettuce big-vein disease in Argentina
2019Co-Authors: Barcala Tabarrozzi, Andrés E., Robles Luna Gabriel, Peña E. J., Dal Bó Elena, Reyes C. A., García, María LauraAbstract:Lettuce big-vein disease (BVD) affects all major lettuce-producing areas of the world. The causal agent is Mirafiori lettuce big-vein virus (MLBVV), an Ophiovirus transmitted by the soil-borne fungus Olpidium brassicae (Lot et al., 2002). MLBVV has been detected in many different areas of the world but never in Argentina. La Plata has about 700 ha of lettuce with a production of about 13 000 tonnes, and with about 70% of the total production from Buenos Aires Province. BVD has been detected in different areas in the north and west of the La Plata horticultural green belt. Many of the plants with BVD symptoms had leaf distortions of moderate severity, which affected their commercial value.Facultad de Ciencias ExactasFacultad de Ciencias Agrarias y Forestale
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Ophioviruses CPsV and MiLBVV movement protein is encoded in RNA 2 and interacts with the coat protein
2019Co-Authors: Robles Luna Gabriel, Borniego, María Belén, Peña, Eduardo José, Heinlein Manfred, García, María LauraAbstract:Citrus psorosis virus (CPsV) and Mirafiori lettuce big-vein virus (MiLBVV), members of the Ophioviridae family, have segmented negative-sense single-stranded RNA genomes. To date no reports have described how Ophioviruses spread within host plants and/or the proteins involved in this process. Here we show that the 54K protein of CPsV is encoded by RNA 2 and describe its subcellular distribution. Upon transient expression in Nicotiana benthamiana epidermal cells the 54K protein, and also its 54K counterpart protein of MiLBVV, localize to plasmodesmata and enhance GFP cell-to-cell diffusion between cells. Both proteins, but not the coat proteins (CP) of the respective viruses, functionally trans-complement cell-to-cell movement-defective Potato virus X (PVX) and Tobacco mosaic virus (TMV) mutants. The 54K and 54K proteins interact with the virus-specific CP in the cytoplasm, suggesting a potential role of CP in Ophiovirus movement. This is the first study characterizing the movement proteins (MP) of Ophioviruses.Facultad de Ciencias ExactasInstituto de Biotecnologia y Biologia Molecula
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Citrus psorosis virus movement protein contains an aspartic protease required for autocleavage and the formation of tubule-like structures at plasmodesmata
'American Society for Microbiology', 2018Co-Authors: Robles Luna Gabriel, Borniego, María Belén, Peña, Eduardo José, Heinlein Manfred, García, María LauraAbstract:Plant virus cell-to-cell movement is an essential step in viral infections. This process is facilitated by specific virus-encoded movement proteins (MPs), which manipulate the cell wall channels between neighboring cells known as plasmodesmata (PD). Citrus psorosis virus (CPsV) infection in sweet orange involves the formation of tubule-like structures within PD, suggesting that CPsV belongs to "tubuleforming" viruses that encode MPs able to assemble a hollow tubule extending between cells to allow virus movement. Consistent with this hypothesis, we show that the MP of CPsV (MPCPsV) indeed forms tubule-like structures at PD upon transient expression in Nicotiana benthamiana leaves. Tubule formation by MPCPsV depends on its cleavage capacity, mediated by a specific aspartic protease motif present in its primary sequence. A single amino acid mutation in this motif abolishes MPCPsV cleavage, alters the subcellular localization of the protein, and negatively affects its activity in facilitating virus movement. The amino-terminal 34-kDa cleavage product (34KCPsV), but not the 20-kDa fragment (20KCPsV), supports virus movement. Moreover, similar to tubule-forming MPs of other viruses, MPCPsV (and also the 34KCPsV cleavage product) can homooligomerize, interact with PD-located protein 1 (PDLP1), and assemble tubule-like structures at PD by a mechanism dependent on the secretory pathway. 20KCPsV retains the protease activity and is able to cleave a cleavage-deficient MPCPsV in trans. Altogether, these results demonstrate that CPsV movement depends on the autolytic cleavage of MPCPsV by an aspartic protease activity, which removes the 20KCPsV protease and thereby releases the 34KCPsV protein for PDLP1-dependent tubule formation at PD. IMPORTANCE Infection by citrus psorosis virus (CPsV) involves a self-cleaving aspartic protease activity within the viral movement protein (MP), which results in the production of two peptides, termed 34KCPsV and 20KCPsV, that carry the MP and viral protease activities, respectively. The underlying protease motif within the MP is also found in the MPs of other members of the Aspiviridae family, suggesting that protease-mediated protein processing represents a conserved mechanism of protein expression in this virus family. The results also demonstrate that CPsV and potentially other Ophioviruses move by a tubule-guided mechanism. Although several viruses from different genera were shown to use this mechanism for cell-to-cell movement, our results also demonstrate that this mechanism is controlled by posttranslational protein cleavage. Moreover, given that tubule formation and virus movement could be inhibited by a mutation in the protease motif, targeting the protease activity for inactivation could represent an important approach for Ophiovirus control.Fil: Robles Luna, Gabriel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; ArgentinaFil: Peña, Eduardo José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; ArgentinaFil: Borniego, María Belén. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; ArgentinaFil: Heinlein, Manfred. Université de Strasbourg; Francia. Centre National de la Recherche Scientifique; FranciaFil: Garcia, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; Argentin
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Identification and characterization of two RNA silencing suppressors encoded by Ophioviruses
2017Co-Authors: Robles Luna Gabriel, Borniego, María Belén, Reyes, Carina A., Peña, Eduardo J., Ocolotobiche Eliana, Baeza Cecilia, Kormelink Richard, García, María LauraAbstract:Citrus psorosis virus and Mirafiori lettuce big-vein virus are two members of the genus Ophiovirus, family Ophioviridae. So far, how these viruses can interfere in the antiviral RNA silencing pathway is not known. In this study, using a local GFP silencing assay on Nicotiana benthamiana, the 24K–25K and the movement protein (MP) of both viruses were identified as RNA silencing suppressor proteins. Upon their co-expression with GFP in N. benthamiana 16c plants, the proteins also showed to suppress systemic RNA (GFP) silencing. The MPCPsV and 24KCPsV proteins bind long (114 nucleotides) but not short-interfering (21 nt) dsRNA, and upon transgenic expression, plants showed developmental abnormalities that coincided with an altered miRNA accumulation pattern. Furthermore, both proteins were able to suppress miRNA-induced silencing of a GFP-sensor construct and the co-expression of MPCPsV and 24KCPsV exhibited a stronger effect, suggesting they act at different stages of the RNAi pathway.
Borniego, María Belén - One of the best experts on this subject based on the ideXlab platform.
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Bioinformatic and mutational analysis of Ophiovirus movement proteins, belonging to the 30K superfamily
2019Co-Authors: Borniego, María Belén, Karlin David, Peña, Eduardo José, Robles Luna Gabriel, García, María LauraAbstract:Ophioviridae is a family of segmented, negative-sense, single-stranded RNA plant viruses. We showed that their cell-to-cell movement protein (MP) is an isolated member of the 30K MP superfamily with a unique structural organization. All 30K MPs share a core domain that contains a nearly-invariant signature aspartate. We examined its role in the MP of Citrus psorosis virus (CPsV) and Mirafiori lettuce big-vein virus (MiLBVV). Alanine substitution of this aspartate prevented plasmodesmata accumulation of MPMiLBVV, while MPCPsV was not affected. The capacity of Ophiovirus MPs to increase the plasmodesmata size exclusion limit and non-cell autonomous protein feature was abolished in both mutants. To investigate the role of the signature aspartate in cell-to-cell movement, we constructed a new movement-deficient Tobacco mosaic virus vector used for trans-complementation assays. We showed that both Ophiovirus MP mutants lack the cell-to-cell movement capacity, confirming that this signature aspartate is essential for viral cell-to-cell movement.Instituto de Biotecnologia y Biologia Molecula
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Ophioviruses CPsV and MiLBVV movement protein is encoded in RNA 2 and interacts with the coat protein
2019Co-Authors: Robles Luna Gabriel, Borniego, María Belén, Peña, Eduardo José, Heinlein Manfred, García, María LauraAbstract:Citrus psorosis virus (CPsV) and Mirafiori lettuce big-vein virus (MiLBVV), members of the Ophioviridae family, have segmented negative-sense single-stranded RNA genomes. To date no reports have described how Ophioviruses spread within host plants and/or the proteins involved in this process. Here we show that the 54K protein of CPsV is encoded by RNA 2 and describe its subcellular distribution. Upon transient expression in Nicotiana benthamiana epidermal cells the 54K protein, and also its 54K counterpart protein of MiLBVV, localize to plasmodesmata and enhance GFP cell-to-cell diffusion between cells. Both proteins, but not the coat proteins (CP) of the respective viruses, functionally trans-complement cell-to-cell movement-defective Potato virus X (PVX) and Tobacco mosaic virus (TMV) mutants. The 54K and 54K proteins interact with the virus-specific CP in the cytoplasm, suggesting a potential role of CP in Ophiovirus movement. This is the first study characterizing the movement proteins (MP) of Ophioviruses.Facultad de Ciencias ExactasInstituto de Biotecnologia y Biologia Molecula
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Citrus psorosis virus movement protein contains an aspartic protease required for autocleavage and the formation of tubule-like structures at plasmodesmata
'American Society for Microbiology', 2018Co-Authors: Robles Luna Gabriel, Borniego, María Belén, Peña, Eduardo José, Heinlein Manfred, García, María LauraAbstract:Plant virus cell-to-cell movement is an essential step in viral infections. This process is facilitated by specific virus-encoded movement proteins (MPs), which manipulate the cell wall channels between neighboring cells known as plasmodesmata (PD). Citrus psorosis virus (CPsV) infection in sweet orange involves the formation of tubule-like structures within PD, suggesting that CPsV belongs to "tubuleforming" viruses that encode MPs able to assemble a hollow tubule extending between cells to allow virus movement. Consistent with this hypothesis, we show that the MP of CPsV (MPCPsV) indeed forms tubule-like structures at PD upon transient expression in Nicotiana benthamiana leaves. Tubule formation by MPCPsV depends on its cleavage capacity, mediated by a specific aspartic protease motif present in its primary sequence. A single amino acid mutation in this motif abolishes MPCPsV cleavage, alters the subcellular localization of the protein, and negatively affects its activity in facilitating virus movement. The amino-terminal 34-kDa cleavage product (34KCPsV), but not the 20-kDa fragment (20KCPsV), supports virus movement. Moreover, similar to tubule-forming MPs of other viruses, MPCPsV (and also the 34KCPsV cleavage product) can homooligomerize, interact with PD-located protein 1 (PDLP1), and assemble tubule-like structures at PD by a mechanism dependent on the secretory pathway. 20KCPsV retains the protease activity and is able to cleave a cleavage-deficient MPCPsV in trans. Altogether, these results demonstrate that CPsV movement depends on the autolytic cleavage of MPCPsV by an aspartic protease activity, which removes the 20KCPsV protease and thereby releases the 34KCPsV protein for PDLP1-dependent tubule formation at PD. IMPORTANCE Infection by citrus psorosis virus (CPsV) involves a self-cleaving aspartic protease activity within the viral movement protein (MP), which results in the production of two peptides, termed 34KCPsV and 20KCPsV, that carry the MP and viral protease activities, respectively. The underlying protease motif within the MP is also found in the MPs of other members of the Aspiviridae family, suggesting that protease-mediated protein processing represents a conserved mechanism of protein expression in this virus family. The results also demonstrate that CPsV and potentially other Ophioviruses move by a tubule-guided mechanism. Although several viruses from different genera were shown to use this mechanism for cell-to-cell movement, our results also demonstrate that this mechanism is controlled by posttranslational protein cleavage. Moreover, given that tubule formation and virus movement could be inhibited by a mutation in the protease motif, targeting the protease activity for inactivation could represent an important approach for Ophiovirus control.Fil: Robles Luna, Gabriel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; ArgentinaFil: Peña, Eduardo José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; ArgentinaFil: Borniego, María Belén. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; ArgentinaFil: Heinlein, Manfred. Université de Strasbourg; Francia. Centre National de la Recherche Scientifique; FranciaFil: Garcia, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; Argentin
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Caracterización mutacional y funcional de las proteínas de movimiento de Ophiovirus
'Universidad Nacional de La Plata', 2017Co-Authors: Borniego, María BelénAbstract:Las plagas de los cultivos son una gran amenaza para la producción agrícola, en particular, las enfermedades virales son responsables de importantes pérdidas económicas en la producción de cultivos de importancia mundial. Los Ophiovirus son virus de plantas causantes de importantes enfermedades en cultivos de cítricos, lechuga, arándanos y plantas ornamentales. La psorosis de los cítricos es una enfermedad viral causada por el Ophiovirus Citrus psorosis virus (CPsV), que produce un deterioro progresivo de los árboles al afectar sus tejidos conductores. Hasta el momento, no se conocen cultivares resistentes a CPsV. La enfermedad se ha reportado en muchas regiones productoras de cítricos a lo largo de todo el mundo. En nuestro país, se disemina de manera natural, reduciendo la producción de cítricos. La situación de la citricultura en todos los países de nuestra región se encuentra en expansión y constituye una actividad de muy alta importancia económica en la mayoría de éstos, con especial impacto en sus economías. El Big-vein de lechuga es otra enfermedad importante causada por Ophiovirus que afecta a las principales áreas de producción de lechuga en nuestra región. El agente etiológico es el Ophiovirus Mirafiori lettuce big vein virus (MiLBVV), el cual es transmitido por zoosporas móviles del hongo Olpidium virulentus. El big-vein causa serios problemas en los cultivos de lechuga durante los períodos más fríos del año y afecta a todos los tipos de lechuga que crecen en suelo al aire libre o bajo cubierta, y a los cultivos hidropónicos. La importancia económica de la enfermedad se debe a los síntoma del follaje, que reducen el valor de mercado, y a los retrasos en la formación de la cabeza y la disminución del tamaño de la planta, que reduce la proporción de plantas cosechables, ya que estas plantas suelen ser descartadas en el momento de la cosecha. Los cultivos de arándanos y variedades ornamentales como tulipán, fresia y ranúnculo también se encuentran afectados por Ophiovirus. Debido a las pérdidas económicas ocasionadas por los Ophiovirus, resulta de interés comprender el proceso infectivo responsable de las enfermedades causadas por estos virus para el diseño de nuevas estrategias que ayuden a controlar su diseminación. En este trabajo de tesis, se realizó un análisis exhaustivo de la secuencia y la predicción de estructura secundaria, junto con estudios funcionales sobre las MPs de esta familia de virus segmentados negativos. Se encontró que la MP de los Ophiovirus es un miembro aislado de la superfamilia 30K, con una organización estructural única. En los Ophiovirus, el ácido aspártico conservado dentro de este dominio es necesario para aumentar el tamaño límite de exclusión molecular de los plasmodesmos, para mantener su capacidad NCAP y para dirigir el movimiento célula a célula de un vector viral de TMV deficiente en tal movimiento. Además, hemos logrado identificar otros posibles dominios y motivos funcionales presentes en la MP de CPsV, involucrados tanto en su localización subcelular como en su actividad como proteína de movimiento viral. Así, se encontró una NLS_BP funcional entre los aminoácidos 255 y 271, y la presencia de una posible señal de localización en plasmodesmos y de un péptido de tránsito a cloroplastos, ambos presentes en la región amino terminal de la MP. Encontramos que la región C-terminal es necesaria para que la proteína conserve la actividad proteasa, sin embargo, no sería necesaria para la función de movimiento célula a célula de la MP. Los ensayos de movimiento sistémico con virus híbridos, nos permitieron mostrar las primeras evidencias que afirman la idea de que el transporte sistémico de los Ophiovirus estaría regulado por sus MP. Por ensayos de coinmunoprecipitación, hemos logrado identificar un número importante de proteínas de la planta que estarían interaccionando con la MP de CPsV. Por búsqueda bibliográfica, se encontró que un alto porcentaje de éstas participan en el transporte celular de macromoléculas y en las distintas etapas de expresión génica. También se identificaron una cantidad significativa de chaperonas, de proteínas asociadas con la respuesta frente a patógenos y hormonas, y de proteínas del sistema ubiquitina-proteasoma. Por último, se confirmó la localización de la proteína 24K en nucléolo, Cajal bodies, microtúbulos y posiblemente D-bodies. Por medio de un análisis bioinformático, seguido del correspondiente análisis mutacional, se encontró la presencia de una posible señal de exportación nuclear que sería requerida para la correcta localización nucleolar de la proteína. Además, se encontró un posible motivo WG/GW de unión a la proteína argonauta que sería necesario para la localización de 24K en D-bodies. La región carboxilo terminal sería requerida para la acumulación de la proteína en nucléolo, mientras que la región amino terminal estaría involucrada en la acumulación de la proteína en agregados en nucleoplasma.Facultad de Ciencias Exacta
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Identification and characterization of two RNA silencing suppressors encoded by Ophioviruses
2017Co-Authors: Robles Luna Gabriel, Borniego, María Belén, Reyes, Carina A., Peña, Eduardo J., Ocolotobiche Eliana, Baeza Cecilia, Kormelink Richard, García, María LauraAbstract:Citrus psorosis virus and Mirafiori lettuce big-vein virus are two members of the genus Ophiovirus, family Ophioviridae. So far, how these viruses can interfere in the antiviral RNA silencing pathway is not known. In this study, using a local GFP silencing assay on Nicotiana benthamiana, the 24K–25K and the movement protein (MP) of both viruses were identified as RNA silencing suppressor proteins. Upon their co-expression with GFP in N. benthamiana 16c plants, the proteins also showed to suppress systemic RNA (GFP) silencing. The MPCPsV and 24KCPsV proteins bind long (114 nucleotides) but not short-interfering (21 nt) dsRNA, and upon transgenic expression, plants showed developmental abnormalities that coincided with an altered miRNA accumulation pattern. Furthermore, both proteins were able to suppress miRNA-induced silencing of a GFP-sensor construct and the co-expression of MPCPsV and 24KCPsV exhibited a stronger effect, suggesting they act at different stages of the RNAi pathway.
J. Hammond - One of the best experts on this subject based on the ideXlab platform.
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first report of freesia sneak virus associated with foliar necrosis of freesia refracta in bulgaria
Plant Disease, 2013Co-Authors: S G Bobev, O I Taphradjiiski, J. Hammond, Anna Maria VairaAbstract:In the early spring of 2011 and 2012, severe necrotic leaf symptoms were observed on freesia (Freesia refracta, family Iridaceae) in several greenhouses around Plovdiv (south central Bulgaria). The disease spread and symptom severity in several cultivars (Medeo, Calvados, and Pink Fountain) led to nearly complete production failure for some growers. Initial symptoms consisted of scattered pale, chlorotic, interveinal lesions that coalesced. Later, irregular brown to black necrotic blotches partially covered the leaves. Flower break was also observed. Diseased plants were collected in late April 2012 from one of the surveyed greenhouses, where >90% of Medeo (white-flowered) and 35 to 40% of Pink Fountain (pink) plants were symptomatic. Total RNA was extracted from three pooled samples of ~10 plants each and analyzed for Freesia sneak virus (3) (FreSV, Ophiovirus, Ophioviridae) infection by RT-PCR. A generic Ophiovirus RT-PCR (4) yielded the diagnostic 136-bp product, while primers FOV1 (TGCTCGAATAGCCGGAACT...
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first report of freesia sneak virus in freesia sp in virginia
Plant Disease, 2009Co-Authors: Anna Maria Vaira, M A Hansen, Michael D Reinsel, C Murphy, J. HammondAbstract:In the spring of 2008, freesia, cvs. Honeymoon and Santana, with striking virus-like symptoms similar to freesia leaf necrosis disease were received by the Virginia Tech Plant Disease Clinic from a cut-flower nursery in Gloucester, VA and forwarded for analysis to the USDA-ARS Floral and Nursery Plants Research Unit in Beltsville, MD. Approximately 25% of the plants had coalescing, interveinal, chlorotic, whitish, necrotic or dark brown-to-purple necrotic spots on leaves. Symptomatic plants were scattered within the planting. Fifteen symptomatic plants were collected between March and May of 2008, and nucleic acid extracts were analyzed for Ophiovirus infection by reverse transcription (RT)-PCR with Ophiovirus-specific degenerate primers (2). The diagnostic 136-bp Ophiovirus product from the RdRp gene was amplified from 14 of 15 freesia plants tested. A partially purified virus preparation was analyzed by transmission electron microscopy and potyvirus- and Ophiovirus-like particles were detected. The poty...
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first report of freesia sneak virus infecting lachenalia cultivars in south africa
Plant Disease, 2007Co-Authors: A M Vaira, R Kleynhans, J. HammondAbstract:Lachenalia (Lachenalia species, family Hyacinthaceae) is a bulbous ornamental plant endemic to southern Africa. In 1998, several lachenalia lines from ARC-Roodeplaat showing virus-like symptoms, and presumed to be infected with Ornithogalum mosaic virus (OrMV), were sent from South Africa under an APHIS permit for examination in Beltsville, MD. In addition to potyvirus-like particles, fine filamentous particles consistent with those of Ophioviruses were observed with electron microscopy in some of the plant samples. Ophiovirus virions are filamentous nucleocapsids approximately 3 nm in diameter forming circularized structures of different lengths and are not easily detectable with electron microscopy. A reverse transcription (RT)-PCR assay using genus-specific degenerate primers that yield a 136-bp fragment from the RdRp gene is currently the best tool for detecting Ophioviruses (3). Complementary DNA was produced from lachenalia total RNA extracts using either random hexamers or Ophiovirus-specific prime...
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Detection of Varicosavirus and Ophiovirus in lettuce associated with lettuce big-vein symptoms in Brazil
Fitopatologia Brasileira, 2005Co-Authors: Addolorata Colariccio, Alexandre L. R. Chaves, Marcelo Eiras, C. M. Chagas, Piero RoggeroAbstract:Em levantamentos realizados entre 1998 e 2003, nas principais regioes produtoras de alface (Lactuca sativa) e escarola (Cichorium endivia) no cinturao verde de Sao Paulo, foram observados sintomas de espessamento de nervuras foliares, clorose, crescimento irregular e ausencia de formacao da cabeca. Por meio de testes biologicos, DAS-Enzyme linked immunosorbent assay (Elisa) e microscopia eletronica de transmissao constatou-se a presenca do Lettuce big-vein associated virus e Mirafiori lettuce virus, responsaveis pela sindrome do espessamento clorotico das nervuras da alface ("lettuce big-vein").
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Transmission by Olpidium brassicae of Mirafiori lettuce virus and Lettuce big-vein virus, and Their Roles in Lettuce Big-Vein Etiology
Phytopathology®, 2002Co-Authors: Hervé Lot, Robert N. Campbell, Sylvie Souche, Robert G. Milne, Piero RoggeroAbstract:Lot, H., Campbell, R. N., Souche, S., Milne, R. G., and Roggero, P. 2002. Transmission by Olpidium brassicae of Mirafiori lettuce virus and Lettuce big-vein virus, and their roles in lettuce big-vein etiology. Phytopathology 92:288-293. Big-vein disease occurs on lettuce worldwide in temperate conditions; the causal agent has been presumed to be Lettuce big-vein virus (LBVV), genus Varicosavirus, vectored by the soilborne fungus Olpidium brassicae. Recently, the role of LBVV in the etiology of big-vein disease has been questioned because a second soilborne virus, Mirafiori lettuce virus (MiLV), genus Ophiovirus, has been found frequently in big-veinaffected lettuce. LBVV and MiLV, detectable and distinguishable by enzyme-linked immunosorbent assay using specific antisera, were tested for their ability to be transmitted from lettuce to lettuce by mechanical inoculation of sap extracts, or by zoospores of O. brassicae, and to cause big-vein disease. Both viruses were mechanically transmissible from lettuce to herbaceous hosts and to lettuce, but very erratically. LBVV was transmitted by O. brassicae but lettuce infected with only this virus never showed symptoms. MiLV was transmitted in the same manner, and lettuce infected with this virus alone consistently developed big-vein symptoms regardless of the presence or absence of LBVV. With repeated mechanical transmission, isolates of both viruses appeared to lose the ability to be vectored, and MiLV appeared to lose the ability to cause bigvein symptoms. The recovery of MiLV (Mendocino isolate, from California) from stored O. brassicae resting spores puts the earliest directly demonstrable existence of MiLV at 1990.