Engineering of Alfalfa mosaic virus RNA 3 into an expression vector

Sánchez‐Navarro, J. A.; Miglino, R.; Ragozzino, A.; Bol, John F.

doi:10.1007/s007050170125

Cited by 47 publications

(55 citation statements)

References 34 publications

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“…Newly synthesized mRNAs egress into the cytoplasm for sgRNA transcription (f) and translation of other viral products, such as MP and CP, that are engaged in virion maturation (g). The presence of viral MP triggers the formation of tubular structures that mediate virion cell-to-cell transfer via a tubule-guided mechanism (99,109,134). ER, endoplasmic reticulum.…”

Section: Rna Replicationmentioning

confidence: 99%

“…These structures were shown to traverse the cell wall through modified plasmodesmata, and they mediate virion transfer via a tubule-guided mechanism (13,41,99,109,134). Recent studies performed with N. benthamiana plants and protoplasts have shown that host factors were engaged in the regulation of the BMV MP localization to the plasmodesmata (50).…”

Section: Virus Movementmentioning

confidence: 99%

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Insights into the Single-Cell Reproduction Cycle of Members of the Family Bromoviridae : Lessons from the Use of Protoplast Systems

Sztuba-Solińska

Bujarski

2008

J Virol

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2The development of single-cell protoplast systems is certainly one of the milestones in the history of plant virology, allowing for the analysis of viral molecular processes at the cellular level. The use of plant cell-based systems in the study of the Bromoviridae family of multipartite single-stranded plant RNA viruses facilitated the discovery and dissection of viral processes engaged in the single-cell reproduction cycle: replication, transcription, protein synthesis, movement, virion assembly, and RNA recombination. This review summarizes the application of protoplast systems to the analysis of consecutive steps of the bromovirus life cycle, emphasizing their temporal and spatial patterns during virus multiplication.Our knowledge of viral infection at the individual cell level determines our understanding of the infection at the entire plant body level. The idea of applying single-cell systems in virology was introduced with the first attempts to transfect Escherichia coli cells with a T4 anti-E. coli bacteriophage (27). E. C. Cocking was the first (1960) to enzymatically isolate plant cells (19), which were then infected with Tobacco mosaic tobamovirus (TMV) (20). The ensuing pioneer work showed that the infection was synchronous and that the uptake of viral particles/RNAs by protoplasts was efficient enough to support virus replication (5, 118). The first viral infection of protoplasts involved the use of poly-L-ornithine (5), but the trials that followed exploited the fusogenic polymers (24), liposomes (28), or electroporation (78,129). By 1980, these procedures had been successfully employed to transfect protoplasts from more than six species of plants, including various members of the Bromoviridae family, e.g., Brome mosaic bromovirus (BMV; 80), Cucumber mosaic cucumovirus (CMV; 69), and Cowpea chlorotic mottle virus (CCMV; 130). The first attempts were based on the transfection of plant cells with whole-virus particles (104). Later on, viral RNA and its chemical modifications were used as the inoculum (62). Further advances in nucleic acid technology, especially the accessibility of infectious transcripts, have broadened the application of protoplast systems to the study of Bromoviridae.The Bromoviridae constitute one of the most important families of plant RNA viruses. They are distributed worldwide, they infect an extensive range of hosts, and some of them (e.g., CMV and Broad bean mottle bromovirus) are responsible for major crop epidemics (111, 63). The family consists of five genera named after their most representative members: Alfamovirus, Bromovirus, Cucumovirus, Ilarvirus, and Oleavirus. All these viruses possess tripartite, single-stranded, positive-sense RNA genomes (Fig. 1). RNA1 and RNA2 encode the RNAdependent RNA polymerase (RdRp) proteins 1a and 2a, respectively. The dicistronic RNA3 encodes the movement protein (MP) and coat protein (CP). The latter is translated from subgenomic RNA 4 (sgRNA4). Many of the family members, such as CMV, BMV, and Alfalfa mosaic alfamovirus (AMV), repres...

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Section: Rna Replicationmentioning

confidence: 99%

Section: Virus Movementmentioning

confidence: 99%

Insights into the Single-Cell Reproduction Cycle of Members of the Family Bromoviridae : Lessons from the Use of Protoplast Systems

Sztuba-Solińska

Bujarski

2008

J Virol

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“…However, an additional number of other promising virus vectors have recently been described. Relatively new or improved additions to the repertoire of viral gene vectors are represented by Tobacco rattle virus (TRV) (13), Wheat streak mosaic virus (WSMV) (24), Tomato bushy stunt virus (TBSV) (2,9,20,25,26), Plum pox virus (PPV) (27), Zucchini yellow mosaic virus (28), Cucumber mosaic virus (29), White clover mosaic virus (18), Clover yellow vein virus (30), Alfalfa mosaic virus (ALMV) (3,19), and Cowpea mosaic virus (CPMV) (4,. 8, 10, 31).…”

Section: Construction Methodologymentioning

confidence: 99%

“…In the complementation approach, a foreign gene is used to replace an essential virus gene. The displaced gene is complemented by a tunctionally analogous gene that is expressed from a co-inoculated viral or subviral component, or from a trans gene (18,19).Most virus vectors are based on viruses that initially were selected as subjects of study because they caused economically impoliant disease problems. In many cases the struggle to combat these viruses continues.…”

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confidence: 99%

Plant Virus Gene Vectors: Biotechnology Applications in Agriculture and Medicine

Mirkov

Scholthof

2002

Genetic Engineering

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INTRODUCTIONPlant virus vectors have been developed as a result of 20 years of advances in molecular biology, which produced infectious plasmid-based clones of RNA and DNA plant viruses and viroids that could be routinely amplified in E. coli. This work built on a century of progress in virology, much of which was based on understanding the biology of Tobacco mosaic virus (TMV) (1). With the availability of the rice and Arabidopis draft genome sequences, expressed sequence tags (ESTs) and microarrays, crystal structures of many plant viruses, and public interest in non-transgenic strategies for crop improvement, the technology of plant virus gene vectors holds much promise for the coming decade (2-4). The intent of this review is to define virus-vector strategies, describe uses for improving crop health, and point out possible beneficial overlaps with veterinary and human medicine.*To whom correspondence should be addressed.(1) Texas Agricultural Experiment Station, Weslaco, TX 78596. DEFINING THE SYSTEMSMethods to regenerate transgenic plants are widely used in plant biology and biotechnology. This involves the use of either Agrobacterium tumefaciens or biolistics (e.g., gene guns) to deliver genetically customized gene expression cassettes into the nucleus of a plant cell. In this case, the DNA becomes integrated into the chromosome and with the use of selectable markers and tissue culture procedures, the cells containing the foreign insert serve as the progenitors for regeneration of whole transgenic plants. This process usually takes at least several months, and mostly longer, to obtain plants with a stable genetic trait.Plant viruses cause harmful diseases that can result in significant reductions in crop yield and economic losses to growers. However, from a biotechnology point of view, viruses collectively can also be regarded as 'molecular tools' (2). One intriguing applicable feature of plant viruses is their capacity to serve as gene vectors. With rare exception (5, 6), plant viruses do not integrate into the plant chromosome but instead transiently express their genes upon infection (7). Because plant viruses replicate to a very high copy number in infected cells, substantial levels of gene expression can be achieved. By standard molecular biology techniques, plant viruses can be supplied with foreign genes and within a few days after inoculation, plant virus gene vectors can provide high levels of transient foreign gene expression (2,4,(8)(9)(10)(11)(12).Many proteins used for biotechnology applications are not biologically active unless they undergo specific post-translational modifications, such as glycosylation or phosphorylation, in eukaryotic cells. Plants are attractive substrates for the synthesis and purification of proteins because they can carry out these modifications and, through the ages of agricultural development, it has become fairly straightforward to grow crop plants that yield a large, uniform biomass. In many cases it is desirable to have the foreign gene expressed in transgenic p...

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“…N. tabacum wild-type, P1, P2, or P12 plants were inoculated with a mixture of capped transcripts corresponding to AMV RNA1 and RNA2, a modified RNA3 that expresses the green fluorescent protein (GFP) (25), and a few micrograms of purified AMV coat protein as described in reference 26. The fluorescence derived from chimeric AMV RNA3 encoding GFP was monitored with a Leica stereoscopic microscope.…”

Section: Dose-response Experimentsmentioning

confidence: 99%

Effects of the Number of Genome Segments on Primary and Systemic Infections with a Multipartite Plant RNA Virus

Sánchez‐Navarro

Zwart

Elena

2013

J Virol

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bMultipartite plant viruses were discovered because of discrepancies between the observed dose response and predictions of the independent-action hypothesis (IAH) model. Theory suggests that the number of genome segments predicts the shape of the dose-response curve, but a rigorous test of this hypothesis has not been reported. Here, Alfalfa mosaic virus (AMV), a tripartite Alfamovirus, and transgenic Nicotiana tabacum plants expressing no (wild type), one (P2), or two (P12) viral genome segments were used to test whether the number of genome segments necessary for infection predicts the dose response. The dose-response curve of wild-type plants was steep and congruent with the predicted kinetics of a multipartite virus, confirming previous results. Moreover, for P12 plants, the data support the IAH model, showing that the expression of virus genome segments by the host plant can modulate the infection kinetics of a tripartite virus to those of a monopartite virus. However, the different types of virus particles occurred at different frequencies, with a ratio of 116:45:1 (RNA1 to RNA2 to RNA3), which will affect infection kinetics and required analysis with a more comprehensive infection model. This analysis showed that each type of virus particle has a different probability of invading the host plant, at both the primary-and systemic-infection levels. While the number of genome segments affects the dose response, taking into consideration differences in the infection kinetics of the three types of AMV particles results in a better understanding of the infection process.

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Engineering of Alfalfa mosaic virus RNA 3 into an expression vector

Cited by 47 publications

References 34 publications

Insights into the Single-Cell Reproduction Cycle of Members of the Family Bromoviridae : Lessons from the Use of Protoplast Systems

Insights into the Single-Cell Reproduction Cycle of Members of the Family Bromoviridae : Lessons from the Use of Protoplast Systems

Plant Virus Gene Vectors: Biotechnology Applications in Agriculture and Medicine

Effects of the Number of Genome Segments on Primary and Systemic Infections with a Multipartite Plant RNA Virus

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