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PubMed1991

Interaction of non-enveloped plant viruses and their viral coat proteins with phospholipid vesicles.

The interaction of the non-enveloped plant viruses TMV (rod-shaped) and CCMV (spherical) and of their coat proteins in several well-defined aggregation states, with artificial membranes was investigated to study the early stages of the cellular infection process. Information about the separate steps in the interaction mechanisms was obtained by employing three assays, performed as a function of vesicle size, net membrane charge, pH and ionic strength. The assays allow to discriminate between aggregation of vesicles (turbidity assay) and membrane destabilization (vesicle leakage assay and lipid mixing assay). The aggregation of the vesicles is a result of electrostatic interactions between the viral material and vesicles surface (cross-linking), while the destabilization of the membrane is a result of penetration or bilayer disruption by hydrophobic protein domains. TMV virions and its coat protein, and CCMV virions, due to their net negative charge, predominantly interact with positively charged membranes. The coat protein of CCMV was found to interact with negatively charged membranes, an interaction that can be assigned to its basical N-terminal sequence. Changing the aggregational state of the viral coat proteins yielded most significant interactions in case of TMV coat protein aggregated in the disk form and CCMV coat protein aggregated in empty capsids with oppositely charged membranes. These protein aggregates are found to be the best compromise between efficiency (capacity of the protein to bridge vesicles and destabilize their membranes) and concentration of protein aggregates. The results are discussed with respect to previously proposed biological models of the early stages of plant virus infection.

Capsid↗
PubMed1989

Logistic and Poisson models for infection by multicomponent plant viruses.

A model for the relationship between virus concentration and infectivity of multicomponent plant viruses is based on a combination of logistic and Poisson equations. Two separate equations are derived from the Poisson distribution assuming, (i) that infections occur only when a set of components containing the complete multicomponent genome is established at an infection site, but that any excess of components present does not reduce the probability of infection (no interference postulate); and (ii) that infection can occur only if a set of components containing the full genome reaches an infection site before it can be preempted by an incomplete set (competitive interference postulate). Postulate (i) affects the form of a dilution series without affecting N, the maximum possible number of infections (lesions), and postulate (ii) changes the value of N but not the form of the dilution series. There is a close correlation between the logit slope of a logistic dilution series and the form of the corresponding multiple Poisson dilution series for viruses with 2, 3 or 4 components. Calibrated by Poisson equations, the logit slope may thus suggest whether or not the virus components have invaded independently and infected similar infection sites. The methods of fitting the combined logistic-Poisson model are demonstrated by applying it to data for cowpea chlorotic mottle virus.

Models, Theoretical↗
PubMed1985

Sindbis virus proteins nsP1 and nsP2 contain homology to nonstructural proteins from several RNA plant viruses.

Although the genetic organization of tobacco mosaic virus (TMV) differs considerably from that of the tripartite viruses (alfalfa mosaic virus [AlMV] and brome mosaic virus [BMV]), all of these RNA plant viruses share three domains of homology among their nonstructural proteins. One such domain, common to the AlMV and BMV 2a proteins and the readthrough portion of TMV p183, is also homologous to the readthrough protein nsP4 of Sindbis virus (Haseloff et al., Proc. Natl. Acad. Sci. U.S.A. 81:4358-4362, 1984). Two more domains are conserved among the AlMV and BMV 1a proteins and TMV p126. We show here that these domains have homology with portions of the Sindbis proteins nsP1 and nsP2, respectively. These results strengthen the view that the four viruses share mechanistic similarities in their replication strategies and may be evolutionarily related. These results also suggest that either the AlMV 1a, BMV 1a, and TMV p126 proteins are multifunctional or Sindbis proteins nsP1 and nsP2 function together as subunits in a single complex.

Amino Acid Sequence↗
PubMed2004

Heterologous expression of plant virus genes that suppress post-transcriptional gene silencing results in suppression of RNA interference in Drosophila cells.

BACKGROUND: RNA interference (RNAi) in animals and post-transcriptional gene silencing (PTGS) in plants are related phenomena whose functions include the developmental regulation of gene expression and protection from transposable elements and viruses. Plant viruses respond by expressing suppressor proteins that interfere with the PTGS system. RESULTS: Here we demonstrate that both transient and constitutive expression of the Tobacco etch virus HC-Pro silencing suppressor protein, which inhibits the maintenance of PTGS in plants, prevents dsRNA-induced RNAi of a lacZ gene in cultured Drosophila cells. Northern blot analysis of the RNA present in Drosophila cells showed that HC-Pro prevented degradation of lacZ RNA during RNAi but that there was accumulation of the short (23nt) RNA species associated with RNAi. A mutant HC-Pro that does not suppress PTGS in plants also does not affect RNAi in Drosophila. Similarly, the Cucumber mosaic virus 2b protein, which inhibits the systemic spread of PTGS in plants, does not suppress RNAi in Drosophila cells. In addition, we have used the Drosophila system to demonstrate that the 16K cysteine-rich protein of Tobacco rattle virus, which previously had no known function, is a silencing suppressor protein. CONCLUSION: These results indicate that at least part of the process of RNAi in Drosophila and PTGS in plants is conserved, and that plant virus silencing suppressor proteins may be useful tools to investigate the mechanism of RNAi.

Animals↗
PubMed1995

Detection of cleavage products from an in vivo transcribed cis hairpin ribozyme in turnips using the CaMV plant virus.

In order to examine ribozyme (Rz) activity in vivo, we have adapted a virus to deliver Rz to plants. DNA fragments that code for both active and mutant cis-hairpin Rz were cloned into the double-stranded DNA plant virus, cauliflower mosaic virus (CaMV). These Rz constructs successfully infected Brassica campestris rapa (turnip). The plants that were infected with the active-Rz construct showed, on average, a one-week delay in the appearance of viral symptoms, when compared to the mutant-Rz control. Since CaMV replicates through reverse transcription of a full-length RNA intermediate, Rz cloned into the CaMV DNA should be transcribed within this viral RNA. If these Rz constructs cleave, the amount of intact virus RNA should be reduced, resulting in attenuated viral symptoms. In addition, lysate RNase protection assays showed fragments corresponding to the sizes of both the 5' and 3' cis cleavage products in the active Rz tissue. No cleavage products were observed from plant tissue infected with the mutant Rz. Both the attenuated systemic viral symptoms and the cleavage products from the protection assay strongly support in vivo transcription and cleavage of this hairpin Rz. This is the first report of an in vivo transcribed Rz showing cleaved products by direct RNA analysis (non-PCR) in plants or animals.

Base Sequence↗
PubMed2000

A century of plant virus management in the Salinas valley of California, 'East of Eden'.

The mild climate of the Salinas Valley, CA lends itself well to a diverse agricultural industry. However, the diversity of weeds, crops and insect and fungal vectors also provide favorable conditions for plant virus disease development. This paper considers the incidence and management of several plant viruses that have caused serious epidemics and been significant in the agricultural development of the Salinas Valley during the 20th century. Beet curly top virus (BCTV) almost destroyed the newly established sugarbeet industry soon after its establishment in the 1870s. A combination of resistant varieties, cultural management of beet crops to provide early plant emergence and development, and a highly coordinated beet leafhopper vector scouting and spray programme have achieved adequate control of BCTV. These programmes were first developed by the USDA and still operate. Lettuce mosaic virus was first recognized as causing a serious disease of lettuce crops in the 1930s. The virus is still a threat but it is controlled by a lettuce-free period in December and a seed certification programme that allows only seed lots with less than one infected seed in 30000 to be grown. 'Virus Yellows' is a term used to describe a complex of yellows inducing viruses which affect mainly sugarbeet and lettuce. These viruses include Beet yellows virus and Beet western yellows virus. During the 1950s, the complex caused significant yield losses to susceptible crops in the Salinas Valley. A beet-free period was introduced and is still used for control. The fungus-borne rhizomania disease of sugarbeet caused by Beet necrotic yellow vein virus was first detected in Salinas Valley in 1983. Assumed to have been introduced from Europe, this virus has now become widespread in California wherever beets are grown and crop losses can be as high as 100%. Movement of infested soil and beets accounts for its spread throughout the beet-growing regions of the United States. Control of rhizomania involves several cultural practices, but the use of resistant varieties is the most effective and is necessary where soils are infested. Rhizomania-resistant varieties are now available that perform almost as well as the non-resistant varieties under non-rhizomania conditions. Another soil-borne disease termed lettuce dieback, caused by a tomato bushy stunt-like tombusvirus, has become economically limiting to romaine and leaf lettuce varieties. The virus has no known vector and it seems to be moved through infested soil and water. Heavy rains in the past 4 years have caused flooding of the Salinas River and lettuce fields along the river have been affected severely by dieback. Studies are now in progress to characterize this new virus and identify sources of resistance. Agriculture in the Salinas Valley continues to grow and diversify, driven by demands for 'clean', high quality food by the American public and for export. The major aspects of plant virus control, including crop-free periods, breeding for resistance, elimination of inoculum sources, and vector control will continue to be vital to this expansion. Undoubtedly, the advances in crop production through genetic manipulation and advances in pest management through biological control will eventually become an important part of agricultural improvement.

Animals↗
PubMed2025

Live-cell RNA imaging with the inactivated endonuclease Csy4 enables new insights into plant virus transport through plasmodesmata.

Plant-infecting viruses spread through their hosts by transporting their infectious genomes through intercellular nano-channels called plasmodesmata. This process is mediated by virus-encoded movement proteins. Whilst the sub-cellular localisations of movement proteins have been intensively studied, live-cell RNA imaging systems have so far not been able to detect viral genomes inside the plasmodesmata. Here, we describe a highly sensitive RNA live-cell reporter based on an enzymatically inactive form of the small bacterial endonuclease Csy4, which binds to its cognate stem-loop with picomolar affinity. This system allows imaging of plant viral RNA genomes inside plasmodesmata and shows that potato virus X RNA remains accessible within the channels and is therefore not fully encapsidated during movement. We also combine Csy4-based RNA-imaging with interspecies movement complementation to show that an unrelated movement protein from tobacco mosaic virus can recruit potato virus X replication complexes adjacent to plasmodesmata. Therefore, recruitment of potato virus X replicase is mediated non-specifically, likely by indirect coupling of movement proteins and viral replicase via the viral RNA or co-compartmentalisation, potentially contributing to transport specificity. Lastly, we show that a 'self-tracking' virus can express the Csy4-based reporter during the progress of infection. However, expression of the RNA-binding protein in cis interferes with viral movement by an unidentified mechanism when cognate stem-loops are present in the viral RNA.

Plasmodesmata↗
PubMed1996

Seed transmission of plant viruses: a lesson in biological complexity.

Seed transmission of plant viruses results from the three-way interplay of the genetic components of the virus, the maternal host and its progeny. The challenge is to use genetic and cell biological analyses, integrated with our knowledge of plant reproduction and embryo development, to dissect this complex and rather poorly understood process.

Genes, Plant↗
PubMed2001

Spherical plant viruses: interactions in solution, phase diagrams and crystallization of brome mosaic virus.

Brome mosaic virus (BMV) is a small icosahedral plant virus of mean diameter 268 A. Interactions between BMV particles in solution were studied by means of small-angle X-ray scattering in order to find crystallization conditions. The interactions between biomacromolecules as large as these viruses have not yet been systematically studied by this method. As it is known that usually proteins crystallize in, or close to, attractive regimes, the interactions between BMV particles in solution were studied as a function of pH, type of salt and size and concentration of polyethylene glycol. An unexpected result of these studies is that the precipitates obtained upon addition of PEG alone or PEG combined with salt were in fact made of microcrystals, which were all characterized by the same series of diffraction peaks, with positions close to those of a centered cubic space group. A phase diagram of the virus as a function of PEG concentration was established by means of microbatch experiments. From the precipitation zones, conditions for crystallization were tested from 5 to 40 mg ml(-1) virus with 3-10%(w/v) PEG 8000 or PEG 20,000. Small crystals were obtained in several conditions after a few days and continued growing for several weeks.

Bromovirus↗
PubMedDate not supplied

Genetic recombination between RNA components of a multipartite plant virus.

Genetic recombination of DNA is one of the fundamental mechanisms underlying the evolution of DNA-based organisms and results in their diversity and adaptability. The importance of the role of recombination is far less evident for the RNA-based genomes that occur in most plant viruses and in many animal viruses. RNA recombination has been shown to promote the evolutionary variation of picornaviruses, it is involved in the creation of defective interfering (DI) RNAs of positive- and negative-strand viruses and is implicated in the synthesis of the messenger RNAs of influenza virus and coronavirus. However, RNA recombination has not been found to date in viruses that infect plants. In fact, the lack of DI RNAs and the inability to demonstrate recombination in mixedly infected plants has been regarded as evidence that plants do not support recombination of viral RNAs. Here we provide the first molecular evidence for recombination of plant viral RNA. For brome mosaic virus (BMV), a plus-stranded, tripartite-genome virus of monocots, we show that a deletion in the 3' end region of a single BMV RNA genomic component can be repaired during the development of infection by recombination with the homologous region of either of the two remaining wild-type BMV RNA components. This result clearly shows that plant viruses have available powerful recombinatory mechanisms that previously were thought to exist only in animal hosts, thus they are able to adapt and diversify in a manner comparable to animal viruses. Moreover, our observation suggests an increased versatility of viruses for use as vectors in introducing new genes into plants.

Base Sequence↗
PubMed1996

Getting it together in plant virus movement: cooperative interactions between bipartite geminivirus movement proteins.

To move cell-to-cell and systemically infect the host, plant viruses must cross the barrier posed by the plant cell wall. Plant viruses accomplish this through strategies that alter the architecture of the infected cell, eliminating this barrier through the action of viral-encoded 'movement proteins'. Detailed studies of a number of cytoplasmically replicating viruses suggest that movement proteins interact with components of the cytoskeleton and transport systems of the plant cell to allow passage of progeny into adjacent cells. Recent work on the two movement proteins encoded by the phloem-restricted geminivirus squash leaf curl virus has defined unique aspects of nuclear transport and protein protein interaction in the movement of this nuclear-replicating virus, and suggests that post-translational phosphorylation may be important in the regulation of movement protein function.

Journal Article↗
PubMed2005

Suppressors of RNA silencing encoded by plant viruses and their role in viral infections.

RNA silencing as a robust host defense mechanism against plant viruses is generally countered by virus-encoded silencing suppressors. This strategy is now increasingly recognized to be used by animal viruses as well. We present here an overview of the common features shared by some of the better studied plant viral silencing suppressors. We then briefly describe the characteristics of the few reported animal viral suppressors, notably their extraordinary ability of cross-kingdom suppression. We next discuss the basis for biased protection of viral RNA and subviral parasites by silencing suppressors, the link between movement and silencing suppression, the influence of temperature on the outcome of viral infection and the effect of viral silencing suppressors on the microRNA pathway.

Gene Silencing↗
PubMed1969

Possibilities for inter- and intracellular translocation of some icosahedral plant viruses.

Southern bean mosaic virus (SBMV) and Tomato ringspot virus (TomRV) were compared with regard to possible ways of inter- and intracellular translocation. The pore complexes in the nuclear membranes of nuclei in leaf palisade and mesophyll cells of several plant species commonly used in plant virus research were studied. The pore structure resembled that earlier described. The diameter of the pores was great enough to allow icosahedral plant viruses between 25 and 30 mmicro wide to move through. SBMV occurred in noncrystalline form in nuclei of infected cells. Although this virus forms paracrystalline structures when partially purified, no virus crystals were seen in the cytoplasm of cells containing high concentrations of SBMV. It was established that this virus could move through nuclear pores. TomRV was found in infected leaf tissue in low concentrations. This virus showed a tendency to crystallize even when present in low concentrations. TomRV was observed only in the cytoplasm, not in nuclei. This virus was present in plasmodesmata, indicating the possibility of cell to cell translocation of whole particles through these structures.

Cytopathogenic Effect, Viral↗
PubMed1986

Sequence homology between the coat proteins of DNA and RNA plant viruses.

The coat proteins of the leafhopper transmitted gemini viruses, viz., maize streak virus and wheat dwarf virus that contain a single circle of DNA in their genomes are shown to bear good amino acid sequence homology with the coat protein of an RNA plant virus, the satellite tobacco necrosis virus. It is suggested that for icosahedral assemblage, the coat protein architecture transcends genomic preferences.

Amino Acid Sequence↗
PubMed2005

Oligonucleotide-based microarray: a new improvement in microarray detection of plant viruses.

Microarrays are one of the new emerging methods in plant virology currently being developed by various laboratories. In this study, a new approach is described on the detection of plant viruses using short synthetic single-stranded oligomers (40 nt) instead of PCR products as capture probes. A microchip detecting potato viruses, PVA, PVS, PVM, PVX, PVY and PLRV, in both single and mixed infections was developed and tested. The chip was also designed to distinguish between the main strains of PVY and PVS. Results of initial tests with PVY(NTN) and PVY(O) strains using several different probes for one virus are presented. Possibilities and advantages of the new oligonucleotide-based microarray approach for plant viral diagnosis are discussed.

Oligonucleotide Array Sequence Analysis↗
PubMed2004

A whole genome perspective on the phylogeny of the plant virus family Tombusviridae.

Most current classifications of viruses are based on single gene analysis of capsid protein or polymerase. The comparison of entire genomes is a more balanced approach that should provide a more complete picture of relatedness. We have used a singular value decomposition (SVD)-based analysis to generate phylogenetic trees using whole genome protein sequences from a family of single-stranded RNA plant viruses. Our dataset includes the 26 species of the family Tombusviridae, 25 of which have complete genome sequences cataloged in GenBank. The resulting phylogenetic tree agrees well with current taxonomic classifications, but with significant exceptions. One previously unassigned virus within this family, Maize necrotic streak virus, is definitively placed within the genus Tombusvirus by this analysis. In addition, the analysis defines two distinct subsets within the genus Necrovirus. Future datasets will be expanded to include other icosahedral positive strand RNA plant viruses, and then perhaps all positive strand RNA plant viruses.

Amino Acid Sequence↗