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At least 19 recordsLinked to original sources
PubMed2001

The multifunctional capsid proteins of plant RNA viruses.

This article summarizes studies of viral coat (capsid) proteins (CPs) of RNA plant viruses. In addition, we discuss and seek to interpret the knowledge accumulated to data. CPs are named for their primary function; to encapsidate viral genomic nucleic acids. However, encapsidation is only one feature of an extremely diverse array of structural, functional, and ecological roles played during viral infection and spread. Herein, we consider the evolution of viral CPs and their multitude of interactions with factors encoded by the virus, host plant, or viral vector (biological transmission agent) that influence the infection and epidemiological facets of plant disease. In addition, applications of today's understanding of CPs in the protection of crops from viral infection and use in the manufacture of valuable compounds are considered.

Biotechnology↗
PubMed2006

Double-stranded RNA binding may be a general plant RNA viral strategy to suppress RNA silencing.

In plants, RNA silencing (RNA interference) is an efficient antiviral system, and therefore successful virus infection requires suppression of silencing. Although many viral silencing suppressors have been identified, the molecular basis of silencing suppression is poorly understood. It is proposed that various suppressors inhibit RNA silencing by targeting different steps. However, as double-stranded RNAs (dsRNAs) play key roles in silencing, it was speculated that dsRNA binding might be a general silencing suppression strategy. Indeed, it was shown that the related aureusvirus P14 and tombusvirus P19 suppressors are dsRNA-binding proteins. Interestingly, P14 is a size-independent dsRNA-binding protein, while P19 binds only 21-nucleotide ds-sRNAs (small dsRNAs having 2-nucleotide 3' overhangs), the specificity determinant of the silencing system. Much evidence supports the idea that P19 inhibits silencing by sequestering silencing-generated viral ds-sRNAs. In this study we wanted to test the hypothesis that dsRNA binding is a general silencing suppression strategy. Here we show that many plant viral silencing suppressors bind dsRNAs. Beet yellows virus Peanut P21, clump virus P15, Barley stripe mosaic virus gammaB, and Tobacco etch virus HC-Pro, like P19, bind ds-sRNAs size-selectively, while Turnip crinkle virus CP is a size-independent dsRNA-binding protein, which binds long dsRNAs as well as ds-sRNAs. We propose that size-selective ds-sRNA-binding suppressors inhibit silencing by sequestering viral ds-sRNAs, whereas size-independent dsRNA-binding suppressors inactivate silencing by sequestering long dsRNA precursors of viral sRNAs and/or by binding ds-sRNAs. The findings that many unrelated silencing suppressors bind dsRNA suggest that dsRNA binding is a general silencing suppression strategy which has evolved independently many times.

Plant Viruses↗
PubMed2000

Improved RNA extraction and one-tube RT-PCR assay for simultaneous detection of control plant RNA plus several viruses in plant extracts.

A procedure was developed for simultaneous detection of plant RNA viruses and of plant RNA, as a control. RT-PCR amplification with primers designed for the detection of the plant mRNAs encoding malate dehydrogenase (MDH) and the large subunit of ribulose bisphosphate carboxylase oxygenase (RubiscoL) was used for the development of a plant extraction procedure that consistently yields extracts that can be amplified. The control amplification was used successfully on extracts from cane, leaf and/or bud tissues from grapevine, apple, raspberry, strawberry, peach, apricot, plum and wheat. Multiplex RT-PCR conditions were established for the simultaneous detection in grapevine extracts of either arabis mosaic virus, rupestris stem pitting associated virus and malate dehydrogenase mRNA, or grapevine virus A, grapevine virus B, grapevine leafroll associated virus-3, and RubiscoL mRNA.

Malate Dehydrogenase↗
PubMed1992

Cap structure of U3 small nucleolar RNA in animal and plant cells is different. gamma-Monomethyl phosphate cap structure in plant RNA.

U3 small nucleolar RNA (snoRNA) is an abundant small RNA involved in the processing of pre-ribosomal RNA of eukaryotic cells. U3 snoRNA has been previously characterized from several sources, including human, rat, mouse, frog, fruit fly, dinoflagellates, slime mold, and yeast; in all these organisms, U3 snoRNA contains trimethylguanosine cap structure. In all instances where investigated, the trimethylguanosine-capped snRNAs including U3 snoRNA, are synthesized by RNA polymerase II. However, in higher plants, the U3 snoRNA is synthesized by RNA polymerase III and contains a cap structure different from trimethylguanosine (Kiss, T., and Solymosy, F. (1990) Nucleic Acids Res. 18, 1941-1949; Marshallsay, C., Kiss, T., and Filipowicz, W. (1990) Nucleic Acids Res. 18, 3451-3458; Kiss, T., Marshallsay, C., and Filipowicz, W. (1991) Cell 65, 517-526). In this study, we present evidence that cowpea and, most likely, tomato plant U3 snoRNA contains a methyl-pppA cap structure. These data show that the same U3 snoRNA contains different cap structures in different species and suggest that the kind of cap structure that an uridylic acid-rich small nuclear RNA contains is dependent on the RNA polymerase responsible for its synthesis. In vitro synthesized plant U3 snoRNA, with pppA or pppG as its 5' end, was converted to methyl-pppA/G cap structure in vitro when incubated with extracts prepared from wheat germ or HeLa cells. These data show that the capping machinery is conserved in organisms as evolutionarily distant as plants and mammals. Nucleotides 1-45 of tomato U3 snoRNA, which are capable of forming a stem-loop structure, are sufficient to direct the methyl cap formation in vitro.

Animals↗
PubMed1994

Plant proteins that bind to the 3'-terminal sequences of the negative-strand RNA of three diverse positive-strand RNA plant viruses.

The replication of positive-strand RNA plant viruses, which involves both virus-encoded and plant-encoded proteins, takes place in two stages: synthesis of a negative-strand RNA using the genomic positive-strand RNA as a template and synthesis of progeny positive-strand RNA using the negative-strand RNA as a template. Using gel mobility shift and photochemical crosslinking assays, we have identified three proteins of M(r) 32K, 50K and 100K in extracts of tobacco and spinach leaves that bind to the 3'-terminal sequences of the negative-strand RNA of three diverse positive-strand RNA plant viruses. The 32K protein was purified to near homogeneity by chromatography on columns of Macro-prep high Q, heparin-sepharose, single-stranded DNA cellulose and poly(U)-sepharose. No binding of any of the three proteins to the 3'-termini of the positive-strand RNA or the 5'-termini of the positive-strand RNA or negative-strand RNA of any of the three viruses, or the 3'-termini of the mRNAs of two chloroplast genes, psbA or petD, could be detected. We propose that 3'-terminal negative-strand RNA binding proteins, which may be widespread in the plant kingdom, could be utilised by at least three different positive-strand RNA plant viruses for the initiation of positive-strand RNA synthesis.

Base Sequence↗
PubMed2006

Mutant clouds and occupation of sequence space in plant RNA viruses.

In nature, RNA viruses of plants often must adapt to ever-changing environments in the form of frequent host switches. This would favor a highly diverse population for transmission. However, most viruses that have been studied have been viruses of monocultural crops. In crop viruses, the mutation frequency of individual viral quasispecies varies greatly, both in experiment evolution studies and in populations of viruses within single field plants. There is some correlation between host range and mutation frequency in experimental evolution studies, but few viruses have been examined at the individual quasispecies level. Many questions about the nature of plant RNA virus populations and factors that affect the effective population sizes, such as genetic bottlenecks and postive and negative selection, have only begun to be studied. Many more analyses are required before generalized patterns can be determined.

Adaptation, Physiological↗
PubMed2005

Computational analysis of plant RNA Pol-II promoters.

Plant promoters have not yet been thoroughly analyzed in terms of their structural and sequence dependent properties like curvature, periodicity and information content and our present study is an attempt in that direction. Results were compared with E. coli and yeast data to get some insight into the promoter organization. Promoters having the TATA box (TATA(+)) and those lacking the same (TATA(-)) were also analyzed separately. It was found that plant promoters have marked differences for all these properties when compared to E. coli and yeast. Bias for A+T was observed in promoters of all the three groups. Compared to E. coli and yeast, plant promoters showed intermediate values for A+T content as well as curvature. Analysis showed that curvature of core promoters is more pronounced than non-promoters. Information theoretic analysis of plant promoters reveal high information content at certain consensus regions such as -30 (TATA box) and +1 transcription start site (TSS); and have moderate values at other positions as well. This factor was taken into account while developing weight matrices. For certain threshold values, these weight matrices could pick up all true positives, and reduce false positives to a great extent in a test set. A new multi-parameterized prediction strategy has been proposed that uses a combination of sequence composition, curvature and position weight matrices for identification of plant promoters. This strategy was tested and validated with experimentally known promoter sequences. Our study is novel in using in silico approaches to study the sequence dependent properties of plant RNA Pol-II promoters and their prediction, and important as there is no dedicated promoter search tool for plants.

Base Sequence↗
PubMed1988

A procedure for the small-scale isolation of plant RNA suitable for RNA blot analysis.

A small-scale method for the isolation of total RNA from plant tissue is described. The method provides RNA of suitable quantity and quality from 0.2 g fresh tissue for the detection of mRNA species by RNA blot analysis. The entire procedure is adapted to 1.5-ml microfuge tubes and takes less than 5 h. This method is well suited for the isolation of RNA from large numbers of samples or from samples of limited quantity.

Blotting, Northern↗
PubMedDate not supplied

Multiple alignment and hierarchical clustering of conserved amino acid sequences in the replication-associated proteins of plant RNA viruses.

We have used multiple alignment computer programs to align and hierarchically cluster the conserved amino acid "signature" sequences found in the replication-associated proteins of all plant RNA viruses sequenced so far. These regions, called "polymerase", "nucleotide-binding" and "N-terminal" are well conserved even between viruses which are only distantly related, and are thus very well suited for this type of analysis. Our results show that the clusterings obtained using these very short amino acid sequences are very robust to computing parameters and are surprisingly well matched with the taxonomic grouping of RNA plant viruses. The possibility of using this system as a new taxonomic criterion is discussed.

Amino Acid Sequence↗
PubMed1989

Evolutionary relationship between luteoviruses and other RNA plant viruses based on sequence motifs in their putative RNA polymerases and nucleic acid helicases.

Comparative studies of sequence motifs in the RNA polymerases and nucleic acid helicases of positive-sense RNA plant viruses have provided a new scheme for the classification of these pathogens. We propose a new luteovirus supergroup which should be added to the already described Sindbisvirus-like and picornavirus-like supergroups. Sequence motifs of nucleic acid helicases and RNA polymerases which previously were considered to be specific for each of the two supergroups now occur together within this new supergroup. We propose that this new viral supergroup provides an evolutionary link between the other two supergroups.

Amino Acid Sequence↗
PubMed2005

A phylogenetic survey of recombination frequency in plant RNA viruses.

The severe economic consequences of emerging plant viruses highlights the importance of studies of plant virus evolution. One question of particular relevance is the extent to which the genomes of plant viruses are shaped by recombination. To this end we conducted a phylogenetic survey of recombination frequency in a wide range of positive-sense RNA plant viruses, utilizing 975 capsid gene sequences and 157 complete genome sequences. In total, 12 of the 36 RNA virus species analyzed showed evidence for recombination, comprising 17% of the capsid gene sequence alignments and 44% of the genome sequence alignments. Given the conservative nature of our analysis, we propose that recombination is a relatively common process in some plant RNA viruses, most notably the potyviruses.

Computational Biology↗
PubMed2006

Genome packaging by spherical plant RNA viruses.

The majority of positive-strand RNA viruses of plants replicate and selectively encapsidate their progeny genomes into stable virions in cytoplasmic compartments of the cell where the opportunity to copackage cellular RNA also exists. Remarkably, highly purified infectious virions contain almost exclusively viral RNA, suggesting that mechanisms exist to regulate preferential packaging of viral genomes. The general principle that governs RNA packaging is an interaction between the structural CP and a specific RNA signal. Mechanisms that enhance selective packaging of viral genomes and formation of infectious virions may involve factors other than CP and nucleic acid sequences. The possible involvement of replicase proteins is an example. Our knowledge concerning genome packaging among spherical plant RNA viruses is still maturing. The main focus of this review is to discuss factors that have limited progress and to evaluate recent technical breakthroughs likely to help unravel the mechanism of RNA packaging among viruses of agronomic importance. A key breakthrough is the development of in vivo systems and comparisons with results obtained in vitro.

Genome, Viral↗
PubMed1997

Formation of plant RNA virus replication complexes on membranes: role of an endoplasmic reticulum-targeted viral protein.

The mechanisms that direct positive-stranded RNA virus replication complexes to plant and animal cellular membranes are poorly understood. We describe a specific interaction between a replication protein of an RNA plant virus and membranes in vitro and in live cells. The tobacco etch virus (TEV) 6 kDa protein associated with membranes as an integral protein via a central 19 amino acid hydrophobic domain. In the presence or absence of other viral proteins, fluorescent fusion proteins containing the 6 kDa protein associated with large vesicular compartments derived from the endoplasmic reticulum (ER). Infection by TEV was associated with a collapse of the ER network into a series of discrete aggregated structures. Viral RNA replication complexes from infected cells were also associated with ER-like membranes. Targeting of TEV RNA replication complexes to membranous sites of replication is proposed to involve post-translational interactions between the 6 kDa protein and the ER.

Amino Acid Sequence↗
PubMed2026

Hybridization capture increases on-target nanopore sequencing of plant RNA tobamovirus- derived cDNA libraries.

High-throughput sequencing (HTS) can support plant virus surveillance, but host nucleic acids often reduce on-target read recovery. We evaluated a targeted hybridization-capture workflow in which barcoded double-stranded cDNA (ds-cDNA) libraries generated from plant RNA extracts spiked with lyophilized tobamovirus-positive controls were enriched before Oxford Nanopore sequencing. Biotinylated probes targeted conserved regions of cucumber green mottle mosaic virus (CGMMV), species Tobamovirus viridimaculae; pepper mild mottle virus (PMMoV), species Tobamovirus capsici; and tobacco mosaic virus (TMV), species Tobamovirus tabaci. Across four pairs per virus, relative target-read abundance increased after capture from 0.76 ± 0.33% to 37.62 ± 15.72% for CGMMV, 8.16 ± 3.86% to 24.68 ± 12.34% for PMMoV, and 15.62 ± 10.40% to 36.83 ± 30.33% for TMV. Exact two-sided Wilcoxon signed-rank tests yielded P = 0.125 for each virus; with four nonzero differences in a common direction, this was the minimum attainable two-sided P value. Genome-coverage breadth was maintained. Retrospective duplex qPCR supported an increased virus-to-18S ratio for CGMMV, showed a variable PMMoV response, and showed a decreased virus-to-18S ratio for TMV because the 18S signal shifted earlier by as much as or more than the TMV signal. The findings provide proof-of-concept evidence for target-dependent library enrichment but do not establish analytical sensitivity, diagnostic performance, or field validity. Validation with naturally infected, low-titer, and mixed-infection samples and comparison with simpler targeted workflows are required.

biosecurity↗
PubMed1991

Phylogeny of capsid proteins of small icosahedral RNA plant viruses.

Statistically significant alignment was generated between the amino acid sequences of the (putative) shell (S) domains of the capsid proteins of small RNA plant viruses with icosahedral capsids in the tombusvirus, carmovirus, dianthovirus, sobemovirus and luteovirus groups. Inspection of the alignment showed good correspondence between the experimentally defined beta-strands and alpha-helices of the capsid proteins of tomato bushy stunt, southern bean mosaic and turnip crinkle viruses, allowing prediction of the secondary structure elements in proteins with unresolved tertiary structure. It is concluded that this set of viral capsid proteins forms a tight evolutionary cluster. Comparison of the alignment of the proteins of this family with the sequences of other capsid proteins of icosahedral RNA viruses revealed more distant similarities to the satellites of tobacco necrosis, panicum mosaic, tobacco mosaic and maize white line mosaic viruses, as well as to nepo- and comoviruses. The tentative phylogenetic tree derived from the capsid protein alignment separated into three main lineages: (I) carmo-, tombus- and dianthoviruses, (II) southern bean mosaic, tobacco necrosis and maize chlorotic mottle viruses, and (III) luteoviruses. Comparison of this tree topology with the tentative evolutionary schemes for the respective virus RNA-dependent RNA polymerases suggested that gene shuffling is the universal trend in the evolution of small RNA plant virus genomes.

Amino Acid Sequence↗
PubMed2004

Selection pressures in the capsid genes of plant RNA viruses reflect mode of transmission.

To determine the selection pressures faced by RNA viruses of plants, patterns of nonsynonymous (dN) and synonymous (dS) substitution in the capsid genes of 36 viruses with differing modes of transmission were analysed. This analysis provided strong evidence that the capsid proteins of vector-borne plant viruses are subject to greater purifying selection on amino acid change than those viruses transmitted by other routes and that virus-vector interactions impose greater selective constraints than those between virus and plant host. This could be explained by specific interactions between capsid proteins and cellular receptors in the insect vectors that are necessary for successful transmission. However, contrary to initial expectations based on phylogenetic relatedness, vector-borne plant viruses are subject to weaker selective constraints than vector-borne animal viruses. The results suggest that the greater complexity involved in the transmission of circulative animal viruses compared with non-circulative plant viruses results in more intense purifying selection.

Animals↗