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Rockmelon is a popular tropical fruit with high nutritional value. Cucumber mosaic virus (CMV), an aphid-transmitted virus, causes severe damage to rockmelon production. Exogenous application of double-stranded RNA (dsRNA) targeting viral sequences has shown promising results in controlling viral infection but has not been reported for CMV in rockmelon. In the current study, the protective effect of exogenous dsRNAs targeting the CMV coat protein (CP) and movement protein (MP) was tested in rockmelon. The effectiveness of dsRNA-mediated protection was measured by disease severity index (DSI) and compound enzyme-linked immunosorbent assay. The individual dsRNA CP and MP treatments each showed protection by reducing the DSI and virus titre, whilst a combination of dsRNA of CP and MP treatment showed much lower DSI (4.31-fold lower) and virus titre (4.91-fold lower) compared to CMV-inoculated plants without dsRNA treatment. Chlorophyll content, relative water content, plant height and number of leaves were not significantly different between virus challenged dsRNA-treated and mock-inoculated plants. Based on the DSI, two applications containing 9000 ng of dsRNA CP and MP in combination showed maximum CMV protection. Taken together, these results indicate that exogenous treatment containing a combination of dsRNA of CP and MP can control CMV infection in rockmelon. [ABSTRACT FROM AUTHOR]

Plant viral movement proteins (MPs) serve a critical role in facilitating intercellular transport, pivotal for propagating infections from afflicted cells to their neighboring counterparts. Leveraging MPs within transient expression systems holds the potential to augment the expanse of protein expression zones. In this investigation, we present evidence showcasing the synergistic effects of fusing a transient expression system vector with the movement protein of tomato mosaic virus (ToMV MP), thus broadening the scope of protein expression within plant stem tissues. Through agroinfiltration, we achieved transient expression of green fluorescent protein (GFP) in tomato stem tissues, which was subsequently confirmed via immunoblotting with an anti-GFP antibody. Quantitative analysis unveiled that the co-expression of ToMV MP alongside GFP led to an over twofold increase in GFP production within stem tissues distal from the primary infection site. This research underscores the efficacy of combining ToMV MP with transient expression systems as a potent strategy for achieving transient expression of target genes within tomato plants. [ABSTRACT FROM AUTHOR]

Biological and molecular properties of five Prune dwarf virus cherry isolates were studied. Double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) proved PDV infection in the respective source trees and absence of six other viruses infecting the cherry. Two of infected source trees were symptomless, and the other three trees showed deformed and elongated leaves with chlorotic lines and spots. The reaction of greenhouse-grown seedlings of 22 herbaceous test plant species was investigated. The woody indicators Prunus tomentosa and Prunus cerasifera were chip-budded. Cucumis sativus cv. Amelia, C. sativus cv. Levina, C. sativus cv. Tessa and Prunus tomentosa appeared as the most suitable test plants for bioassay of studied PDV isolates. The nucleotide sequences corresponding to the full-length coat protein (CP) of 657 bp were obtained. The identity among studied isolates in CP was 95.5–100% and 97.4–100% at nucleotide and amino acid levels, respectively. The movement protein (MP) sequences were 753 nucleotides long, translating 251 amino acids and shared 94.1–98.4% nucleotide and 96.8–99.2% amino acid identity. Constructed trees with the nucleotide sequences of both MP and CP genes of studied isolates and referent PDV cherry variants showed the formation of two phylogroups in the two genomic regions. All studied isolates separated into two clusters in group I. No correlation between the phylogenetic grouping and the biological properties, as well the geographical origin, was observed. [ABSTRACT FROM AUTHOR]

Background: Strawberry crinkle virus (SCV) is a member of the genus Cytorhabdovirus, family Rhabdovirida, and order Mononegavirales. SCV affects the production of various strawberry cultivars. In this study we investigated the genetic diversity of SCV in strawberry fields based on P3 (movement protein) gene. Methods and results: The samples were collected from strawberry fields in the Kurdistan Province, Iran. P3 gene from 20 SCV isolates, representing 18 nucleic acid haplotypes, is composed of 729 nucleotides, encoding a protein with 243 amino acids. SCV-P3 sequences shared 98.77%–99.86% nucleotide and 97.5%–100% amino acid sequence identity. Phylogenetic analyses of the new P3 sequences with two previously published SCV-P3 sequences from the Czech Republic showed that there are two major phylogroups (I and II) and three minor phylogroups in the body of the phylogeny, I-1, I-2, II-1. Comparisons of P3 gene sequences revealed a mutational bias, with more differences being transitions than transversions. The ratio of non-synonymous/synonymous nucleotide changes was < 1, indicating that SCV-P3 gene is under predominantly negative selection. Conclusions: Phylogenetic and sequence identity analyses showed that SCV isolates from Iran are closely related and have not diverged more than 2% based on P3 gene despite geographical separation and strawberry cultivar. This is the first report of the genetic diversity of SCV worldwide. [ABSTRACT FROM AUTHOR]

The tobacco N gene encodes the N resistance factor that recognizes a tobacco mosaic virus elicitor called p50 to induce a cell-death-associated hypersensitive reaction, which confines virus infections to local areas. Coexpression of N and p50 cDNAs in Nicotiana benthamiana induces resistance against virus infection without visible cell death. In this study, we examined how this N-mediated resistance in the heterologous plant inhibits virus infection. After inoculation with GFP-encoding tomato mosaic virus (ToMV), plants coexpressing N and p50 had fewer and smaller infection sites as the levels of coexpressed proteins of N and p50 increased. This antiviral response was associated with production of reactive oxygen species but not ion leakage or expression of PR1a (encoding a salicylic acid signaling marker). In addition, it was likely that GFP-tagged movement protein (MP) of ToMV seemed to have failed to localize at plasmodesmata during virus infection in leaves coexpressing N and p50. These findings indicate that the cell death-independent, N-mediated antiviral response in N. benthamiana may involve interference with the intracellular transport function of MP. [ABSTRACT FROM AUTHOR]

RNA interference (RNAi) is a sequence-specific, gene silencing mechanism, induced by double-stranded RNA (dsRNA). It is a defense mechanism that protects eukaryotic cells from invasive nucleic acids such as viruses and transposons. In this study, we used a non-transgenic strategy in order to activate the antiviral RNAi mechanism against Sesbania mosaic virus (SeMV) in its natural host Sesbania grandiflora. DsRNA molecules from SeMV coat protein (CP) and movement protein (MP) (sobemovirus MP is considered a silencing suppressor) genes were produced by a two-step PCR approach followed by in vitro transcription and exogenously applied on sesbania plants along with SeMV. DsRNA for CP and MP conferred 53% and 64% resistance against SeMV, respectively based on the disease incidence data. The effect of dsRNA molecules against SeMV infection on sesbania plants was confirmed by direct antigen coating ELISA (DAC-ELISA). The strategy employed demonstrated the applicability of the RNA-based vaccination method, for the first time, to control sobemoviruses in a simple, highly specific and environmentally safe way. [ABSTRACT FROM AUTHOR]

The nonstructural protein pc6 encoded by rice grassy stunt virus (RGSV) plays a significant role in viral cell-to-cell movement, presumably by transport through plasmodesmata (PD). We confirmed the association of pc6 with PD, and also elucidated the mechanisms of protein targeting to PD. Several inhibitor treatments showed conclusively that pc6 is targeted to PD via the ER-to-Golgi secretory system and actin filaments. In addition, VIII-1 myosin was also found to be involved in pc6 PD targeting. Deletion mutants demonstrated that C-terminal amino acid residues 209-229 (transmembrane domain 2; TM2) are essential for pc6 to move through PD. [ABSTRACT FROM AUTHOR]

Apple mosaic virus (ApMV) infects over 65 plants species of Rosaceae family including hazelnut. The virus causes chlorotic or yellow patterns, rings and mosaic symptoms on the leaves and losses in yield of nuts. The occurrence and genetic diversity of ApMV were evaluated in 14 plantations of hazelnut in Poland. A total of 13 of 125 hazelnut samples tested positive for ApMV using ELISA and reverse transcriptase-polymerase chain reaction. Several primer pairs were used for RT-PCR to amplify the coat protein (CP) and movement protein (MP) genes as well as complete RNA-3 of the ApMV. Genetic diversity was observed in full-length sequences of RNA-3 of the ApMV isolates from hazelnut. The similarity of nucleotide sequence of the CP and MP genes of the isolates was 90.3-99.9, and 87-99.7 %, respectively. Phylogenetic analysis showed that ApMV isolates from hazelnut grouped separately from the virus strains found in other plant species. [ABSTRACT FROM AUTHOR]

Maize rayado fino virus (MRFV) possesses an open reading frame (ORF43) predicted to encode a 43 kDa protein (p43) that has been postulated to be a viral movement protein. Using a clone of MRFV (pMRFV-US) from which infectious RNA can be produced, point mutations were introduced to either prevent initiation from three potential AUG initiation codons near the 5′-end of ORF43 or prematurely terminate translation of ORF43. Inoculation of maize seed via vascular puncture inoculation (VPI) resulted in plants exhibiting symptoms typical of MRFV infection for all mutants tested. Furthermore, corn leafhoppers ( Dalbulus maidis) transmitted the virus mutants to healthy plants at a frequency similar to that for wild-type MRFV-US. Viral RNA recovered from plants infected with mutants both prior to and after leafhopper transmission retained mutations blocking ORF43 expression. The results indicate that ORF43 of MRFV is dispensable for both systemic infection of maize and transmission by leafhoppers. [ABSTRACT FROM AUTHOR]

Grapevine fanleaf virus (GFLV), a nematode-transmitted virus belonging to the genus Nepovirus, is the major pathogen responsible for fanleaf degeneration, one of the most widespread and damaging viral diseases of grapevine. The virus is characterized by a genome constituted by two single positive sense RNAs (RNA1 and RNA2), coding for two polyproteins. Here we investigated the genetic variability of the movement and coat protein genes (2B and 2C) of Italian GFLV isolates by sequencing analyses. The presence of high molecular heterogeneity between our isolates suggests that GFLV comprises a family of sequence variants. [ABSTRACT FROM AUTHOR]

During plant infection by tobamoviruses such as Tomato mosaic virus (ToMV), viral replication complexes containing a viral 126-kDa replicase component (126K) are generated, temporarily in association with a viral movement protein (MP) that is indispensable for viral intercellular movement. The tobamovirus 126K protein has been shown to play an important role not only in replication but also in intercellular transport of viral RNAs. Transient expression of the 126K protein has recently been reported to result in formation of cytoplasmic bodies with shapes similar to those of the replication complexes. In this study, to investigate the roles of ToMV MP in the formation, localization, and dynamics of cytoplasmic bodies of the 126K protein, we carried out confocal microscopy studies of fluorescently tagged 126K and MP that were transiently and simultaneously expressed in Nicotiana benthamiana. Without MP expression, 126K was evenly distributed in the cytoplasm, with occasional formation of small 126K bodies. These small 126K bodies moved in the cytoplasmic stream and along the endoplasmic reticulum network. In contrast, co-expression of MP markedly promoted the formation of larger MP-associated 126K bodies, which were generally static but moved occasionally in the cytoplasmic stream. In addition, we found that small, mobile 126K bodies were captured at the terminal part of MP-derived filamentous structures, leading to the formation of 126K-MP complexes and their translocation to another cellular site. [ABSTRACT FROM AUTHOR]

Plant viruses use plasmodesmata (PD) to spread infection between cells and systemically. Dependent on viral species, movement through PD can occur in virion or non-virion form, and requires different mechanisms for targeting and modification of the pore. These mechanisms are supported by viral movement proteins and by other virus-encoded factors that interact among themselves and with plant cellular components to facilitate virus movement in a coordinated and regulated fashion. [ABSTRACT FROM AUTHOR]

Plasmodesmata (PD) are plasma membrane-lined cytoplasmic channels that cross the cell wall and establish symplasmic continuity between neighboring cells in plants. Recently, a wide range of cellular RNAs (including mRNAs and small RNAs (sRNAs)) have been reported to move from cell to cell through PD trafficking pathways. sRNAs are key molecules that function in transcriptional and post-transcriptional RNA silencing, which is a gene expression regulatory mechanism that is conserved among eukaryotes and is important for protection against invading nucleic acids (such as viruses and transposons) and for developmental and physiological regulation. One of the most intriguing aspects of RNA silencing is that it can function either cell autonomously or non-cell autonomously in post-transcriptional RNA silencing pathways. Although the mechanisms underlying cell-to-cell trafficking of RNA and RNA silencing signals are not fully understood, the movement of specific RNAs seems to play a critical role in cell-to-cell and long-distance regulation of gene expression, thereby coordinating growth and developmental processes, gene silencing, and stress responses. In this review, we summarize the current knowledge regarding cell-to-cell trafficking of RNA molecules (including small RNAs), and we discuss potential molecular mechanisms of cell-to-cell trafficking that are mediated by complex networks. [ABSTRACT FROM AUTHOR]

In addition to small molecules such as sugars and amino acids, phloem sap contains macromolecules, including mRNA and proteins. It is generally assumed that all molecules in the phloem sap are on the move from source to sink, but recent evidence suggests that the macromolecules' direction of movement can be controlled by endogenous plant mechanisms. To test the hypothesis that the phloem-sap protein profile is affected by local metabolic activities, we analyzed the phloem-sap proteome in young and mature tissues of melon plants. We also examined the effect of cucumber mosaic virus (CMV) infection and expression of CMV movement protein in transgenic melon plants on the phloem protein profile. Sap collected from cut sections of young stems or petioles contained specific proteins that were absent from sap collected from mature stems or petioles. Most of these proteins were involved in defense response and protection from oxidative stress, suggesting that they play a role in maintaining safe activity of the sieve tubes in young tissues. Phloem sap collected from CMV-infected plants and transgenic plants expressing the CMV movement protein contained only a few additional proteins with molecular masses of 18 to 75 kDa. Here again, most of the additional proteins were associated with stress responses. Our study indicated that the proteome of phloem sap is dynamic and under developmental control. Entry and exit of proteins from the sieve tube can be regulated at the tissue level. Moreover, the plant can maintain regulation of protein trafficking from companion cells to sieve elements under viral infection or other perturbations in plasmodesmal function. [ABSTRACT FROM AUTHOR]

The KELP protein of Arabidopsis thaliana (AtKELP), which binds to the Tomato mosaic virus (ToMV) movement protein in vitro, can interfere with cell-to-cell movement of the virus under transient overexpression conditions. In this study, we constructed expression plasmids encoding a fluorescent protein fused to AtKELP or to its homolog from Brassica, Nicotiana, Solanum, or Oryza species. When expressed transiently, all the AtKELP homologs inhibited the cell-to-cell movement of ToMV with different efficiencies. The results of domain swapping between two selected AtKELP homologs suggest the importance of combination between the N- and C-terminal half regions for high inhibitory function. [ABSTRACT FROM AUTHOR]

A 25-kD movement protein (25K protein) encoded by the first gene of the potexvirus Potato virus X triple gene block of transport genes is essential for the viral movement in infected plants. The 25K protein belongs to superfamily 1 of NTPase/helicases and exhibits in vitro RNA helicase, Mg2+-dependent NTPase, and RNA-binding activities. In the present work, the ability of 25K protein for homologous interactions was studied using the yeast two-hybrid system, protein chemical cross-linking in the presence of glutaraldehyde, far-Western blotting, and ultracentrifugation in sucrose density gradients. The 25K protein was shown to form homodimers and homooligomers. Sites of homologous protein-protein interactions were found in both the N- and C-terminal portions of the protein. [ABSTRACT FROM AUTHOR]

In DAS-ELISAs of 86 grapevine samples from northwestern Iran, Grapevine fanleaf virus (GFLV) was detected in 18 samples. RT-PCR with two primer pairs (M2/M4 or M0/M4) corresponding to GFLV movement protein (MP) amplified the expected 854- and/or 1,489-bp fragment(s), respectively, from all ELISA-positive samples. Four smaller and three larger PCR products were cloned and sequenced, which revealed that the MP region of the isolates was 1,044 nucleotides (nt) long, corresponding to the GFLV MP. There were 83–86% nucleotide and 93–94% amino acid identities deduced between the MPs of the sequenced isolates. Nucleotide sequence identities of 81–87 and 75–79% were found between the MP regions of these isolates and that of previously published GFLV and Arabis mosaic virus (ArMV) strains/isolates, respectively. On a consensus parsimony tree based on the nucleotide sequences, isolates La208 and X300 remained distinct from previously reported GFLVs. This is the first molecular characterization of GFLV MP isolates from Iran. [ABSTRACT FROM AUTHOR]

An approach that enables the increase of the quantity of a specific amino acid in crop plants is reported.Oleosingene fromArabidopsis thalianaor 30Kmovement proteingene ofTobacco mosaic virus(TMV; genusTobamovirus) were cloned under the control of napin or hybrid promoters, and in fusion to synthetic poly-histidine (poly-His) sequences for transformation into spring turnip rape (Brassica rapasubsp.oleifera; synonym toB. campestris). The most stable expression cassettes for the poly-His production prior to the plant transformation were selected by analyzing the protein expression inin vitrotranslation and in transient plant expression systems using GFP as marker. Expression of the poly-His-constructs in transgenicBrassica rapaplants was analyzed using dot and western blotting and PCR. The constructs were stably expressed in the third generation of the transgenic plant lines. Histidine content was measured from the seeds of the transgenic plants, and some plant lines had more than 20% increase in histidine content compared to wild type. The methodology may be widely applicable to increase the content of any amino acid in crop plants including those encoded by rare codons. [ABSTRACT FROM AUTHOR]

Tobacco mosaic virus (TMV) and Tomato mosaic virus (ToMV) are two closely related viruses in the genus Tobamovirus, but they induce obviously different sizes of necrotic lesions in tobacco plants containing the N gene. Comparison of the symptoms produced by TMV, ToMV and a chimaeric virus (T/OMP), in which the TMV movement protein (MP) gene was replaced by the ToMV MP gene, showed T/OMP caused necrotic lesions that were similar in size to those of ToMV in tobacco plants containing the N gene. The coat protein and MP of the three viruses accumulated in planta with similar levels, and the replication level of TMV and T/OMP in protoplasts also had no difference. Comparison of the activities of defense-related enzymes (PAL, POD and PPO) induced by the three viruses also showed that the variability of enzyme activity induced by T/OMP was similar to that induced by TMV, but different from that induced by ToMV. The results indicate that the size difference of necrotic lesions induced by TMV and ToMV in tobacco plants containing the N gene results from the functional difference of their MP genes. [ABSTRACT FROM AUTHOR]

The nucleotide sequences of the RNA 3 of fifteen isolates of Prunus necrotic ringspot virus (PNRSV) varying in the symptomatology they cause in six different Prunus spp. were determined. Analysis of the molecular variability has allowed, in addition to study the phylogenetic relationships among them, to evaluate the minimal requirements for the synthesis of the subgenomic RNA in Ilarvirus genus and their comparison to other members of the Bromoviridae family. Computer assisted comparisons led recently to Jaspars (Virus Genes 17, 233–242, 1998) to propose that a hairpin structure in viral minus strand RNA is required for subgenomic promoter activity of viruses from at least two, and possibly all five, genera in the family of Bromoviridae. For PNRSV and Apple mosaic virus two stable hairpins were proposed whereas for the rest of Ilarviruses and the other four genera of the Bromoviridae family only one stable hairpin was predicted. Comparative analysis of this region among the fifteen PNRSV isolates characterized in this study revealed that two of them showed a 12-nt deletion that led to the disappearance of the most proximal hairpin to the initiation site. Interestingly, the only hairpin found in these two isolates is very similar in primary and secondary structure to the one previously shown in Brome mosaic virus to be required for the synthesis of the subgenomic RNA. In this hairpin, the molecular diversity was concentrated mostly at the loop whereas compensatory mutations were observed at the base of the stem strongly suggesting its functional relevance. The evolutionary implications of these observations are discussed. [ABSTRACT FROM AUTHOR]

The 3a protein of Cucumber mosaic virus is essential for the cell-to-cell movement of the viral RNA through plasmodesmata. We have introduced an epitope peptide before the stop codon of the 3a protein and cloned the tagged ORF into a binary vector for Agrobacterium-mediated transformation. The established transgenic tobacco lines produced the 3a protein, which was specifically detected with anti-3a and anti-epitope antisera. Metabolic labeling and subsequent immunoprecipitation revealed that [32P]-orthophosphate was incorporated into the 3a protein. The phosphoamino acid analysis indicated that the 3a protein contained phosphoserine but not phosphothreonine or phosphotyrosine. This is the first demonstration of the 3a protein phosphorylation in planta. [ABSTRACT FROM AUTHOR]

Nicotiana benthamiana plants were transformed with the movement protein (MP) gene of tobacco mosaic virus (TMV), using Agrobacterium-mediated transformation. Plants regenerated from the transformed cells accumulated 30-kDa MP and complemented the activity of TMV MP when infected with chimeric TMVs containing defective MR These transgenic plants displayed stunting, pale-green leaves, and starch accumulations, indicating that TMV MP altered the carbon partitioning for leaves involved in TMV cell-to-cell movement. [ABSTRACT FROM AUTHOR]

Interactions of plant cells with pathogens or other biotic or abiotic environmental factors can give rise to systemic defense responses that rely upon the cell-to-cell and systemic transport of specific signals. A novel type of systemic signaling was revealed by recent evidence indicating the existence of RNA species that travel cell to cell and through the vasculature. The most compelling evidence for intercellular and systemic transport of RNA in plants is provided by viroids and viruses that apparently use the endogenous transport machinery to spread infection. The cell to cell movement of plant viruses occurs through small pores in the cell wall known as plasmodesmata and depends on virus-encoded 'movement proteins'. This review summarizes current knowledge of Tobacco mosaic virus infection with emphasis on the mechanism by which this virus targets its RNA genome from sites of replication to plasmodesmata to achieve intercellular spread. [ABSTRACT FROM AUTHOR]

Transgenic tobacco ( Nicotiana tabacum L. cv. Xanthi) plants expressing wild-type or mutant forms of the 30-kDa movement protein of tobacco mosaic virus (TMV-MP) were employed to study the effects of the TMV-MP on carbon metabolism in source leaves. Fully expanded source leaves of transgenic plants expressing the TMV-MP were found to retain more newly fixed C compared with control plants. Analysis of C-export from young leaves of TMV-MP plants, where the MP is yet to influence plasmodesmal size exclusion limit, indicated a similar pattern, in that daytime C export was slower in TMV-MP plants as compared to equivalent-aged leaves on control plants. Pulse-chase experiments were used to monitor radioactivity present in the different carbohydrate fractions, at specified intervals following CO labeling. These studies established that the-TMV-MP can cause a significant adjustment in short-term -C-photosynthate storage and export. That these effects of the TMV-MP on carbon metabolism and phloem function were not attributable to the effect of this protein on plasmodesmal size exclusion limits, per se, was established using transgenic tobacco plants expressing temperature-sensitive and C-terminal deletion mutant forms of the TMV-MP. Collectively, these studies establish the pleiotropic nature of the TMV-MP in transgenic tobacco, and the results are discussed in terms of potential sites of interaction between the TMV-MP and endogenous processes involved in regulating carbon metabolism and export. [ABSTRACT FROM AUTHOR]

Transgenic tobacco ( Nicotiana tabacum L.) plants expressing the 30-kDa movement protein of tobacco mosaic virus (TMV-MP) were employed to investigate the influence of a localized change in mesophyll-bundle sheath plasmodesmal size exclusion limit on photosynthetic performance and on carbon metabolism and allocation. Under conditions of saturating irradiance, tobacco plants expressing the TMV-MP were found to have higher photosynthetic CO-response curves compared with vector control plants. However, this difference was significant only in the presence of elevated CO levels. Photosynthetic measurements made in the green-house, under endogenous growth conditions, revealed that there was little difference between TMV-MP-expressing and control tobacco plants. However, analysis of carbon metabolites within source leaves where a TMV-MP-induced increase in plasmodesmal size exclusion limit had recently taken place established that the levels of sucrose, glucose, fructose and starch were considerably elevated above those present in equivalent control leaves. Although expression of the TMV-MP did not alter total plant biomass, it reduced carbon allocation to the lower region of the stem and roots. This difference in biomass distribution was clearly evident in the lower root-to-shoot ratios for the TMV-MP transgenic plants. Microinjection (dye-coupling) studies established that the TMV-MP-associated reduction in photosynthate delivery (allocation) to the roots was not due to a direct effect on root cortical plasmodesmata. Rather, this change appeared to result from an alteration in phloem transport from young source leaves in which the TMV-MP had yet to exert its influence over plasmodesmal size exclusion limits. These results are discussed in terms of the rate-limiting steps involved in sucrose movement into the phloem. [ABSTRACT FROM AUTHOR]

Hydrolytic activities of leaf extracts from normal and transgenic plants, with (+ MP) and without (-MP) the movement protein of tobacco mosaic virus, were examined. In the + MP transgenic plants, as compared with non-transgenic and - MP plants, higher hydrolytic activities were found on the following substrates: bis-(nitrophenyl)-phosphate (BPNPP, phosphodiesterase), p-nitrophenyl-(phenyl)-phosphate (PNPPP, nucleotidephosphodiesterase) and thymidine-3′-monophosphate p-nitrophenyl ester (TMPP; 3′nucleotide phosphodiesterase.) The + MP plant lines, as compared with other transgenic plants, exhibited higher nucleotide-phosphodiesterase activity in the soluble as well as in the membrane fraction. Substrate concentration kinetic studies revealed the presence of a nucleotide-phospho-diesterase with a high substrate affinity in the +MP extracts in addition to the enzyme with a relatively low substrate affinity present also in the - MP transgenic plants. This 'high affinity' enzyme could be removed from the soluble fraction by precipitation with anti-MP serum, indicating its possible association with the movement protein. [ABSTRACT FROM AUTHOR]

Transgenic Nicotiana tabacum plants expressing RNA sequences of the tomato spotted wilt virus NSM gene, which encodes the putative viral movement protein, were found to be highly resistant to infection with the virus. Expression of untranslatable as well as anti-sense RNA of the NSM gene resulted in resistance levels as high as those in plants expressing translatable RNA sequences. For all three types of transgenic plants resistance levels of up to 100% were reached in the S2 progeny. These results indicate that the resistance mediated by the NSM gene is accomplished by expression of transcripts rather than protein in transgenic plants, similar to previously observed N gene-mediated resistance. Protoplast inoculations revealed that resistant plants expressing NSM are, in contrast to N transgenic resistant plants, not resistant at the cellular level. This suggests the RNA-mediated resistance mechanism against TSWV targets viral mRNAs rather than the viral genome. [ABSTRACT FROM AUTHOR]

The partial nucleotide sequence of the 3′-terminal region of the Korean isolate of odontoglossum ringspot tobamovirus (ORSV-Cy) from cool-growing Cymbidium was determined. The sequence contained a full length open reading frame (ORF) coding for the viral cell-to-cell movement protein (MP). The ORF was located upstream of the coat protein gene and 105 nucleotides longer than that of tobacco mosaic virus (TMV). The ORF predicts a polypeptide chain of 303 amino acids with a molecular weight of 33573. The ORF contained a similar region of conserved sequence motif of tobamoviruses and putative assembly origin of the viral RNA was located at about 1,100 nucleotides away from the 3′ end. The predicted amino acid sequence for the MP gene of ORSV-Cy is more closely related to pepper mild mottle virus (PMMV), TMV-vulgare and TMV-Rakkyo than to tobacco mild green mosaic virus (TMGMV), TMV-L, cowpea strain of TMV (SHMV), and cucumber green mottle mosaic virus (CGMMV). [ABSTRACT FROM AUTHOR]

Cell-to-cell communication in plants occurs through plasmodesmata, cytoplasmic channels that traverse the cell wall between neighboring cells. Plasmodesmata are also exploited by many viruses as an avenue for spread of viral progeny. In the case of tobacco mosaic virus (TMV), a virally-encoded movement protein (MP) enables the virus to move through plasmodesmata during infection. We have used thin section electron microscopy and immunocytochemistry to examine the structure of plasmodesmata in transgenic tobacco plants expressing the TMV MP. We observed a change in structure of the plasmodesmata as the leaves age, both in control and MP expressing [MP(+)] plants. In addition, the plasmodesmata of older cells of MP(+) plants accumulate a fibrous material in the central cavity. The presence of the fibers is correlated with the ability to label plasmodesmata with anti-MP antibodies. The developmental stage of leaf tissue at which this material is observed is the stage at which an increase in the size exclusion limit of the plasmodesmata can be measured in MP(+) plants. Using cell fractionation and aqueous phase partitioning studies, we identified the plasma membrane and cell wall as the compartments with which the MP stably associates. The nature of the interaction between the MP and the plasma membrane was studied using sodium carbonate and Triton X-100 washes. The MP behaves as an integral membrane protein. Identifying the mechanism by which the MP associates with plasma membrane and plasmodesmata will lead to a better understanding of how the MP alters the function of the plasmodesmata. [ABSTRACT FROM AUTHOR]