Virus-induced changes in root volatiles attract soil nematode vectors to infected plants.

Plant-derived volatiles mediate interactions among plants, pathogenic viruses, and viral vectors. These volatile-dependent mechanisms have not been previously demonstrated belowground, despite their likely significant role in soil ecology and agricultural pest impacts. We investigated how the plant virus, tobacco rattle virus (TRV), attracts soil nematode vectors to infected plants.

Dynamic biospeckle analysis, a new tool for the fast screening of plant nematicide selectivity.

Background: Plant feeding, free-living nematodes cause extensive damage to plant roots by direct feeding and, in the case of some trichodorid and longidorid species, through the transmission of viruses. Developing more environmentally friendly, target-specific nematicides is currently impeded by slow and laborious methods of toxicity testing. Here, we developed a bioactivity assay based on the dynamics of light 'speckle' generated by living cells and we demonstrate its application by assessing chemicals' toxicity to different nematode trophic groups.

New live screening of plant-nematode interactions in the rhizosphere.

Free living nematodes (FLN) are microscopic worms found in all soils. While many FLN species are beneficial to crops, some species cause significant damage by feeding on roots and vectoring viruses. With the planned legislative removal of traditionally used chemical treatments, identification of new ways to manage FLN populations has become a high priority.

ICTV Virus Taxonomy Profile: Virgaviridae.

The family Virgaviridae is a family of plant viruses with rod-shaped virions, a ssRNA genome with a 3'-terminal tRNA-like structure and a replication protein typical of alpha-like viruses. Differences in the number of genome components, genome organization and the mode of transmission provide the basis for genus demarcation. Tobacco mosaic virus (genus Tobamovirus) was the first virus to be discovered (in 1886); it is present in high concentrations in infected plants, is extremely stable and has been extensively studied.

Molecular and Biochemical Examination of Spraing Disease in Potato Tuber in Response to Tobacco rattle virus Infection.

Field-grown tubers of potato were examined for infection by Tobacco rattle virus (TRV) and consequent production of corky ringspot or spraing symptoms. A microarray study identified genes that are differentially expressed in tuber tissue in response to TRV infection and to spraing production, suggesting that hypersensitive response (HR) pathways are activated in spraing-symptomatic tubers. This was confirmed by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) of a selected group of HR-related genes and by histochemical staining of excised tuber tissue with spraing symptoms.

RNA-dependent RNA polymerase 1 in potato (Solanum tuberosum) and its relationship to other plant RNA-dependent RNA polymerases.

Cellular RNA-dependent RNA polymerases (RDRs) catalyze synthesis of double-stranded RNAs that can serve to initiate or amplify RNA silencing. Arabidopsis thaliana has six RDR genes; RDRs 1, 2 and 6 have roles in anti-viral RNA silencing. RDR6 is constitutively expressed but RDR1 expression is elevated following plant treatment with defensive phytohormones.

The complete nucleotide sequence of pelargonium leaf curl virus.

Investigation of a tombusvirus isolated from tulip plants in Scotland revealed that it was pelargonium leaf curl virus (PLCV) rather than the originally suggested tomato bushy stunt virus. The complete sequence of the PLCV genome was determined for the first time, revealing it to be 4789 nucleotides in size and to have an organization similar to that of the other, previously described tombusviruses. Primers derived from the sequence were used to construct a full-length infectious clone of PLCV that recapitulates the disease symptoms of leaf curling in systemically infected pelargonium plants.

Cassava Ivorian bacilliform virus is a member of the genus Anulavirus.

The complete genomic sequence of Cassava Ivorian bacilliform virus (CIBV) is described. The virus has a genomic organization similar to that of pelargonium zonate spot virus (PZSV), the type member of the genus Anulavirus, but it is most closely related to a second, recently described, anulavirus, Amazon lily mild mottle virus (ALiMMV).

Regulation of RNA-dependent RNA polymerase 1 and isochorismate synthase gene expression in Arabidopsis.

Background: RNA-dependent RNA polymerases (RDRs) function in anti-viral silencing in Arabidopsis thaliana and other plants. Salicylic acid (SA), an important defensive signal, increases RDR1 gene expression, suggesting that RDR1 contributes to SA-induced virus resistance. In Nicotiana attenuata RDR1 also regulates plant-insect interactions and is induced by another important signal, jasmonic acid (JA).

Dynamic localization of two tobamovirus ORF6 proteins involves distinct organellar compartments.

ORF6 is a small gene that overlaps the movement and coat protein genes of subgroup 1a tobamoviruses. The ORF6 protein of tomato mosaic virus (ToMV) strain L (L-ORF6), interacts in vitro with eukaryotic elongation factor 1α, and mutation of the ORF6 gene of tobacco mosaic virus (TMV) strain U1 (U1-ORF6) reduces the pathogenicity in vivo of TMV, whereas expression of this gene from two other viruses, tobacco rattle virus (TRV) and potato virus X (PVX), increases their pathogenicity. In this work, the in vivo properties of the L-ORF6 and U1-ORF6 proteins were compared to identify sequences that direct the proteins to different subcellular locations and also influence virus pathogenicity.

Raspberry leaf blotch virus, a putative new member of the genus Emaravirus, encodes a novel genomic RNA.

A new, segmented, negative-strand RNA virus with morphological and sequence similarities to other viruses in the genus Emaravirus was discovered in raspberry plants exhibiting symptoms of leaf blotch disorder, a disease previously attributed to the eriophyid raspberry leaf and bud mite (Phyllocoptes gracilis). The virus, tentatively named raspberry leaf blotch virus (RLBV), has five RNAs that each potentially encode a single protein on the complementary strand. RNAs 1, 2 and 3 encode, respectively, a putative RNA-dependent RNA polymerase, a glycoprotein precursor and the nucleocapsid.

RNA2 of TRV SYM breaks the rules for tobravirus genome structure.

Currently, all of the RNA2 molecules described for all of the more than thirty sequenced isolates of the three tobraviruses, Tobacco rattle virus (TRV), Pea early-browning virus (PEBV) and Pepper ringspot virus (PepRSV), have the virus coat protein (CP) gene located in the 5' proximal position. However, sequencing of the RNA2 of the SYM isolate of TRV revealed that this isolate has a unique genome structure in which the virus CP gene is located in the central region of RNA2 downstream of three completely novel open reading frames (ORFN1, ORFN2 and ORFN3). An infectious clone of SYM RNA2 was constructed and mutations were introduced separately into each of the novel genes to interrupt their translation.

Tobraviruses--plant pathogens and tools for biotechnology.

The tobraviruses, Tobacco rattle virus (TRV), Pea early-browning virus (PEBV) and Pepper ringspot virus (PepRSV), are positive-strand RNA viruses with rod-shaped virus particles that are transmitted between plants by trichodorid nematodes. As a group, these viruses infect many plant species, with TRV having the widest host range. Recent studies have begun to dissect the interaction of TRV with potato, currently the most commercially important crop disease caused by any of the tobraviruses.

Testing of transmission of tobraviruses by nematodes.

Virus diseases often are spread between plants by vector organisms, some of which live below ground (e.g., fungi and nematodes) and feed on the plant root system.

Genome activation by raspberry bushy dwarf virus coat protein.

Two sets of infectious cDNA clones of raspberry bushy dwarf virus (RBDV) have been constructed, enabling either the synthesis of infectious RNA transcripts or the delivery of infectious binary plasmid DNA by infiltration of Agrobacterium tumefaciens. In whole plants and in protoplasts, inoculation of RBDV RNA1 and RNA2 transcripts led to a low level of infection, which was greatly increased by the addition of RNA3, a subgenomic RNA coding for the RBDV coat protein (CP). Agroinfiltration of RNA1 and RNA2 constructs did not produce a detectable infection but, again, inclusion of a construct encoding the CP led to high levels of infection.

Yeast two-hybrid assay to identify host-virus interactions.

The small size of most plant virus genomes and their very limited coding capacities requires that plant viruses are dependent on proteins expressed by the host plant for all stages of their life cycle. Identification of these host proteins is essential if we are to understand in any meaningful way the interactions that exist between virus and plant. A variety of methods are now available to isolate and study interacting proteins, however, the yeast two-hybrid (Y2H) assay system, which was one of the earliest mass analysis methods to be developed [Nature 340:245-246, 1989] remains one of the most popular and amenable approaches in current use.

Translocation of Tomato bushy stunt virus P19 protein into the nucleus by ALY proteins compromises its silencing suppressor activity.

The P19 protein of Tomato bushy stunt virus is a potent suppressor of RNA silencing and, depending on the host species, is required for short- and long-distance virus movement and symptom production. P19 interacts with plant ALY proteins and relocalizes a subset of these proteins from the nucleus to the cytoplasm. Here we showed that coexpression by agroinfiltration in Nicotiana benthamiana of P19 and the subset of ALY proteins that are not relocalized from the nucleus interfered with the ability of P19 to suppress RNA silencing.

Virus-induced gene silencing as a tool for functional genomics in a legume species.

Virus-induced gene silencing (VIGS) is an attractive reverse-genetics tool for studies of gene function. However, efficient VIGS has only been accomplished in a few plant species. In order to extend the application of VIGS, we examined whether a VIGS vector based on Pea early browning virus (PEBV) would produce recognizable phenotypes in Pisum sativum.

ORF6 of Tobacco mosaic virus is a determinant of viral pathogenicity in Nicotiana benthamiana.

Tobacco mosaic virus (TMV) contains a sixth open reading frame (ORF6) that potentially encodes a 4.8 kDa protein. Elimination of ORF6 from TMV attenuated host responses in Nicotiana benthamiana without alteration in virus accumulation.

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.

Relocalization of nuclear ALY proteins to the cytoplasm by the tomato bushy stunt virus P19 pathogenicity protein.

The P19 protein of tomato bushy stunt virus (TBSV) is a multifunctional pathogenicity determinant involved in suppression of posttranscriptional gene silencing, virus movement, and symptom induction. Here, we report that P19 interacts with the conserved RNA-binding domain of an as yet uncharacterized family of plant ALY proteins that, in animals, are involved in export of RNAs from the nucleus and transcriptional coactivation. We show that the four ALY proteins encoded by the Arabidopsis genome and two ALY proteins from Nicotiana benthamiana are localized to the nucleus.

Molecular determinants of the transmission of plant viruses by nematodes.

SUMMARY Understanding the mechanisms of transmission of plant viruses is an important part of devising effective and sustainable strategies to protect crop plants against plant virus diseases. There are many difficulties associated with the study of virus transmission by nematodes, particularly as these vector organisms live below ground in the soil feeding on plant roots and cannot be maintained in pure culture. Nevertheless, with recent advances in molecular cloning techniques many details of the transmission process have begun to be revealed, especially with regard to the virus proteins that are required for successful transmission.

Analysis of the involvement of an inducible Arabidopsis RNA-dependent RNA polymerase in antiviral defense.

RNA-dependent RNA polymerases (RdRPs) have been implicated in posttranscriptional gene silencing (PTGS) and antiviral defense. An Arabidopsis RdRP (SDE1/SGS2) has been previously shown to be required for transgene-induced PTGS but has no general role in antiviral defense. On the other hand, we have recently shown that transgenic tobacco deficient in an inducible RdRP (NtRdRP1) activity became more susceptible to both Tobacco mosaic virus and Potato virus X.

Functional replacement of the tobacco rattle virus cysteine-rich protein by pathogenicity proteins from unrelated plant viruses.

Mutation of the 16K gene encoded by RNA1 of Tobacco rattle virus (TRV) greatly reduced the levels of viral RNA that accumulated in both infected protoplasts and plants, showing that the 16K cysteine-rich protein (CRP) is required for efficient multiplication of TRV. Overexpression of the 16K protein, either from an additional copy of the gene carried on TRV RNA2 or from a PVX vector, led to an increase in the severity of disease symptoms, suggesting that the protein has a role in the pathogenicity of the virus. Mutation of the 16K gene could be overcome by expression from RNA2 of the Cucumber mosaic virus 2b gene, the Soil-borne wheat mosaic virus 19K gene, or the Barley stripe mosaic virus gammab gene, indicating that the proteins encoded by these diverse genes may have similar functions.

Substitution of a single amino acid in the 2b protein of Pea early-browning virus affects nematode transmission.

The 2b protein of Pea early-browning virus (PEBV) is required for transmission of the virus by nematodes. Comparison of the 2b proteins of highly transmissible (TpA56) and poorly transmissible (SP5) isolates of PEBV identified two amino acid substitutions (G90S and G177R) that might be responsible for the poor transmission of isolate SP5. Hybrid viruses were created in which the TpA56 2b protein carried SP5-specific substitutions at residue 90 or 177, and in which the SP5 2b protein carried TpA56-specific substitutions at these positions.