In vitro synthesis of biologically active beet necrotic yellow vein virus RNA

Quillet, Laurent; Guilley, H.; Jonard, G.; Richards, K.

doi:10.1016/0042-6822(89)90131-1

Cited by 109 publications

(99 citation statements)

References 22 publications

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“…These mutations were designed to test the requirement for intact CP during the infectivity cycle of PMV. CP deletion mutants of beet necrotic yellow vein virus (Quillet et al, 1989), barley stripe mosaic virus (BSMV; Petty & Jackson, 1990), TCV (Hacker et al, 1992), TMV (Takamatsu et al, 1987;Dawson et al, 1988) and tobacco rattle virus (TRV) RNA2 (Angenent et al, 1989) were still infectious despite the lack of a functional CP. RNA1 of the bipartite tobravirus TRV can replicate in the absence of RNA2 which encodes the CP gene (Harrison & Robinson, 1986;Hamilton & Baulcombe, 1989).…”

Section: Discussionmentioning

confidence: 99%

Infectious RNA transcripts derived from cloned cDNA of papaya mosaic virus: effect of mutations to the capsid and polymerase proteins

Sit

AbouHaidar

1993

Journal of General Virology

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Genomic length cDNAs of papaya mosaic virus (PMV)RNA were generated utilizing reverse transcriptase (RNase H-) for first strand synthesis, Sequenase for second strand synthesis and primers specific for the 5' and 3' termini of the viral genome. These cDNAs were cloned into plasmid pUC18 and infectious RNA transcripts were synthesized in vitro from a bacteriophage T7 RNA polymerase promoter incorporated into the 5' specific primer. The infectivity of transcripts was 16 % that of native PMV RNA. Increasing the poly(A) tail length from A24 to A71 produced a 43 % increase in infectivity. Transcripts synthesized with or without an m7GpppG cap structure were biologically active although uncapped transcripts were much less infectious. The addition of up to 2434 non-viral nucleotides at the 3' end of transcripts decreased but did not abolish infectivity. Insertions of two amino acid residues within the polymerase coding region inactivated viral transcripts. A single amino acid deletion within the capsid protein (CP) produced local lesions of a reduced size as compared to native PMV RNA. Viral particles could not be observed in crude extracts from lesions produced by this deletion mutant suggesting that it exists as a naked RNA species within the host. Mutations to the CP suggest that it is required not only for viral assembly but also for some other unidentified function(s) during the replication cycle.

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Section: Discussionmentioning

confidence: 99%

Infectious RNA transcripts derived from cloned cDNA of papaya mosaic virus: effect of mutations to the capsid and polymerase proteins

Sit

AbouHaidar

1993

Journal of General Virology

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“…However, when BNYVV is mechanically inoculated into experimental hosts such as C. quinoa, RNA1 and -2 are sufficient to induce and maintain infection (Quillet et al, 1989). At 6 days p.i., the inoculated plants displayed chlorotic local lesions of medium size (Fig.…”

Section: P0mentioning

confidence: 99%

“…Clones of RNA1, RNA2 and RNA3-derived replicon (rep0) used in this study have been described previously (Quillet et al, 1989;Jupin et al, 1992;Erhardt et al, 2000). The RNA2DP14 mutant carrying a mutation in the P14 ORF (+2 frameshift mutation) has been described by Hehn et al (1995).…”

Section: Virus Aa* Sequencementioning

confidence: 99%

“…To assess the capacity of P0 BC and P0 BM to cross-complement the RSS activity of BNYVV P14, we used infectious transcripts comprising RNA1+2DP14 of BNYVV strain F2 (Quillet et al, 1989), an artificial strain deficient in P14 synthesis (Hehn et al, 1995). C. quinoa leaves inoculated with this strain developed small necrotic local lesions at 5 days p.i.…”

Section: P0mentioning

confidence: 99%

“…In parallel experiments, total RNA was extracted from inoculated C. quinoa leaves at 7 days p.i. RNAs were detected using specific 32 P-labelled probes corresponding to BNYVV RNA1, -2 and -3 (Quillet et al, 1989).…”

Section: Virus Aa* Sequencementioning

confidence: 99%

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P0 proteins of European beet-infecting poleroviruses display variable RNA silencing suppression activity

Kozlowska-Makulska

Guilley

Szyndel

et al. 2009

Journal of General Virology

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Post-transcriptional gene silencing (PTGS), or RNA silencing, is one of the key mechanisms of antiviral defence used by plants. To counter this defence response, viruses produce suppressor proteins that are able to inhibit the PTGS pathway or to interfere with some of its function. The aim of this study was to evaluate the RNA silencing suppressor (RSS) activity of P0 proteins from selected European isolates of the beet-infecting poleroviruses beet chlorosis virus (BChV) and beet mild yellowing virus (BMYV) using two different experimental systems: (i) agro-infiltration of Nicotiana benthamiana green fluorescent protein-positive plants and (ii) mechanical inoculation of Chenopodium quinoa using a beet necrotic yellow vein virus (BNYVV, genus Benyvirus) RNA3-based replicon. The results demonstrated that P0 of most BMYV isolates exhibited RSS activity, although at various efficiencies among isolates. Conversely, P0 of BChV isolates displayed no RSS activity in either of the two systems under the experimental conditions used. These results are the first reported evidence that P0 proteins of two closely related beet poleroviruses show strain-specific differences in their effects on RNA silencing.

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Two proteins encoded by beet necrotic yellow vein virus RNA 3 influence symptom phenotype on leaves.

et al. 1992

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RNA 3 of the beet necrotic yellow vein virus (BNYVV) quadripartite RNA genome is not essential for virus multiplication on leaves of Tetragonia expansa but has dramatic effects on symptom expression. Virus isolates containing RNA 3 produce bright yellow local lesions while isolates lacking RNA 3 produce much milder symptoms. Using directed mutagenesis of cDNA clones followed by in vitro synthesis of biologically active transcripts, a 25 kDa open reading frame (ORF) of RNA 3 was shown to be responsible for the yellow local lesion phenotype. In addition, two deletion mutants of RNA 3 were found to elicit the appearance of severe necrotic local lesions. Analysis of one of these mutants revealed that necrosis was due to the overexpression of a second short ORF, N, overlapping the 3′‐terminal portion of the 25 kDa ORF. As shown by gene fusion studies, gene N is not detectably expressed from full‐length RNA 3 but is translationally activated by deletion of upstream sequences. Introduction of gene N into the genome of the unrelated DNA virus, cauliflower mosaic virus, elicits a necrotic response instead of the typical mosaic symptoms, demonstrating that gene N can induce necrosis outside of the context of a BNYVV infection.

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In vitro synthesis of biologically active beet necrotic yellow vein virus RNA

Cited by 109 publications

References 22 publications

Infectious RNA transcripts derived from cloned cDNA of papaya mosaic virus: effect of mutations to the capsid and polymerase proteins

Infectious RNA transcripts derived from cloned cDNA of papaya mosaic virus: effect of mutations to the capsid and polymerase proteins

P0 proteins of European beet-infecting poleroviruses display variable RNA silencing suppression activity

Two proteins encoded by beet necrotic yellow vein virus RNA 3 influence symptom phenotype on leaves.

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