The 5'-terminal region of a tombusvirus genome determines the origin of multivesicular bodies

Burgyán, József; Rubino, Luisa; Russo, Marcello

doi:10.1099/0022-1317-77-8-1967

Cited by 85 publications

(67 citation statements)

References 23 publications

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“…Vesicular bodies of various sizes and shapes are cytopathological structures associated with infections of many different types of plant viruses, and in some cases these have been shown to contain viral dsRNAs and\or RNA-dependent RNA polymerases (Burgyan et al, 1996 ;Martelli et al, 1984 ;De Graaff & Jaspars, 1994 ;Lesemann, 1988). These observations and the fact that LIYV RNA 1 is alone competent for replication and primarily encodes proteins predicted to be involved in replication (Klaassen et al, 1995), further support the idea that the vesicles induced by LIYV are associated with LIYV RNA replication.…”

supporting

confidence: 65%

Specific inclusion bodies are associated with replication of lettuce infectious yellows virus RNAs in Nicotiana benthamiana protoplasts.

Medina

Tian

Wierzchos

et al. 1998

Journal of General Virology

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Nicotiana benthamiana mesophyll protoplasts, either mock-inoculated or inoculated using in vitro transcripts derived from lettuce infectious yellows virus (LIYV) RNA 1-and/or RNA 2-cloned cDNAs were analysed by transmission electron microscopy (TEM) and, in some cases, also by immunogold labelling. TEM revealed the main cytopathological effects of LIYV infections in N. benthamiana protoplasts infected with RNAs 1 and 2 : (a) typical closterovirus-induced (beet yellows virus-type) accumulations of vesiculated cytoplasmic membranes as inclusion bodies, sometimes with associated virions ; (b) scattered aggregations of virions within the cytoplasm ; and (c) electron-dense plasmalemma deposits. These were not seen in mock-inoculated protoplasts. Protoplasts inoculated only with LIYV RNA 1 contained vesiculated cytoplasmic inclusion bodies, but not virions or plasmalemma deposits. Thus, infection by only LIYV RNA 1 is sufficient to induce characteristic closterovirus vesiculated cytoplasmic inclusion bodies. However, both LIYV RNAs 1 and 2 are needed for production of virions and plasmalemma deposits.Lettuce infectious yellows virus (LIYV) is the type member of the genus Crinivirus of the family Closteroviridae. The LIYV genome is bipartite, divided among two ssRNAs of 8118 and 7193 nucleotides (RNAs 1 and 2 respectively), giving a total of 15 311 nucleotides (Klaassen et al., 1995). On LIYV RNA 2 is the ' hallmark closterovirus gene array ' including five genes encoding : a small hydrophobic protein of as yet unknown function (ORF 1) ; an HSP70 homologue (ORF 2) ; a protein of unknown function (p59, ORF 3) ; the capsid protein (CP,

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supporting

confidence: 65%

Specific inclusion bodies are associated with replication of lettuce infectious yellows virus RNAs in Nicotiana benthamiana protoplasts.

Medina

Tian

Wierzchos

et al. 1998

Journal of General Virology

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“…Preparation of protoplasts from fully expanded leaves, transfection and RNA analysis were done as previously described (Dalmay et al, 1993a;Rubino et al, 2001). Protoplasts were transfected with full-length CIRV genomic RNA (Burgyan et al, 1996) or deletion mutants DB/H (see above) or DB/N (obtained by restriction of the CIRV full-length clone with BamHI and NcoI at nt 3870, at the beginning of ORF4), together with DI (Burgyan et al, 1992) or sat (Rubino et al, 1990) RNA in vitro transcripts.…”

Section: Methodsmentioning

confidence: 99%

“…To do this, two deletion mutants of the full-length infectious clone of CIRV (Burgyan et al, 1996) were constructed (Fig. 5a).…”

Section: The Replication Of Cymrsv Sat Rna With Defective Helper Genomesmentioning

confidence: 99%

“…Preparation of protoplasts from fully expanded leaves, transfection and RNA analysis were done as previously described (Dalmay et al, 1993a; Rubino et al, 2001). Protoplasts were transfected with full-length CIRV genomic RNA (Burgyan et al, 1996) tail. When the same transformants were grown in the GAL1-repressive medium (containing 0?1 % glucose), these bands were no longer present (Fig.…”

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confidence: 99%

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Expression of tombusvirus open reading frames 1 and 2 is sufficient for the replication of defective interfering, but not satellite, RNA

Rubino¹,

Pantaleo²,

Navarro³

et al. 2004

Journal of General Virology

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Yeast cells co-expressing the replication proteins p36 and p95 of Carnation Italian ringspot virus (CIRV) support the RNA-dependent replication of several defective interfering (DI) RNAs derived from either the genome of CIRV or the related Cymbidium ringspot virus (CymRSV), but not the replication of a satellite RNA (sat RNA) originally associated with CymRSV. DI, but not sat RNA, was replicated in yeast cells co-expressing both DI and sat RNA. Using transgenic Nicotiana benthamiana plants constitutively expressing CymRSV replicase proteins (p33 and p92), or transiently expressing either these proteins or CIRV p36 and p95, it was shown that expression of replicase proteins alone was also not sufficient for the replication of sat RNA in plant cells. However, it was also shown that replicating CIRV genomic RNA deletion mutants encoding only replicase proteins could sustain replication of sat RNA in plant cells. These results suggest that sat RNA has a replication strategy differing from that of genomic and DI RNAs, for it requires the presence of a cis-replicating genome acting as a trans-replication enhancer. INTRODUCTIONTombusviruses are small isometric plant viruses belonging to the genus Tombusvirus, family Tombusviridae. Their genome is a monopartite, single-stranded, positive-sense RNA of 4800 nt containing five ORFs. ORF1 and -2 encode the replicase proteins of 33 or 36 kDa (p33 or p36), depending on the species, and the corresponding readthrough products, p92 and p95. ORF3 encodes the coat protein (CP), and two nested ORFs (ORF4 and -5) encode two proteins involved in virus movement and pathogenesis Silhavy et al., 2002). Tombusvirus infections are often associated with small parasitic RNAs, which can be either defective interfering (DI) or satellite (sat) RNAs. DI RNAs are shortened forms of genomic RNA deprived of all viral genes required for replication, encapsidation and movement of the viral genome. They are generated by errors of the viral replicase, which bypasses local intramolecular base-paired regions of genomic RNA, thus leading to the synthesis of deletion mutants. All described tombusvirus DI RNA molecules contain segments of viral RNA, derived from the 59 leader sequence, the replicase and movement protein genes and the 39 noncoding sequence. Tombusviruses can support the replication of homologous or heterologous DI RNA in plant (Havelda et al., 1998) or yeast cells (Pantaleo et al., 2003(Pantaleo et al., , 2004Panavas & Nagy, 2003). Tombusvirus DI RNAs have been widely used for the analysis of cis-acting factors directing the replication of viral RNA both in vivo Chang et al., 1995;Ray & White, 1999White & Morris, 1994;Wu & White, 1998;Wu et al., 2001) and in vitro (Nagy & Pogany, 2000; Panavas et al., 2002a, b;Panavas & Nagy, 2003).The molecular biology of tombusvirus sat RNAs has been much less studied. So far, the best characterized sat RNA is the 621 nt sat RNA of Cymbidium ringspot virus (CymRSV) whose cis-acting sequence requirements essential for replication and the significance and ...

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“…The full-length infectious cDNA clone of CIRV has been described previously (Burgyan et al 1996). All mutants were constructed by standard recombinant DNA cloning techniques and PCR-based oligonucleotide-mediated mutagenesis (Sambrook et al 1989).…”

Section: Plasmid Constructionmentioning

confidence: 99%

Tombusvirus recruitment of host translational machinery via the 3′ UTR

Nicholson¹,

Wu²,

Chevtchenko³

et al. 2010

RNA

136

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RNA viruses recruit the host translational machinery by different mechanisms that depend partly on the structure of their genomes. In this regard, the plus-strand RNA genomes of several different pathogenic plant viruses do not contain traditional translation-stimulating elements, i.e., a 59-cap structure and a 39-poly(A) tail, and instead rely on a 39-cap-independent translational enhancer (39CITE) located in their 39 untranslated regions (UTRs) for efficient synthesis of viral proteins. We investigated the structure and function of the I-shaped class of 39CITE in tombusviruses-also present in aureusviruses and carmoviruses-using biochemical and molecular approaches and we determined that it adopts a complex higher-order RNA structure that facilitates translation by binding simultaneously to both eukaryotic initiation factor (eIF) 4F and the 59 UTR of the viral genome. The specificity of 39CITE binding to eIF4F is mediated, at least in part, through a direct interaction with its eIF4E subunit, whereas its association with the viral 59 UTR relies on complementary RNA-RNA base-pairing. We show for the first time that this tripartite 59 UTR/39CITE/eIF4F complex forms in vitro in a translationally relevant environment and is required for recruitment of ribosomes to the 59 end of the viral RNA genome by a mechanism that shares some fundamental features with cap-dependent translation. Notably, our results demonstrate that the 39CITE facilitates the initiation step of translation and validate a molecular model that has been proposed to explain how several different classes of 39CITE function. Moreover, the virus-host interplay defined in this study provides insights into natural host resistance mechanisms that have been linked to 39CITE activity.

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The 5'-terminal region of a tombusvirus genome determines the origin of multivesicular bodies

Cited by 85 publications

References 23 publications

Specific inclusion bodies are associated with replication of lettuce infectious yellows virus RNAs in Nicotiana benthamiana protoplasts.

Specific inclusion bodies are associated with replication of lettuce infectious yellows virus RNAs in Nicotiana benthamiana protoplasts.

Expression of tombusvirus open reading frames 1 and 2 is sufficient for the replication of defective interfering, but not satellite, RNA

Tombusvirus recruitment of host translational machinery via the 3′ UTR

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