Molecular analysis of the pothos latent virus genome.

Rubino, Luisa; Russo, Marcello

doi:10.1099/0022-1317-78-6-1219

Cited by 33 publications

(29 citation statements)

References 23 publications

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“…Similarly, L5 lengths were variable and ranged from 3 to 12 nt, with an apparent bias for pyrimidines. The SL5 comparison was also extended to the genus Aureusvirus (family Tombusviridae), whose members are most closely related to tombusviruses (Miller et al 1997;Rubino and Russo 1997). Again, conservation of SL5 and its general features was also observed (Fig.…”

Section: Rna Secondary Structure Of the 3 Half Of The Tbsv 5 Utrmentioning

confidence: 79%

A second functional RNA domain in the 5′ UTR of the Tomato bushy stunt virus genome: Intra- and interdomain interactions mediate viral RNA replication

Ray

Wu²,

White³

2003

RNA

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The 5 untranslated regions (UTRs) of (+)-strand RNA viruses play a variety of roles in the reproductive cycles of these infectious agents. Tomato bushy stunt virus (TBSV) belongs to this class of RNA virus and is the prototype member of the genus Tombusvirus. Previous studies have demonstrated that a T-shaped domain (TSD) forms in the 5 half of the TBSV 5 UTR and that it plays a central role in viral RNA replication. Here we have extended our structure-function analysis to the 3 half of the 5 UTR. Investigation of this region in the context of a model viral replicon (i.e., a TBSV-derived defective interfering [DI] RNA) revealed that this segment contains numerous functionally relevant structural features. In vitro solution structure probing along with comparative and computer-aided RNA secondary structure analyses predicted the presence of a simple stem loop (SL5) followed by a more complex downstream domain (DSD). Both structures were found to be essential for efficient DI RNA accumulation when tested in a plant protoplast system. For SL5, maintenance of the base of its stem was the principal feature required for robust in vivo accumulation. In the DSD, both helical and unpaired regions containing conserved sequences were necessary for efficient DI RNA accumulation. Additionally, optimal DI RNA accumulation required a TSD-DSD interaction mediated by a pseudoknot. Modifications that reduced accumulation did not appreciably affect DI RNA stability in vivo, indicating that the DSD and SL5 act to facilitate viral RNA replication.

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Section: Rna Secondary Structure Of the 3 Half Of The Tbsv 5 Utrmentioning

confidence: 79%

A second functional RNA domain in the 5′ UTR of the Tomato bushy stunt virus genome: Intra- and interdomain interactions mediate viral RNA replication

Ray

Wu²,

White³

2003

RNA

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“…Due to this remarkable specificity of these suppressors, they can be expressed in all stages of infections. Interestingly, the expression of P14 and TCV CP size-independent silencing suppressors in infected plants might be downregulated by the virus (49,71). The level of Sg2 RNA, from which P14 is translated, declines after 2 to 3 d.p.i.…”

Section: Discussionmentioning

confidence: 99%

“…The level of Sg2 RNA, from which P14 is translated, declines after 2 to 3 d.p.i. (39,49,50). In TCV-infected cells, the formation of capsid reduces available suppressor levels.…”

Section: Discussionmentioning

confidence: 99%

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Double-Stranded RNA Binding May Be a General Plant RNA Viral Strategy To Suppress RNA Silencing

Mérai

Kerényi

Kertész

et al. 2006

J Virol

278

255

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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 ␥B, 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.RNA silencing (termed RNA interference [RNAi] in animals) is an RNA-based eukaryotic gene regulatory system that plays essential roles in many biological processes. RNA silencing is induced by accumulation of double-stranded RNAs (dsRNAs). dsRNAs are first processed by an RNase III-like nuclease called DICER (in plants it is termed DICER-LIKE [DCL]) into (21-to 25-nucleotide [nt]) small dsRNAs (ds-sRNAs) having 2-nt 3Ј overhangs, and then these sRNAs incorporate into different silencing effector complexes. In the active effector complexes sRNAs are present as single-stranded molecules, which guide these complexes to the complementary nucleic acids for suppression (2, 3, 22, 61).In plants, different dsRNA precursors are processed by distinct DCLs into functionally different short (21-to 22-nt) and long (23-to 25-nt) sRNAs (24, 25, 67). Short sRNAs guide a multicomponent nuclease (RNA-induced silencing complex [RISC]) to homologous mRNAs for suppression. RISC cleaves targeted mRNA in the case of (near) perfect base pairing between mRNA and guide RNA. When the guide RNA is only partially complementary to the mRNA, RISC mediates translational repression. Short sRNAs could also provide ...

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“…9A). In addition, members of Aureusvirus (15,24)-the genus in the family Tombusviridae whose members are most closely related to tombusviruses-also possess potential TSDs with S1s that are 10-bp long (Fig. 9B).…”

Section: Discussionmentioning

confidence: 99%

Structural Properties of a Multifunctional T-Shaped RNA Domain That Mediate Efficient Tomato Bushy Stunt Virus RNA Replication

Ray

Hong

White

2004

J Virol

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In positive-strand RNA viruses, 5 untranslated regions (5 UTRs) mediate many essential viral processes, including genome replication. Previously, we proposed that the 5-terminal portion of the genomic leader sequence of Tomato bushy stunt virus (TBSV) forms an RNA structure containing a 3-helix junction, termed the T-shaped domain (TSD). In the present study, we have carried out structure-function analysis of the proposed TSD and have confirmed an important role for this domain in mediating efficient viral RNA amplification. Using a model TBSV defective interfering RNA replicon and a protoplast system, we demonstrated that various TSD subelements contribute to the efficiency of viral RNA replication. In particular, the stabilities of all three stems (S1, S2, and S4) forming the 3-helix junction are important, while stem-loop 3-a terminal extension of S2-is largely dispensable. Additionally, some of the sequences forming the 3-helix junction are required in an identity-dependent manner. Thus, both secondary structure and nucleotide identity are important for TSDmediated viral RNA replication. Importantly, these results are fully consistent with the dual functions we defined previously for the sequences corresponding to loops 3 and 4, respectively, in facilitating 5 cap-and 3 poly(A) tail-independent translation of the genome by forming a loop-loop interaction with the 3-proximal translational enhancer and in mediating viral RNA replication through formation of a pseudoknot with the adjacent downstream RNA domain. Also, since comparable TSDs and associated interactions are predicted in the 5 UTRs of all sequenced Aureusvirus genomes, members of at least one other genus in the family Tombusviridae appear to utilize this type of multifunctional RNA domain.Genome replication is an essential feature of virus reproduction. For positive-strand RNA viruses, efficient genome replication proceeds through a negative-strand RNA intermediate and is regulated by viral RNA elements, viral proteins, and host factors (2). The 5Ј untranslated regions (5Ј UTRs) in viral genomes contain primary, secondary, and tertiary RNA structures that are involved in genome replication. Accordingly, structural and functional assessments of these 5Ј UTRs have led to important advances in our understanding of viral genome replication and related regulatory mechanisms (2).Tomato bushy stunt virus (TBSV), the prototype member of the genus Tombusvirus in the family Tombusviridae, is a monopartite positive-strand RNA virus (25). Its 4.8-kb genome encodes five viral proteins with known functions (10) (Fig. 1A). Viral RNA replication requires only p33 and its translational readthrough product p92 (the RNA-dependent RNA polymerase, RdRp) (17). These two proteins are also able to replicate defective interfering (DI) RNAs, which are small deleted forms of the genome that maintain cis-acting viral RNA replication elements (e.g., promoters and enhancers) (28). As such, tombusvirus DI RNAs have been very useful for defining RNA sequences and structures involv...

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Molecular analysis of the pothos latent virus genome.

Cited by 33 publications

References 23 publications

A second functional RNA domain in the 5′ UTR of the Tomato bushy stunt virus genome: Intra- and interdomain interactions mediate viral RNA replication

A second functional RNA domain in the 5′ UTR of the Tomato bushy stunt virus genome: Intra- and interdomain interactions mediate viral RNA replication

Double-Stranded RNA Binding May Be a General Plant RNA Viral Strategy To Suppress RNA Silencing

Structural Properties of a Multifunctional T-Shaped RNA Domain That Mediate Efficient Tomato Bushy Stunt Virus RNA Replication

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