Localization of cis-acting sequences essential for cymbidium ringspot tombusvirus defective interfering RNA replication

Havelda, Zoltán; Dalmay, Tamás; Burgyán, József

doi:10.1099/0022-1317-76-9-2311

Cited by 27 publications

(25 citation statements)

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(11 reference statements)

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“…We also demonstrated that the poor target activity of short DI RNAs was not caused by either the low level of the DI RNAspecific guide RNAs or the size of the target molecule. Short DI RNAs, which contain almost exclusively cis -acting sequences required for replication (Havelda et al, 1995), may bind efficiently to the replicase complex and thus are not accessible to the RISC degradation complex of PTGS. Alternatively, the folding of DI RNAs may prevent PTGS-mediated degradation.…”

Section: Ptgs Selects For DI Rnas Containing Poor Ptgs Target Regionsmentioning

confidence: 99%

“…DI RNAs of Cymbidium ringspot tombusvirus (CymRSV) are among the most extensively studied DI RNA molecules. They are incapable of autonomous replication but contain all of the necessary cis -acting elements for replication (Havelda et al, 1995). During replication of CymRSV genomic RNA, a series of DI RNAs of ‫ف‬ 0.4 to 0.7 kb are generated de novo (Burgyán et al, 1991).…”

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Short Defective Interfering RNAs of Tombusviruses Are Not Targeted but Trigger Post-Transcriptional Gene Silencing against Their Helper Virus

et al. 2002

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Post-transcriptional gene silencing (PTGS) is INTRODUCTIONPost-transcriptional gene silencing (PTGS), first identified in plants, is now thought to be an ancient self-defense mechanism acting against molecular parasites (Waterhouse et al., 2001). Introduction of double-stranded RNA (dsRNA) into plant cells triggers PTGS, resulting in the degradation of dsRNA and cognate mRNAs (Schweizer et al., 2000). A similar mechanism appears to operate in a wide variety of organisms, including filamentous fungi, nematodes, Drosophila , mice, and cultured HeLa cells, and generally is referred to as RNA interference (RNAi) (Cogoni and Macino, 1999a;Fire, 1999;Grant, 1999;Sharp and Zamore, 2000;Elbashir et al., 2001a;Svoboda et al., 2000). Recently, homologous genes required for PTGS were identified from different organisms, demonstrating the conservation of the gene-silencing machinery Macino, 1999a, 1999b;Ketting et al., 1999;Tabara et al., 1999;Catalanotto et al., 2000;Dalmay et al., 2000Dalmay et al., , 2001Domeier et al., 2000;Fagard et al., 2000;Mourrain et al., 2000;Smardon et al., 2000;Wu-Scharf et al., 2000). The accumulation of 21-to 25-nucleotide RNAs corresponding to both sense and antisense strands of target RNA occurs during PTGS in plant and animal cells (Hamilton and Baulcombe, 1999;Hammond et al., 2000;Parrish et al., 2000). These 21-to 25-nucleotide RNAs are generated by an RNase III-like enzyme (DICER) as the initiation step of RNAi, providing the specificity of a second RNase complex (RISC) that targets the cognate single-stranded (ss) RNAs (Bernstein et al., 2001).In plants, PTGS has evolved as an antiviral system. PTGS is triggered efficiently by dsRNA intermediates of cytoplasmically replicating viruses. The RNA genome of the invading virus is targeted and eliminated specifically when this natural antiviral mechanism is activated (Waterhouse et al., 1998(Waterhouse et al., , 1999Baulcombe, 1999;Smith et al., 2000;. In higher plants, PTGS is not limited to the cells in which it is activated, because mobile signals produced by PTGS can spread and confer sequence-specific RNA degradation in distant tissues (Palauqui et al., 1997;Voinnet and Baulcombe, 1997).Consistent with the importance of PTGS as an antiviral response, many viruses encode gene-silencing suppressor proteins (Anandalakshmi et al., 1998;Beclin et al., 1998;Brigneti et al., 1998;Kasschau and Carrington, 1998;Voinnet et al., 1999Voinnet et al., , 2000. However, not all viruses are able to suppress PTGS, and some virus-infected plants recover after the development of the first systemic viral symptoms (e.g., 1 These authors contributed equally to this work. 2 To whom correspondence should be addressed. E-mail burgyan@ abc.hu; fax 36-28-430-416. Article, publication date, and citation information can be found at www.plantcell.org/cgi/doi/10.1105/tpc.010366. 360The Plant Cell nepovirus-infected tobacco plants; Ratcliff et al., 1997). Upper leaves of recovered plants lack symptoms (or show attenuated symptoms), and the virus content in these leav...

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Section: Ptgs Selects For DI Rnas Containing Poor Ptgs Target Regionsmentioning

confidence: 99%

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confidence: 99%

Short Defective Interfering RNAs of Tombusviruses Are Not Targeted but Trigger Post-Transcriptional Gene Silencing against Their Helper Virus

et al. 2002

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“…DI RNAs are incapable of autonomous replication but contain all the necessary cis-acting elements for replication (Havelda et al, 1995). Trans-acting products required for CymRSV DI RNA replication are expressed from ORFs 1 and 2 of the CymRSV genome Kolla!…”

Section: Mutational Analysis Of Defective Interfering (Di)mentioning

confidence: 99%

Secondary structure-dependent evolution of Cymbidium ringspot virus defective interfering RNA.

Havelda

Dalmay

Burgyán

1997

Journal of General Virology

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Mutational analysis of defective interfering (DI)RNAs of Cymbidium ringspot virus (CymRSV) was used to study the mechanism of DI RNA evolution. It was shown that a highly base-paired structure in the 3h region of the longer DI RNA directed the formation of smaller DI RNA molecules. Mutations which increased the stability of the computerpredicted, highly structured 3h region of the longest DI RNA of CymRSV significantly enhanced the generation and accumulation of the smaller derivatives. Sequence analysis of smaller progeny molecules revealed that the highly base-paired region was deleted from the precursor DI RNA. Moreover, sites of recombination were found in other regions of the DI RNA progenies due to transposition of the highly base-paired structure. It is likely that the deletion event was structure-and not sequencespecific, and operated when a foreign sequence containing a 37-nt-long base-paired stem was inserted at the appropriate position of DI RNA.Defective interfering (DI) RNAs are shortened forms of viral genomes that have frequently lost all the essential viral genes required for movement, replication and encapsidation. DI RNAs require the presence of a helper virus for trans-acting factors necessary for replication and, in general, accumulate at the expense of the helper virus from which they are derived. Interference with the helper genome often results in remarkable symptom attenuation (Roux et al., 1991). DI RNAs are commonly found associated with both animal and plant virus infections ; however, the presence of DI RNAs in plant virus systems is less prevalent. The family Tombusviridae may be an exception because an increasing number of DI RNAs are found associated with it. The most extensively studied plant virus DI Author for correspondence : Jo! zsef Burgya! n.Fax j36 28 430 482. e-mail burgyan!hubi.abc.huThe nucleotide sequence of TBSV-P genomic RNA has been deposited in the EMBL and GenBank nucleotide databases under accession number U80935.

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“…1) (33,(45)(46)(47). These features are also common for DIs derived from other tombusviruses, such as Cymbidium ringspot virus (CymRSV) and Cucumber necrosis virus (CNV) (9,12,13,16,17).Regions I, II, and IV of DIs represent sequences essential for RNA amplification (3,33,45,49), and while region III is dispensable for DI replication of TBSV, it is required for CymRSV DI accumulation (3,17,33). The length of TBSV DI region III varies from 46 to 81 nucleotides (nt), depending on the DI isolate (3,20,47).…”

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confidence: 99%

“…The tombusvirus coat protein is translated from p41 on subgenomic RNA1 (sgRNA1), and P22 and P19 are expressed from p22 and p19 on sgRNA2 (21)(22)(23)36). TBSV P22 is required for cell-to-cell movement (7,10,42), and P19 is involved in host-specific systemic invasion and symptom development (6,38,41,42) and is active as a suppressor of gene silencing (31,32,44).Tombusviruses can serve as helper viruses for satellite RNAs (1, 2), and they are notorious for the accumulation of defective interfering RNAs (DIs) (13,17,20,24). DIs are spontaneous deletion mutants derived from the helper genome, and they are successfully amplified by trans-acting helper replicase proteins (20, 24).…”

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confidence: 99%

Tomato Bushy Stunt Virus Genomic RNA Accumulation Is Regulated by Interdependent cis -Acting Elements within the Movement Protein Open Reading Frames

Park

Desvoyes

Scholthof

2002

J Virol

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This study on Tomato bushy stunt virus identified and defined three previously unknown regulatory sequences involved in RNA accumulation that are located within the 3-proximal nested movement protein genes p22 and p19. The first is a 16-nucleotide (nt) element termed III-A that is positioned at the very 3 end of p22 and is essential for RNA accumulation. Approximately 300 nt upstream of III-A resides an ϳ80-nt inhibitory element (IE) that is obstructive to replication only in the absence of a third regulatory element of ϳ30 nt (SUR-III) that is positioned immediately upstream of III-A. Inspection of the nucleotide sequences predicted that III-A and SUR-III can form looped hairpins. A comparison of different tombusviruses showed, in each case, conservation for potential base pairing between the two predicted hairpin-loops. Insertion of a spacer adjacent to the predicted hairpins had no or a minimal effect on RNA accumulation, whereas an insertion in the putative III-A loop abolished genomic RNA multiplication. We conclude that the sequences composing the predicted III-A and SUR-III hairpin-loops are crucial for optimal RNA accumulation and that the inhibitory effect of IE surfaces when the alleged interaction between SUR-III and III-A is disturbed.Tomato bushy stunt virus (TBSV), the type member of the genus Tombusvirus in the family Tombusviridae, is a small icosahedral virus with a positive-sense single-stranded RNA genome of approximately 4.8 kb that encodes five major open reading frames (ORFs) (Fig. 1) (19). The genomic RNA (gRNA) functions as an mRNA for the translation of two 5Ј-proximal ORFs, p33 and p92, encoding two replicationassociated proteins, P33 and its readthrough product P92 (27,43). The tombusvirus coat protein is translated from p41 on subgenomic RNA1 (sgRNA1), and P22 and P19 are expressed from p22 and p19 on sgRNA2 (21)(22)(23)36). TBSV P22 is required for cell-to-cell movement (7,10,42), and P19 is involved in host-specific systemic invasion and symptom development (6,38,41,42) and is active as a suppressor of gene silencing (31,32,44).Tombusviruses can serve as helper viruses for satellite RNAs (1, 2), and they are notorious for the accumulation of defective interfering RNAs (DIs) (13,17,20,24). DIs are spontaneous deletion mutants derived from the helper genome, and they are successfully amplified by trans-acting helper replicase proteins (20, 24). DIs have been valuable tools with which to identify RNA elements involved in viral accumulation (3,16,17). TBSV DIs are composed of four noncontiguous RNA regions (I through IV) from the helper genome ( Fig. 1) (45). Regions I and IV correspond to the 5Ј-and 3Ј-untranslated regions of the TBSV genome, and regions II and III represent internal regions of p92 and a segment at the 3Ј end of the overlapping p19 and p22 ORFs, respectively ( Fig. 1) (33,(45)(46)(47). These features are also common for DIs derived from other tombusviruses, such as Cymbidium ringspot virus (CymRSV) and Cucumber necrosis virus (CNV) (9,12,13,16,17).Regions I, II, and IV...

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Localization of cis-acting sequences essential for cymbidium ringspot tombusvirus defective interfering RNA replication

Cited by 27 publications

References 7 publications

Short Defective Interfering RNAs of Tombusviruses Are Not Targeted but Trigger Post-Transcriptional Gene Silencing against Their Helper Virus

Short Defective Interfering RNAs of Tombusviruses Are Not Targeted but Trigger Post-Transcriptional Gene Silencing against Their Helper Virus

Secondary structure-dependent evolution of Cymbidium ringspot virus defective interfering RNA.

Tomato Bushy Stunt Virus Genomic RNA Accumulation Is Regulated by Interdependent cis -Acting Elements within the Movement Protein Open Reading Frames

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