Sequence analysis of rice hoja blanca virus RNA 3

Miranda, Joachim de; Hernández, Míriam Albert; Hull, Roger; Espinoza, Ana M.

doi:10.1099/0022-1317-75-8-2127

Cited by 38 publications

(12 citation statements)

References 25 publications

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“…Like for TSWV, the largest RNA segment of RHBV (RNA 1) is of complete negative polarity and encodes the putative viral polymerase. The other three RNA segments have an ambisense coding strategy, thus encoding a further six proteins, of which only the function of the nucleoprotein encoded on RNA 3 has been indicated (14,15,41,42). The genome arrangements of TSWV and RHBV are indicated in Fig.…”

mentioning

confidence: 99%

Negative-Strand Tospoviruses and Tenuiviruses Carry a Gene for a Suppressor of Gene Silencing at Analogous Genomic Positions

Bucher¹,

Sijen

Haan³

et al. 2003

J Virol

220

166

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Posttranscriptional silencing of a green fluorescent protein (GFP) transgene in Nicotiana benthamiana plants was suppressed when these plants were infected with Tomato spotted wilt virus (TSWV), a plant-infecting member of the Bunyaviridae. Infection with TSWV resulted in complete reactivation of GFP expression, similar to the case for Potato virus Y, but distinct from that for Cucumber mosaic virus, two viruses known to carry genes encoding silencing suppressor proteins. Agrobacterium-based leaf injections with individual TSWV genes identified the NS S gene to be responsible for the RNA silencing-suppressing activity displayed by this virus. The absence of short interfering RNAs in NS S -expressing leaf sectors suggests that the tospoviral NS S protein interferes with the intrinsic RNA silencing present in plants. Suppression of RNA silencing was also observed when the NS3 protein of the Rice hoja blanca tenuivirus, a nonenveloped negative-strand virus, was expressed. These results indicate that plant-infecting negative-strand RNA viruses carry a gene for a suppressor of RNA silencing.RNA silencing involves a sequence-specific degradation which is induced by overabundant RNA and by doublestranded RNA (dsRNA) molecules and which can target transgenes as well as homologous endogenous genes. RNA silencing was first described for plants (35,50) and over recent years has been described for other organisms, where it is also referred to as cosuppression, posttranscriptional gene silencing (17), or RNA-mediated virus resistance (3,11,30) in plants, quelling in fungi (9), or RNAi in animals (19). Building blocks of the gene-silencing pathway proved to have remarkable similarities in the different organisms and hence suggest an ancient role of gene silencing in pathogen resistance or development (10,25,53). One of the key intermediary elements in the RNA silencing pathway is dsRNA, which is recognized by a dsRNAspecific nuclease (5) to yield small (21 to 23 nucleotides) short interfering RNAs (siRNAs) (21). These siRNAs subsequently serve as guides for cleavage of homologous RNA molecules. In plants, versions of transgenes that produce dsRNA molecules have been shown to be very potent activators of RNA silencing (47). As all RNA viruses replicate through formation of dsRNA intermediates, these are potential targets of the RNA silencing mechanism. Indeed, antiviral RNA silencing has been shown to occur in nature and has been proposed as a natural defense mechanism protecting plants against viruses, resulting in resistance (1, 43).To counteract the RNA silencing mechanism of their host, plant viruses have developed ways to evade or neutralize this response. Over recent years, RNA silencing-inhibiting proteins have been identified in several plant viruses. Among the best-studied examples are the helper component-proteinase (HC-Pro) of the potyvirus Potato virus Y (PVY) and the 2b protein of Cucumber mosaic virus (CMV) (7, 40). Other plus-strand RNA (and some DNA) viruses also have been found to suppress gene silencing, and ...

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mentioning

confidence: 99%

Negative-Strand Tospoviruses and Tenuiviruses Carry a Gene for a Suppressor of Gene Silencing at Analogous Genomic Positions

Bucher¹,

Sijen

Haan³

et al. 2003

J Virol

220

166

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“…The IRs of ambisense viruses have been suggested to adopt a hairpin structure which would play an important role in transcription termination (9). vRNA4 can adopt such a structure (27), but vRNA3 cannot (7). It is possible that another secondary structure and/or signal participates in transcription termination of vRNA3.…”

Section: Discussionmentioning

confidence: 99%

“…Its tetrapartite genome has been characterized (26). The three smallest RNA segments have been sequenced; they are ambisense in genome organization (7,8,27). Similarly to RSV, the RHBV v-and vcRNA2 code for the NS2 (23 kDa) and NSvc2 (94 kDa) proteins, the v-and vcRNA3 code for the NS3 (23 kDa) and NC (35 kDa) proteins, and the v-and vcRNA4 code for the NS4 (20 kDa) and NSvc4 (32 kDa) proteins.…”

Section: Rice Hoja Blanca Virus (Rhbv) Is a Species Of The Genusmentioning

confidence: 99%

Characterization of the in vitro activity of the RNA-dependent RNA polymerase associated with the ribonucleoproteins of rice hoja blanca tenuivirus

Nguyen

Ramirez

Goldbach

et al. 1997

J Virol

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An RNA-dependent RNA polymerase (RdRp) activity associated with the ribonucleoproteins of rice hoja blanca tenuivirus (RHBV) was detected and analyzed. Conditions for in vitro RNA synthesis and for coupled RNA synthesis-translation of RHBV were established. In both cases, synthesis of the viral and viral complementary genomic and subgenomic RNA3 and RNA4 were observed, demonstrating that both transcription and replication occurred. Though coupling of RNA synthesis to translation allowed efficient translation of the newly synthesized subgenomic RNAs, studies of the effect of various inhibitors of protein synthesis revealed that RNA synthesis was independent of translation. Primer extension experiments demonstrated that in the presence of capped exogenous RNAs, a stretch of 10 to 16 nonviral nucleotides was added to the 5 end of a population of newly synthesized viral complementary RNA4. It appears that in addition to RdRp activity, RHBV-associated protein(s) also possessed cap-snatching capacity. by guest http://jvi.asm.org/ Downloaded from RNA purification. When total RNAs were purified from RHBV RNPs by proteinase K digestion followed by CsCl centrifugation (26), they are referred to as CsCl-purified RHBV RNAs. When they were obtained, after proteinase K treatment, by phenol extraction and ethanol precipitation of the RHBV RNPs, they are referred to as phenol-extracted RNAs.Preparation of cDNA subclones. cDNAs of CsCl-purified RHBV RNA3 and -4 were cloned into pBluescript II KSϩ or SKϪ (Stratagene) as described previously (24, 27). To avoid nonspecific hybridization signals due to the IR contained in the transcripts of the RHBV cDNA clones, a subcloning step deleting the IR was carried out in three of the clones. Deletions were performed by digestion of plasmids RHB4C-341, R3-1, and RHBC-8 by pairs of restriction enzymes, i.e., StuI-PstI, StuI-KpnI, and BglII-PstI, respectively. The plasmids were blunt ended by using T4 DNA polymerase (Boehringer Mannheim) and religated with T4 DNA ligase (Boehringer Mannheim) as indicated by the manufacturer. The subclones are designated by the names of their parental plasmids followed by the suffix sc (Table 1). Plasmid R4-26 was previously subcloned to yield R4-26B (27) designated here R4-26sc (Table 1).Preparation of transcripts and/or riboprobes. Plasmids containing the ORFs of proteins encoded by v-or vcRNA3 or -4 were linearized with appropriate restriction enzymes prior to transcription using T3 or T7 RNA polymerase. Plasmids R3-1 and RHBC-8 (24) were linearized with KpnI in the multiple cloning site (KpnI MCS ) to yield the NS3 and NC ORFs, respectively. Plasmid R4-26 was linearized with XbaI MCS to yield the NS4 ORF, and plasmid RHB4C-341 was linearized with BamHI MCS to yield the NSvc4 ORF. In addition, transcripts of about 245 nucleotides and devoid of any ORF (referred to as SKϪ RNAs) were obtained by digestion of pBluescript II SKϪ with PvuII and transcription by T3 RNA polymerase.The transcripts were synthesized essentially as described by the manufacturer (Strata...

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“…The p3 protein encoded in the virion-sense of RNA-4 of RSV and RHBV functions as the viral silencing suppressor (Bucher et al, 2003; Hemmes et al, 2007; Schnettler et al, 2008; Xiong et al, 2009). The pC3 protein encoded in the virion-complementary sense of RNA-3 of RSV and RHBV and pC5 encoded in the virion-complementary sense of RGSV RNA-5 are nucleocapsid proteins and major components of the thin filamentous particles (Zhu et al, 1991; de Miranda et al, 1994; Toriyama et al, 1997). The p4 protein encoded in the virion-sense strand of RNA-4 of RSV and RHBV and p6 encoded by the virion-sense of the RGSV RNA-6 accumulate in large amounts and form crystalline inclusion bodies in virus-infected rice plants, but their functions are unknown (Koganezawa, 1977; Falk et al, 1987; Hayano et al, 1990; Miranda and Koganezawa, 1995).…”

Section: Conferring Resistance Against Rice-infecting Tenuivirusesmentioning

confidence: 99%

Transgenic strategies to confer resistance against viruses in rice plants

et al. 2014

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Rice (Oryza sativa L.) is cultivated in more than 100 countries and supports nearly half of the world’s population. Developing efficient methods to control rice viruses is thus an urgent necessity because viruses cause serious losses in rice yield. Most rice viruses are transmitted by insect vectors, notably planthoppers and leafhoppers. Viruliferous insect vectors can disperse their viruses over relatively long distances, and eradication of the viruses is very difficult once they become widespread. Exploitation of natural genetic sources of resistance is one of the most effective approaches to protect crops from virus infection; however, only a few naturally occurring rice genes confer resistance against rice viruses. Many investigators are using genetic engineering of rice plants as a potential strategy to control viral diseases. Using viral genes to confer pathogen-derived resistance against crops is a well-established procedure, and the expression of various viral gene products has proved to be effective in preventing or reducing infection by various plant viruses since the 1990s. RNA interference (RNAi), also known as RNA silencing, is one of the most efficient methods to confer resistance against plant viruses on their respective crops. In this article, we review the recent progress, mainly conducted by our research group, in transgenic strategies to confer resistance against tenuiviruses and reoviruses in rice plants. Our findings also illustrate that not all RNAi constructs against viral RNAs are equally effective in preventing virus infection and that it is important to identify the viral “Achilles’ heel” gene to target for RNAi attack when engineering plants.

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Sequence analysis of rice hoja blanca virus RNA 3

Cited by 38 publications

References 25 publications

Negative-Strand Tospoviruses and Tenuiviruses Carry a Gene for a Suppressor of Gene Silencing at Analogous Genomic Positions

Negative-Strand Tospoviruses and Tenuiviruses Carry a Gene for a Suppressor of Gene Silencing at Analogous Genomic Positions

Characterization of the in vitro activity of the RNA-dependent RNA polymerase associated with the ribonucleoproteins of rice hoja blanca tenuivirus

Transgenic strategies to confer resistance against viruses in rice plants

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