A four-nucleotide translation enhancer in the 3′-terminal consensus sequence of the nonpolyadenylated mRNAs of rotavirus

Chizhikov, Vladimir; Patton, John T.

doi:10.1017/s1355838200992264

Cited by 43 publications

(42 citation statements)

References 37 publications

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“…RV mRNAs are unique in that they possess a 5Ј cap structure but lack 3Ј poly(A) tails (26,27), and several studies have revealed the importance of these structures for efficient genome replication and translation (28)(29)(30). In this study, each of the authentic 5Ј and 3Ј end structures were planned to be obtained with vaccinia virus-encoded capping enzyme and the HDV ribozyme sequences inserted in the transcription plasmids (Fig.…”

Section: Discussionmentioning

confidence: 99%

Reverse genetics system for introduction of site-specific mutations into the double-stranded RNA genome of infectious rotavirus

Komoto

Sasaki

Taniguchi

2006

Proc. Natl. Acad. Sci. U.S.A.

136

135

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We describe here the successful establishment of a reverse genetics system for rotavirus (RV), a member of the Reoviridae family whose genome consists of 10 -12 segmented dsRNA. The system is based on the recombinant vaccinia virus T7 RNA polymerase-driven procedure for supplying artificial viral mRNA in the cytoplasm. With the aid of helper virus (human RV strain KU) infection, intracellularly transcribed full-length VP4 mRNA of simian RV strain SA11 resulted in the rescue of the KU-based transfectant virus carrying the SA11 VP4 RNA segment derived from cDNA. In addition to the rescued transfectant virus with the authentic SA11 VP4 gene, three more infectious RV transfectants, into which silent mutation(s) were introduced to destroy both or one of the two restriction enzyme sites as gene markers in the SA11 VP4 genome, were also rescued with this method. The ability to artificially manipulate the RV genome will greatly increase the understanding of the replication and the pathogenicity of RV and will provide a tool for the design of attenuated vaccine vectors.rescue of transfectant virus ͉ viral selection system ͉ Reoviridae R otavirus (RV) is the leading etiological agent of severe gastroenteritis in infants and young children worldwide and is estimated to cause 440,000 deaths and 140 million episodes of diarrhea each year (1, 2). As a member of the Reoviridae family, RV is a dsRNA virus that possesses an 11-segment genome (3). Most positive-and negative-stranded RNA viruses (reviewed in refs. 4-8) can be altered through site-specific mutagenesis by using cloned cDNA. Such reverse genetics systems allow artificial manipulation of viral genomes at the cDNA level by site-directed mutagenesis, deletion͞insertion, and rearrangement and have led to the accumulation of significant new knowledge relating to the replication, biological characteristics, and pathogeneses of these viral genera and families (5, 9). For dsRNA viruses, which comprise three families, the Reoviridae, Birnaviridae, and Cystoviridae, such achievements have so far been restricted to the low-numbered segmented dsRNA viruses: two segmented birnaviruses (10, 11) and three segmented 6 bacteriophage of the Cystoviridae (12). The Reoviridae viruses that possess 10-12 segmented genomes have been proven to be very refractory to this approach, except for the reoviruses. Roner and Joklik (13-15) developed a unique but complicated reovirus reverse genetics system involving temperature-sensitive mutants and transformed cells that stably express a particular viral protein encoded by the gene segment to be manipulated. However, there have been no reports on the performance of this method in other laboratories or its application to other Reoviridae members so far.Since the first development of an RV template-dependent in vitro replication system in 1994 (16) in which the RV open core can direct the synthesis of genomic dsRNA from viral mRNA in a cell-free system, no infectious RV transfectants have been rescued at all as far as we know, despite intensive attem...

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Section: Discussionmentioning

confidence: 99%

Reverse genetics system for introduction of site-specific mutations into the double-stranded RNA genome of infectious rotavirus

Komoto

Sasaki

Taniguchi

2006

Proc. Natl. Acad. Sci. U.S.A.

136

135

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“…Considering that RbcS mRNAs encode photosynthetic proteins that accumulate to very high levels in the chloroplasts of C 4 as well as C 3 plants (Edwards and Huber, 1981;Miziorko and Lorimer, 1983;Sage et al, 1987), it might be expected that these transcripts would contain sequences capable of acting as strong translational enhancers. Translational enhancement is primarily known from some plant and animal viral RNAs (Gallie, 1993(Gallie, , 2002De La Luna et al, 1995;Chizhikov and Patton, 2000;Salvatore et al, 2002). There are also several examples of plant cellular transcripts showing enhanced translation capabilities (Pitto et al, 1992;Gallie and Young, 1994;Bate et al, 1996;Ling et al, 2000).…”

Section: Discussionmentioning

confidence: 99%

Untranslated Regions from C4 Amaranth AhRbcS1 mRNAs Confer Translational Enhancement and Preferential Bundle Sheath Cell Expression in Transgenic C4Flaveria bidentis

et al. 2004

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Many aspects of photosynthetic gene expression are posttranscriptionally regulated in C4 plants. To determine if RbcS mRNA untranslated regions (UTRs) in themselves could confer any characteristic C4 expression patterns, 5′- and 3′-UTRs of AhRbcS1 mRNA from the C4 dicot amaranth were linked to a gusA reporter gene. These were constitutively transcribed from a cauliflower mosaic virus promoter and assayed for posttranscriptional expression patterns in transgenic lines of the C4 dicot Flaveria bidentis. Three characteristic C4 expression patterns were conferred by heterologous AhRbcS1 UTRs in transgenic F. bidentis. First, the AhRbcS1 UTRs conferred strong translational enhancement of gusA expression, relative to control constructs lacking these UTRs. Second, while the UTRs did not appear to confer tissue-specific expression when analyzed by β-glucuronidase activity assays, differences in gusA mRNA accumulation were observed in leaves, stems, and roots. Third, the AhRbcS1 UTRs conferred preferential gusA expression (enzyme activity and gusA mRNA accumulation) in leaf bundle sheath cells. AhRbcS1 UTR-mediated translational enhancement was also observed in transgenic C3 plants (tobacco [Nicotiana tabacum]) and in in vitro translation extracts. These mRNAs appear to be translated with different efficiencies in C4 versus C3 plants, indicating that processes determining overall translational efficiency may vary between these two categories of higher plants. Our findings suggest that the AhRbcS1 5′-UTR functions as a strong translational enhancer in leaves and other tissues, and may work synergistically with the 3′-UTR to modulate overall levels of Rubisco gene expression in different tissues and cell types of C4 plants.

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“…RNA viruses that lack a cap and/or a poly(A) tail have developed alternative strategies for translation regulation involving the 3Ј UTR (9,22). The roles of the 3Ј UTR in translational enhancement of viral RNAs have been studied with human and animal viral systems, including HCV (13) and rotavirus (4). Translation enhancer elements within the 3Ј UTRs of plant viruses that lack both the cap and the poly(A) tail have been implicated as determinants for translation efficiency in tomato bushy stunt virus (46), satellite necrosis virus (22), BYDV (1,40), and TCV (29).…”

Section: Discussionmentioning

confidence: 99%

A Six-Nucleotide Segment within the 3′ Untranslated Region of Hibiscus Chlorotic Ringspot Virus Plays an Essential Role in Translational Enhancement

Koh

Liu

Wong

2002

J Virol

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RNA plant viruses use various translational regulatory mechanisms to control their gene expression.Translational enhancement of viral mRNAs that leads to higher levels of protein synthesis from specific genes may be essential for the virus to successfully compete for cellular translational machinery. The control elements have yet to be analyzed for members of the genus Carmovirus, a small group of plant viruses with positive-sense RNA genomes. In this study, we examined the 3 untranslated region (UTR) of hibiscus chlorotic ringspot virus (HCRSV) genomic RNA (gRNA) and subgenomic RNA (sgRNA) for its role in the translational regulation of viral gene expression. The results showed that the 3 UTR of HCRSV significantly enhanced the translation of several open reading frames on gRNA and sgRNA and a viral gene in a bicistronic construct with an inserted internal ribosome entry site. Through deletion and mutagenesis studies of both the bicistronic construct and full-length gRNA, we demonstrated that a six-nucleotide sequence, GGGCAG, that is complementary to the 3 region of the 18S rRNA and a minimal length of 180 nucleotides are required for the enhancement of translation induced by the 3 UTR.The 3Ј untranslated regions (UTRs) of plant virus RNAs are likely to function in a manner similar to that of the corresponding regions of cellular mRNA in regulating translation and maintaining RNA stability. Due to their great variety in terminal structures, the 3Ј UTRs of plus-strand viral RNAs play varied roles in replication and translation (5). For example, the turnip yellow mosaic virus RNA contains a 3Ј tRNA mimic believed to regulate minus-strand RNA synthesis (6). For brome mosaic virus, the 3Ј UTR harbors a unique promoter element that directs minus-strand RNA synthesis (23). The 3Ј UTR of tobacco mosaic virus contains a translation enhancer (8).Hibiscus chlorotic ringspot virus (HCRSV), a member of the genus Carmovirus, is an isometric monopartite plant virus which measures 28 nm in diameter. The virus is found worldwide where hibiscus is cultivated (16,41,42). The symptoms on HCRSV-infected plants range from generalized mottle to chlorotic ringspots and vein-banding patterns, severe stunting, and flower distortion (42,45). HCRSV possesses a singlestranded positive-sense RNA that is not polyadenylated at the 3Ј terminus. The genomic RNA (gRNA) is 3,911 nucleotides (nt) long and has the potential to encode seven open reading frames (ORFs), including two novel ORFs, p23 and p25 (12). Two 3Ј-coterminated subgenomic RNA (sgRNA) species have been identified.In this study, we examined the 3Ј UTR of HCRSV gRNA and sgRNA for its role in the regulation of translation. Through deletion studies and elimination of putative translation enhancer elements, we were able to identify the prerequisites required for the translational enhancement induced by the 3Ј UTR of HCRSV. The results showed that the 3Ј UTR of HCRSV differentially enhanced the translation of ORFs on gRNA and sgRNA and an HCRSV gene in a bicistronic construct by 1.5-t...

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A four-nucleotide translation enhancer in the 3′-terminal consensus sequence of the nonpolyadenylated mRNAs of rotavirus

Abstract: The 59 cap and poly(A) tail of eukaryotic mRNAs work synergistically to enhance translation through a process that requires interaction of the cap-associated eukaryotic initiation factor, eIF-4G, and the poly (

Cited by 43 publications

References 37 publications

Reverse genetics system for introduction of site-specific mutations into the double-stranded RNA genome of infectious rotavirus

Reverse genetics system for introduction of site-specific mutations into the double-stranded RNA genome of infectious rotavirus

Untranslated Regions from C4 Amaranth AhRbcS1 mRNAs Confer Translational Enhancement and Preferential Bundle Sheath Cell Expression in Transgenic C4Flaveria bidentis

A Six-Nucleotide Segment within the 3′ Untranslated Region of Hibiscus Chlorotic Ringspot Virus Plays an Essential Role in Translational Enhancement

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