The Kissing-Loop T-Shaped Structure Translational Enhancer of Pea Enation Mosaic Virus Can Bind Simultaneously to Ribosomes and a 5′ Proximal Hairpin

Gao, Feng; Gulay, Suna P.; Kasprzak, Wojciech K.; Dinman, Jonathan D.; Shapiro, Bruce A.; Simon, Anne

doi:10.1128/jvi.02005-13

Cited by 33 publications

(58 citation statements)

References 56 publications

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“…In vitro translation and trans inhibition assays were performed as previously described (28). Briefly, 3 pmol of in vitro-synthesized RNA transcripts from designated translation reporter constructs were used for a 15-l translation reaction using wheat germ extracts (WGE; Promega) according to the manufacturer's instructions.…”

Section: Methodsmentioning

confidence: 99%

“…The kl-TSS is a two hairpin, three-way branched element that forms a three-dimensional (3-D) T-shaped structure similar to the three hairpin/two pseudoknot TCV TSS. Unlike the TCV TSS, the kl-TSS binds to 40S ribosomal subunits in addition to 60S and 80S ribosomes and can engage in a simultaneous long-distance kissing-loop interaction with a 5= end coding sequence hairpin (13,28). It has been proposed that the kl-TSS binds to ribosomal subunits following translation termination to facilitate reinitiation at the 5= end.…”

Section: Importancementioning

confidence: 99%

“…In PEMV, the adjacent kl-TSS and PTE are 3=CITEs thought to function together to efficiently attract ribosomes to the kl-TSS (28). As described above, the kl-TSS can bind to ribosomes and simultaneously engage in a long-distance kissing-loop interaction between its 5= side hairpin (3H1) and a stable coding region hairpin (5H2) near the gRNA 5= end (28). Major questions therefore are whether the PEMV 3=TSS functions as a third 3=CITE and, if so, whether an additional kissing-loop interaction connects the 3=TSS with the 5= end.…”

Section: Figmentioning

confidence: 99%

“…PEMV contains two adjacent 3=CITEs in the center of its 703-nt 3=UTR: eIF4E-binding PTE and the ribosome-binding, kissingloop TSS (kl-TSS) (13,28). Unlike PTEs in carmoviruses (12), the PEMV PTE is not associated with any long-distance RNA-RNA interaction and instead likely functions to assist in ribosomes binding to the adjacent, upstream kl-TSS (28). The kl-TSS is a two hairpin, three-way branched element that forms a three-dimensional (3-D) T-shaped structure similar to the three hairpin/two pseudoknot TCV TSS.…”

Section: Importancementioning

confidence: 99%

“…Two overlapping proteins (p26 and p27) are expressed from ORF3 and ORF4 via at least one subgenomic RNA (26,27). PEMV contains two adjacent 3=CITEs in the center of its 703-nt 3=UTR: eIF4E-binding PTE and the ribosome-binding, kissingloop TSS (kl-TSS) (13,28). Unlike PTEs in carmoviruses (12), the PEMV PTE is not associated with any long-distance RNA-RNA interaction and instead likely functions to assist in ribosomes binding to the adjacent, upstream kl-TSS (28).…”

Section: Importancementioning

confidence: 99%

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The 3′ Untranslated Region of Pea Enation Mosaic Virus Contains Two T-Shaped, Ribosome-Binding, Cap-Independent Translation Enhancers

Gao

Kasprzak

Szarko

et al. 2014

J Virol

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Many plant viruses without 5=caps or 3= poly(A) tails contain 3= proximal, cap-independent translation enhancers (3=CITEs) that bind to ribosomal subunits or translation factors thought to assist in ribosome recruitment. Most 3=CITEs participate in a longdistance kissing-loop interaction with a 5= proximal hairpin to deliver ribosomal subunits to the 5= end for translation initiation. Pea enation mosaic virus (PEMV) contains two adjacent 3=CITEs in the center of its 703-nucleotide 3= untranslated region (3=UTR), the ribosome-binding, kissing-loop T-shaped structure (kl-TSS) and eukaryotic translation initiation factor 4E-binding Panicum mosaic virus-like translation enhance (PTE). We now report that PEMV contains a third, independent 3=CITE located near the 3= terminus. This 3=CITE is composed of three hairpins and two pseudoknots, similar to the TSS 3=CITE of the carmovirus Turnip crinkle virus (TCV). As with the TCV TSS, the PEMV 3=TSS is predicted to fold into a T-shaped structure that binds to 80S ribosomes and 60S ribosomal subunits. A small hairpin (kl-H) upstream of the 3=TSS contains an apical loop capable of forming a kissing-loop interaction with a 5= proximal hairpin and is critical for the accumulation of full-length PEMV in protoplasts. Although the kl-H and 3=TSS are dispensable for the translation of a reporter construct containing the complete PEMV 3=UTR in vitro, deleting the normally required kl-TSS and PTE 3=CITEs and placing the kl-H and 3=TSS proximal to the reporter termination codon restores translation to near wild-type levels. This suggests that PEMV requires three 3=CITEs for proper translation and that additional translation enhancers may have been missed if reporter constructs were used in 3=CITE identification. IMPORTANCEThe rapid life cycle of viruses requires efficient translation of viral-encoded proteins. Many plant RNA viruses contain 3= capindependent translation enhancers (3=CITEs) to effectively compete with ongoing host translation. Since only single 3=CITEs have been identified for the vast majority of individual viruses, it is widely accepted that this is sufficient for a virus's translational needs. Pea enation mosaic virus possesses a ribosome-binding 3=CITE that can connect to the 5= end through an RNA-RNA interaction and an adjacent eukaryotic translation initiation factor 4E-binding 3=CITE. We report the identification of a third 3=CITE that binds weakly to ribosomes and requires an upstream hairpin to form a bridge between the 3= and 5= ends. Although both ribosome-binding 3=CITEs are critical for virus accumulation in vivo, only the CITE closest to the termination codon of a reporter open reading frame is active, suggesting that artificial constructs used for 3=CITE identification may underestimate the number of CITEs that participate in translation.

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Section: Methodsmentioning

confidence: 99%

Section: Importancementioning

confidence: 99%

Section: Figmentioning

confidence: 99%

Section: Importancementioning

confidence: 99%

Section: Importancementioning

confidence: 99%

See 3 more Smart Citations

The 3′ Untranslated Region of Pea Enation Mosaic Virus Contains Two T-Shaped, Ribosome-Binding, Cap-Independent Translation Enhancers

Gao

Kasprzak

Szarko

et al. 2014

J Virol

Self Cite

View full text Add to dashboard Cite

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Fatal attraction: The roles of ribosomal proteins in the viral life cycle

2020

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Upon viral infection of a host cell, each virus starts a program to generate many progeny viruses. Although viruses interact with the host cell in numerous ways, one critical step in the virus life cycle is the expression of viral proteins, which are synthesized by the host ribosomes in conjunction with host translation factors. Here we review different mechanisms viruses have evolved to effectively seize host cell ribosomes, the roles of specific ribosomal proteins and their posttranslational modifications on viral RNA translation, or the cellular response to infection. We further highlight ribosomal proteins with extra‐ribosomal function during viral infection and put the knowledge of ribosomal proteins during viral infection into the larger context of ribosome‐related diseases, known as ribosomopathies. This article is categorized under: Translation > Translation Mechanisms Translation > Translation Regulation

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Genome-Wide Approaches for RNA Structure Probing

Silverman

Berkowitz

Gosai

2016

Advances in Experimental Medicine and Biology

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RNA molecules of all types fold into complex secondary and tertiary structures that are important for their function and regulation. Structural and catalytic RNAs such as ribosomal RNA (rRNA) and transfer RNA (tRNA) are central players in protein synthesis, and only function through their proper folding into intricate three-dimensional structures. Studies of messenger RNA (mRNA) regulation have also revealed that structural elements embedded within these RNA species are important for the proper regulation of their total level in the transcriptome. More recently, the discovery of microRNAs (miRNAs) and long non-coding RNAs (lncRNAs) has shed light on the importance of RNA structure to genome, transcriptome, and proteome regulation. Due to the relatively small number, high conservation, and importance of structural and catalytic RNAs to all life, much early work in RNA structure analysis mapped out a detailed view of these molecules. Computational and physical methods were used in concert with enzymatic and chemical structure probing to create high-resolution models of these fundamental biological molecules. However, the recent expansion in our knowledge of the importance of RNA structure to coding and regulatory RNAs has left the field in need of faster and scalable methods for high-throughput structural analysis. To address this, nuclease and chemical RNA structure probing methodologies have been adapted for genome-wide analysis. These methods have been deployed to globally characterize thousands of RNA structures in a single experiment. Here, we review these experimental methodologies for high-throughput RNA structure determination and discuss the insights gained from each approach.

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The Kissing-Loop T-Shaped Structure Translational Enhancer of Pea Enation Mosaic Virus Can Bind Simultaneously to Ribosomes and a 5′ Proximal Hairpin

Cited by 33 publications

References 56 publications

The 3′ Untranslated Region of Pea Enation Mosaic Virus Contains Two T-Shaped, Ribosome-Binding, Cap-Independent Translation Enhancers

The 3′ Untranslated Region of Pea Enation Mosaic Virus Contains Two T-Shaped, Ribosome-Binding, Cap-Independent Translation Enhancers

Fatal attraction: The roles of ribosomal proteins in the viral life cycle

Genome-Wide Approaches for RNA Structure Probing

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