Coding Changes in the Poliovirus Protease 2A Compensate for 5′NCR Domain V Disruptions in a Cell-Specific Manner

Rowe, Alison; Ferguson, Geraldine; Minor, Philip D.; Macadam, Andrew

doi:10.1006/viro.2000.0244

Cited by 23 publications

(19 citation statements)

References 42 publications

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“…It had been suggested initially that the 2A protease may directly interact with the IRES element (27), but the internal location of some of the residues modified in the revertant viruses appears to make this unlikely. It has now been suggested that these mutations may disrupt a protein-protein interaction (32). Indeed, on the basis of our results, these substitutions, which served to overcome the deficit in translational efficiency imposed by the IRES mutation, may function by modifying the interaction of the PV 2A protease with some cellular component involved in the activation of IRES activity.…”

Section: Discussionmentioning

confidence: 52%

“…Two out of 10 such mutants analyzed had substitutions in residues adjacent to the catalytic triad, and the other substitutions were predicted to occur in two main clusters on the protease surface. Recently additional mutations in 2A that have this effect have been identified (32). It had been suggested initially that the 2A protease may directly interact with the IRES element (27), but the internal location of some of the residues modified in the revertant viruses appears to make this unlikely.…”

Section: Discussionmentioning

confidence: 99%

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An Attenuating Mutation in the 2A Protease of Swine Vesicular Disease Virus, a Picornavirus, Regulates Cap- and Internal Ribosome Entry Site-Dependent Protein Synthesis

Sakoda

Ross-Smith

Inoue

et al. 2001

J Virol

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Section: Discussionmentioning

confidence: 52%

Section: Discussionmentioning

confidence: 99%

An Attenuating Mutation in the 2A Protease of Swine Vesicular Disease Virus, a Picornavirus, Regulates Cap- and Internal Ribosome Entry Site-Dependent Protein Synthesis

Sakoda

Ross-Smith

Inoue

et al. 2001

J Virol

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“…Notably, one of the mutations which gave rise to reversions in 2A was the first nucleotide of the bulge under study here. Interestingly, the compensatory effect of 2A changes was found to be cell specific (33).…”

Section: Discussionmentioning

confidence: 96%

Poliovirus Internal Ribosome Entry Segment Structure Alterations That Specifically Affect Function in Neuronal Cells: Molecular Genetic Analysis

Malnou

Pöyry

Jackson

et al. 2002

J Virol

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Translation of poliovirus RNA is driven by an internal ribosome entry segment (IRES) present in the 5 noncoding region of the genomic RNA. This IRES is structured into several domains, including domain V, which contains a large lateral bulge-loop whose predicted secondary structure is unclear. The primary sequence of this bulge-loop is strongly conserved within enteroviruses and rhinoviruses: it encompasses two GNAA motifs which could participate in intrabulge base pairing or (in one case) could be presented as a GNRA tetraloop. We have begun to address the question of the significance of the sequence conservation observed among enterovirus reference strains and field isolates by using a comprehensive site-directed mutagenesis program targeted to these two GNAA motifs. Mutants were analyzed functionally in terms of (i) viability and growth kinetics in both HeLa and neuronal cell lines, (ii) structural analyses by biochemical probing of the RNA, and (iii) translation initiation efficiencies in vitro in rabbit reticulocyte lysates supplemented with HeLa or neuronal cell extracts. Phenotypic analyses showed that only viruses with both GNAA motifs destroyed were significantly affected in their growth capacities, which correlated with in vitro translation defects. The phenotypic defects were strongly exacerbated in neuronal cells, where a temperature-sensitive phenotype could be revealed at between 37 and 39.5°C. Biochemical probing of mutated domain V, compared to the wild type, demonstrated that such mutations lead to significant structural perturbations. Interestingly, revertant viruses possessed compensatory mutations which were distant from the primary mutations in terms of sequence and secondary structure, suggesting that intradomain tertiary interactions could exist within domain V of the IRES.Poliovirus (PV) is often considered the prototype of the Enterovirus genus of the Picornaviridae family. As such, its genome is a single-stranded RNA molecule of positive polarity. It is now well established that the long 5Ј untranslated region (5Ј-UTR; 742 nucleotides [nt] for PV type 1 Mahoney strain [PV1(M)]) plays different important roles in the PV life cycle. The first hundred nucleotides form a cloverleaf structure involved in RNA replication (3), acting at the initiation of negative-strand RNA synthesis (4). Similarly, on negativestrand replication intermediates, the complement of the 5Ј-UTR is involved in the synthesis of positive-strand viral RNA (32). Furthermore, the 5Ј-UTR contains an internal ribosome entry segment (IRES) which promotes translation initiation by a cap-and 5Ј-end-independent mechanism, following direct entry of the 40S ribosomal subunit at the 3Ј end of the IRES (for a review, see reference 6). The PV IRES is approximately 450 nt long and has a predicted secondary structure comprised of five complex stem-loops (Fig. 1A), one of which has been reported to be dispensable for efficient translation at least ex vivo (9). The original predicted secondary structure of the PV 5Ј-UTR was initially b...

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“…All these modifications observed in the 5 0 NTR and in the 2A gene, allow us to conclude that the virus has adapted to the cell, as it has been observed for the poliovirus [37]. In addition, a study of the impact of the modified 2A protein on the Dev cell line is necessary to complete our knowledge about the role of the mutated 2A pro in the viral persistence.…”

Section: Discussionmentioning

confidence: 85%

Cumulative Mutations in the Genome of Echovirus 6During Establishment of a Chronic Infection in Precursors of Glial Cells

Beaulieux¹,

Zreik²,

Deleage³

et al. 2005

Virus Genes

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Although Enteroviruses are mainly described as responsible for acute diseases, their role in severe chronic pathology has been also established. Echovirus 6-like sequences have been detected by PCR analysis in central nervous system specimens from patients presenting with Amyotrophic Lateral Sclerosis. These findings suggested a persistent infection with viruses that underwent, genetic changes precluding viral particle release. To support this hypothesis, we developed a model system of Echovirus 6 chronic infection in precursors of glial cells. The nucleotide sequences of the 5'non-translated region (5'NTR), 2A and 3C regions of the virus developing persistent genome were analysed during establishment of the chronic phenotype. This study revealed that at day 160 of chronic infection, several mutations were observed: one mutation at nucleotide 108 upstream the domain II of the internal ribosome entry site (IRES) structure, one mutation at nucleotide 30 in the cloverleaf, and two mutations in the 2A region (translated in His48 to Tyr, and Ile 123 to Met). No mutations were detected in the 3C region. The impact of these mutations on viral replication have been analysed in a rabbit reticulocyte lysate (RRL) translation assay supplemented with HeLa cell lysate, and by plaque assay. Viruses with these mutations displayed a phenotype with a significant reduction of replication, while in vitro translation was not affected by the nucleotide 108 mutation. This model allowed the description of molecular changes observed in the genome of Echovirus 6 during the establishment of a chronic infection phenotype, and may be helpful for the understanding of the mechanisms leading Enteroviruses to develop chronic infections in man.

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Coding Changes in the Poliovirus Protease 2A Compensate for 5′NCR Domain V Disruptions in a Cell-Specific Manner

Cited by 23 publications

References 42 publications

An Attenuating Mutation in the 2A Protease of Swine Vesicular Disease Virus, a Picornavirus, Regulates Cap- and Internal Ribosome Entry Site-Dependent Protein Synthesis

An Attenuating Mutation in the 2A Protease of Swine Vesicular Disease Virus, a Picornavirus, Regulates Cap- and Internal Ribosome Entry Site-Dependent Protein Synthesis

Poliovirus Internal Ribosome Entry Segment Structure Alterations That Specifically Affect Function in Neuronal Cells: Molecular Genetic Analysis

Cumulative Mutations in the Genome of Echovirus 6During Establishment of a Chronic Infection in Precursors of Glial Cells

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