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Identification of a cis-acting replication element within the poliovirus coding region
I Goodfellow1, Y Chaudhry, A Richardson
1Division of Virology, Institute of Biomedical and Life Sciences, University of Glasgow, Glasgow, United Kingdom.
Journal of Virology
|April 25, 2000
Summary
Researchers identified a novel RNA secondary structure in poliovirus essential for replication. This structure acts as a cis-acting replication element (CRE), crucial for synthesizing the negative-strand template required for viral genome copying.
Area of Science:
- Virology
- Molecular Biology
- RNA Biology
Background:
- Poliovirus, a positive-stranded RNA virus, requires specific RNA sequences for replication.
- The synthesis of the negative-stranded RNA template is a critical step.
- RNA secondary structures are known to play roles in picornavirus replication.
Purpose of the Study:
- To identify and characterize novel cis-acting replication elements (CREs) in the poliovirus genome.
- To investigate the role of RNA secondary structures in poliovirus replication.
Main Methods:
- Bioinformatic prediction of local RNA secondary structures within the poliovirus genome.
- Site-directed mutagenesis to disrupt predicted stem-loop structures.
- Analysis of viral genome viability and replication kinetics.
- Reversion analysis to assess the importance of structural integrity.
- Subgenomic replicon assays to confirm functional activity.
Main Results:
- A conserved 61-nucleotide stem-loop structure was identified in the 2C protein-coding region.
- Disruption of this stem-loop abolished poliovirus genome viability and negative-strand synthesis.
- Mutations were critical for function in the positive sense, and structural integrity was essential.
- Replication competence was restored in subgenomic replicons by adding a second copy of the wild-type sequence.
Conclusions:
- A novel poliovirus cis-acting replication element (CRE) has been identified, comprising an RNA secondary structure.
- This CRE is essential for the synthesis of the negative-strand template during viral replication.
- The identified poliovirus CRE shares functional similarities but differs in sequence, structure, and location from elements in other picornaviruses.