Analyzing Coxsackievirus B3 Genomic RNA by Site Directed Mutagenesis and SHAPE Analysis
Abstract
Virulent Coxsackievirus B3 (CVB3) is linked to myocarditis, pancreatitis, and type I diabetes. Avirulent CVB3 doesn’t cause disease. CVB3 possesses a positive, single-stranded ribonucleic acid (RNA) genome containing 7,400 nucleotides, organized into four sections: a 5’ untranslated region (5’ UTR), a single open reading frame, a 3’ untranslated region, and a poly-A tail. The 5’ UTR is 743 nucleotides divided into 7 domains. Mutations within the 5’ UTR can alter RNA structure, and thus alter virulence. Nucleotide associations responsible for higher level RNA folding patterns and structure haven’t been defined. We are investigating the 5’ UTR by selective 2’ hydroxyl acylation analyzed by primer extension (SHAPE) to learn about the flexibility of the RNA backbone, thus determining the relationship between RNA structure and virulence. New data generated through SHAPE will be compared to known models of the CVB3 RNA genome. This will provide quantitative data regarding flexibility in the RNA structure. Previously, the virulent and avirulent strains were characterized by base-specific mutagenesis and the structure qualitatively analyzed. My work will determine the nucleotide associations responsible for virulence.