Abstract
Influenza A virus is a significant public health threat, but little is understood about the viral RNA structure and function. Current vaccines and therapeutic options to control influenza A virus infections are mostly protein-centric and of limited effectiveness. Here, we report using an ensemble defect approach to design mutations to misfold regions of conserved mRNA structures in influenza A virus segments 7 and 8. Influenza A mutant viruses inhibit pre-mRNA splicing and attenuate viral replication in cell culture, thus providing evidence for functions of the targeted regions. Targeting these influenza A viral RNA regions provides new possibilities for designing vaccines and therapeutics against this important human respiratory pathogen. The results also demonstrate that the ensemble defect approach is an efficient way to test for function of RNA sequences.
Highlights
Influenza A virus is a segmented, single-stranded, negative sense RNA virus [1]
Little is known about the biological function of RNA structure in influenza A virus [37], and no current vaccines or therapeutics are designed to target viral RNA structures
Conserved RNA secondary structures throughout protein coding regions of influenza A mRNAs were predicted by bioinformatics using a combination of thermodynamics and comparative sequence analysis, including suppression of synonymous codon usage [18, 19]
Summary
Influenza A virus is a segmented, single-stranded, negative sense RNA virus [1]. Every year, 15–20% of the world’s population is infected, and 250 to 500 thousand people are killed by influenza A virus [2]. Structure targeted mutations in mRNA can potentially be used as a new approach to develop weakened virus for LAIV. Two classes of therapeutics have been approved by the Food and Drug Administration (FDA) for use against influenza: adamantanes and neuraminidase inhibitors [13]. Both target essential proteins coded by influenza, the ion channel protein M2, and the neuraminidase protein, respectively [14]. Conserved viral RNA structures important for function are potentially new therapeutic targets. Based largely on sequence comparison and predicted thermodynamics, conserved RNA structures were identified near or containing splice sites in segments 7 and 8 mRNAs [18, 19]. The results demonstrate that the ensemble defect approach can rapidly reveal function for regions predicted to fold into stable secondary structures
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