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Trans-dominant inhibition of RNA viral replication can slow growth of drug-resistant viruses

Abstract

The high error rates of viral RNA-dependent RNA polymerases create heterogeneous viral populations whose disparate RNA genomes affect each other's survival. We systematically screened the poliovirus genome and identified four sets of dominant mutations. Mutated alleles in capsid- and polymerase-coding regions resulted in dominant negative phenotypes, probably due to the proteins' oligomeric properties. We also identified dominant mutations in an RNA element required for priming RNA synthesis (CRE) and in the protein primer (VPg), suggesting that nonproductive priming intermediates are inhibitory. Mutations that inhibit the activity of viral proteinase 2A were dominant, arguing that inhibition of its known intramolecular activity creates a toxic product. Viral products that, when defective, dominantly interfere with growth of nondefective viruses will probably be excellent drug targets because drug-sensitive viruses should be dominant over drug-resistant variants. Accordingly, a virus sensitive to anticapsid compound WIN51711 dominantly inhibited the intracellular growth of a drug-resistant virus. Therefore, dominant inhibitor screening should validate or predict targets for antiviral therapy with reduced risk for drug resistance.

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Figure 1: Dominant inhibitor screen for capsid-coding genome regions.
Figure 2: Effect of mutations in 3D polymerase on the yield of wild-type virus during cotransfection.
Figure 3: Dominant inhibitor screen in the CRE- and 3B-coding regions.
Figure 4: Dominant inhibitor screen for 2A proteinase and VP1-2A cleavage site mutations.
Figure 5: RNA replication or translation requirements for dominance of mutated poliovirus alleles.
Figure 6: Superinfections of wild-type and temperature-sensitive polioviruses.
Figure 7: Coinfections of drug-sensitive and drug-resistant viruses.

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Acknowledgements

We thank S. Dutcher, P. Sarnow and J. Theriot for reading the manuscript; P. Harbury for editing; J.M. Lyle for experimental insights; and A. Mosser and A. Macadam for advice and reagents. This work was supported by the US National Institutes of Health (National Research Service Award to S.C.), the Hutchison Program in Translational Medicine and the Ellison Medical Foundation.

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Correspondence to Karla Kirkegaard.

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Supplementary information

Supplementary Table 1

Phenotypes of temperature-sensitive polioviruses generated by hydrophobic mutations. (PDF 2509 kb)

Supplementary Table 2

Dominant inhibitor screen results of mutant 2B, 2C, 3A, and 3C alleles. (PDF 4946 kb)

Supplementary Table 3

Summary of dominant negative alleles of poliovirus. (PDF 386 kb)

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Crowder, S., Kirkegaard, K. Trans-dominant inhibition of RNA viral replication can slow growth of drug-resistant viruses. Nat Genet 37, 701–709 (2005). https://doi.org/10.1038/ng1583

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