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Journal of Virology, April 2006, p. 3259-3272, Vol. 80, No. 7
0022-538X/06/$08.00+0     doi:10.1128/JVI.80.7.3259-3272.2006

Modulation of Poliovirus Replicative Fitness in HeLa Cells by Deoptimization of Synonymous Codon Usage in the Capsid Region

Cara Carthel Burns,* Jing Shaw, Ray Campagnoli, Jaume Jorba, Annelet Vincent, Jacqueline Quay,{dagger} and Olen Kew

Division of Viral and Rickettsial Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia 30333

Received 20 September 2005/ Accepted 2 January 2006

We replaced degenerate codons for nine amino acids within the capsid region of the Sabin type 2 oral poliovirus vaccine strain with corresponding nonpreferred synonymous codons. Codon replacements were introduced into four contiguous intervals spanning 97% of the capsid region. In the capsid region of the most highly modified virus construct, the effective number of codons used (NC) fell from 56.2 to 29.8, the number of CG dinucleotides rose from 97 to 302, and the G+C content increased from 48.4% to 56.4%. Replicative fitness in HeLa cells, measured by plaque areas and virus yields in single-step growth experiments, decreased in proportion to the number of replacement codons. Plaque areas decreased over an ~10-fold range, and virus yields decreased over an ~65-fold range. Perhaps unexpectedly, the synthesis and processing of viral proteins appeared to be largely unaltered by the restriction in codon usage. In contrast, total yields of viral RNA in infected cells were reduced ~3-fold and specific infectivities of purified virions (measured by particle/PFU ratios) decreased ~18-fold in the most highly modified virus. The replicative fitness of both codon replacement viruses and unmodified viruses increased with the passage number in HeLa cells. After 25 serial passages (~50 replication cycles), most codon replacements were retained, and the relative fitness of the modified viruses remained well below that of the unmodified virus. The increased replicative fitness of high-passage modified virus was associated with the elimination of several CG dinucleotides. Potential applications for the systematic modulation of poliovirus replicative fitness by deoptimization of codon usage are discussed.

* Corresponding author. Mailing address: Respiratory and Enteric Viruses Branch, G-10, Division of Viral and Rickettsial Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, 1600 Clifton Rd., N.E., Atlanta, GA 30333. Phone: (404) 639-5499. Fax: (404) 639-4011. E-mail: cburns{at}cdc.gov.

{dagger} Present address: Office of Technology Development, The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599.

Journal of Virology, April 2006, p. 3259-3272, Vol. 80, No. 7
0022-538X/06/$08.00+0     doi:10.1128/JVI.80.7.3259-3272.2006




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