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A codon-shuffling method to prevent reversion during production of replication-defective herpesvirus stocks: Implications for herpesvirus vaccines

Herpesviruses establish life-long chronic infections that place infected hosts at risk for severe disease. Herpesvirus genomes readily undergo homologous recombination (HR) during productive replication, often leading to wild-type (WT) reversion during complementation of replication-defective and at...

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Autores principales: Li, Gang, Ward, Charles, Yeasmin, Rukhsana, Skiena, Steven, Krug, Laurie T., Forrest, J. Craig
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5347388/
https://www.ncbi.nlm.nih.gov/pubmed/28287622
http://dx.doi.org/10.1038/srep44404
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author Li, Gang
Ward, Charles
Yeasmin, Rukhsana
Skiena, Steven
Krug, Laurie T.
Forrest, J. Craig
author_facet Li, Gang
Ward, Charles
Yeasmin, Rukhsana
Skiena, Steven
Krug, Laurie T.
Forrest, J. Craig
author_sort Li, Gang
collection PubMed
description Herpesviruses establish life-long chronic infections that place infected hosts at risk for severe disease. Herpesvirus genomes readily undergo homologous recombination (HR) during productive replication, often leading to wild-type (WT) reversion during complementation of replication-defective and attenuated viruses via HR with the helper gene provided in trans. To overcome this barrier, we developed a synthetic-biology approach based on a technique known as codon shuffling. Computer-assisted algorithms redistribute codons in a helper gene, thereby eliminating regions of homology, while enabling manipulation of factors such as codon-pair bias and CpG content to effectively titrate helper-gene protein levels. We apply this technique to rescue the replication of a murine gammaherpesvirus engineered with a mutation in the major immediate-early transactivator protein RTA. Complementation with codon-shuffled RTA constructs did not yield any WT revertant virus, a sharp contrast to WT virus contamination frequently observed during complementation with an unmodified helper gene. We further demonstrate the importance of eliminating WT virus contamination in an animal model of gammaherpesvirus lethality. We propose complementation by codon shuffling as a means to produce replication-defective or attenuated viruses. This method has immediate utility for investigating roles of essential genes in viral replication and will better enable future development of herpesvirus vaccines.
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spelling pubmed-53473882017-03-14 A codon-shuffling method to prevent reversion during production of replication-defective herpesvirus stocks: Implications for herpesvirus vaccines Li, Gang Ward, Charles Yeasmin, Rukhsana Skiena, Steven Krug, Laurie T. Forrest, J. Craig Sci Rep Article Herpesviruses establish life-long chronic infections that place infected hosts at risk for severe disease. Herpesvirus genomes readily undergo homologous recombination (HR) during productive replication, often leading to wild-type (WT) reversion during complementation of replication-defective and attenuated viruses via HR with the helper gene provided in trans. To overcome this barrier, we developed a synthetic-biology approach based on a technique known as codon shuffling. Computer-assisted algorithms redistribute codons in a helper gene, thereby eliminating regions of homology, while enabling manipulation of factors such as codon-pair bias and CpG content to effectively titrate helper-gene protein levels. We apply this technique to rescue the replication of a murine gammaherpesvirus engineered with a mutation in the major immediate-early transactivator protein RTA. Complementation with codon-shuffled RTA constructs did not yield any WT revertant virus, a sharp contrast to WT virus contamination frequently observed during complementation with an unmodified helper gene. We further demonstrate the importance of eliminating WT virus contamination in an animal model of gammaherpesvirus lethality. We propose complementation by codon shuffling as a means to produce replication-defective or attenuated viruses. This method has immediate utility for investigating roles of essential genes in viral replication and will better enable future development of herpesvirus vaccines. Nature Publishing Group 2017-03-13 /pmc/articles/PMC5347388/ /pubmed/28287622 http://dx.doi.org/10.1038/srep44404 Text en Copyright © 2017, The Author(s) http://creativecommons.org/licenses/by/4.0/ This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/
spellingShingle Article
Li, Gang
Ward, Charles
Yeasmin, Rukhsana
Skiena, Steven
Krug, Laurie T.
Forrest, J. Craig
A codon-shuffling method to prevent reversion during production of replication-defective herpesvirus stocks: Implications for herpesvirus vaccines
title A codon-shuffling method to prevent reversion during production of replication-defective herpesvirus stocks: Implications for herpesvirus vaccines
title_full A codon-shuffling method to prevent reversion during production of replication-defective herpesvirus stocks: Implications for herpesvirus vaccines
title_fullStr A codon-shuffling method to prevent reversion during production of replication-defective herpesvirus stocks: Implications for herpesvirus vaccines
title_full_unstemmed A codon-shuffling method to prevent reversion during production of replication-defective herpesvirus stocks: Implications for herpesvirus vaccines
title_short A codon-shuffling method to prevent reversion during production of replication-defective herpesvirus stocks: Implications for herpesvirus vaccines
title_sort codon-shuffling method to prevent reversion during production of replication-defective herpesvirus stocks: implications for herpesvirus vaccines
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5347388/
https://www.ncbi.nlm.nih.gov/pubmed/28287622
http://dx.doi.org/10.1038/srep44404
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