Optimized P2A for reporter gene insertion into Nipah virus results in efficient ribosomal skipping and wild-type lethality

The Journal of General Virology
Arnold ParkBenhur Lee

Abstract

Incorporation of reporter genes within virus genomes is an indispensable tool for interrogation of virus biology and pathogenesis. In previous work, we incorporated a fluorophore into a viral ORF by attaching it to the viral gene via a P2A ribosomal skipping sequence. This recombinant Nipah virus, however, was attenuated in vitro relative to WT virus. In this work, we determined that inefficient ribosomal skipping was a major contributing factor to this attenuation. Inserting a GSG linker before the P2A sequence resulted in essentially complete skipping, significantly improved growth in vitro, and WT lethality in vivo. To the best of our knowledge, this represents the first time a recombinant virus of Mononegavirales with integration of a reporter into a viral ORF has been compared with the WT virus in vivo. Incorporating the GSG linker for improved skipping efficiency whenever functionally important is a critical consideration for recombinant virus design.

References

Aug 10, 2000·Molecular Therapy : the Journal of the American Society of Gene Therapy·H MizuguchiT Hayakawa
Feb 21, 2006·Nature Methods·Jeff HolstDario A A Vignali
Oct 21, 2006·Proceedings of the National Academy of Sciences of the United States of America·Misako YonedaChieko Kai
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Apr 1, 2014·Antiviral Research·Michael K LoChristina F Spiropoulou

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Citations

Sep 9, 2016·Molecular Therapy. Methods & Clinical Development·Arnold ParkBenhur Lee
Mar 10, 2018·PLoS Neglected Tropical Diseases·Stephen R WelchCésar G Albariño
Oct 31, 2019·The Journal of Infectious Diseases·Stephen R WelchJessica R Spengler
May 17, 2018·Scientific Reports·Brian E DawesAlexander N Freiberg

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