Reversible Gene Regulation in Mammalian Cells Using Riboswitch-Engineered Vesicular Stomatitis Virus Vector

ACS Synth Biol. 2019 Sep 20;8(9):1976-1982. doi: 10.1021/acssynbio.9b00177. Epub 2019 Aug 15.


Synthetic riboswitches based on small molecule-responsive self-cleaving ribozymes (aptazymes) embedded in the untranslated regions (UTRs) allow chemical control of gene expression in mammalian cells. In this work, we used a guanine-responsive aptazyme to control transgene expression from a replication-incompetent vesicular stomatitis virus (VSV) vector. VSV is a nonsegmented, negative-sense, cytoplasmic RNA virus that replicates without DNA intermediates, and its applications for vaccines and oncolytic viral therapy are being explored. By inserting the guanine-activated ribozyme in the 3' UTRs of viral genes and transgenes, GFP expression from the VSV vector in mammalian cells was repressed by as much as 26.8-fold in the presence of guanine. Furthermore, we demonstrated reversible regulation of a transgene (secreted NanoLuc) by adding and withdrawing guanine from the medium over the course of 12 days. In summary, our riboswitch-controlled VSV vector allows robust, long-term, and reversible regulation of gene expression in mammalian cells without the risk of undesirable genomic integration.

Keywords: RNA replicon; RNA virus; aptazyme; riboswitch.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 3' Untranslated Regions
  • Animals
  • Cell Line
  • Cricetinae
  • Gene Expression Regulation
  • Genetic Vectors / genetics
  • Genetic Vectors / metabolism
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • Guanine / metabolism
  • Metabolic Engineering / methods*
  • RNA, Catalytic / genetics
  • RNA, Catalytic / metabolism
  • Riboswitch / genetics*
  • Vesiculovirus / genetics*


  • 3' Untranslated Regions
  • RNA, Catalytic
  • Riboswitch
  • Green Fluorescent Proteins
  • Guanine