A Dual Promoter System to Monitor IFN-γ Signaling in vivo at Single-cell Resolution

Cell Struct Funct. 2021 Dec 22;46(2):103-111. doi: 10.1247/csf.21052. Epub 2021 Nov 6.


IFN-γ secreted from immune cells exerts pleiotropic effects on tumor cells, including induction of immune checkpoint and antigen presentation, growth inhibition, and apoptosis induction. We combined a dual promoter system with an IFN-γ signaling responsive promoter to generate a reporter named the interferon sensing probe (ISP), which quantitates the response to IFN-γ by means of fluorescence and bioluminescence. The integration site effect of the transgene is compensated for by the PGK promoter-driven expression of a fluorescent protein. Among five potential IFN-γ-responsive elements, we found that the interferon γ-activated sequence (GAS) exhibited the best performance. When ISP-GAS was introduced into four cell lines and subjected to IFN-γ stimulation, dose-dependency was observed with an EC50 ranging from 0.2 to 0.9 ng/mL, indicating that ISP-GAS can be generally used as a sensitive biosensor of IFN-γ response. In a syngeneic transplantation model, the ISP-GAS-expressing cancer cells exhibited bioluminescence and fluorescence signals in an IFN-γ receptor-dependent manner. Thus, ISP-GAS could be used to quantitatively monitor the IFN-γ response both in vitro and in vivo.Key words: in vivo imaging, tumor microenvironment, interferon-gamma, dual promoter system.

Keywords: dual promoter system; in vivo imaging; interferon-gamma; tumor microenvironment.

MeSH terms

  • Interferon-gamma* / genetics
  • Promoter Regions, Genetic / genetics
  • RNA, Messenger
  • Signal Transduction
  • Transcription, Genetic*


  • RNA, Messenger
  • Interferon-gamma