Mouse STING-KI Dual Reporter THP-1 Cells

Murine STING knockin dual reporter monocytes

THP1-Dual™ KI-mSTING cells stably express the wild-type murine STING (stimulator of interferon genes, mSTING also known as TMEM173) gene. They were generated by knockin (KI) of the mSTING intronless coding sequence (from ATG to TGA) into THP1-Dual™ KO-STING cells, which derive from the human THP-1 monocytes.

STING is a cytosolic pattern recognition receptor (PRR) that acts as a direct sensor of cyclic dinucleotides (CDNs) [1]. The binding of CDNs to STING leads to TANK-binding-kinase-I (TBK1)-mediated interferon regulatory factor (IRF3) activation and type I interferon (IFN) production [1] and to NF-κB-dependent inflammatory cytokine production [2].

THP1-Dual™ KI-mSTING cells feature two reporter genes allowing the simultaneous study of the IRF pathway, by monitoring the activity of an inducible secreted Lucia luciferase, and the NF-κB pathway by monitoring the activity of an inducible SEAP (secreted embryonic alkaline phosphatase). Lucia luciferase and SEAP activities are readily assessable in the supernatant using QUANTI-Luc™ 4 Lucia/Gaussia and QUANTI-Blue™ Solution detection reagents, respectively.

As expected, THP1-Dual™ KI-mSTING cells exhibit a robust response to STING agonists such as 2’3’-cGAMP and 3’3’-cGAMP. In addition, these cells respond to DMXAA (5,6-Dimethylxanthenone 4-acetic acid), a potent activator of murine STING-dependent IFN responses, that does not activate human STING [3]. These cells are resistant to Blasticidin and Zeocin™.

Features of THP1-Dual™ KI-mSTING Cells:

• Knockin of the mSTING gene
• No functional hSTING expression
• IRF activation readily assessable using the Lucia luciferase reporter gene
• NF-κB activation readily assessable using the SEAP reporter gene

Validation of THP1-Dual™ KI-mSTING Cells:

• Murine STING knockin verified by PCR, DNA sequencing, and Western blot
• Functionally tested

References:

 1. Sun L. et al., 2013. Cyclic GMP-AMP synthase is a cytosolic DNA sensor that activates the type I interferon pathway. Science. 339:786-91.
2. Ishikawa H. & Barber G.N. 2008. STING is an endoplasmic reticulum adaptor that facilitates innate immune signalling. Nature 455:674-8.
3. Zhang H. et al., 2015. Rat and human STINGs profile similarly towards anticancer/antiviral compounds. Sci Rep. 5:18035.