Figure 5: The fusion-activated STING pathway is essential for full innate control of RNA viruses.
From: Influenza A virus targets a cGAS-independent STING pathway that controls enveloped RNA viruses
(a,b) Illustration of hSTING dimer (monomers shown in yellow and cyan) depicting the position of Arg169 relative to the position of cGAMP. (c–j) WT mSTING, R168A mSTING or eGFP were stably expressed in MEFs or BMDCs from Tmem173gt/gt mice. Scale bar, 5 μm. (c) Transduction efficiency was assessed in BMDCs by RT–qPCR analysis of measuring eGFP transcripts. (d) Laser-scanning microscopy image of mSTING expression in WT and R168A STING MEFs. (e) WT and R168A STING MEFs were stimulated with cGAMP and analysed for interferon-α/-β by RT–qPCR. (f) BMDCs were stimulated with fusogenic liposomes (Lipo, 2 μg ml−1), cGAMP (10 nM) or interferon-γ (1 ng ml−1). After 6 h mCxcl10 mRNA was analysed by RT–qPCR. (g) MEFs expressing either WT or R168A mSTING were infected overnight with VSV (MOI 0.01). Virus production was determined by end point titration. The western blot below the graph show the levels of STING and β-actin in the cells used for experiments. (h) BMDCs were pretreated with FP where indicated and infected with NDV. Supernatants were collected after 20 h and analysed for interferon-α/-β bioactivity. (i,j) MEFs expressing either WT or R168A mSTING were infected overnight with EMCV (MOI 0.01). Virus production was determined by end point titration. WT or Stinggt/gt cells infected with EMCV (MOI 0.1) for 6 h were lysed and total RNA was analysed for levels of interferon-β mRNA, normalized to β-actin and presented as normalized ratios relative to uninfected controls. (k) MEFs from C57BL/6 and STINGgt/gt mice were pretreated with 10 μM of FP and infected VSV (MOI 0.01). Supernatants were isolated 20 h later and viral titres were quantified by end point titration. The numerical data are presented as means of four biological replicates (±s.d.). *P<0.05 by Mann–Whitney compared-ranks test. IFN, interferon; HA, hemagglutinin. All graphs represent data from one of at least two independent experiments. b.d. indicates measurements below the detection limit.
