Figure 6.

Effects of IFI16 and cGAS KO on HSV-1 stimulated cGAMP production and cGAMP stimulated and STING activated gene transcription. (A) ISRE reporter cells in a 24-well plate (2 × 10⁵ cells/well) were transfected with 0.5 μg of pEGFP-pSTING, and 24 h later, the transfected cells were collected by trypsin digestion and cell number was calculated. Simultaneously, PK15 cGAS KO, IFI16 KO, and control stable cells in a 24-well plate (3 × 10⁵ cells/well) were stimulated with HSV-1 at the indicated MOIs for 12 h, then the stimulated cells harvested and cell number calculated. Next, 0.3 × 10⁵ pSTING transfected ISRE reporter cells were incubated with 1.2 × 10⁵ of each type of PK15 stable cells (1:4) in a 24-well plate for 12 h. The incubated pool cells were examined for Fluc and Rluc activities. (B–D) The PK15 cGAS KO, IFI16 KO, and control stable cells in a 24-well plate (3 × 10⁵ cells/well) were stimulated with 10 μg/ml 2′3′-cGAMP by transfection for 24 h, and the stimulated PK15 cells were examined for downstream gene transcription by RT-qPCR. NS denotes mock stimulation. ^*p < 0.05, ^**p < 0.01, ^***p < 0.001, ns, not significant.