Figure 5. Vaccina virus triggers STING-dependent antiviral immunity in bystander cells.

a, b, HEK cGAS^low cells or HEK cells were infected with MVA–GFP, washed and then loaded onto HEK STING cells for 8 h. a, Confocal microscopy of HEK STING cells co-incubated with MVA-infected HEK cGAS cells (arrows, STING activation in bystander cells). b, Quantification of STING activation in response to MVA-infected HEK cGAS^low cells or MVA-infected HEK cells. c, HEK cells or HEK cGAS^low cells were MVA-infected or left untreated, added onto HEK cells or HEK STING cells and studied for IRF3 phosphorylation (viral particles per ml: ++ = 3.2 × 10⁷; + = 1.6 × 10⁷). d, e, Experiments as in a in the presence or absence of CBX 150 μM (d) or using HEK STING CX43/45^WT and HEK STING CX43/45^DKO cells as responder cells (e). f, HEK cGAS* cells were co-incubated with MEFs and after 14 h mouse Ifnb mRNA, mouse Cxcl10 mRNA and mouse Irf7 mRNA were assessed by qPCR. g, h, HEK cGAS* cells were co-incubated with MEFs and after 12 h vaccinia virus was added (multiplicity of infection (m.o.i.): 2-0.5). Twenty-four hours later cell survival was analysed. Visual fields of one representative experiment are depicted (g, right panel) and mean and s.e.m. of three independent experiments are summarized (h). VV, vaccinia virus. One representative experiment out of two (a, b, e) or three (c, d) independent experiments are shown or mean and s.e.m. of n = 5 independent experiments is presented (f). *P < 0.05, **P < 0.01, ***P < 0.001.