STING specifies IRF3 phosphorylation by TBK1 in the cytosolic DNA signaling pathway. Yasuo Tanaka and Zhijian J. Chen investigated how STING activates IRF3 through TBK1. They used an in vitro reconstitution system to show that STING stimulates IRF3 phosphorylation by TBK1. They identified a carboxyl terminal region of STING that is necessary and sufficient for this activation. STING interacts with both TBK1 and IRF3, and mutations in STING that disrupt its binding to IRF3 abrogate IRF3 phosphorylation without impairing TBK1 activation. These results suggest that STING functions as a scaffold to specify and promote IRF3 phosphorylation by TBK1. The scaffolding function of STING and other adaptors may explain why IRF3 is activated only in a subset of signaling pathways that activate TBK1. STING activates IRF3 in an in vitro reconstitution system by binding to TBK1 and recruiting IRF3 to TBK1 to activate the IRF3 pathway. STING's C-terminus is important for IRF3 activation in vitro. STING directly activates TBK1 in vitro. Ser366 and Leu374 of STING are important for IRF3 activation. STING recruits IRF3 to TBK1 to induce IRF3 activation. STING is phosphorylated in response to stimulation by cytosolic DNA. STING's phosphorylation is dependent on TBK1. STING's C-terminal fragment is sufficient to support IRF3 phosphorylation by TBK1. STING's C-terminal fragment forms high molecular weight aggregates, which are necessary for IRF3 activation. STING's C-terminal fragment is required for IRF3 activation. STING's C-terminal fragment is necessary for IRF3 activation. STING's C-terminal fragment is sufficient to support IRF3 phosphorylation by TBK1. STING's C-terminal fragment is necessary for IRF3 activation. STING's C-terminal fragment is sufficient to support IRF3 phosphorylation by TBK1. STING's C-terminal fragment is necessary for IRF3 activation. STING's C-terminal fragment is sufficient to support IRF3 phosphorylation by TBK1. STING's C-terminal fragment is necessary for IRF3 activation. STING's C-terminal fragment is sufficient to support IRF3 phosphorylation by TBK1. STING's C-terminal fragment is necessary for IRF3 activation. STING's C-terminal fragment is sufficient to support IRF3 phosphorylation by TBK1. STING's C-terminal fragment is necessary for IRF3 activation. STING's C-terminal fragment is sufficient to support IRF3 phosphorylation by TBK1. STING's C-terminal fragment is necessary for IRF3 activation. STING's C-terminal fragment is sufficient toSTING specifies IRF3 phosphorylation by TBK1 in the cytosolic DNA signaling pathway. Yasuo Tanaka and Zhijian J. Chen investigated how STING activates IRF3 through TBK1. They used an in vitro reconstitution system to show that STING stimulates IRF3 phosphorylation by TBK1. They identified a carboxyl terminal region of STING that is necessary and sufficient for this activation. STING interacts with both TBK1 and IRF3, and mutations in STING that disrupt its binding to IRF3 abrogate IRF3 phosphorylation without impairing TBK1 activation. These results suggest that STING functions as a scaffold to specify and promote IRF3 phosphorylation by TBK1. The scaffolding function of STING and other adaptors may explain why IRF3 is activated only in a subset of signaling pathways that activate TBK1. STING activates IRF3 in an in vitro reconstitution system by binding to TBK1 and recruiting IRF3 to TBK1 to activate the IRF3 pathway. STING's C-terminus is important for IRF3 activation in vitro. STING directly activates TBK1 in vitro. Ser366 and Leu374 of STING are important for IRF3 activation. STING recruits IRF3 to TBK1 to induce IRF3 activation. STING is phosphorylated in response to stimulation by cytosolic DNA. STING's phosphorylation is dependent on TBK1. STING's C-terminal fragment is sufficient to support IRF3 phosphorylation by TBK1. STING's C-terminal fragment forms high molecular weight aggregates, which are necessary for IRF3 activation. STING's C-terminal fragment is required for IRF3 activation. STING's C-terminal fragment is necessary for IRF3 activation. STING's C-terminal fragment is sufficient to support IRF3 phosphorylation by TBK1. STING's C-terminal fragment is necessary for IRF3 activation. STING's C-terminal fragment is sufficient to support IRF3 phosphorylation by TBK1. STING's C-terminal fragment is necessary for IRF3 activation. STING's C-terminal fragment is sufficient to support IRF3 phosphorylation by TBK1. STING's C-terminal fragment is necessary for IRF3 activation. STING's C-terminal fragment is sufficient to support IRF3 phosphorylation by TBK1. STING's C-terminal fragment is necessary for IRF3 activation. STING's C-terminal fragment is sufficient to support IRF3 phosphorylation by TBK1. STING's C-terminal fragment is necessary for IRF3 activation. STING's C-terminal fragment is sufficient to support IRF3 phosphorylation by TBK1. STING's C-terminal fragment is necessary for IRF3 activation. STING's C-terminal fragment is sufficient to