Cyclic dinucleotide stimulation of Nematostella vectensis

In mammals, the cGAS-cGAMP-STING pathway is crucial for sensing viral infection and initiating an anti-viral type I interferon response. cGAS and STING are highly conserved genes that originated in bacteria and are present in most animals. By contrast, interferons only emerged in vertebrates; thus, the function of STING in invertebrates is unclear. Here, we use the STING ligand 2'3'-cGAMP to activate immune responses in a model cnidarian invertebrate, the starlet sea anemone Nematostella vectensis. Using RNA-Seq, we found that 2'3'-cGAMP induces robust transcription of both anti-viral and anti-bacterial genes, including the conserved transcription factor NF-?B. Knockdown experiments identified a role for NF-?B in specifically inducing anti-bacterial genes downstream of 2'3'-cGAMP, and some of these genes were also found to be induced during Pseudomonas aeruginosa infection. Furthermore, we characterized the protein product of one of the putative anti-bacterial genes, the N. vectensis homolog of Dae4, and found that it has conserved anti-bacterial activity. This work describes an unexpected role of a cGAMP sensing pathway in anti-bacterial immunity and suggests that a broad transcriptional response is an evolutionarily ancestral output of 2'3'-cGAMP signaling in animals. Overall design: Examine the response of N. vectensis polyps with cyclic dinucleotides (untreated, c-di-AMP, c-di-GMP, 2'3'-cGAMP, 3'3'-cGAMP). To determine factors required for the response to 2'3'-cGAMP, microinjected N. vectensis embryos with shRNAs targeting GFP (control), nvSTING, nvSTAT, and nvNF-kB and treated with 2'3'-cGAMP.