No abstract

We previously reported that an ortholog of STING regulates infection by picorna-like viruses in Drosophila. In mammals, STING is activated by the cyclic dinucleotide 2′3′-cGAMP produced by cGAS, which acts as a receptor for cytosolic DNA. Here, we showed that injection of flies with 2′3′-cGAMP induced the expression of dSTING-regulated genes. Coinjection of 2′3′-cGAMP with a panel of RNA or DNA viruses resulted in substantially reduced viral replication. This 2′3′-cGAMP–mediated protection was still observed in flies with mutations in Atg7 and AGO2, genes that encode key components of the autophagy and small interfering RNA pathways, respectively. By contrast, this protection was abrogated in flies with mutations in the gene encoding the NF-κB transcription factor Relish. Transcriptomic analysis of 2′3′-cGAMP–injected flies revealed a complex response pattern in which genes were rapidly induced, induced after a delay, or induced in a sustained manner. Our results reveal that dSTING regulates an NF-κB–dependent antiviral program that predates the emergence of interferons in vertebrates.

28We recently reported that an orthologue of STING regulates infection by picorna-like 29 viruses in drosophila. In mammals, STING is activated by the cyclic dinucleotide 2'3'-30 cGAMP produced by cGAS, which acts as a receptor for cytosolic DNA. Here, we 31show that injection of flies with 2'3'-cGAMP can induce expression of dSTING-32 regulated genes. Co-injection of 2'3'-cGAMP with a panel of RNA or DNA viruses 33 results in significant reduction of viral replication. This 2'3'-cGAMP-mediated 34 protection is still observed in flies mutant for the genes Atg7 and AGO2, which 35encode key components of the autophagy and small interfering RNA pathways, 36 respectively. By contrast, it is abrogated in flies mutant for the NF-κB transcription 37 factor Relish. Analysis of the transcriptome of 2'3'-cGAMP injected flies reveals a 38 complex pattern of response, with early and late induced genes. Our results reveal 39 that dSTING regulates an NF-κB -dependent antiviral program, which predates the 40 emergence of Interferon Regulatory Factors and interferons in vertebrates. 41 42 43 44 45 46 47 48 49 50 51 52 Introduction 53Insects, like all animals, are plagued by viral infections, which they oppose 54 through their innate immune system. Induction of transcription of antiviral genes upon 55 sensing of infection is a common antiviral response observed across kingdoms. In 56 insects, inducible responses contribute to defense against viruses, together with RNA 57 interference (RNAi) and constitutively expressed restriction factors (reviewed in 1 ). 58Apart from RNAi, these mechanisms are still poorly characterized and appear to be 59 largely virus-specific 2-4 . Combining genetics and transcriptomic analysis, we recently 60 showed that the evolutionarily conserved factor dSTING participates together with the 61 kinase IKKβ and the NF-κB transcription factor Relish in a novel pathway controlling 62 infection by the picorna-like viruses Drosophila C virus (DCV) and Cricket Paralysis 63Virus (CrPV) in the model organism Drosophila melanogaster 5 . 64In mammals, STING is a central component of the mammalian cytosolic DNA 65 sensing pathway, where it acts downstream of the receptor cyclic GMP-AMP 66 synthase (cGAS) 6 . Upon binding DNA, cGAS synthesizes 2'3'-cGAMP, a cyclic 67 dinucleotide (CDN) secondary messenger that binds to and activates STING 7-10 . 68Bacteria also synthesize CDNs such as c-di-AMP, c-di-GMP and 3'3'-cGAMP, which 69 can be sensed by STING (reviewed in 11 ). Upon activation, STING recruits through its 70 C-terminal tail (CTT) region the kinase TBK1, which phosphorylates and activates the 71 transcription factor Interferon Regulatory Factor (IRF) 3 to trigger interferon (IFN) 72 production 12 . STING can also activate NF-κB and autophagy independently from its 73 CTT domain in mammalian cells [13][14][15] . 74The identification of STING in animals devoid of interferons, such as insects, 75 raises the question of the ancestral function of this molecule. Since most 76 invertebrates lack the IRF family of tran...