Induction of vast transcriptional programs is a central event of innate host responses to viral infections. Here we report a transcriptional program with potent antiviral activity, driven by E74-like ETS transcription factor 1 (ELF1). Using microscopy to quantify viral infection over time, we found that ELF1 inhibits eight diverse RNA and DNA viruses after multi-cycle replication. Elf1 deficiency results in enhanced susceptibility to influenza A virus infections in mice. ELF1 does not feed-forward to induce interferons, and ELF1’s antiviral effect is not abolished by the absence of STAT1 or by inhibition of JAK phosphorylation. Accordingly, comparative expression analyses by RNA-seq revealed that the ELF1 transcriptional program is distinct from interferon signatures. Thus, ELF1 provides an additional layer of the innate host response, independent from the action of type I interferons.

31 32 Keywords: ETS transcription factor, interferon, RNA virus, DNA virus, innate immunity 33 34 35 Seifert and Si et al., page 3 36 ABSTRACT 37The transcription of interferon-stimulated genes (ISGs) is classically triggered via 38 activation of the JAK-STAT pathway, and together, ISGs raise a multifaceted antiviral 39 barrier. An increasing body of evidence reports the existence of additional, non-canonical 40 pathways and transcription factors that coordinate ISG expression. Detailed knowledge 41 of how heterogenous mechanisms regulate ISG expression is crucial for the rational 42 design of drugs targeting the type I interferon response. Here, we characterize the first 43 ETS transcription factor family member as a regulator of non-canonical ISG expression:44 E74-like ETS transcription factor 1 (ELF1). Using high-content microscopy to quantify viral 45 infection over time, we found that ELF1, itself an ISG, inhibits eight diverse RNA and DNA 46 viruses uniquely at multi-cycle replication. ELF1 did not regulate expression of type I or II 47 interferons, and ELF1's antiviral effect was not abolished by the absence of STAT1 or by 48 inhibition of JAK phosphorylation. Accordingly, comparative expression analyses by 49 RNAseq revealed that the ELF1 transcriptional program is distinct from, and delayed with 50 respect to, the immediate interferon response. Finally, knockdown experiments 51 demonstrated that ELF1 is a critical component of the antiviral interferon response in vitro 52 and in vivo. Our findings reveal a previously overlooked mechanism of non-canonical ISG 53 regulation that both amplifies and prolongs the initial interferon response by expressing 54 broadly antiviral restriction factors. 55 Seifert and Si et al., page 456 AUTHOR SUMMARY 57 Over 60 years after their discovery, we still struggle to understand exactly how interferons 58 inhibit viruses. Our gap in knowledge stems, on one hand, from the sheer number of 59 interferon-stimulated effector genes, of which only few have been characterized in 60 mechanistic detail. On the other hand, our knowledge of interferon-regulated gene 61 transcription is constantly evolving. We know that different regulatory mechanisms greatly 62 influence the quality, magnitude, and timing of interferon-stimulated gene expression, all 63 of which may contribute to the antiviral mechanism of interferons. Deciphering these 64 regulatory mechanisms is indispensable for understanding this critical first line of host 65 defense, and for harnessing the power of interferons in novel antiviral therapies. Here, 66 we report a novel mechanism of interferon-induced gene regulation by an interferon-67 stimulated gene, which, paradoxically, inhibits viruses in the absence of additional 68 interferon signaling: E74-like ETS transcription factor 1 (ELF1) raises an unusually 69 delayed antiviral program that potently restricts propagation of all viruses tested in our 70 study. Reduced levels of ELF1 significantly diminished interferon-mediated host defenses 71 against influenza A viru...