AIP inhibition of IRF7 and innate antiviral signaling

ABSTRACT Interferon regulatory factor 7 (IRF7) is a transcription factor known as the master regulator of the type I interferon (IFN) response; however, it remains unclear how IRF7 is negatively regulated to restore immune homeostasis after viral infections are resolved. We have identified aryl hydrocarbon receptor interacting protein (AIP) as a negative regulator of IRF7 that plays essential roles in the inhibition of IRF7 activation. AIP regulates the aryl hydrocarbon receptor (AhR), a ligand-activated receptor of the host xenobiotic response to environmental toxins such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCCD). We found that AIP interacts with IRF7 and this interaction is further enhanced by virus infection. Aip–/– murine embryonic fibroblasts (MEFs) produce significantly more type I IFN compared to wild-type cells infected with virus, thus rendering these cells highly resistant to virus infection. AIP antagonizes the nuclear localization of IRF7 to prevent the transcriptional activation of type I IFN genes. However, key questions remain regarding the roles of AIP and AhR in the regulation of IRF7 and innate immune responses. What is the molecular mechanism underlying specificity for AIP regulation of IRF7 and antiviral innate immunity? Given the established functional links between AIP and AhR, does AhR inhibit IRF7, and is AIP required for the inhibition of antiviral signaling by AhR ligands? In preliminary studies, we have identified AIP as a novel substrate of the noncanonical IkB kinase TBK1 (TANK- binding kinase 1), and three putative phosphorylation sites were mapped by mass spectrometry. Furthermore, treatment of cells with the AhR agonist L-kynurenine (L-Kyn) significantly enhanced the replication of vesicular stomatitis virus, providing evidence that AhR signaling inhibits the innate antiviral response. AhR interacts with IRF7 suggesting that IRF7 may be a target of AhR in innate immune signaling. Indoleamine 2,3-dioxygenase (IDO) is the rate-limiting enzyme in the generation of immune regulatory tryptophan metabolites, including the AhR ligand L-Kyn. IDO is induced by IFN during virus infection suggesting that IDO participates in the negative regulation of innate immune signaling. The central hypothesis driving these investigations is that AIP and AhR cooperate as a functional unit to inhibit IRF7 and virus-induced type I IFN by a two-step negative feedback mechanism instigated by TBK1 and the IFN axis. We will test our hypothesis experimentally with the following Specific Aims: 1) Define the role of AIP phosphorylation in the inhibition of IRF7 and innate signaling, and 2) Determine the role of AIP in the regulation of AhR during virus infection. Completion of these studies may provide new mechanistic insight into how environmental factors can influence innate immunity to virus infection and may provide rationale for the use of AhR agonists or antagonists as an approach to fine-tune host innate immune responses in pathogenic settings such as autoimmunity or virus-triggered cytokine storms (e.g. influenza A or SARS-CoV2/COVID-19).