AIP inhibition of IRF7 and innate antiviral signaling

摘要 干扰素调节因子7(Irf7)是一种转录因子，被认为是I型干扰素的主要调节因子。 干扰素反应；然而，目前尚不清楚IRF7是如何被负调控以恢复免疫的 病毒感染解决后的体内平衡。我们已经鉴定出芳烃受体相互作用蛋白 (AIP)作为IRF7的负性调节因子，在抑制IRF7激活过程中起着至关重要的作用。AIP监管 芳香烃受体(AhR)是宿主异种生物反应的配体激活受体 环境毒素，如2，3，7，8-四氯二苯并对二恶英(TCD)。我们发现AIP与 IRF7，这种相互作用因病毒感染而进一步增强。AIP-/-小鼠胚胎成纤维细胞 与感染病毒的野生型细胞相比，产生更多的I型干扰素，从而使这些细胞 高度抵抗病毒感染。AIP拮抗IRF7的核定位以阻止转录 I型干扰素基因的激活。然而，关于AIP和AhR在 IRF7和先天免疫反应的调节。其特异性背后的分子机制是什么？ AIP对IRF7的调节与抗病毒天然免疫？鉴于AIP和AIP之间建立的功能链接 AHR，AhR抑制IRF7吗？AhR配体是否需要AIP来抑制抗病毒信号？在……里面 初步研究表明，AIP是一种新的非规范IKB激酶TBK1底物(TANK-1)。 结合激酶1)，并通过质谱学绘制了三个可能的磷酸化位点。此外， 用AhR激动剂L-犬尿氨酸(L-京)处理细胞可显著增强囊泡的复制 口腔炎病毒，提供证据表明AhR信号抑制固有的抗病毒反应。AHR与 IRF7提示IRF7可能是AhR先天免疫信号转导的靶点。吲哚胺2，3-双加氧酶 (IDO)是产生免疫调节色氨酸代谢物的限速酶，包括 AHR配体L-京。干扰素在病毒感染过程中诱导IDO，提示IDO参与了病毒感染的阴性反应 先天免疫信号的调节。推动这些调查的中心假设是AIP和AhR 作为一个功能单位通过两步负反馈抑制IRF7和病毒诱导的I型干扰素 Tbk1和干扰素轴的作用机制。我们将通过以下实验来验证我们的假设 具体目的：1)确定AIP磷酸化在抑制IRF7和天然信号转导中的作用；2) 确定AIP在病毒感染过程中对AhR的调节作用。完成这些研究可能 为环境因素如何影响对病毒感染的先天免疫提供新的机制洞察力 并且可以为使用AhR激动剂或拮抗剂作为微调宿主固有的方法提供理论基础 致病环境中的免疫反应，如自身免疫或病毒触发的细胞因子风暴(例如 甲型流感或SARS-CoV2/新冠肺炎)。

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).