STING is essential for TLR8 induced IFN

The innate immune system is equipped with multiple receptors to detect microbial nucleic acids and induce type I interferon (IFN) to restrict viral replication. When dysregulated these receptor pathways induce inflammation in response to host nucleic acids and promote development and persistence of autoimmune diseases like Systemic Lupus Erythematosus (SLE). IFN production is regulated by the Interferon Regulatory Factor (IRF) transcription factor family of proteins that function downstream of several innate immune receptors such as Toll-like receptors (TLRs) and Stimulator of Interferon Genes (STING). Although both TLRs and STING activate the same downstream molecules, the pathway by which TLRs and STING activate IFN response are thought to be independent. Here we show that STING plays a previously undescribed role in human TLR8 signaling. Stimulation with the TLR8 ligands induced IFN secretion in primary human monocytes, and inhibition of STING reduced IFN secretion from primary monocytes from 8 healthy donors. We demonstrate that TLR8-induced IRF activity was reduced by STING inhibitors. Moreover, TLR8-induced IRF activity was blocked by inhibition or loss of IKKe, but not TBK1. Bulk RNA transcriptomic analysis supported a model where TLR8 induces transcriptional responses associated with SLE that can be downregulated by inhibition of STING. These data demonstrate that STING is required for full TLR8-to-IRF signaling and provide evidence for a new framework of crosstalk between cytosolic and endosomal innate immune receptors, which could be leveraged to treat IFN driven autoimmune diseases. Overall design: 2x10^6 THP-1 WT or STING cells were treated with DMSO or 500ng/mL H151 for 1 hour, then stimulated with media or 10ug/mL TL8 for 8 hours. After 8 hours RNA was collected using trizol.

固有免疫系统（innate immune system）配备多种受体以识别微生物核酸，并诱导I型干扰素（type I interferon, IFN）以限制病毒复制。当此类受体通路失调时，会响应宿主核酸引发炎症，并促进系统性红斑狼疮（Systemic Lupus Erythematosus, SLE）等自身免疫疾病的发生与持续进展。干扰素的产生受干扰素调节因子（Interferon Regulatory Factor, IRF）蛋白转录因子家族调控，该家族在Toll样受体（Toll-like receptors, TLRs）、干扰素基因刺激因子（Stimulator of Interferon Genes, STING）等多种固有免疫受体的下游发挥功能。尽管TLRs与STING均可激活相同的下游分子，但学界普遍认为二者激活干扰素应答的通路相互独立。本研究揭示了STING在人类TLR8信号通路中此前未被报道的功能。使用TLR8配体刺激可诱导原代人单核细胞分泌干扰素，而抑制STING则可降低8名健康供体的原代人单核细胞的干扰素分泌量。我们证实，STING抑制剂可降低TLR8诱导的IRF活性。此外，TLR8诱导的IRF活性可通过IKKe的抑制或缺失阻断，但不受TBK1影响。批量RNA转录组学分析支持如下模型：TLR8诱导的转录应答与SLE相关，且可通过抑制STING得以下调。上述数据表明，STING是完整的TLR8-IRF信号通路所必需的，并为胞质与内体固有免疫受体间的全新串扰框架提供了证据，该框架可被用于治疗干扰素驱动的自身免疫疾病。
整体实验设计：将2×10^6个THP-1野生型或STING细胞用二甲基亚砜（DMSO）或500ng/mL的H151处理1小时，随后分别用培养基或10μg/mL的TL8刺激8小时。刺激结束后，使用Trizol试剂（Trizol）收集RNA。