Janus kinases (JAKs) bind to class I and II cytokine receptors, activating signaling and regulating gene transcription through signal transducer and activator of transcription (STAT) proteins. Type I interferons (IFNs) require the JAK members TYK2 and JAK1, which bind to the receptor subunits IFNAR1 and IFNAR2, respectively. We investigated the role of JAKs in regulating IFNAR signaling activity. Synthetic IFNARs in which the extracellular domains of IFNAR1 and IFNAR2 are replaced with nanobodies had near-native type I IFN signaling, whereas the homomeric variant of IFNAR2 initiated much weaker signaling, despite harboring docking sites for JAKs and STATs. Cells with JAK1 and TYK2 knockout (KO) showed residual signaling, suggesting partial complementation by the remaining JAKs, particularly when they were overexpressed. Live-cell micropatterning experiments confirmed the promiscuous binding of JAK1, JAK2, and TYK2 to IFNAR1 and IFNAR2, and their recruitment correlated with their relative cellular abundances. However, each JAK had a different efficacy in inducing cross-phosphorylation and downstream signaling. JAK binding was also promiscuous for other cytokine receptors, including IFN-L1, IL-10R beta, TPOR, and GHR, but not for EPOR, which activated different downstream signaling pathways. These findings suggest that competitive binding of JAKs to cytokine receptors together with the varying absolute and relative abundances of the JAKs in different cell types can account for the cell type-dependent signaling pleiotropy of cytokine receptors.

Promiscuous binding of different Janus kinases (JAKs) to class I/II cytokine receptors has been reported, yet its role in signaling is unclear. To systematically explore JAK pairing in type I interferon (IFN-I) signaling, we generated an artificial IFN-I receptor (AIR) by replacing the extracellular domains of IFNAR1 and IFNAR2 with anti mEGFP and mCherry nanobodies. The heterodimeric AIR restored near-native IFN-I activity, while the homomeric variant of IFNAR2 (AIR-dR2) initiated much weaker signaling despite harboring docking sites for signal transducer and activator of transcription (STAT) proteins. AIR-dR1 was signaling inactive, yet, pulldown uncovered its ICD to bind both TYK2 and JAK1. To further investigate the roles of JAKs on the receptors, knockout (KO) JAK1, JAK2, TYK2, and JAK2/TYK2 were generated. JAK1 KO led to complete loss of IFN-I signaling, which was partially restored by TYK2 overexpression. TYK2 KO cells retained partial activity, which was elevated by JAK1 overexpression, suggesting both JAKs to partially substitute each other. Conversely, JAK2 KO only moderately impacted the biological activity of IFN-Is, even in JAK2/TYK2 KO cells. Live cell micropatterning confirmed promiscuous binding of JAK1, JAK2 and TYK2 to IFNAR1 and IFNAR2, in line with an AlphaFold model that shows JAKs interchangeability on IFNAR ICDs. Similar promiscuity of JAK binding was observed for TPOR and GHR but not EPOR, accompanied by different downstream signaling activity. The competitive binding of JAKs to cytokine receptors together with the highly diverse absolute and relative JAK expression levels can account for cell type-dependent signaling pleiotropy observed for cytokine receptors. One Sentence Summary Promiscuous and interchangeable binding of JAKs to cytokine receptors enables cell type-specific pleiotropic signaling.

Alternative splicing of pre-mRNA increases transcriptome and proteome diversity by generating distinct isoforms that encode functionally diverse proteins, thus affecting many biological processes, including innate immunity. cGAS-STING signaling pathway, whose key molecules also undergo alternative splicing, plays a crucial role in regulating innate immunity. Protein isoforms of key components in the cGAS-STING-TBK1-IRF3 axis have been detected in a variety of species. A chain of evidence showed that these protein isoforms exhibit distinct functions compared to their normal counterparts. The mentioned isoforms act as positive or negative modulators in interferon response via distinct mechanisms. Particularly, we highlight that alternative splicing serves a vital function for the host to avoid the overactivation of the cGAS-STING signaling pathway and that viruses can utilize alternative splicing to resist antiviral response by the host. These findings could provide insights for potential alternative splicing-targeting therapeutic applications.

The RIG-I receptor induces the innate antiviral responses upon sensing RNA viruses. The mechanisms through which RIG-I optimizes the strength of the downstream signaling remain incompletely understood. In this study, we identified that NSUN5 could potentiate the RIG-I innate signaling pathway. Deficiency of NSUN5 enhanced RNA virus proliferation and inhibited the induction of the downstream antiviral genes. Consistently, NSUN5-deficient mice were more susceptible to RNA virus infection than their wild-type littermates. Mechanistically, NSUN5 bound directly to both viral RNA and RIG-I, synergizing the recognition of dsRNA by RIG-I. Collectively, to our knowledge, this study characterized NSUN5 as a novel RIG-I coreceptor, playing a vital role in restricting RNA virus infection.

识别病毒核酸可触发宿主针对病毒的固有免疫反应.维甲酸诱导基因Ⅰ(retinoic acid-inducible geneⅠ,RIG-Ⅰ)和黑色素瘤分化相关基因5 (melanoma differentiation-associated gene 5,MDA5)存在于多种类型细胞中,参与识别入侵细胞质的病毒RNA,诱导下游抗病毒效应分子产生,在细胞抗病毒免疫反应中发挥重要作用.RIG-Ⅰ或MDA5与病毒RNA结合进而被激活的过程在时空上受到精密复杂的调控.近年来研究发现,辅助受体在RIG-Ⅰ和MDA5激活的过程中发挥重要辅助功能.文章概述了目前已知的RIG-Ⅰ和MDA5辅助受体研究进展,重点阐述了其在所介导的抗病毒信号通路中的调控机制和作用.这有助于深刻理解RIG-Ⅰ和MDA5所介导的抗病毒信号通路,并为相关疾病的治疗提供新策略.