EIF2AK2 inhibits type I interferon production by inhibiting RIG-I activation and disrupting mitochondria function via recruiting VAMP8 and p-MLKL.

Type I interferons (IFN-I) are central to antiviral immunity, but their excessive or sustained production can result in immunopathological damage. RIG-I-like receptor (RLR) signaling is pivotal in regulating RNA virus-induced IFN-I responses and requires precise modulation to maintain immune homeostasis. Here, we report that viral infection induced elevated expression of Eukaryotic translation initiation factor 2 alpha kinase 2 (EIF2AK2), which is an interferon-stimulated gene (ISG) with unclear role in the innate immunity. Using EIF2AK2-deficient mice and cells, we demonstrated that the loss of EIF2AK2 specifically enhances RNA virus-induced IFN-I production in macrophages and suppresses the replication of RNA virus vesicular stomatitis virus (VSV), and that this function is tightly associated with the N-terminal dsRNA binding domain of EIF2AK2. Mechanistically, EIF2AK2 competes with RIG-I for binding viral RNA, thereby inhibiting RIG-I activation. In addition, EIF2AK2 promotes the translocation of p-MLKL to mitochondria via recruiting VAMP8, leading to disruption of mitochondrial membrane potential and dysfunction of MAVS, ultimately inhibiting IFN-I production. These findings identify EIF2AK2 as a critical negative regulator of RLR-mediated innate immune response to RNA viruses via dual inhibitory mechanisms, and suggest its potential as a therapeutic target for controlling dysregulated IFN-I responses.