Japanese encephalitis virus orchestrates GLUT4-mediated glucose metabolism to potentiate viral replication via insulin receptor signaling.

Flaviviruses intricately rewire host metabolic networks to establish a replication-permissive environment; however, the role of glucose transporter-mediated uptake, particularly via glucose transporter 4 (GLUT4), remains insufficiently defined. Japanese encephalitis virus (JEV) infection induces extensive remodeling of glucose metabolism, exemplified by the coordinated upregulation of critical metabolic effectors. Pharmacological blockade of glucose metabolic pathways markedly attenuates JEV replication, whereas exogenous glucose supplementation enhances viral propagation in a concentration-dependent manner. A targeted screen of 111 metabolism-oriented compounds identified selective GLUT4 inhibitors with potent antiviral efficacy. Notably, GLUT4 expression is consistently upregulated during JEV infection across multiple cell types, albeit to varying degrees, and is similarly induced by duck Tembusu virus (DTMUV), suggesting a potentially conserved mechanism shared by these two flaviviruses. However, broader validation across additional members of the Flavivirus genus remains warranted. Mechanistically, the viral nonstructural protein 3 (NS3) engages insulin receptor substrate 1 (IRS1), thereby activating the IRS1-PI3K-Akt-mTORC1-SREBP-1c signaling axis to transcriptionally drive GLUT4 expression. Concurrently, JEV infection induces PI3K-Akt-dependent phosphorylation of AS160, promoting GLUT4 vesicular trafficking via the coordinated action of Rab8 and Rab10. Collectively, these findings delineate a previously unrecognized mechanism whereby JEV commandeers host insulin signaling to orchestrate GLUT4 biosynthesis and membrane translocation, thereby ensuring continuous metabolic substrate availability to sustain replication. This GLUT4-centric metabolic circuitry represents a mechanistically tractable target for host-directed antiviral strategies against Flavivirus.