Calycosin targets the MAPK-EGR1 signaling axis to inhibit enterovirus 71 replication in cells and human colonic organoids.

The downstream effectors that mediate the pro‑viral function of the MAPK pathway during enterovirus 71 (EV71) infection remain poorly defined. Here we show that early growth response 1 (EGR1) is independently regulated by each of the three classical MAPK branches including extracellular signal-regulated kinase (ERK), p38 and c-Jun N-terminal kinase (JNK), and serves as a common downstream target of these pathways to promote EV71 replication and virus-induced cell death. Furthermore, we found that the natural isoflavone calycosin (CA) from Astragalus membranaceus acts as a regulatory probe of the MAPK-EGR1 axis. CA suppresses EV71-induced expression and phosphorylation of ERK, p38, and JNK, as well as the subsequent upregulation of EGR1, thereby inhibiting both viral replication and virus-induced cell death in conventional cell lines and human colonic organoid infection models. This pro-viral axis is conserved among several enteroviruses, including coxsackievirus B3 and enterovirus D68, and CA exhibits broad-spectrum activity against multiple enteroviruses by targeting this axis. Collectively, our findings establish the MAPK‑EGR1 axis as a key host‑dependency node for EV71 and other enteroviruses, and identify CA as a valuable chemical probe for interrogating this axis. This work not only deepens our understanding of the interplay between enteroviruses such as EV71 and their host, but also provides a mechanistic basis for host‑directed antiviral strategies.