Targeting viral Fe-S clusters with TEMPOL confers broad-spectrum anti-coronavirus activity

PEDV infection induces intracellular iron accumulation in host cells. • Iron promotes CoV replication by enhancing Fe-S cluster-dependent RdRp activity. • TEMPOL suppresses CoV replication by disrupting viral Fe-S cluster integrity. • TEMPOL exhibits broad-spectrum anti-CoV activity in vitro and in vivo. • TEMPOL effectively counteracts iron-exacerbated viral pathogenesis in vivo. Coronaviruses (CoVs) can cause severe respiratory and enteric diseases, posing major threats to human health and the livestock industry. However, the host metabolic mechanisms underlying viral replication remain unclear. Using porcine epidemic diarrhea virus (PEDV) as the primary model, this study investigated the role of iron metabolism in CoV replication and its mechanisms. Our results showed that PEDV infection markedly induced intracellular iron accumulation, while exogenous iron supplementation enhanced viral replication and iron chelation suppressed it. Mechanistically, iron enhances viral replication by promoting the formation of iron-sulfur (Fe-S) clusters, essential cofactors for the viral RNA-dependent RNA polymerase (RdRp). Based on these findings, we further evaluated the antiviral activity of TEMPOL, a small molecule targeting viral Fe-S clusters. TEMPOL reduced RdRp activity and effectively inhibited PEDV replication. Moreover, TEMPOL exhibited broad-spectrum antiviral activity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), infectious bronchitis virus (IBV), and porcine deltacoronavirus (PDCoV). Importantly, TEMPOL effectively reversed FAC-promoted viral pathogenesis, alleviating tissue lesions and clinical symptoms across multiple CoV animal models. Collectively, this study reveals a key role of iron metabolism in CoV replication and suggests that targeting viral Fe-S clusters may provide a novel strategy for anti-CoV therapy against diverse CoVs.