Abstract
Severe acute respiratory syndrome coronavirus 2 is an enveloped virus that encodes four structural proteins, including the small transmembrane envelope (E) protein. While E is known to function in viral assembly and egress, it contributes to host cell dysfunction and disease severity. We demonstrate that severe acute respiratory syndrome coronavirus 2 E localizes to host cell mitochondria and alters mitochondrial structure, metabolism, and redox homeostasis. Using fluorescence microscopy, we observed that E forms tubular cytoplasmic structures that colocalize with mitochondria and ceramide-rich domains. Lipidomic analysis revealed that E expression leads to reductions in cardiolipin, phosphatidylcholine, and lysophospholipids. Mitochondrial membrane potential was decreased in E-expressing cells, consistent with disrupted electron transport chain activity, which was further supported by mitochondria stress testing via Seahorse. Despite increased mitochondrial reactive oxygen species, E did not trigger apoptosis, suggesting containment of oxidative stress within the organelle. Metabolomic profiling revealed decreased levels of key glycolytic and tricarboxylic acid cycle intermediates, along with altered GSH and sulfur metabolism. Notably, glutamine levels increased, potentially to compensate for reduced 2-oxoglutarate. Together, these findings suggest that E protein localizes to the mitochondria, perturbs lipid and metabolic homeostasis, and promotes reactive oxygen species retention without inducing cell death. This mitochondrial dysfunction may support a shift toward aerobic glycolysis, facilitating viral replication. Our study highlights an underappreciated role for E in modulating host metabolism.