Zhao, Y.; Kergoat, L.; Dias de Melo, G.; Larrous, F.; Drumont, G.; Hernandez Camacho, J. D.; Vimont, E.; Le Seac'h, E.; Saunders, N.; Kaechele, M.; Kornobis, E.; Giai Gianetto, Q.; Matondo, M.; Tardieux, I.; Schwartz, O.; Bourhy, H.; Wai, T.

Mitochondrial outer membrane proteins are exploited by diverse intracellular pathogens, to modulate cell metabolism and innate sensing pathways. Here, we demonstrate that TOMM70-dependent mitochondrial recruitment is required to protect SARS-CoV-2 ORF9b from proteasomal degradation, a dependency conserved across ORF9b homologs from related coronaviruses. ORF9b mitochondrial recruitment requires the E477 residue of TOMM70, a surface distinct from that used by the parasite Toxoplasma gondii to engage host mitochondria. We further show that TOMM70 is not a passive scaffold: its depletion activates interferon-stimulated gene expression independently of infection and remodels host immunity distinctly from ORF9b, establishing the receptor and viral protein as mechanistically separable. Using ORF9b-deficient SARS-CoV-2, we demonstrate that ORF9b is dispensable for viral replication and pathological responses in human respiratory epithelial cells and lungs of infected golden Syrian hamsters. Omics profiling of SARS-CoV-2 infected lungs revealed an induction of pathways related to COVID-19 in the absence of ORF9b. Notably, ORF9b-deficient virus-infected lungs show elevated expression of C15ORF48, a nuclear-encoded mitochondrial protein that substitutes for Complex IV subunit NDUFA4 to attenuate inflammation. Collectively, we propose that ORF9b is a receptor-gated viral protein whose principal measurable consequence during authentic infection is a restraint on inflammatory respiratory-chain remodeling.