SARS-CoV-2 Orf6 Triggers MTCH1-Dependent Mitochondrial Dysfunction and Necroptosis in Human Neurons

The molecular basis of neuronal damage in SARS-CoV-2 infection remains elusive. Here, we identify the viral accessory protein Orf6 as a key driver of necroptotic neuronal death. Through a systematic screen of 22 structural and non-structural SARS-CoV-2 proteins, Orf6 emerged to be the most cytotoxic to human cells. In primary human neurons, Orf6 selectively activates the necroptotic pathway by inducing phosphorylation of RIPK3 and MLKL, without engaging apoptosis, pyroptosis, or autophagy. These findings were corroborated in SARS-CoV-2-infected neurons and in hippocampal brain sections from COVID-19 patients, where elevated necroptosis markers were observed. Mechanistically, Orf6 was found to directly interact with the mitochondrial membrane protein Mitochondrial Carrier Homolog 1 (MTCH1), triggering mitochondrial calcium overload, membrane potential loss, and oxidative stress. Genetic knockdown of MTCH1 or pharmacological inhibition of RIPK3 effectively rescued neurons from Orf6-induced necroptosis and mitochondrial dysfunction. Collectively, our findings reveal a novel, mitochondria-dependent mechanism of viral neuropathogenesis and implicate Orf6-mediated necroptosis as a contributor to long-term neurological sequelae in COVID-19.