Enhancement of -GlcNAcylation on Mitochondrial Proteins with 2-(4-Methoxyphenyl)ethyl-2-acetamido-2-deoxy-β-d-pyranoside, Contributes to the Mitochondrial Network, Cellular Bioenergetics and Stress Response in Neuronal Cells under Ischemic-like Conditions.

-GlcNAcylation is a nutrient-driven post-translational modification known as a metabolic sensor that links metabolism to cellular function. Recent evidences indicate that the activation of -GlcNAc pathway is a potential pro-survival pathway and that acute enhancement of this response is conducive to the survival of cells and tissues. 2-(4-Methoxyphenyl)ethyl-2-acetamido-2-deoxy-β-d-pyranoside (SalA-4g), is a salidroside analogue synthesized in our laboratory by chemical structure-modification, with a phenyl ring containing a para-methoxy group and a sugar ring consisting of -acetylglucosamine. We have previously shown that SalA-4g elevates levels of protein -GlcNAc and improves neuronal tolerance to ischemia. However, the specific target of SalA-4g regulating -GlcNAcylation remains unknown. To address these questions, in this study, we have focused on mitochondrial network homeostasis mediated by -GlcNAcylation in SalA-4g's neuroprotection in primary cortical neurons under ischemic-like conditions. -GlcNAc-modified mitochondria induced by SalA-4g demonstrated stronger neuroprotection under oxygen glucose deprivation and reoxygenation stress, including the improvement of mitochondrial homeostasis and bioenergy, and inhibition of mitochondrial apoptosis pathway. Blocking mitochondrial protein -GlcNAcylation with OSMI-1 disrupted mitochondrial network homeostasis and antagonized the protective effects of SalA-4g. Collectively, these data demonstrate that mitochondrial homeostasis mediated by mitochondrial protein -GlcNAcylation is critically involved in SalA-4g neuroprotection.