Hydrolyzing NAD to NMN and AMP by SelO is Required for Mitochondria Homeostasis

The regulation of mitochondrial nicotinamide adenine dinucleotide (mNAD) is crucial for numerous life processes. However, until now, no metabolic reaction responsible for downregulating mNAD has been identified. Through in silico screening of potential NAD-binding proteins, we discovered a previously unrecognized reaction where NAD+ (or NADH) is hydrolyzed to NMN (or NMNH) and AMP in mitochondria. This reaction is catalyzed by the selenoprotein SelO, with Mn2+ serving as an essential cofactor. Selenocysteine 667 on the SelO protein plays a critical role in its NAD-hydrolyzing activity. SelO directly interacts with the mitochondrial trifunctional complex, and inhibits lipid β-oxidation, where NAD+ is required as an essential cofactor. This reaction is enhanced by a gradual increase in pH, which signals heightened mitochondrial catabolism, and it downregulates matrix pH as a negative feedback mechanism to maintain mitochondrial homeostasis. The NAD-hydrolyzing activity of SelO and its role in lipid oxidation are conserved across distant species, from mammalian cells to bacteria. Furthermore, SelO has been identified as a direct target of scutellarein, a clinically used agent for various ailments. These findings reveal a conserved mechanism for spatiotemporal NAD regulation, and highlight its physiological significance across both prokaryotes and eukaryotes.