NAD+ is an important cofactor regulating metabolic homeostasis and a rate-limiting substrate for sirtuin deacylases. We show that NAD+ levels are reduced in aged mice and Caenorhabditis elegans and that decreasing NAD+ levels results in a further reduction in worm lifespan. Conversely, genetic or pharmacological restoration of NAD+ prevents age-associated metabolic decline and promotes longevity in worms. These effects are dependent upon the protein deacetylase sir-2.1 and involve the induction of mitonuclear protein imbalance as well as activation of stress signaling via the mitochondrial unfolded protein response (UPRmt) and the nuclear translocation and activation of FOXO transcription factor DAF-16. Our data suggest that augmenting mitochondrial stress signaling through the modulation of NAD+ levels may be a target to improve mitochondrial function and prevent or treat age-associated decline. Display omitted •Enhancing NAD+ availability extends lifespan in C. elegans through sir-2.1•The NAD+/sir-2.1-dependent activation of mitochondrial UPR is crucial for longevity•Higher NAD+ bioavailability protects against ROS by enhancing daf-16 activity•UPRmt is a conserved target of the NAD+/SIRT1 axis in mammalian cells Perturbations that increase the levels of NAD+, a substrate for sirtuin deacylases, extend lifespan in worms by activating both the mitochondrial unfolded protein response and DAF-16/FOXO-dependent antioxidant defenses.