The role of nicotinamide riboside kinases in mammalian NAD⁺ metabolism

NAD+ has emerged as a central metabolic node and an important co-substrate for the activity of various enzymes, including the protein deacetylase SIRT1. Different strategies to increase NAD+ bioavailability have been shown to boost SIRT1 activity, which results in protection against metabolic disease, neurodegenerative disorders and certain types of cancer. Nicotinamide riboside (NR) and nicotinamide mononucleotide (NMN) have recently been discovered as new NAD+ precursors. Recent studies have shown that dietary supplementation with NR or NMN prevented against high-fat diet-induced body weight gain. However, the physiological role of NR in mammalian metabolism remains largely unexplored. In this project, we focused on the first step of NR metabolism â the phosphorylation of NR by nicotinamide riboside kinases (NRK1 and NRK2). We have investigated the role of NRKs in the metabolism of NAD+ precursors and mammalian physiology in vivo. The results obtained indicate that NRKs are the rate-limiting and essential step for NR- and NMN-driven NAD+ synthesis in hepatocytes and that NMN has to be extracellularly converted to NR for its use as an NAD+ precursor. Accordingly, animals lacking NRK1 have defects in NR/NMN utilization. We demonstrated that effective NR utilization is key for the maintenance of metabolic homeostasis as NRK1 whole-body KO mice display numerous alterations in glucose and lipid management. Using NRK1 liver-specific KO mice, we demonstrate that NRK1 is required to maintain hepatic NAD+ levels and mitochondrial function upon high-fat feeding. Consequently, NRK1LKO mice are sensitized to diet-induced glucose intolerance and insulin resistance. Finally, we show that both isoforms of NRK are redundant in mediating NR/NMN effects in muscle. Hence, this work identifies NRKs as a novel valuable target in the fight against metabolic disease and indicates that the regulation of NRK will enable us to find novel ways of enhancing NAD+ availability. Moreover, it has unveiled a critical role of NRK1 for the metabolism of diverse forms of Vitamin B3 and highlights how circulating NR is key to preserve metabolic health upon dietary challenges.

Endogenous nicotinamide riboside metabolism protects against diet-induced liver damage

Carlos Canto Alvarez, Sofia Moco, Joanna Ratajczak, Miriam Valera Alberni

Supplementation with the NAD(+) precursor nicotinamide riboside (NR) ameliorates and prevents a broad array of metabolic and aging disorders in mice. However, little is known about the physiological role of endogenous NR metabolism. We have previously shown that NR kinase 1 (NRK1) is rate-limiting and essential for NR-induced NAD(+) synthesis in hepatic cells. To understand the relevance of hepatic NR metabolism, we generated whole body and liver-specific NRK1 knockout mice. Here, we show that NRK1 deficiency leads to decreased gluconeogenic potential and impaired mitochondrial function. Upon high-fat feeding, NRK1 deficient mice develop glucose intolerance, insulin resistance and hepatosteatosis. Furthermore, they are more susceptible to diet-induced liver DNA damage, due to compromised PARP1 activity. Our results demonstrate that endogenous NR metabolism is critical to sustain hepatic NAD(+) levels and hinder diet-induced metabolic damage, highlighting the relevance of NRK1 as a therapeutic target for metabolic disorders.

2019

Targeting mitochondrial bioenergetics and NAD+ metabolism during sarcopenia and regeneration of skeletal muscle

Tanja Sonntag

The progressive decline of muscle mass and function called sarcopenia is a multi-factorial process associated to frailty, disability, and low quality of life in the elderly. The co-factor nicotinamide adenine dinucleotide (NAD+) becomes a limiting factor in the course of aging and other pathological conditions. It has emerged as a major regulator of cellular metabolism, which is modulated by dietary precursors of the vitamin B3 family. So far, no direct link between sarcopenia and alterations in NAD+ metabolism has been reported. In this thesis, I have taken a multi-disciplinary approach combining molecular profiling in humans and dietary and genetic manipulations in rodents to characterize the skeletal muscle NAD+ metabolism in the context of muscle plasticity and aging. We have performed a multi-center study to characterize the molecular signature of human sarcopenia compared to age-matched controls. Sarcopenic participants of three cohorts in Singapore, the United Kingdom and Jamaica presented a prominent transcriptional signature of mitochondrial dysfunction, which translated into functional bioenergetic deficiency with lower mitochondrial protein expression and activity. Furthermore, we established a concurrent decrease in NAD+ content of sarcopenic muscle, which correlated to lower muscle strength and function in these patients. Supplementation with the dietary NAD+ precursor nicotinamide riboside (NR) was shown to improve pathological muscle conditions and revert age-related mitochondrial dysfunction and stem cell senescence in mice. To test whether boosting NAD+ through NR can be beneficial in the context of sarcopenia, we investigated the acute response to NR supplementation in young adult and old sarcopenic rats. Acute NR treatment efficiently increased NAD+ levels and normalized specific age-related gene expression signatures in old rats, in particular related to fibrosis. Interestingly, our transcriptional profiling also revealed that the molecular response to NR is partially blunted in aging and suggests that aged muscle could be in a state of partial NAD+ resistance. To investigate the role of endogenous NR as NAD+ precursor in the muscle, we studied mice deficient for NR kinases 1 and 2 (NRK1/2 dKO), the rate-limiting enzymes for NR salvage. NRK1/2 dKO mice develop normally, but exhibit elevated salvage of the NAD+ precursor nicotinamide (NAM) to maintain tissue NAD+. Thus, our results demonstrate for the first time that an endogenous NRK-dependent flux actively converts NR into NAD+ in healthy skeletal muscle. Moreover, we demonstrated that NRKs are required for muscle regeneration, as dKO mice failed to efficiently regenerate the NAD+ pool during the active phases of myofiber remodeling, which coincided with a transient delay of myofiber maturation. Conversely, dietary NR improved the activation of muscle stem cells and accelerated recovery of NAD+ levels in regenerating myofibers of wild-type mice. Altogether, this work demonstrates that both in humans and preclinical models, sarcopenia is tightly linked to mitochondrial bioenergetics and NAD+ metabolism. Dietary manipulation of NAD+ levels is a promising therapeutic strategy for the management of age-related muscle dysfunction for which we uncovered important mechanistic insights linking metabolic fluxes through the NAD+ pathway to physiological adaptations.

EPFL2019

Targeting mitochondrial bioenergetics and NAD+ metabolism during sarcopenia and regeneration of skeletal muscle

Tanja Sonntag

EPFL2019