Elevation of Cellular NAD Levels by Nicotinic Acid and Involvement of Nicotinic Acid Phosphoribosyltransferase in Human Cells

Abstract

NAD plays critical roles in various biological processes through the function of SIRT1. Although classical studies in mammals showed that nicotinic acid (NA) is a better precursor than nicotinamide (Nam) in elevating tissue NAD levels, molecular details of NAD synthesis from NA remain largely unknown. We here identified NA phosphoribosyltransferase (NAPRT) in humans and provided direct evidence of tight link between NAPRT and the increase in cellular NAD levels. The enzyme was abundantly expressed in the small intestine, liver, and kidney in mice and mediated [ 14 C]NAD synthesis from [ 14 C]NA in human cells. In cells expressing endogenous NAPRT, the addition of NA but not Nam almost doubled cellular NAD contents and decreased cytotoxicity by H 2 O 2 . Both effects were reversed by knockdown of NAPRT expression. These results indicate that NAPRT is essential for NA to increase cellular NAD levels and, thus, to prevent oxidative stress of the cells. Kinetic analyses revealed that NAPRT, but not Nam phosphoribosyltransferase (NamPRT, also known as pre-B-cell colony-enhancing factor or visfatin), is insensitive to the physiological concentration of NAD. Together, we conclude that NA elevates cellular NAD levels through NAPRT function and, thus, protects the cells against stress, partly due to lack of feedback inhibition of NAPRT but not NamPRT by NAD. The ability of NA to increase cellular NAD contents may account for some of the clinically observed effects of the vitamin and further implies a novel application of the vitamin to treat diseases such as those associated with the depletion of cellular NAD pools.

Identification of a New Urate and High Affinity Nicotinate Transporter, hOAT10 (SLC22A13)

“…Moreover, in additional experiments on brush border membrane vesicles of human jejunum, they confirmed a strict pH dependence of nicotinate reabsorption, being higher at low pH. This is consistent with our observation of a pH dependence of hOAT10, Nicotinate may serve not only for NAD ϩ synthesis, which is important for energetic processes, signal transduction pathways, or even the activation of the NAD ϩ -dependent histone deacetylase SIRT1 and therefore life extension (42), but also for reabsorption of the purine metabolite urate.…”

Inhibition of Nicotinamide Phosphoribosyltransferase (NAMPT) Activity by Small Molecule GMX1778 Regulates Reactive Oxygen Species (ROS)-mediated Cytotoxicity in a p53- and Nicotinic Acid Phosphoribosyltransferase1 (NAPRT1)-dependent Manner

“…In higher eukaryotes, two major biochemical pathways, the salvage and the de novo pathway, are involved in the biosynthesis of NAD ϩ . The salvage pathway operates via the two major pathways using nicotinamide phosphoryltransferase (NAMPT) and nicotinamide phosphoribosyltransferase 1 (NAPRT1), which use nicotinamide and nicotinic acid (NA), respectively, as the substrate for NAD ϩ recycling (18). Although both pathways are employed to generate NAD ϩ , in cells expressing endogenous NAPRT1, only the NA added in the salvage pathway can increase cellular levels of NAD ϩ and reduce cytotoxicity by an oxidizing agent (18,19).…”

“…The salvage pathway operates via the two major pathways using nicotinamide phosphoryltransferase (NAMPT) and nicotinamide phosphoribosyltransferase 1 (NAPRT1), which use nicotinamide and nicotinic acid (NA), respectively, as the substrate for NAD ϩ recycling (18). Although both pathways are employed to generate NAD ϩ , in cells expressing endogenous NAPRT1, only the NA added in the salvage pathway can increase cellular levels of NAD ϩ and reduce cytotoxicity by an oxidizing agent (18,19). GMX1778 (CHS 828; teglarinad) is a potent inhibitor of NAMPT that exerts a cytotoxic effect by decreasing the cellular level of NAD ϩ (20,21).…”

Erythropoietin and Oxidative Stress

“…These agents can involve superoxide free radicals, hydrogen peroxide, singlet oxygen, nitric oxide (NO), and peroxynitrite (Chong, et al, 2005e). Most species are produced at low levels during normal physiological conditions and are scavenged by endogenous antioxidant systems that include superoxide dismutase, glutathione peroxidase, catalase, and small molecule substances such as vitamins C and E. Other closely linked pathways to oxidative stress may be tempered by different vitamins, such as vitamin D 3 (Regulska, et al, 2007) and the amide form of niacin or vitamin B 3 , nicotinamide (Chlopicki, et al, 2007, Chong, et al, 2002d, Hara, et al, 2007, Ieraci and Herrera, 2006.Throughout the body, cell survival and lifespan is tied to the presence of oxidative stress and the subsequent induction of apoptotic cell injury , De Felice, et al, 2007, Lin and Maiese, 2001). It has recently been shown that genes involved in the apoptotic process are replicated early during processes that involve cell replication and transcription, suggesting a much broader role for these genes than originally anticipated (Cohen, et al, 2007).…”