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NAD+ metabolism and its roles in cellular processes during ageing - PubMed

Review

NAD+ metabolism and its roles in cellular processes during ageing

Anthony J Covarrubias et al. Nat Rev Mol Cell Biol. 2021 Feb.

Abstract

Nicotinamide adenine dinucleotide (NAD+) is a coenzyme for redox reactions, making it central to energy metabolism. NAD+ is also an essential cofactor for non-redox NAD+-dependent enzymes, including sirtuins, CD38 and poly(ADP-ribose) polymerases. NAD+ can directly and indirectly influence many key cellular functions, including metabolic pathways, DNA repair, chromatin remodelling, cellular senescence and immune cell function. These cellular processes and functions are critical for maintaining tissue and metabolic homeostasis and for healthy ageing. Remarkably, ageing is accompanied by a gradual decline in tissue and cellular NAD+ levels in multiple model organisms, including rodents and humans. This decline in NAD+ levels is linked causally to numerous ageing-associated diseases, including cognitive decline, cancer, metabolic disease, sarcopenia and frailty. Many of these ageing-associated diseases can be slowed down and even reversed by restoring NAD+ levels. Therefore, targeting NAD+ metabolism has emerged as a potential therapeutic approach to ameliorate ageing-related disease, and extend the human healthspan and lifespan. However, much remains to be learnt about how NAD+ influences human health and ageing biology. This includes a deeper understanding of the molecular mechanisms that regulate NAD+ levels, how to effectively restore NAD+ levels during ageing, whether doing so is safe and whether NAD+ repletion will have beneficial effects in ageing humans.

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Conflict of interest statement

Competing interests

E.V. is a scientific co-founder of Napa Therapeutics and serves on the scientific advisory board of Seneque. E.V., A.J.C. and R.P. receive research support from Napa Therapeutics. E.V. and A.G. receive research support from BaReCia. A.G serves as Chief Scientific Officer for Seneque USA and is one of the inventors on a patent (PCT/US18/46233) for the SLC12A8 nicotinamide mononucleotide transporter, whose applicant is Washington University in St. Louis and which has been licensed by Teijin Limited (Japan).

Figures

Fig. 1 |
Fig. 1 |. NAD+ metabolism.

a | Nicotinamide adenine dinucleotide (NAD+) biosynthetic pathways. NAD+ levels are maintained by three independent biosynthetic pathways. The kynurenine pathway (or de novo synthesis pathway) uses the dietary amino acid tryptophan to generate NAD+. Tryptophan enters the cell via the transporters SLC7A5 and SLC36A4. Within the cell, tryptophan is converted to N-formylkynurenine by the rate-limiting enzyme indoleamine 2,3-dioxygenase (IDO) or the rate-limiting enzyme tryptophan 2,3-dioxygenase (TDO). N-Formylkynurenine is transformed into

l

-kynurenine, which is further converted to 3-hydroxykynurenine (3-HK) by kynurenine 3-monooxygenase (KMO) and to 3-hydroxyanthranilic acid (3-HAA) by tryptophan 2,3-dioxygenase (KYNU). The next step is performed by 3-hydroxyanthranilic acid oxygenase (3HAO) to generate α-amino-β-carboxymuconate ε-semialdehyde (ACMS). This compound can spontaneously condense and rearrange into quinolinic acid, which is transformed by quinolinate phosphoribosyltransferase (QPRT) into nicotinamide mononucleotide (NAMN), at which point it converges with the Preiss–Handler pathway. The Preiss–Handler pathway uses dietary nicotinic acid (NA), which enters the cell via SLC5A8 or SLC22A13 transporters, and the enzyme nicotinic acid phosphoribosyltransferase (NAPRT) to generate NAMN, which is then transformed into nicotinic acid adenine dinucleotide (NAAD) by nicotinamide mononucleotide adenylyltransferases (NMNAT1, NMNAT2 and NMNAT3). The process is completed by the transformation of NAAD into NAD+ by NAD+ synthetase (NADS). The NAD+ salvage pathway recycles the nicotinamide (NAM) generated as a by-product of the enzymatic activities of NAD+-consuming enzymes (sirtuins, poly(ADP-ribose) polymerases (PARPs) and the NAD+ glycohydrolase and cyclic ADP-ribose synthases CD38, CD157 and SARM1). Initially, the intracellular nicotinamide phosphoribosyltransferase (iNAMPT) recycles NAM into nicotinamide mononucleotide (NMN), which is then converted into NAD+ via the different NMNATs. NAM can be alternatively methylated by the enzyme nicotinamide N-methyltransferase (NNMT) and secreted via the urine. In the extracellular space, NAM is generated as a by-product of the ectoenzymes CD38 and CD157 and can be converted to NMN by extracellular NAMPT (eNAMPT). NMN is then dephosphorylated by CD73 to nicotinamide riboside (NR), which is transported into the cell via an unknown nucleoside transporter (question mark). NMN can be imported into the cell via an NMN-specific transporter (SLC12A8 in the small intestine). Intracellularly, NR forms NMN via nicotinamide riboside kinases 1 and 2 (NRK1 and NRK2). NMN is then converted to NAD+ by NMNAT1, NMNAT2 and NMNAT3. b | NAD+ metabolism in different subcellular compartments. The NAD+ homeostasis is a balance of synthesis, consumption and regeneration in different subcellular compartments, which are regulated by subcellular-specific NAD+-consuming enzymes, subcellular transporters and redox reactions. NAD+ precursors enter the cell via the three biosynthetic pathways (part a). In the cytoplasm, NAM is converted to NMN by intracellular NAMPT (iNAMPT). NMN is then converted to NAD+ by NMNAT2, which is the cytosol-specific isoform of this enzyme. NAD+ is utilized during glycolysis, generating NADH, which is transferred to the mitochondrial matrix via the malate/aspartate shuttle and the glyceraldehyde 3-phosphate shuttle. The mitochondrial NADH imported via the malate/aspartate shuttle is oxidized by complex I in the electron transport chain (ETC), whereas the resulting reduced flavin adenine dinucleotide (FADH2) from the glyceraldehyde 3-phosphate shuttle is oxidized by complex II. Recently the mammalian NAD+ mitochondrial transporter SLC25A51 was identified, and it has been demonstrated to be responsible for intact NAD+ uptake in the organelle. The salvage pathway for NAD+ in mitochondria has not been fully resolved, but the role of a specific NMNAT isoform (NMNAT3) has been proposed. In the mitochondria, NAD+ is consumed by NAD+-dependent mitochondrial SIRT3–SIRT5, generating NAM. It is still unknown whether NAM can be converted back to NMN or can be converted to NAD+ within the mitochondrion, or whether other precursors can be transported through the mitochondrial membrane to fuel NAD+ synthesis. The nuclear NAD+ pool probably equilibrates with the cytosolic one by diffusion through the nuclear pore; however, the full dynamics are still largely unexplored. A nuclear-specific NMNAT isoform (NMNAT1) has been described and is part of the nuclear NAM salvage NAD+ pathway. MNAM, N1-methylnicotinamide; TCA, tricarboxylic acid.

Fig. 2 |
Fig. 2 |. Three main classes of NAD+-consuming enzymes.

a | Sirtuins remove acyl groups from lysine residues on target proteins using nicotinamide adenine dinucleotide (NAD+) as a co-substrate. NAD+ is cleaved, generating nicotinamide (NAM) and ADP-ribose, where ADP-ribose serves as an acyl group acceptor, generating acetyl-ADP-ribose (acetyl-ADPR). b | Poly(ADP-ribose) polymerases (PARP1–PARP3) use NAD+ as a co-substrate to mono(ADP-ribosyl)ate or poly(ADP-ribosyl)ate target proteins, generating NAM as a by-product. c | Reactions of NAD+ glycohydrolases and cyclic ADP-ribose (cADPR) synthases (CD38, CD157 and SARM1). The main catalytic activity of this group of proteins is the hydrolysis of NAD+ to NAM and ADP-ribose. To a lesser extent, CD38, CD157 and SARM1 have ADP-ribosyl cyclase activity, generating NAM and cADPR from NAD+. In acidic conditions, CD38 can also perform a base-exchange reaction, swapping the NAM of NAD(P)+ for nicotinic acid (NA), generating nicotinic acid adenine dinucleotide (phosphate) (NAAD(P)). CD38 and CD157 are reported to be able to use alternative substrates in their catalytic reactions. CD38 can degrade NMN to NAM and ribose monophosphate (RMP), while CD157 can degrade NR, generating NAM and ribose (R). NR, nicotinamide riboside.

Fig. 3 |
Fig. 3 |. NAD+ metabolism in ageing.

A | Decreased nicotinamide adenine dinucleotide (NAD+) levels have been implicated in various processes associated with ageing (see also Supplementary Box 2). Aa | Ageing is associated with aberrant proinflammatory immune cell activation or ‘inflammageing’, leading to sustained low-grade inflammation. This is caused in part by the accumulation of senescent cells, which via a senescence-associated secretory phenotype (SASP) promote the phenotypic polarization of macrophages towards a proinflammatory M1 state, thereby driving inflammation. There is evidence that in response to the SASP in these macrophages expression of the NAD+-consuming enzymes CD38 and poly(ADP-ribose) polymerases (PARPs) increases, leading to NAD+ level decline, and that these mechanisms importantly contribute to the decrease of NAD+ levels in ageing. In addition, it has been shown that in aged T cells mitochondrial function declines, and this leads to increased secretion of proinflammatory cytokines that promote the state of inflammation and also induce senescence. Ab | Axonal degeneration, which is a precursor to many age-related neuronal disorders, is characterized by rapid NAD+ depletion. During normal physiological conditions, NAD+ biosynthetic enzymes, nicotinamide mononucleotide adenylyltransferases (NMNATs), are protective against axonal degeneration, and their expression supports maintenance of axons and prevents neurodegeneration. In particular, NMNAT2 is an important survival factor in axons, to which it needs to be constantly delivered from the soma — where it is synthesized — to account for its rapid turnover, and these transport processes are disturbed during axonal degeneration. Moreover, the NAD+-consuming enzyme SARM1 is activated by axonal injury and mediates axonal degeneration by promoting NAD+ degradation. Ac | Autophagy is a key cellular catabolic process that allows cells to adapt to variable nutrient availability and serves in cellular quality control, allowing removal of defective organelles and proteins. Autophagy is regulated downstream of NAD+ levels via sirtuins (mostly SIRT1). Decline of NAD+ levels reduces overall autophagic flux as well as selective removal of mitochondria via mitophagy, suggesting that defective autophagy can be a consequence of NAD+ depletion during ageing, contributing to cell dysfunction. B | Because NAD+ is a cofactor for various enzymes, loss of NAD+ impacts a plethora of cellular processes. For example, NAD+ is required for the activity of epigenetic regulators such as SIRT1, and decline in its level causes changes in histone modifications, thereby affecting chromatin organization and function in gene expression. There is evidence that the ageing-associated loss of NAD+ is related to increased expression of PARPs, which can be caused by increased levels of DNA damage and the need for DNA repair during ageing (panel Ba). NAD+ also affects transcriptional activity of the core clock components CLOCK and BMAL, thereby regulating circadian expression of important metabolic genes as well as nicotinamide phosphoribosyltransferase (NAMPT), which in turn is required for circadian oscillation in NAD+ levels (panel Bb). Decreased NAD+ levels also interfere with the activity of PARPs and sirtuins in DNA repair, leading to genomic instability: a hallmark of ageing and cancer (panel Bc). ADPR, ADP-ribose; ATG, autophagy-related protein; FOXO, forkhead box protein O; ROS, reactive oxygen species.

Fig. 4 |
Fig. 4 |. Therapeutic approaches to restore NAD+ levels and their impact on health.

Ageing is associated with decreased nicotinamide adenine dinucleotide (NAD+) levels that promote or exacerbate ageing-related diseases. Thus, restoring NAD+ levels has emerged as a therapeutic approach to prevent and treat ageing-related diseases and to restore health and vigour during the ageing process (part a). Some potential strategies that boost NAD+ levels include lifestyle changes, such as increasing exercise, reducing caloric intake, eating a healthy diet and following a consistent daily circadian rhythm pattern by conforming to healthy sleeping habits and mealtimes. Another approach is the use of small-molecule inhibitors or activators to boost NAD+ biosynthesis and the use of dietary supplements, including NAD+ precursors, such as nicotinamide mononucleotide and nicotinamide riboside. All of these approaches promote tissue NAD+ levels and are beneficial for health. These include improved tissue and organ function, protection from cognitive decline, improved metabolic health, reduced inflammation and increased physiological benefits, such as increased physical activity, which may collectively extend patient healthspan and potentially lifespan (part b).

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