fig1

Figure 1. NAD⁺ metabolism in cells and its compartmentalization^[99,103]. Human cells produce NAD⁺ through three major pathways: the Kynurenine pathway, Preiss-Handler pathway, and Salvage pathway. In the Kynurenine pathway, which is a de novo pathway, the precursor molecule, tryptophan (Trp), after entering the cells via the transporters SLC7A5 and SLC36A4, is converted to N-formyl kynurenine (FK) by the rate-limiting enzyme indoleamine 2,3- dioxygenase (IDO) or the rate-limiting enzyme tryptophan 2,3- dioxygenase (TDO) and then FK is converted to kynurenine. The kynurenine aminotransferases (KATs) convert kynurenine to kynurenic acid, further converted to quinaldic acid. In addition to this, kynurenine 3-monooxygenase (KMO) converts kynurenine to 3-hydroxykynurenine (3-HK), which is further transformed to 3- hydroxy anthranilic acid (3-HAA) by tryptophan 2,3- dioxygenase (KYNU). The 3-HAA gives rise to α- amino- β- carboxy muconate ε- semialdehyde (ACMS) by the enzyme 3-hydroxyanthranilic acid oxygenase (3HAO). Finally, ACMS is transformed to picolinic acid by α-amino-β-carboxy muconate-ε-semialdehyde decarboxylase (ACMSD) or quinolinic acid. In the Preiss-Handler pathway, the precursor molecule nicotinic acid (NA) first enters the cells via SLC5A8 or SLC22A3 transporters. It is then converted to nicotinic acid mononucleotide (NAMN) by the enzyme nicotinic acid phosphoribosyltransferase (NAPRT), which is then converted into nicotinic acid adenine dinucleotide (NAAD) by the enzymes called nicotinamide mononucleotide adenylyl transferases (NMNAT1, NMNAT2, and NMNAT3). Next, NAD⁺ synthase (NADS) transforms NAAD to NAD⁺. The NAD⁺ can be directly phosphorylated by NAD⁺ kinase (NADK) to produce NADP(H). In the Salvage pathway, the intracellular nicotinamide (NAM) is recycled back to NAD⁺ via the formation of nicotinamide mononucleotide (NMN) by intracellular nicotinamide phosphoribosyltransferase (iNAMPT). The NAM is the byproduct generated by the NAD⁺ consuming enzymes, sirtuins, poly (ADP- ribose) polymerases (PARPs), CD38, CD157, and SARM1. The Salvage pathway also uses nicotinamide riboside (NR) to produce the NMN via the enzyme nicotinamide riboside kinases 1 and 2 (NRK1 and NRK2). The cellular NAD⁺ level is balanced by biosynthesis and consumption in different subcellular compartments. For example, in the cytoplasm, the intracellular NAMPT (iNAMPT) converts NAM to NMN, further transformed to NAD⁺ by another cytoplasm-specific enzyme, NMNAT2. The NADH, generated from the NAD⁺ in the cytoplasm and utilized by Glycolysis, is transported to the mitochondria via the malate/aspartate shuttle. Via the electron transport chain (ETC), the NADH is oxidized to NAD⁺ by mitochondria specific complex I, while by tricarboxylic acid (TCA) cycle, the NAD⁺ is transformed to NADH. The mitochondrial SIRT 3, 4, 5 convert NAD⁺ to NAM. The NADH can enter the mitochondria via the glyceraldehyde 3- phosphate shuttle and results in reduced flavin adenine dinucleotide (FADH2), which is converted to the FADH mitochondrial complex II. The mitochondrial transporter SLC25A51 can also help the direct mitochondrial entry of NAD⁺. The nuclear NAD⁺ pool equilibrates with the cytosolic NAD⁺ pool by diffusion through the unidentified nuclear pore^[99,103]. The nuclear enzymes SIRT 1, 6, 7, and PARPs, CD38, SARM1, consume NAD⁺ and regulate the NAD⁺ homeostasis in the nucleus.