Nmnat3 deficiency in hemolytic anemia exacerbates malaria infection.

Malaria is an infectious disease caused by Plasmodium parasites and has high mortality rates, especially among children in African and Southeast Asian countries. Patients with hemolytic anemia are suggested to adapt protective measures against malarial infection. Nicotinamide adenine dinucleotide (NAD+) is a crucial cofactor associated with numerous biological processes that maintain homeostasis in all living organisms. In a previous study, we had demonstrated that the deficiency of nicotinamide mononucleotide adenylyltransferase 3 (Nmnat3), an enzyme catalyzing NAD+ synthesis, causes hemolytic anemia accompanied by a drastic decline in the NAD+ levels in the erythrocytes. It is well known that hemolytic anemia is linked to a reduced risk of malarial infections. In the present study, we investigated whether hemolytic anemia caused by Nmnat3 deficiency is beneficial against malarial infections. We found that Nmnat3 deficiency exacerbated malarial infection and subsequently caused death. Moreover, we demonstrated that the NAD+ levels in malaria-infected Nmnat3 red blood cells significantly increased and the glycolytic flow was largely enhanced to support the rapid growth of malarial parasites. Our results revealed that hemolytic anemia induced by the deletion of Nmnat3 was harmful rather than protective against malaria.

Metabolism and biochemical properties of nicotinamide adenine dinucleotide (NAD) analogs, nicotinamide guanine dinucleotide (NGD) and nicotinamide hypoxanthine dinucleotide (NHD).

Nicotinamide adenine dinucleotide (NAD) is an important coenzyme that regulates various metabolic pathways, including glycolysis, ß-oxidation, and oxidative phosphorylation. Additionally, NAD serves as a substrate for poly(ADP-ribose) polymerase (PARP), sirtuin, and NAD glycohydrolase, and it regulates DNA repair, gene expression, energy metabolism, and stress responses. Many studies have demonstrated that NAD metabolism is deeply involved in aging and aging-related diseases. Previously, we demonstrated that nicotinamide guanine dinucleotide (NGD) and nicotinamide hypoxanthine dinucleotide (NHD), which are analogs of NAD, are significantly increased in Nmnat3-overexpressing mice. However, there is insufficient knowledge about NGD and NHD in vivo. In the present study, we aimed to investigate the metabolism and biochemical properties of these NAD analogs. We demonstrated that endogenous NGD and NHD were found in various murine tissues, and their synthesis and degradation partially rely on Nmnat3 and CD38. We have also shown that NGD and NHD serve as coenzymes for alcohol dehydrogenase (ADH) in vitro, although their affinity is much lower than that of NAD. On the other hand, NGD and NHD cannot be used as substrates for SIRT1, SIRT3, and PARP1. These results reveal the basic metabolism of NGD and NHD and also highlight their biological function as coenzymes.

Overexpression of Nmnat3 efficiently increases NAD and NGD levels and ameliorates age-associated insulin resistance.

Nicotinamide adenine dinucleotide (NAD) is an important cofactor that regulates various biological processes, including metabolism and gene expression. As a coenzyme, NAD controls mitochondrial respiration through enzymes of the tricarboxylic acid (TCA) cycle, ß-oxidation, and oxidative phosphorylation and also serves as a substrate for posttranslational protein modifications, such as deacetylation and ADP-ribosylation by sirtuins and poly(ADP-ribose) polymerase (PARP), respectively. Many studies have demonstrated that NAD levels decrease with aging and that these declines cause various aging-associated diseases. In contrast, activation of NAD metabolism prevents declines in NAD levels during aging. In particular, dietary supplementation with NAD precursors has been associated with protection against age-associated insulin resistance. However, it remains unclear which NAD synthesis pathway is important and/or efficient at increasing NAD levels in vivo. In this study, Nmnat3 overexpression in mice efficiently increased NAD levels in various tissues and prevented aging-related declines in NAD levels. We also demonstrated that Nmnat3-overexpressing (Nmnat3 Tg) mice were protected against diet-induced and aging-associated insulin resistance. Moreover, in skeletal muscles of Nmnat3 Tg mice, TCA cycle activity was significantly enhanced, and the energy source for oxidative phosphorylation was shifted toward fatty acid oxidation. Furthermore, reactive oxygen species (ROS) generation was significantly suppressed in aged Nmnat3 Tg mice. Interestingly, we also found that concentrations of the NAD analog nicotinamide guanine dinucleotide (NGD) were dramatically increased in Nmnat3 Tg mice. These results suggest that Nmnat3 overexpression improves metabolic health and that Nmnat3 is an attractive therapeutic target for metabolic disorders that are caused by aging.

Deficiency of nicotinamide mononucleotide adenylyltransferase 3 (nmnat3) causes hemolytic anemia by altering the glycolytic flow in mature erythrocytes.

NAD biosynthesis is of substantial interest because of its important roles in regulating various biological processes. Nicotinamide mononucleotide adenylyltransferase 3 (Nmnat3) is considered a mitochondria-localized NAD synthesis enzyme involved in de novo and salvage pathways. Although the biochemical properties of Nmnat3 are well documented, its physiological function in vivo remains unclear. In this study, we demonstrated that Nmnat3 was localized in the cytoplasm of mature erythrocytes and critically regulated their NAD pool. Deficiency of Nmnat3 in mice caused splenomegaly and hemolytic anemia, which was associated with the findings that Nmnat3-deficient erythrocytes had markedly lower ATP levels and shortened lifespans. However, the NAD level in other tissues were not apparently affected by the deficiency of Nmnat3. LC-MS/MS-based metabolomics revealed that the glycolysis pathway in Nmnat3-deficient erythrocytes was blocked at a glyceraldehyde 3-phosphate dehydrogenase (GAPDH) step because of the shortage of the coenzyme NAD. Stable isotope tracer analysis further demonstrated that deficiency of Nmnat3 resulted in glycolysis stall and a shift to the pentose phosphate pathway. Our findings indicate the critical roles of Nmnat3 in maintenance of the NAD pool in mature erythrocytes and the physiological impacts at its absence in mice.