Spatiotemporal compartmentalization of hepatic NADH and NADPH metabolism

Abstract

Compartmentalization is a fundamental design principle of eukaryotic metabolism. Here, we review the compartmentalization of NAD⁺/NADH and NADP⁺/NADPH with a focus on the liver, an organ that experiences the extremes of biochemical physiology each day. Historical studies of the liver, using classical biochemical fractionation and measurements of redox-coupled metabolites, have given rise to the prevailing view that mitochondrial NAD(H) pools tend to be oxidized and important for energy homeostasis, whereas cytosolic NADP(H) pools tend to be highly reduced for reductive biosynthesis. Despite this textbook view, many questions still remain as to the relative size of these subcellular pools and their redox ratios in different physiological states, and to what extent such redox ratios are simply indicators versus drivers of metabolism. By performing a bioinformatic survey, we find that the liver expresses 352 known or predicted enzymes composing the hepatic NAD(P)ome, i.e. the union of all predicted enzymes producing or consuming NADP(H) or NAD(H) or using them as a redox co-factor. Notably, less than half are predicted to be localized within the cytosol or mitochondria, and a very large fraction of these genes exhibit gene expression patterns that vary during the time of day or in response to fasting or feeding. A future challenge lies in applying emerging new genetic tools to measure and manipulate in vivo hepatic NADP(H) and NAD(H) with subcellular and temporal resolution. Insights from such fundamental studies will be crucial in deciphering the pathogenesis of very common diseases known to involve alterations in hepatic NAD(P)H, such as diabetes and fatty liver disease.

Keywords: NAD biosynthesis; NAD(P)ome; cell metabolism; hepatic metabolism; hepatocyte; intermediary metabolism; liver; liver metabolism; nicotinamide adenine dinucleotide (NAD); nicotinamide adenine dinucleotide (NADH); oxidation-reduction (redox).

Figure 1.

Chemical, spatial, and temporal compartmentalization of metabolism.

Figure 2.

Compartmentalization of NAD(P)(H) in hepatocytes. A, classical measurements of the cytosolic and mitochondrial NAD⁺/NADH and NADP⁺/NADPH ratios in rat livers. B, classical feeding/fasting changes of compartment free NAD(P)⁺ ratios. Data based on Veech et al. (16). nd, not determined; ns, not significant.

Figure 3.

Compartmentalization of the hepatic NAD(P)ome. A, hepatic NAD(P)ome is defined as the 352 members of the human NAD(P)ome that show expression in liver. B, tissue expression is shown across 15 human tissues selected from the GTEx atlas. Expression is shown as RPKM mapped reads. C, relative distribution of hepatic enzymes using NAD(P)⁺ as cofactors in redox reactions, as substrates, or in NAD(P)⁺ biosynthesis. D, subcellular distribution of the hepatic NAD(P)ome with data available from the Human Protein Atlas (HPA). E, temporal compartmentalization during fasting/feeding is shown for 105 hepatic NADPome genes showing significant differential expression changes based on the Montagner et al. (54) study in the mouse (Student's t test p value < 0.05 after Bonferroni correction). F, gene expression is shown for the 83 genes showing significant circadian periodicity from Hughes et al. (57). Gray bars indicate periods of darkness. G, circadian expression of NADK and HMGCR based on data from Hughes et al. (57).

Figure 4.

Relative hepatic expression and feeding/fasting changes of the NAD(P) biosynthesis/salvage pathway. A, histogram of liver-specific expression of all genes, hepatic NAD(P)ome genes, and the NAD(P) biosynthesis/salvage pathway based on GNFv3 mouse tissue atlas (71). B, schematic diagram of NAD(P) biosynthesis/salvage genes showing liver-specific expression (Z-score in the GNFv3 mouse tissue atlas (71)) and in fasting versus refeeding (54), where arrows indicate significant changes based on Student's t test, after Bonferroni correction.