Differential role of nicotinamide adenine dinucleotide deficiency in acute and chronic kidney disease

Background. Nicotinamide adenine dinucleotide (NAD(+)) is a ubiquitous coenzyme involved in electron transport and a cosubstrate for sirtuin function. NAD(+) deficiency has been demonstrated in the context of acute kidney injury (AKI). Methods. We studied the expression of key NAD(+) biosynthesis enzymes in kidney biopsies from human allograft patients and patients with chronic kidney disease (CKD) at different stages. We used ischaemia-reperfusion injury (IRI) and cisplatin injection to model AKI, urinary tract obstruction [unilateral ureteral obstruction (UUO)] and tubulointerstitial fibrosis induced by proteinuria to investigate CKD in mice. We assessed the effect of nicotinamide riboside (NR) supplementation on AKI and CKD in animal models. Results. RNA sequencing analysis of human kidney allograft biopsies during the reperfusion phase showed that the NAD(+) de novo synthesis is impaired in the immediate post-transplantation period, whereas the salvage pathway is stimulated. This decrease in de novo NAD(+) synthesis was confirmed in two mouse models of IRI where NR supplementation prevented plasma urea and creatinine elevation and tubular injury. In human biopsies from CKD patients, the NAD(+) de novo synthesis pathway was impaired according to CKD stage, with better preservation of the salvage pathway. Similar alterations in gene expression were observed in mice with UUO or chronic proteinuric glomerular disease. NR supplementation did not prevent CKD progression, in contrast to its efficacy in AKI. Conclusion. Impairment of NAD(+) synthesis is a hallmark of AKI and CKD. NR supplementation is beneficial in ischaemic AKI but not in CKD models.

Role of ACMSD in regulation of NAD+ metabolism and mitochondrial function

Elena Katsyuba

Discovered in the beginning of the 20th century, nicotinamide adenine dinucleotide (NAD+) has evolved from a simple oxidoreductase cofactor to being an essential cosubstrate for a wide range of regulatory proteins that include most notoriously the sirtuin family of NAD+-dependent protein deacylases. Altered NAD+ metabolism is associated with many pathological conditions, including aging and obesity, while beneficial effects of increased NAD+ levels and subsequent sirtuin activation have been established across many different species. Due to their capacity to ameliorate mitochondrial homeostasis, organismal metabolism and lifespan, strategies aiming to boost NAD+ content possess a high therapeutic potential. This thesis describes a novel approach to increase NAD+ levels by stimulating its de novo biosynthesis from the amino acid tryptophan. The enzyme Aminocarboxymuconate Semialdehyde Decarboxylase (ACMSD) converts its substrate, ACMS, into the side branch of the pathway leading to its complete oxidation, consequently limiting the proportion of ACMS capable to generate NAD+. Since reducing ACMSD activity would prevent the flux of metabolites into the side branch and channel them towards NAD+ formation instead, the ultimate goal of this thesis was to establish the role of ACMSD in the control of the intracellular NAD+ content. First, by using genetic tools I found that reduction of ACMSD activity indeed boosts de novo NAD+ synthesis through a mechanism that is conserved from the worm C. elegans to the mouse. Beneficial effects associated with the observed increase in cellular NAD+ levels promoted mitochondrial biogenesis involving sirtuin activation. Secondly, I participated in the development of a panel of potent and selective ACMSD inhibitors. Given the restricted expression of ACMSD in the kidney and liver, its pharmacological inhibition can be a valuable strategy to increase NAD+ content in these tissues and to protect them from injury. After primary characterisation of the ACMSD inhibitors in cell models, I explored the therapeutic potential of the lead compounds to protect from non-alcoholic fatty liver disease and acute kidney injury in mice. Collectively, the data reported in this thesis describe ACMSD as an evolutionary conserved modulator of cellular NAD+ levels, sirtuin activity and mitochondrial homeostasis. Through its impact on cellular NAD+ levels ACMSD inhibition is able to preserve the hepatic and renal function from injury.

EPFL2017

Circadian and Feeding Rhythms Orchestrate the Diurnal Liver Acetylome

Florian Atger, Carlos Canto Alvarez, Sebastiano Collino, Loïc Dayon, Frédéric Bruno Martin Gachon, Martin Kussmann, Daniel Alain Dominique Marie Mauvoisin, Felix Naef, Joanna Ratajczak, Jingkui Wang

Lysine acetylation is involved in various biological processes and is considered a key reversible post-translational modification in the regulation of gene expression, enzyme activity, and subcellular localization. This post-translational modification is therefore highly relevant in the context of circadian biology, but its characterization on the proteome-wide scale and its circadian clock dependence are still poorly described. Here, we provide a comprehensive and rhythmic acetylome map of the mouse liver. Rhythmic acetylated proteins showed subcellular localization-specific phases that correlated with the related metabolites in the regulated pathways. Mitochondrial proteins were over-represented among the rhythmically acetylated proteins and were highly correlated with SIRT3-dependent deacetylation. SIRT3 activity being nicotinamide adenine dinucleotide (NAD)(+) level-dependent, we show that NAD(+) is orchestrated by both feeding rhythms and the circadian clock through the NAD(+) salvage pathway but also via the nicotinamide riboside pathway. Hence, the diurnal acetylome relies on a functional circadian clock and affects important diurnal metabolic pathways in the mouse liver.

Elsevier2017

Nicotinamide riboside kinases display redundancy in mediating nicotinamide mononucleotide and nicotinamide riboside metabolism in skeletal muscle cells

Carlos Canto Alvarez, Joanna Ratajczak

Objective: Augmenting nicotinamide adenine dinucleotide (NAD(+)) availability may protect skeletal muscle from age-related metabolic decline. Dietary supplementation of NAD(+) precursors nicotinamide mononucleotide (NMN) and nicotinamide riboside (NR) appear efficacious in elevating muscle NAD(+). Here we sought to identify the pathways skeletal muscle cells utilize to synthesize NAD(+) from NMN and NR and provide insight into mechanisms of muscle metabolic homeostasis. Methods: We exploited expression profiling of muscle NAD(+) biosynthetic pathways, single and double nicotinamide riboside kinase 1/2 (NRK1/2) loss-of-function mice, and pharmacological inhibition of muscle NAD(+) recycling to evaluate NMN and NR utilization. Results: Skeletal muscle cells primarily rely on nicotinamide phosphoribosyltransferase (NAMPT), NRK1, and NRK2 for salvage biosynthesis of NAD(+). NAMPT inhibition depletes muscle NAD(+) availability and can be rescued by NR and NMN as the preferred precursors for elevating muscle cell NAD(+) in a pathway that depends on NRK1 and NRK2. Nrk2 knockout mice develop normally and show subtle alterations to their NAD(+) metabolome and expression of related genes. NRK1, NRK2, and double KO myotubes revealed redundancy in the NRK dependent metabolism of NR to NAD(+). Significantly, these models revealed that NMN supplementation is also dependent upon NRK activity to enhance NAD(+) availability. Conclusions: These results identify skeletal muscle cells as requiring NAMPT to maintain NAD(+) availability and reveal that NRK1 and 2 display overlapping function in salvage of exogenous NR and NMN to augment intracellular NAD(+) availability. (C) 2017 The Authors. Published by Elsevier GmbH.

Elsevier Science Bv2017

Role of ACMSD in regulation of NAD+ metabolism and mitochondrial function

Elena Katsyuba

EPFL2017