Sources and Importance of Mitochondrial NAD

Abstract Nicotinamide adenine dinucleotide (NAD) levels fall with age or disease, and rise with exercise or caloric restriction. Moreover, the demonstration that supplemental NAD precursors drive beneficial effects in rodent models has driven a resurgence in interest in the basic biology of this molecule. Although NAD is present in the mitochondrial matrix and critical to the function of the organelle, the source of mitochondrial NAD has been debated. We recently used isotopic labeling to demonstrate that direct uptake of intact NAD is one mechanism by which mitochondria are able to obtain this nucleotide. Here, we show that this activity is sufficient to restore respiratory capacity in NAD-deficient isolated mitochondria, and identify SLC25A51 as a carrier that can mediate the transport of NAD across mitochondrial membranes. Understanding the compartment-specific regulation of NAD will be crucial to understanding how cells and tissues adapt their metabolism to changes in NAD availability. Funding: DK098656 to J.A.B., GM126897 to L.A.C.

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Nicotinamide adenine dinucleotide is transported into mammalian mitochondria

eLife ◽

10.7554/elife.33246 ◽

2018 ◽

Vol 7 ◽

Cited By ~ 54

Author(s):

Antonio Davila ◽

Ling Liu ◽

Karthikeyani Chellappa ◽

Philip Redpath ◽

Eiko Nakamaru-Ogiso ◽

...

Keyword(s):

Oxidative Phosphorylation ◽

Nicotinamide Adenine Dinucleotide ◽

Mitochondrial Matrix ◽

Pyridine Nucleotides ◽

Nicotinamide Riboside ◽

Isolated Mitochondria ◽

Mitochondrial Inner Membrane ◽

Fuel Selection ◽

Nicotinamide Mononucleotide ◽

Mammalian Mitochondria

Mitochondrial NAD levels influence fuel selection, circadian rhythms, and cell survival under stress. It has alternately been argued that NAD in mammalian mitochondria arises from import of cytosolic nicotinamide (NAM), nicotinamide mononucleotide (NMN), or NAD itself. We provide evidence that murine and human mitochondria take up intact NAD. Isolated mitochondria preparations cannot make NAD from NAM, and while NAD is synthesized from NMN, it does not localize to the mitochondrial matrix or effectively support oxidative phosphorylation. Treating cells with nicotinamide riboside that is isotopically labeled on the nicotinamide and ribose moieties results in the appearance of doubly labeled NAD within mitochondria. Analogous experiments with doubly labeled nicotinic acid riboside (labeling cytosolic NAD without labeling NMN) demonstrate that NAD(H) is the imported species. Our results challenge the long-held view that the mitochondrial inner membrane is impermeable to pyridine nucleotides and suggest the existence of an unrecognized mammalian NAD (or NADH) transporter.

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Beneficial effects of methanolic extract of Indigofera linnaei Ali. on the inflammatory and nociceptive responses in rodent models

Brazilian Journal of Pharmaceutical Sciences ◽

10.1590/s1984-82502016000100013 ◽

2016 ◽

Vol 52 (1) ◽

pp. 113-123

Author(s):

Raju Senthil Kumar ◽

Balasubramanian Rajkapoor ◽

Perumal Perumal ◽

Sekar Vinoth Kumar ◽

Arunachalam Suba Geetha

Keyword(s):

Methanol Extract ◽

Latency Period ◽

Dependent Manner ◽

Rodent Models ◽

Hot Plate ◽

Concentration Dependent Manner ◽

Standard Drug ◽

Beneficial Effects ◽

Nociceptive Responses

ABSTRACT Indigofera linnaei Ali. (Tamil Name: Cheppu Nerinjil) belongs to the family Fabaceae, used for the treatment of various ailments in the traditional system of medicine. In the present study, the beneficial effects of methanol extract of whole plant of I. linnaei (MEIL) were evaluated on inflammation and nociception responses in rodent models. In vitro nitric oxide (NO), lipoxygenase (LOX) and cyclooxygense (COX) inhibitory activities were also performed to understand the mode of action. MEIL at the dose of 200 & 400 mg/kg, p.o. significantly inhibited carrageenan induced rat paw volume and reduced the weight of granuloma in cotton pellet granuloma model. The results obtained were comparable with the standard drug aceclofenac. The anti-nociceptive effect of MEIL in mice was evaluated in hot plate and acetic acid induced writhing model. The plant extract significantly reduced the number of writhes and the analgesic effect was higher than that of the standard drug aspirin. However, the extract fails to increase the latency period in hot plate method suggesting that the extract produce nociception by peripheral activity. The extract produced inhibitory effect on NO, LOX and COX in concentration dependent manner. The extract exhibited pronounced and selective COX-2 inhibition. Altogether, these results suggested that the methanol extract of Indigofera linnaei could be considered as a potential anti-inflammatory and analgesic agent.

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Beneficial Effects of Canagliflozin in Combination with Pioglitazone on Insulin Sensitivity in Rodent Models of Obese Type 2 Diabetes

PLoS ONE ◽

10.1371/journal.pone.0116851 ◽

2015 ◽

Vol 10 (1) ◽

pp. e0116851 ◽

Cited By ~ 21

Author(s):

Yoshinori Watanabe ◽

Keiko Nakayama ◽

Nobuhiko Taniuchi ◽

Yasushi Horai ◽

Chiaki Kuriyama ◽

...

Keyword(s):

Type 2 Diabetes ◽

Insulin Sensitivity ◽

Rodent Models ◽

Beneficial Effects

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Mitochondrial mislocalization and altered assembly of a cluster of Barth syndrome mutant tafazzins

The Journal of Cell Biology ◽

10.1083/jcb.200605043 ◽

2006 ◽

Vol 174 (3) ◽

pp. 379-390 ◽

Cited By ~ 104

Author(s):

Steven M. Claypool ◽

J. Michael McCaffery ◽

Carla M. Koehler

Keyword(s):

Point Mutations ◽

Mutant Gene ◽

Barth Syndrome ◽

Membrane Association ◽

Intermembrane Space ◽

Mitochondrial Matrix ◽

Mitochondrial Membranes ◽

Membrane Bilayer ◽

Conserved Residues ◽

Tm Domain

None of the 28 identified point mutations in tafazzin (Taz1p), which is the mutant gene product associated with Barth syndrome (BTHS), has a biochemical explanation. In this study, endogenous Taz1p was localized to mitochondria in association with both the inner and outer mitochondrial membranes facing the intermembrane space (IMS). Unexpectedly, Taz1p does not contain transmembrane (TM) segments. Instead, Taz1p membrane association involves a segment that integrates into, but not through, the membrane bilayer. Residues 215–232, which were predicted to be a TM domain, were identified as the interfacial membrane anchor by modeling four distinct BTHS mutations that occur at conserved residues within this segment. Each Taz1p mutant exhibits altered membrane association and is nonfunctional. However, the basis for Taz1p dysfunction falls into the following two categories: (1) mistargeting to the mitochondrial matrix or (2) correct localization associated with aberrant complex assembly. Thus, BTHS can be caused by mutations that alter Taz1p sorting and assembly within the mitochondrion, indicating that the lipid target of Taz1p is resident to IMS-facing leaflets.

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Functions of selected biochemical systems from the exercised-trained dog heart

Journal of Applied Physiology ◽

10.1152/jappl.1977.42.3.426 ◽

1977 ◽

Vol 42 (3) ◽

pp. 426-431 ◽

Cited By ~ 17

Author(s):

L. A. Sordahl ◽

G. K. Asimakis ◽

R. T. Dowell ◽

H. L. Stone

Keyword(s):

Sarcoplasmic Reticulum ◽

Calcium Transport ◽

Calcium Uptake ◽

Atp Synthesis ◽

Myocardial Cell ◽

Ruthenium Red ◽

Mitochondrial Membranes ◽

Sedentary Control ◽

Isolated Mitochondria ◽

Biochemical Systems

Mitochondria and sarcoplasmic reticulum (SR) fractions were isolated from exercised-trained (E-T) and sedentary control dog hearts. Measurements of mitochondrial respiratory functions indicated no changes in energy-producing (ATP synthesis) capacity in mitochondria from E-T compared to control dog hearts. However, the ability of isolated mitochondria from E-T hearts to retain accumulated calcium was markedly decreased compared to controls. Inhibition of mitochondrial rates of calcium uptake with the inhibitor, ruthenium red, revealed fewer binding and/or transport sites in mitochondrial membranes from exercised-trained heart preparations. ATP-dependent binding (- oxalate) and uptake (+ oxalate) of calcium by SR preparations from E-T hearts were unchanged compared to controls. In contrast, significant differences in the rates of release of bound calcium were found in SR isolated from E-T hearts. Total myocardial protein, nucleic acids, and connective tissue levels were unchanged in E-T hearts compared to controls. The results suggest subtle changes are occurring in the energy-utilizing mechanism(s) involving calcium transport of the myocardial cell during exercise training. These changes may be related to alterations in the performance of the exercised-trained heart.

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MORPHOLOGY AND ATP-ASE OF ISOLATED MITOCHONDRIA

The Journal of Cell Biology ◽

10.1083/jcb.1.2.127 ◽

1955 ◽

Vol 1 (2) ◽

pp. 127-138 ◽

Cited By ~ 50

Author(s):

Robert F. Witter ◽

Michael L. Watson ◽

Mary A. Cottone

Keyword(s):

Electron Microscope ◽

Liver Mitochondria ◽

Rat Liver Mitochondria ◽

Substantial Part ◽

Mitochondrial Matrix ◽

Isolated Mitochondria ◽

The Matrix ◽

First Time ◽

Methods Of Isolation

Changes in the morphology of rat liver mitochondria brought about by different methods of isolation and the concomitant changes in ATP-ase activity were studied. The morphology was investigated with the electron microscope. It was found that the ATP-ase activity of the isolated mitochondria cannot be readily correlated with the morphology of the mitochondria. The ATP-ase found in these preparations was latent, resembling the enzyme described in mitochondria prepared in 0.25 M sucrose. In confirmation of earlier results the use of 0.88 M sucrose yielded preparations with a higher initial ATP-ase than did other methods. Preparation in 0.25 M sucrose resulted in round, swollen mitochondria of which 30 to 40 per cent appeared to have lost a substantial part of the mitochondrial matrix. Preparations in 0.44 to 0.88 M sucrose contained mainly rod-shaped mitochondria plus a small amount of another type of swollen mitochondria. The matrix of mitochondria isolated in 0.88 M sucrose was highly condensed. By the use of 0.44 M sucrose adjusted to pH 6.2 with citric acid, it was possible to isolate, for the first time, mitochondria closely resembling those in situ and containing latent ATP-ase.

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A Regulatory Role of NAD Redox Status on Flavin Cofactor Homeostasis inS. cerevisiaeMitochondria

Oxidative Medicine and Cellular Longevity ◽

10.1155/2013/612784 ◽

2013 ◽

Vol 2013 ◽

pp. 1-16 ◽

Cited By ~ 6

Author(s):

Teresa Anna Giancaspero ◽

Vittoria Locato ◽

Maria Barile

Keyword(s):

Saccharomyces Cerevisiae ◽

Nicotinamide Adenine Dinucleotide ◽

Flavin Adenine Dinucleotide ◽

Redox Balance ◽

Redox Status ◽

Cellular Redox ◽

Isolated Mitochondria ◽

Nudix Hydrolases ◽

Order Of Magnitude

Flavin adenine dinucleotide (FAD) and nicotinamide adenine dinucleotide (NAD) are two redox cofactors of pivotal importance for mitochondrial functionality and cellular redox balance. Despite their relevance, the mechanism by which intramitochondrial NAD(H) and FAD levels are maintained remains quite unclear inSaccharomyces cerevisiae. We investigated here the ability of isolated mitochondria to degrade externally added FAD and NAD (in both its reduced and oxidized forms). A set of kinetic experiments demonstrated that mitochondrial FAD and NAD(H) destroying enzymes are different from each other and from the already characterized NUDIX hydrolases. We studied here, in some detail, FAD pyrophosphatase (EC 3.6.1.18), which is inhibited by NAD+and NADH according to a noncompetitive inhibition, withKivalues that differ from each other by an order of magnitude. These findings, together with the ability of mitochondrial FAD pyrophosphatase to metabolize endogenous FAD, presumably deriving from mitochondrial holoflavoproteins destined to degradation, allow for proposing a novel possible role of mitochondrial NAD redox status in regulating FAD homeostasis and/or flavoprotein degradation inS. cerevisiae.

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Tissue-specific regulation of sirtuin and nicotinamide adenine dinucleotide biosynthetic pathways identified in C57Bl/6 mice in response to high-fat feeding

The Journal of Nutritional Biochemistry ◽

10.1016/j.jnutbio.2016.07.013 ◽

2016 ◽

Vol 37 ◽

pp. 20-29 ◽

Cited By ~ 14

Author(s):

Janice E. Drew ◽

Andrew J. Farquharson ◽

Graham W. Horgan ◽

Lynda M. Williams

Keyword(s):

Nicotinamide Adenine Dinucleotide ◽

Nicotinamide Adenine ◽

Biosynthetic Pathways ◽

High Fat ◽

Tissue Specific ◽

Specific Regulation ◽

Fat Feeding

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MEMBRANE JUNCTIONS IN THE INTERMEMBRANE SPACE OF MITOCHONDRIA FROM MAMMALIAN TISSUES

The Journal of Cell Biology ◽

10.1083/jcb.60.3.653 ◽

1974 ◽

Vol 60 (3) ◽

pp. 653-663 ◽

Cited By ~ 24

Author(s):

Akitsugu Saito ◽

Murray Smigel ◽

Sidney Fleischer

Keyword(s):

Skeletal Muscle ◽

Cross Sections ◽

Rat Kidney ◽

Respiratory Control ◽

Heart Tissue ◽

Intermembrane Space ◽

Mitochondrial Membranes ◽

Fresh Tissue ◽

Isolated Mitochondria ◽

Mammalian Tissues

There have been several reports describing paracrystalline arrays in the intermembrane space of mitochondria. On closer inspection these structures appear to be junctions of two adjoining membranes. There are two types. They can be formed between the outer and inner mitochondrial membranes (designated outer-inner membrane junctions) or between two cristal membranes (intercristal membrane junctions). In rat heart, adjoining membranes appeared associated via a central dense midline approximately 30 Å wide. In rat kidney, the junction had a ladder-like appearance with electron-dense "bridges" approximately 80 Å wide, spaced 130 Å apart, connecting the adjoining membranes. We have investigated the conditions which favor the visualization of such structures in mitochondria. Heart mitochondria isolated rapidly from fresh tissue (within 30 min of death) contain membrane junctions in approximately 10–15% of the cross sections. This would indicate that the percentage of membrane junctions in the entire mitochondrion is far greater. Mitochondria isolated from heart tissue which was stored for 1 h at 0°–4°C showed an increased number of membrane junctions, so that 80% of the mitochondrial cross sections show membrane junctions. No membrane junctions are observed in mitochondria in rapidly fixed fresh tissue or in mitochondria isolated from tissue disrupted in fixative. Thus, the visualization of junctions in the intermembrane space of mitochondria appears to be dependent upon the storage of tissue after death. Membrane junctions can also be observed in mitochondria from other stored tissues such as skeletal muscle, kidney, and interstitial cells from large and small intestine. In each case, no such junctions are observed in these tissues when they are fixed immediately after removal from the animal. It would appear that most studies in the literature in which isolated mitochondria from tissues such as heart or kidney were used were carried out on mitochondria which contained membrane junctions. The presence of such structures does not significantly affect normal mitochondrial function in terms of respiratory control and oxidative phosphorylation.

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