Nicotinic Acid Adenine Dinucleotide Phosphate Synthesis Research Articles

Interleukin-8 drives CD38 to form NAADP from NADP+ and NAAD in the endolysosomes to mobilize Ca2+ and effect cell migration.

Nicotinic acid adenine dinucleotide phosphate (NAADP) is the most potent Ca2+ mobilizing second messenger whose formation has remained elusive. In vitro, CD38-mediated NAADP synthesis requires an acidic pH and a nonphysiological concentration of nicotinic acid (NA). We discovered that CD38 catalyzes synthesis of NAADP by exchanging the nicotinamide moiety of nicotinamide adenine dinucleotide phosphate (NADP+ ) for the NA group of nicotinic acid adenine dinucleotide (NAAD) inside endolysosomes of interleukin 8 (IL8)-treated lymphokine-activated killer (LAK) cells. Upon IL8 stimulation, cytosolic NADP+ is transported to acidified endolysosomes via connexin 43 (Cx43) and gated by cAMP-EPAC-RAP1-PP2A signaling. CD38 then performs a base-exchange reaction with the donor NA group deriving from NAAD, produced by newly described endolysosomal activities of NA phosphoribosyltransferase (NAPRT) and NMN adenyltransferase (NMNAT) 3. Thus, the membrane organization of endolysosomal CD38, a signal-mediated transport system for NADP+ and luminal NAD+ biosynthetic enzymes integrate signals from a chemokine and cAMP to specify the spatiotemporal mobilization of Ca2+ to drive cell migration.

Synthesis of the Ca2+-mobilizing messengers NAADP and cADPR by intracellular CD38 enzyme in the mouse heart: Role in β-adrenoceptor signaling

Nicotinic acid adenine dinucleotide phosphate (NAADP) and cyclic ADP-ribose (cADPR) are Ca2+-mobilizing messengers important for modulating cardiac excitation–contraction coupling and pathophysiology. CD38, which belongs to the ADP-ribosyl cyclase family, catalyzes synthesis of both NAADP and cADPR in vitro. However, it remains unclear whether this is the main enzyme for their production under physiological conditions. Here we show that membrane fractions from WT but not CD38−/− mouse hearts supported NAADP and cADPR synthesis. Membrane permeabilization of cardiac myocytes with saponin and/or Triton X-100 increased NAADP synthesis, indicating that intracellular CD38 contributes to NAADP production. The permeabilization also permitted immunostaining of CD38, with a striated pattern in WT myocytes, whereas CD38−/− myocytes and nonpermeabilized WT myocytes showed little or no staining, without striation. A component of β-adrenoreceptor signaling in the heart involves NAADP and lysosomes. Accordingly, in the presence of isoproterenol, Ca2+ transients and contraction amplitudes were smaller in CD38−/− myocytes than in the WT. In addition, suppressing lysosomal function with bafilomycin A1 reduced the isoproterenol-induced increase in Ca2+ transients in cardiac myocytes from WT but not CD38−/− mice. Whole hearts isolated from CD38−/− mice and exposed to isoproterenol showed reduced arrhythmias. SAN4825, an ADP-ribosyl cyclase inhibitor that reduces cADPR and NAADP synthesis in mouse membrane fractions, was shown to bind to CD38 in docking simulations and reduced the isoproterenol-induced arrhythmias in WT hearts. These observations support generation of NAADP and cADPR by intracellular CD38, which contributes to effects of β-adrenoreceptor stimulation to increase both Ca2+ transients and the tendency to disturb heart rhythm.

Intracellular CD38 Mediates Cardiac Synthesis of NAADP and CADPR

Previously, we have reported that nicotinic acid adenine dinucleotide phosphate (NAADP) and cyclic ADP-ribose (cADPR) are calcium mobilising messengers that have significant roles in excitation-contraction coupling and disease development of the heart. The enzyme responsible for producing these molecules is a potential therapeutic target, but its identity remains unclear. CD38 is an ADP-ribosyl cyclase (ARC) that catalyses synthesis of NAADP and cADPR in various tissues, with one caveat: the targets of NAADP and cADPR are intracellular while the active site of CD38 is generally thought to be located on the extracellular surface, making the physiological relevance of this ecto-enzyme questionable. Permeabilised cardiac myocytes showed greater ARC activity than intact cells, suggesting the presence of intracellular ARCs. Immunostaining experiments showed that cell permeabilisation was critical for labelling CD38 with a striated staining pattern. We further showed that sarcoplasmic reticulum enriched membrane preparations from sheep hearts exhibited ARC activity with CD38-like properties. Genetic knock-out of CD38 in mice suppressed NAADP and cADPR synthesis. Isolated ventricular myocytes from CD38-/- mice showed reduced peak calcium transients and contractions, both under basal conditions and when stimulated by the β-adrenoceptor agonist, isoprenaline. CD38-/- hearts were more resistant to arrhythmias during over-stimulation of β-adrenoceptor pathway. A novel ARC inhibitor, SAN4825, suppressed the in vitro synthesis of NAADP and cADPR, and in blind docking simulations was shown to target the active site of CD38. The observations are consistent with an important role for CD38 in producing cADPR and NAADP intracellularly under basal conditions; the production of both these substances is increased by β-adrenoceptor stimulation and the elevated levels enhance arrhythmogenicity. In conclusion, we provide evidence that CD38 generates NAADP and cADPR in situ intracellularly and this enzymatic activity plays a role in both physiology and pathophysiology of the heart.

Synthesis of caged NAADP.

Caged derivatives of Ca²⁺-mobilizing messengers, such as nicotinic acid adenine dinucleotide phosphate (NAADP), are particularly useful for establishing the effects of these messengers on Ca²⁺ signaling. Caged NAADP is no longer commercially available but can be synthesized in house, as described here. In brief, a stable precursor of the caging reagent is made and converted to an unstable reactive reagent immediately before addition to the compound to be caged.

Autocrine/Paracrine Function of Nicotinic Acid Adenine Dinucleotide Phosphate (NAADP) for Glucose Homeostasis in Pancreatic β-Cells and Adipocytes

Nicotinic acid adenine dinucleotide phosphate (NAADP) is a second messenger for mobilizing Ca(2+) from intracellular stores in various cell types. Extracellular application of NAADP has been shown to elicit intracellular Ca(2+) signals, indicating that it is readily transported into cells. However, little is known about the functional role of this NAADP uptake system. Here, we show that NAADP is effectively transported into selected cell types involved in glucose homeostasis, such as adipocytes and pancreatic β-cells, but not the acinar cells, in a high glucose-dependent manner. NAADP uptake was inhibitable by Ned-19, a NAADP mimic; dipyridamole, a nucleoside inhibitor; or NaN3, a metabolic inhibitor or under Ca(2+)-free conditions. Furthermore, NAADP was found to be released from pancreatic islets upon stimulation by high glucose. Consistently, administration of NAADP to type 2 diabetic mice improved glucose tolerance. We propose that NAADP is functioning as an autocrine/paracrine hormone important in glucose homeostasis. NAADP is thus a potential antidiabetic agent with therapeutic relevance.

Ca2+ releasing process and nicotinic acid adenine dinucleotide phosphate

The paper is devoted to recognizing of nicotinic acid adenine dinucleotide phosphate (NAADP) as releaser of intracellular calcium from intracellular stores. It was revealed the current concepts of the mechanisms of NAADP synthesis inside the cell resuming involve enzyme CD38. The effect of NAADP was described in several cells and tissue preparation, as well as in sea urchin eggs. Briefly, it was characterized the acidic store of cells, that is sensitive to NAADP as it had been shown by different authors. Assumed mechanisms of Ca 2+ accumulating in acidic store predict involve proton gradient across membrane of acidic store. The channels structure of acidic store and endoplasmatic reticulum are considered as receptors to NAADP. Potential mechanisms for NAADP-induced calcium release was considered: direct model, trigger Ca 2+ -induced Ca 2+ -releasing model, promiscuous coupling model, conformational coupling model and also unifying hypothesis, that explains the differences between other mechanisms. Physiological processes in which NAADP is involved are described. Keywords : NAADP, acidic store, Ca 2+ , ryanodine receptor, two-pore channels.

NAADP Mediates Insulin-Stimulated Glucose Uptake and Insulin Sensitization by PPARγ in Adipocytes

Insulin stimulates glucose uptake through the membrane translocation of GLUT4 and GLUT1. Peroxisome proliferator-activated receptor γ (PPARγ) enhances insulin sensitivity. Here, we demonstrate that insulin stimulates GLUT4 and GLUT1 translocation, and glucose uptake, by activating the signaling pathway involving nicotinic acid adenine dinucleotide phosphate (NAADP), a calcium mobilizer, in adipocytes. We also demonstrate that PPARγ mediates insulin sensitization by enhancing NAADP production through upregulation of CD38, the only enzyme identified for NAADP synthesis. Insulin produced NAADP by both CD38-dependent and -independent pathways, whereas PPARγ produced NAADP by CD38-dependent pathway. Blocking the NAADP signaling pathway abrogated both insulin-stimulated and PPARγ-induced GLUT4 and GLUT1 translocation, thereby inhibiting glucose uptake. CD38 knockout partially inhibited insulin-stimulated glucose uptake. However, CD38 knockout completely blocked PPARγ-induced glucose uptake in adipocytes and PPARγ-mediated amelioration of glucose tolerance in diabetic mice. These results demonstrated that the NAADP signaling pathway is a critical molecular target for PPARγ-mediated insulin sensitization.

CD38: A NAADP degrading enzyme

The role of the multifunctional enzyme CD38 in formation of the Ca 2+-mobilizing second messenger nicotinic acid adenine dinucleotide phosphate (NAADP) was investigated. Gene silencing of CD38 did neither inhibit NAADP synthesis in intact Jurkat T cells nor in thymus or spleen obtained from CD38 knock out mice. In vitro, both NAADP formation by base-exchange and degradation to 2-phospho adenosine diphosphoribose were efficiently decreased. Thus in vivo CD38 appears to be a NAADP degrading rather than a NAADP forming enzyme, perhaps avoiding desensitizing NAADP levels in intact cells.

The Ecto-enzyme CD38 Is a Nicotinic Acid Adenine Dinucleotide Phosphate (NAADP) Synthase That Couples Receptor Activation to Ca2+ Mobilization from Lysosomes in Pancreatic Acinar Cells

Nicotinic acid adenine dinucleotide phosphate (NAADP) is the most potent Ca(2+)-mobilizing intracellular messenger and is linked to a variety of stimuli and cell surface receptors. However, the enzyme responsible for endogenous NAADP synthesis in vivo is unknown, and it has been proposed that another enzyme differing from ADP-ribosyl cyclase family members may exist. The ecto-enzyme CD38, involved in many functions as diverse as cell proliferation and social behavior, represents an important alternative. In pancreatic acinar cells, the hormone cholecystokinin (CCK) stimulates NAADP production evoking Ca(2+) signals by discharging acidic Ca(2+) stores and leading to digestive enzyme secretion. From cells derived from CD38(-/-) mice, we provide the first physiological evidence that CD38 is required for endogenous NAADP generation in response to CCK stimulation. Furthermore, CD38 expression in CD38-deficient pancreatic AR42J cells remodels Ca(2+)-signaling pathways in these cells by restoring Ca(2+) mobilization from lysosomes during CCK-induced Ca(2+) signaling. In agreement with an intracellular site for messenger synthesis, we found that CD38 is expressed in endosomes. These CD38-containing vesicles, likely of endosomal origin, appear to be proximal to lysosomes but not co-localized with them. We propose that CD38 is an NAADP synthase required for coupling receptor activation to NAADP-mediated Ca(2+) release from lysosomal stores in pancreatic acinar cells.

Generation of Cyclic ADP-ribose and Nicotinic Acid Adenine Dinucleotide Phosphate by CD38 for Ca2+ Signaling in Interleukin-8-treated Lymphokine-activated Killer Cells

We have previously demonstrated that cyclic ADP-ribose (cADPR) is a calcium signaling messenger in interleukin 8 (IL-8)-induced lymphokine-activated killer (LAK) cells. In this study we examined the possibility that IL-8 activates CD38 to produce another messenger, nicotinic acid adenine dinucleotide phosphate (NAADP), in LAK cells, and we showed that IL-8 induced NAADP formation after cADPR production. These calcium signaling messengers were not produced when LAK cells prepared from CD38 knock-out mice were treated with IL-8, indicating that the synthesis of both NAADP and cADPR is catalyzed by CD38 in LAK cells. Application of cADPR to LAK cells induced NAADP production, whereas NAADP failed to increase intracellular cADPR levels, confirming that the production of cADPR precedes that of NAADP in IL-8-treated LAK cells. Moreover, NAADP increased intracellular Ca(2+) signaling as well as cell migration, which was completely blocked by bafilomycin A1, suggesting that NAADP is generated in lysosome-related organelles after cADPR production. IL-8 or exogenous cADPR, but not NAADP, increased intracellular cAMP levels. cGMP analog, 8-(4-chlorophenylthio)-guanosine 3',5'-cyclic monophosphate, increased both cADPR and NAADP production, whereas the cAMP analog, 8-(4-chlorophenylthio)-cAMP, increased only NAADP production, suggesting that cAMP is essential for IL-8-induced NAADP formation. Furthermore, activation of Rap1, a downstream molecule of Epac, was required for IL-8-induced NAADP formation in LAK cells. Taken together, our data suggest that IL-8-induced NAADP production is mediated by CD38 activation through the actions of cAMP/Epac/protein kinase A/Rap1 in LAK cells and that NAADP plays a key role in Ca(2+) signaling of IL-8-induced LAK cell migration.

The Calcium-mobilizing Messenger Nicotinic Acid Adenine Dinucleotide Phosphate Participates in Sperm Activation by Mediating the Acrosome Reaction

Before a sperm can fertilize an egg it must undergo a final activation step induced by the egg termed the acrosome reaction. During the acrosome reaction a lysosome-related organelle, the acrosome, fuses with the plasma membrane to release hydrolytic enzymes and expose an egg-binding protein. Because NAADP (nicotinic acid adenine dinucleotide phosphate) releases Ca2+ from acidic lysosome-related organelles in other cell types, we investigated a possible role for NAADP in mediating the acrosome reaction. We report that NAADP binds with high affinity to permeabilized sea urchin sperm. Moreover, we used Mn2+ quenching of luminal fura-2 and 45Ca2+ to directly demonstrate NAADP regulation of a cation channel on the acrosome. Additionally, we show that NAADP synthesis occurs through base exchange and is driven by an increase in Ca2+. We propose a new model for acrosome reaction signaling in which Ca2+ influx initiated by egg jelly stimulates NAADP synthesis and that this NAADP acts on its receptor/channel on the acrosome to release Ca2+ to drive acrosomal exocytosis.

NAADP: A Universal Ca 2+ Trigger

Cells possess multiple calcium ion (Ca2+) stores and multiple messenger molecules to mobilize them. These include d-myo-inositol 1,4,5-trisphosphate (IP(3)), cyclic adenosine diphosphoribose (cADPR), and the most recently identified Ca2+-mobilizing messenger, nicotinic acid adenine dinucleotide phosphate (NAADP), which acts on a wide spectrum of cells, from plant cells to mammalian cells. Accumulating evidence indicates that NAADP targets both acidic (lysosome-like) Ca2+ stores and endoplasmic reticular stores. Recent studies in invertebrate and mammalian cells suggest that NAADP provides an initiating Ca2+ signal, which is amplified by cADPR- or IP(3)-dependent mechanisms (or both) through Ca2+-induced Ca2+ release. Diverse stimuli activate a rapid rise of endogenous NAADP concentration, resulting in severalfold increases of NAADP over basal values within seconds. The enzyme CD38 can catalyze both the synthesis and hydrolysis of NAADP, making it ideal for effecting the rapid metabolism of NAADP. The crystal structure of CD38 and the structures of its various substrate complexes have now been determined, clarifying the mechanism of its multifunctional catalysis. We anticipate that these advances will lead to the unmasking of all the key components of the Ca2+ signaling pathway mediated by NAADP.

Acidic Residues at the Active Sites of CD38 and ADP-ribosyl Cyclase Determine Nicotinic Acid Adenine Dinucleotide Phosphate (NAADP) Synthesis and Hydrolysis Activities

Nicotinic acid adenine dinucleotide phosphate (NAADP) is a novel metabolite of NADP that has now been established as a Ca(2+) messenger in many cellular systems. Its synthesis is catalyzed by multifunctional enzymes, CD38 and ADP-ribosyl cyclase (cyclase). The degradation pathway for NAADP is unknown and no enzyme that can specifically hydrolyze it has yet been identified. Here we show that CD38 can, in fact, hydrolyze NAADP to ADP-ribose 2'-phosphate. This activity was low at neutrality but greatly increased at acidic pH. This novel pH dependence suggests that the hydrolysis is determined by acidic residues at the active site. X-ray crystallography of the complex of CD38 with one of its substrates, NMN, showed that the nicotinamide moiety was in close contact with Glu(146) at 3.27 A and Asp(155) at 2.52 A. Changing Glu(146) to uncharged Gly and Ala, and Asp(155) to Gln and Asn, by site-directed mutagenesis indeed eliminated the strong pH dependence. Changing Asp(155) to Glu, in contrast, preserved the dependence. The specificity of the two acidic residues was further demonstrated by changing the adjacent Asp(147) to Val, which had minimal effect on the pH dependence. Crystallography confirmed that Asp(147) was situated and directed away from the bound substrate. Synthesis of NAADP catalyzed by CD38 is known to have strong preference for acidic pH, suggesting that Glu(146) and Asp(155) are also critical determinants. This was shown to be case by mutagensis. Likewise, using similar approaches, Glu(98) of the cyclase, which is equivalent to Glu(146) in CD38, was found to be responsible for controlling the pH dependence of NAADP synthesis by the cyclase. Based on these findings, a catalytic model is proposed.

NAADP as a second messenger: neither CD38 nor base-exchange reaction are necessary for in vivo generation of NAADP in myometrial cells

Nicotinic acid adenine dinucleotide phosphate (NAADP) has recently been shown to act as a second messenger controlling intracellular Ca(2+) responses in mammalian cells. Many questions remain regarding this signaling pathway, including the role of the ryanodine receptor (RyR) in NAADP-induced Ca(2+) transients. Furthermore, the exact metabolic pathway responsible for the synthesis of NAADP in vivo has not been determined. Here, we demonstrate that the NAADP mediated Ca(2+) release system is present in human myometrial cells. We also demonstrate that human myometrial cells use the NAADP second messenger system to generate intracellular Ca(2+) transients in response to histamine. It has been proposed in the past that the NAADP system in mammalian cells is dependent on the presence of functional RyRs. Here, we observed that the histamine-induced Ca(2+) transients are dependent on both the NAADP and inositol 1,4,5-trisphosphate signaling pathways but are independent of RyRs. The enzyme CD38 has been shown to catalyze the synthesis of NAADP in vitro by the base-exchange reaction. Furthermore, it has been proposed that this enzyme is responsible for the intracellular generation of NAADP in vivo. Using CD38 knockout mice, we observed that both the basal and histamine stimulated levels of NAADP are independent of CD38 and the base-exchange reaction. Our group is the first to demonstrate that NAADP is a second messenger for histamine-elicited Ca(2+) transients in human myometrial cells. Furthermore, the NAADP mediated mechanism in mammalian cells can be independent of RyRs and CD38. Our data provides novel insights into the understanding of the mechanism of action and metabolism of this new second messenger system.

Role of NAADP and cADPR in the Induction and Maintenance of Agonist-Evoked Ca2+ Spiking in Mouse Pancreatic Acinar Cells

Nicotinic acid adenine dinucleotide phosphate (NAADP) and cyclic adenosine diphosphate ribose (cADPR) were first demonstrated to mobilize Ca2+ in sea urchin eggs. In the absence of direct measurements of these messengers, pharmacological studies alone have implicated these molecules as intracellular second messengers for specific cell surface receptor agonists. We now report that in mouse pancreatic acinar cells, cholecystokinin, but not acetylcholine, evokes rapid and transient increases in NAADP levels in a concentration-dependent manner. In contrast, both cholecystokinin and acetylcholine-mediated production of cADPR followed a very different time course. The rapid and transient production of NAADP evoked by cholecystokinin precedes the onset of the Ca2+ signal and is consistent with a role for NAADP in the initiation of the Ca2+ response. Continued agonist-evoked Ca2+ spiking is maintained by prolonged elevations of cADPR levels through sensitization of Ca2+ -induced Ca2+ -release channels. This study represents the first direct comparison of NAADP and cADPR measurements, and the profound differences observed in their time courses provide evidence in support of distinct roles of these Ca2+ -mobilizing messengers in shaping specific Ca2+ signals during agonist stimulation.

Channelling of substrate promiscuity of the skeletal-muscle ADP-ribosyl cyclase isoform.

The novel Ca2+-mobilizing second messengers cADPr (cyclic ADP-ribose) and NAADP (nicotinic acid-adenine dinucleotide phosphate) are both synthesized by ADP-ribosyl cyclases. Using HSR (heavy sarcoplasmic reticulum) fractions from rabbit skeletal muscle, NAADP-induced Ca2+ release was observed. In the present paper, we show in HSR membranes the formation of authentic cADPr, cGDPr (cyclic GDP-ribose) and NAADP. The cyclization reaction to form cADPr and cGDPr as well as the base-exchange reaction to form NAADP were strictly dependent on pH. Although the formation of cGDPr is optimized at pH 6, the synthesis of NAADP was most pronounced at a pH below 5. A novel regulation mechanism is provided for nicotinic acid, the co-substrate for NAADP synthesis. Nicotinic acid had virtually no influence on the cyclization reaction, but increased the affinity of NADP at an acidic pH and had the opposite effect at alkaline pH. Nicotinamide, the side product of cADPr synthesis, is an inhibitor of the cyclization reaction (IC50, 0.7+/-0.1 mM) and was 30-fold more potent at suppressing the base-exchange reaction. Although the synthesis of NAADP was highly sensitive to nicotinamide inhibition, this was not via a competition with the nicotinic-acid-binding site. In contrast with the ecto-ADP-ribosyl cyclase (CD38), the cyclization and base-exchange reaction of the skeletal muscle isoform was inhibited by Cu2+ and Zn2+, while other bivalent cations such as Ca2+, Mg2+ and Mn2+ had virtually no effect. These findings allow for the prediction of a novel ADP-ribosyl cyclase isoform in skeletal muscle HSR, other than CD38. Hence the enzymic prerequisite for cADPr- and NAADP-mediated Ca2+ signalling is present.

A novel cycling assay for nicotinic acid-adenine dinucleotide phosphate with nanomolar sensitivity.

Nicotinic acid-adenine dinucleotide phosphate (NAADP) is a novel nucleotide derived from NADP that has now been shown to be active in releasing Ca(2+) from intracellular stores in a wide variety of cells ranging from plant to human. Despite the obvious importance of monitoring its cellular levels under various physiological conditions, no assay has been reported for NAADP to date. In the present study, a widely applicable assay for NAADP with high sensitivity is described. NAADP was first dephosphorylated to nicotinic acid-adenine dinucleotide by treatment with alkaline phosphatase. The conversion was shown to be stoichiometric. NMN-adenylyltransferase was then used to convert nicotinic acid-adenine dinucleotide into NAD in the presence of high concentrations of NMN. The resultant NAD was amplified by a cycling assay involving alcohol dehydrogenase and diaphorase. Each time NAD cycled through these coupled reactions, a molecule of highly fluorescent resorufin was generated. The reaction could be performed for hours, resulting in more than a 1000-fold amplification. Concentrations of NAADP over the 10-20 nM range could be routinely measured. This novel cycling assay was combined with an enzymic treatment to provide the necessary specificity for the assay. NAADP was found to be resistant to NADase and apyrase. Pretreatment of samples with a combination of the hydrolytic enzymes completely eliminated the interference from common nucleotides. The versatility of the cycling assay can also be extended to measure nicotinic acid, which is a substrate in the synthesis of NAADP catalysed by ADP-ribosyl cyclase, over the micromolar range. All the necessary reagents for the cycling assay are widely available and it can be performed using a multi-well fluorescence plate reader, providing a high-throughput method. This is the first assay reported for NAADP and nicotinic acid, which should be valuable in elucidating the messenger functions of NAADP.

Nicotinic acid-adenine dinucleotide phosphate (NAADP) elicits specific microsomal Ca2+ release from mammalian cells.

Nicotinic acid-adenine dinucleotide phosphate (NAADP), a molecule derived from beta-NADP, has been shown to promote intracellular calcium release in sea urchin eggs. However, there is little information regarding the role of NAADP in the regulation of intracellular calcium fluxes in mammalian cells. We found recently that several mammalian tissues have a high capacity for NAADP synthesis, as assessed by sea urchin egg bioassay. To determine the functional significance of NAADP production by mammalian tissues, we sought to determine whether NAADP is capable of inducing calcium release from microsomes prepared from cultured cells. We found that NAADP, but not beta-NADP, activates a specific microsomal calcium release system in mesangial cells isolated from rat kidney; NAADP was without effect in renal tubular epithelial cells. NAADP-induced calcium release is not affected by inhibitors of the inositol 1,4,5-trisphosphate or ryanodine channels. However, NAADP-elicited calcium release was inhibited by L-type calcium channel blockers and by alkaline phosphatase treatment of NAADP. NAADP also promotes specific microsomal calcium release in rat vascular smooth muscle cells, cardiac myocytes, fibroblasts and a human leukaemia cell line, indicating that the capacity for NAADP-induced calcium release is widespread in mammalian cells. We propose that NAADP may be an important regulator of intracellular calcium in many mammalian tissues.

Nicotinic acid–adenine dinucleotide phosphate (NAADP) elicits specific microsomal Ca2+ release from mammalian cells

Nicotinic acid–adenine dinucleotide phosphate (NAADP), a molecule derived from β-NADP, has been shown to promote intracellular calcium release in sea urchin eggs. However, there is little information regarding the role of NAADP in the regulation of intracellular calcium fluxes in mammalian cells. We found recently that several mammalian tissues have a high capacity for NAADP synthesis, as assessed by sea urchin egg bioassay. To determine the functional significance of NAADP production by mammalian tissues, we sought to determine whether NAADP is capable of inducing calcium release from microsomes prepared from cultured cells. We found that NAADP, but not β-NADP, activates a specific microsomal calcium release system in mesangial cells isolated from rat kidney; NAADP was without effect in renal tubular epithelial cells. NAADP-induced calcium release is not affected by inhibitors of the inositol 1,4,5-trisphosphate or ryanodine channels. However, NAADP-elicited calcium release was inhibited by L-type calcium channel blockers and by alkaline phosphatase treatment of NAADP. NAADP also promotes specific microsomal calcium release in rat vascular smooth muscle cells, cardiac myocytes, fibroblasts and a human leukaemia cell line, indicating that the capacity for NAADP-induced calcium release is widespread in mammalian cells. We propose that NAADP may be an important regulator of intracellular calcium in many mammalian tissues.

Nicotinic acid adenine dinucleotide phosphate: a new Ca2+ releasing agent in kidney.

Nicotinic acid adenine dinucleotide phosphate (NAADP), a molecule derived from beta-NADP, has been shown to trigger Ca2+ release from intracellular stores of invertebrate eggs and mammalian cell microsomes. NAADP-induced Ca2+ release occurs through a mechanism distinct from that of inositol-1,4,5-trisphosphate- or cyclic ADP-ribose-elicited Ca2+ release. This study investigated whether NAADP can be synthesized in rat kidney. Extracts from glomeruli, mesangial cells, and papilla have high NAADP synthetic capacities. Conversely, synthesis of NAADP in kidney cortex was almost undetectable. Furthermore, 9-cis-retinoic acid significantly up-regulated NAADP synthesis in mesangial cells. Authenticity of NAADP biosynthesis in glomeruli was affirmed by HPLC analysis. NAADP stimulated Ca2+ release from mesangial cell microsomes through a pathway distinct from that of inositol-1,4,5-trisphosphate or cyclic ADP-ribose. NAADP-triggered Ca2+ release may play an important role in regulation of renal function.