Free Radical Biology & Medicine, Vol. 29, No. 5, pp. 416 – 424, 2000 Copyright © 2000 Elsevier Science Inc. Printed in the USA. All rights reserved 0891-5849/00/$–see front matter
PII S0891-5849(00)00320-8
Review Article NADPH OXIDASE: A UNIVERSAL OXYGEN SENSOR? RICHARD D. JONES,* JOHN T. HANCOCK,†
and
ALYN H. MORICE*
*Section of Respiratory Medicine, Academic Department of Medicine, The University of Hull, Hull, UK; and †Department of Biological and Biomedical Sciences, The University of the West of England, Bristol, UK (Received 10 February 2000; Revised 13 April 2000; Accepted 5 May 2000)
Abstract—NADPH oxidase is classically regarded as a key enzyme of neutrophils, where it is involved in the pathogenic production of reactive oxygen species. However, NADPH oxidase-like enzymes have recently been identified in non-neutrophil cells, supporting a separate role for NADPH-oxidase derived oxygen species in oxygen sensitive processes. This article reviews the current literature surrounding the potential role of NADPH oxidase in the oxygen sensing processes which underlie hypoxic pulmonary vasoconstriction, systemic vascular smooth muscle proliferation, carotid and airways chemoreceptor activation, erythropoietin gene expression, and oxytropic responses of plant cells. © 2000 Elsevier Science Inc. Keywords—NADPH oxidase, Pulmonary circulation, Systemic circulation, Erythropoietin, Airways, Carotid body, Plants, Free radicals
INTRODUCTION
icals is vital for defense against infection, highlighted by sufferers of chronic granulomatous disease (CGD) who are susceptible to bacterial and fungal infection due to the lack of a functional enzyme [3]. Neutrophil NADPH oxidase utilizes NADPH as its substrate and catalyzes the one-electron reduction of oxygen to superoxide (Fig. 1). It is a membrane associated multisubunit enzyme, primarily located in the plasma membranes of cells. It consists in part of a flavocytochrome that is in fact a heterodimer. The smaller or ␣ subunit is approximately 22 kDa, referred to as p22-phox, while the larger  subunit is approximately 91 kDa (gp91-phox) but is heavily glycosylated [4,5]. In quiescent neutrophils no other polypeptides are associated with the membrane, but on activation three further polypeptides of 67 kDa (p67-phox), 47 kDa (p47phox), and 40 kDa (p40-phox) are recruited. It is speculated that these cytosolic factors aid formation of the correct conformation of the flavocytochrome and are important for substrate binding and electron flow through the complex [6,7]. Activity of the complex also requires the involvement of at least two G proteins. These are proteins that act as molecular switches, alternating between binding GDP in the inactive state and GTP in the active state. The G protein Rap1A has been found to copurify with the flavocytochrome [8], while the G protein p21rac2 translocates to associate with the complex in a manner similar to that of the other cytosolic polypeptides [9,10]. Normally p21rac2 re-
NADPH oxidase is recognized as one of the primary enzyme complexes involved in the antipathogen action of neutrophils and other phagocytic white cells. This action is mediated by the production of toxic forms of oxygen species, commonly referred to as reactive oxygen species (ROS) or active oxygen species (AOS), which are released into the forming phagosome (Fig. 1). The ability of NADPH oxidase to produce toxic oxygen radRichard Jones is a Postdoctoral Research Fellow in Respiratory Medicine at the University of Hull, where he heads the pulmonary circulation research group. He obtained his PhD from the University of Sheffield in 1999. John Hancock is a Senior Lecturer in Molecular Cell Biology at the University of the West of England. He obtained his PhD in 1987 from the University of Bristol, moving to University of the West of England in 1993. He currently heads the free radical research group, whose interests include the generation and role of oxygen species in animals and plants. Alyn Morice is Professor of Respiratory Medicine and Head of the Academic Department of Medicine, University of Hull. Previously a Lecturer in Medicine at the University of Cambridge, and Senior Lecturer then Reader in Medicine at The University of Sheffield, his main research interest is the role of hypoxia in control of pulmonary arterial tone. Address correspondence to: Dr. Richard D. Jones, The University of Hull, Section of Respiratory Medicine, Academic Department of Medicine, Castle Hill Hospital, Cottingham, East Yorkshire, HU16 5JQ, UK; Tel: ⫹44 1482 624067; Fax: ⫹44 1482 624068; E-Mail:
[email protected]. 416
NADPH oxidase: a universal oxygen sensor?
Fig. 1. Generation of superoxide radicals (O2 •⫺) by NADPH oxidase [1,2]. Superoxide radicals are released into the phagosome during phagocytosis and due to their toxicity facilitate bacterial and other nonhost cell destruction. Superoxide radicals are spontaneously converted into hydrogen peroxide (H2O2) and also via the action of the enzyme superoxide dismutase. Further interaction with superoxide radicals leads to the formation of toxic hydroxyl radicals (OH•). Both these additional products are also involved in bacterial cell lysis.
sides in the cytoplasm in association with the guanine nucleotide exchange inhibitor rhoGDI, but activation is catalyzed by GDS (a GDP dissociation stimulator protein) and GDP/GTP exchange. In some cell types the G protein is another member of the G protein superfamily, p21rac1. The redox activity of the NADPH oxidase complex appears to reside in the gp91-phox subunit. Here a FAD moeity is found along with two heme groups of different
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midpoint potentials ⫺225mV and ⫺265mV [11]. This cytochrome was originally assigned a single midpoint potential and is commonly referred to as cytochrome b⫺245 [12], although this would appear to be somewhat misleading in light of recent research. Alternate nomenclature refers to the cytochrome as cytochrome b558 due to light absorbance of its ␣ band. Due to the catalytic mechanism of NADPH oxidase, electrons are passed through the enzyme complex from intracellular NADPH to extracellular molecular oxygen [13]. Consequently activation of NADPH oxidase is associated with a rapid depolarization of the membrane potential, attributed to the net outward movement of electrons [14]. The coproduced protons are initially retained intracellularly, which would result in further depolarization and a fall in pH if this occurred indefinitely. However after about a minute of stimulation, membrane depolarization reaches a steady state, even though superoxide is still being produced, indicating the movement of a compensatory charge into the external media. This is attributed to the delayed release of protons via an associated proton channel, conductance of which is voltage gated [15,16]. The overall mechanism of superoxide generation by NADPH oxidase is shown in Fig. 2. NADPH OXIDASE AS AN OXYGEN-SENSING ENZYME
Oxygen sensing is a ubiquitous process, vital throughout the plant and animal kingdoms. Plants display growth
Fig. 2. Diagrammatic representation of NADPH oxidase showing the route of passage for the protons (H⫹) and electrons (e⫺) generated from the oxidation of NADPH. The enzyme consists of an ␣ and  subunit (p22-phox and gp91-phox respectively), the latter being a FAD-containing flavoprotein (FAD-FP), two molecules of heme (cytochrome b558, Cyt b), and an associated proton channel. Full activity of the enzyme requires the translocation of polypeptide activating factors p67-phox (p67), p47-phox (p47) and possibly p40-phox (p40), and the G proteins p21rac1/2 (p21) and possibly Rap1A. Inhibitors of the enzyme complex include diphenyleneiodonium (DPI), which competes with NADPH for binding at the flavoprotein-FAD complex, and cadmium and zinc ions (Cd2⫹/Zn2⫹), which reversibly block the proton channel.
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patterns that are determined in part by oxygen tension, and lower organisms and many mammalian systems also respond to the concentration of available oxygen. Hypoxic pulmonary vasoconstriction (HPV) is the physiological process by which pulmonary arteries and capillaries constrict in response to hypoxia, ensuring correct ventilation-perfusion matching. Although HPV is the most widely reported vascular response to hypoxia, other vascular beds are sensitive to oxygen tension. A reduction in arterial PO2 usually results in vasodilation in the systemic vasculature, increasing blood flow in response to an increase in metabolic need. Other oxygen-sensitive processes include such diverse responses as systemic smooth muscle proliferation, carotid body hypoxic neural discharge, hypoxic erythropoietin gene induction, hypoxic depolarization of pulmonary neuroepithelial bodies, and oxytropic responses of plant cells. The mechanism of oxygen sensing that underlies the vast variety of these biological responses remains unknown, and is likely to be complex, requiring both an oxygen sensor to detect hypoxia, and a transduction mechanism to elicit a cellular change. There is extensive evidence of potential effector mechanisms for these processes, but what is unclear is how hypoxia is detected. Recent evidence suggests that an NADPH oxidase, similar to the neutrophil enzyme, could constitute an oxygen-sensing mechanism for such responses. The major subunit of neutrophil NADPH oxidase, gp91phox, has recently been identified in a variety of mammalian cells closely associated with oxygen-sensing processes, and has the ability to produce ROS regulated by changes in oxygen concentration, the primary characteristic of any potential oxygen sensor. Furthermore, pharmacological evidence of NADPH oxidase inhibitors altering such responses has led to speculation that they may be mediated by the reactive oxygen products generated by this enzyme. Recent evidence would suggest that the classical gp91phox subunit is not involved since oxygen sensing is preserved in both gp91phox-deficient sufferers of CGD and gp91phox-knockout mice [17]. However several homologues of gp91phox are likely to exist in humans; indeed several human gene sequences highly homologous to gp91phox have recently been deposited in GenBank. Consequently it is likely that a gp91phox homologue will be responsible for the oxygen-sensing processes described below. NADPH OXIDASE IN THE PULMONARY CIRCULATION
An NADPH oxidase has been proposed as the oxygen sensor postulated to control the contractile response of the pulmonary circulation to hypoxia. HPV ensures ventilation is matched with perfusion in the lung; vasoconstriction occurs in areas with low alveolar oxygen ten-
sions, thus avoiding systemic hypoxemia. However HPV is also responsible for the increase in pulmonary vascular resistance present in numerous disease states characterized by global alveolar hypoxia, such as chronic obstructive airways disease and acute respiratory distress syndrome. Under pathophysiological conditions, sustained HPV leads to vascular remodeling and development of pulmonary hypertension. Although first reported in 1946 [18], the underlying mechanism of HPV has yet to be fully elucidated. Initial studies centered around release of an endogenous vasoactive agent, most recently endothelin-1 [19 –22], as the trigger for HPV. More sophisticated mechanisms, whereby the level of oxygen tension in the pulmonary arterial smooth muscle cell controls membrane potential or calcium homeostasis, have since been proposed [23]. Variation in the production of ROS over the physiological oxygen tension range has been suggested as a potential effector mechanism, and recent speculation focuses on the role of an enzyme similar to neutrophil NADPH oxidase. The presence of a superoxide-generating enzyme similar to neutrophil NADPH oxidase has recently been identified in cultured calf pulmonary artery smooth muscle cells [24]. Immunoblotting identified gp91phox in both small and large pulmonary arteries and low-temperature spectrophotometry indicated the presence of cytochrome b558, uniquely associated with NADPH oxidase. In the same paper, superoxide generation was shown to be dependent on oxygen tension, being increased under hypoxic conditions. This is in contrast to earlier observations of ROS production in isolated lungs being reduced as the oxygen availability falls [23]. However, these two observations are not necessarily mutually exclusive; ROS production at the level of the whole lung may not mirror ROS production in individual pulmonary artery smooth muscle cells. Other contradictory observations surround the vasoreactivity of NADPH oxidase– derived ROS within the lung and also the efficacy of NADPH oxidase inhibitors. These opposing findings generate two potential mechanisms whereby an NADPH oxidase enzyme could act as an oxygen sensor in the lung. Further research will be required to determine which of these proposed roles is correct.
Hypothesis 1: vasoconstrictive ROS are produced under hypoxia Generation of vasoconstrictive ROS under hypoxia by an NADPH oxidase is a possible mechanism by which pulmonary vasoconstriction could be triggered in response to hypoxia. The evidence of Marshall et al. [24] that ROS are produced from an NADPH oxidase–like enzyme under hypoxia supports this hypothesis, as do the
NADPH oxidase: a universal oxygen sensor?
observations of ROS having a pulmonary vascular reactivity similar to HPV. Superoxide radicals produced via xanthine–xanthine oxidase have been shown to cause pulmonary constriction [25,26] and indirect properties of superoxide also support a vasoconstrictive role. The redox potential of the pulmonary artery smooth muscle cell has long been recognized to be altered by levels of ROS [27], and has been shown to be an important determinant of membrane channel function; cellular redox potentials generated by reducing agents such as superoxide promote potassium channel closure [28 –30] with subsequent membrane depolarization and Ca2⫹ entry via activation of voltagesensitive calcium channels. Similarly hydrogen peroxide (H2O2) has been shown to elicit pulmonary vasoconstriction [25,26,31– 40] although this occurs to supra physiological doses. The mechanism of this H2O2-induced pulmonary vasoconstriction has not been fully characterized, although it is not thought to be due to its redox properties; H2O2-mediated pulmonary vasoconstriction has been reported to be inhibited following protein kinase C [25,26] or phospholipase C and serine esterase [37] inhibition. Alternatively, recent evidence suggests H2O2 causes constriction via activation of tyrosine kinase [39]. Further evidence supportive of this hypothesis is provided by inhibitors of NADPH oxidase exhibiting efficacy against HPV. Iodonium compounds such as diphenyleneiodonium (DPI) or iodonium diphenyl (ID) produce noncompetitive inhibition of NADPH oxidase via binding covalently to FAD when the enzyme is activated [41]. An alternative mechanism of inhibition is provided by cadmium and zinc ions, which act as blockers of the proton channel of NADPH oxidase [16,42]. Thomas et al. [43] showed DPI to completely inhibit HPV in the isolated perfused rat lung. This was relatively specific since contractions to the thromboxane analogue U46619 were unaffected. DPI has also been show to inhibit HPV in isolated rat [44] and cat [24] pulmonary artery rings and in the isolated perfused rabbit lung [45]. In addition Zhang et al. [46] showed DPI to inhibit the shortening (contraction) of cultured pulmonary arterial smooth muscle cells associated with hypoxia. Observations from our group also show ID and cadmium and zinc ions to specifically inhibit HPV in isolated rat pulmonary arteries [47,48]. No effect was seen upon contraction to prostaglandin F2␣, which occurs in this preparation via extracellular calcium entry via membranous calcium channels. However iodonium compounds are also associated with several nonspecific actions, inhibiting nitric oxide synthase (NOS) [49], NADH dehydrogenase [50], and notably both potassium and calcium currents in isolated
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pulmonary artery smooth muscle cells [51]. Consequently, these alternative effects could also be interpreted to explain the observed inhibition of HPV, although distinct separation between such co-effects and NADPH oxidase inhibition with DPI has been demonstrated [45].
Hypothesis 2: a reduced production of vasodilatory ROS occurs under hypoxia Alternative mechanisms of pulmonary oxygen sensing via an NADPH oxidase have been proposed. These arise due to the fact that NADPH oxidase– derived ROS are not consistently reported to elicit pulmonary vasoconstriction, and are supported by the previously mentioned reports of ROS production in the lung occurring in proportion to the available oxygen tension [23]. One of the sources of this ROS production has been proposed to be an NAD(P)H oxidase [40,52,53]. Burke and Wolin [54] demonstrated that H2O2 produced concentration-dependent dilatation of preconstricted bovine pulmonary arteries, which could be reversed by catalase and inhibited by the soluble guanylate cyclase inhibitor methylene blue. Since cGMP levels were further elevated by, and closely linked to, the administration of catalase (but not other H2O2-metabolizing enzymes), an H2O2/catalase complex was proposed to induce pulmonary arterial dilation. Oxygen-induced pulmonary arterial dilation and HPV were subsequently demonstrated to be a function of H2O2/catalase–induced activation of guanylate cyclase [55] and the following oxygen-sensing mechanism was proposed: under normoxia large amounts of H2O2 are generated and metabolized by catalase producing the oxidized species of the enzyme, Compound I. Compound I then activates guanylate cyclase triggering vasodilatation that contributes to the low arterial tone, inherent to the pulmonary circulation. Under hypoxia, production of H2O2 and, hence, generation of Compound I is reduced, the vasodilatory control is removed, and a contraction ensues. In addition observations of NADPH oxidase– derived ROS inhibiting HPV also exist. H2O2 [56] and t-butyl hydroperoxide [31] have been shown to reverse HPV and H2O2 has also been shown to inhibit HPV via a mechanism proposed to involve inhibition of intracellular calcium release [33]. Such findings suggest that a similar mechanism of oxygen sensing to that proposed by Burke-Wolin and Wolin [55] may occur: under normoxic conditions production of H2O2 would be high and thus inhibit smooth muscle contraction, whereas under hypoxic conditions H2O2 production would be reduced and HPV allowed to occur.
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NADPH OXIDASE IN THE SYSTEMIC CIRCULATION
While in the pulmonary circulation exposure to hypoxia results in vasoconstriction, in the systemic circulation a reduction in oxygen tension triggers vasodilation in order to increase blood flow to a hypoxic area. Although a large number of references concern the mechanism of HPV, the mechanism of systemic dilatation to acute hypoxia has received little attention. To our knowledge, to date no reports have studied the possible involvement of an NADPH oxidase in this systemic response. However, reports do propose that an NADPH oxidase–like enzyme is involved in pathogenic systemic vascular smooth muscle cell proliferation, a process that is triggered by ROS production: H2O2 has been shown to stimulate vascular smooth muscle cell proliferation [57], a phenomenon that can be inhibited by administration of antioxidants [58 – 60]. Evidence for the existence of an enzyme similar to neutrophil NADPH oxidase in vascular smooth muscle cells was first shown by Fukui et al. [61] who identified the presence of p22phox in rat aortic smooth muscle cells. ROS production via this enzyme is postulated to be involved in vascular smooth muscle cell proliferation since p22phox mRNA expression is increased in rat aorta following infusion of the hypertrophic agent angiotensin II [62]. In addition angiotensin II also stimulates ROS production in vascular smooth muscle cells, which is sensitive to DPI [63,64] and dependent upon expression of p22phox [65]. Angiotensin II–induced ROS production via an NADPH oxidase–like enzyme is also reported in rabbit aortic adventitial fibroblasts [66]. Evidence suggests that ROS-induced vascular smooth muscle cell proliferation may play a key role in the development of hypertension and atherosclerosis. Hypertensive rats have been shown to have increased NADPH oxidase activity [67] and treatment of spontaneously hypertensive rats and angiotensin II hypertensive rats with superoxide dismutase results in a lowering of blood pressure [68,69]. A pathogenic role in atherosclerosis is suggested by the observation that rabbits fed on a high cholesterol diet have elevated aortic ROS production compared to controls [70]. NADPH OXIDASE AND THE CAROTID BODY
The carotid body is the structure responsible for the central detection of hypoxia. As with other hypoxic responses, the underlying mechanism whereby the carotid body is able to detect and transduce changes in PO2 remains unknown. Data also supports an NADPH oxidase as a potential oxygen sensor in the carotid body. Acker et al. [71] deduced from light absorbance spectra that an enzyme similar to neutrophil NADPH oxidase
was present in the carotid body and showed that the NADPH oxidase inhibitor DPI blocked the change in light absorbance associated with hypoxia-induced activation. In addition, Cross et al. [72] showed the same concentration of DPI to inhibit the hypoxia-induced increase in nervous chemoreceptor discharge and reduction of FAD and NADP. More recently, Kummer and Acker [73] identified the presence of p22phox, gp91phox, p47phox, and p67phox in type I cells of human, guinea pig, and rat carotid bodies via immunohistochemistry. Clearly this is strong evidence to suggest that an NADPH oxidase is involved in the oxygen-sensing process in the carotid body. However the hypoxic excitation of the carotid body is dependent on Ca2⫹ influx into type I cells of the carotid body, and evidence suggests that DPImediated inhibition may be via nonspecific blockade of these Ca2⫹ currents [74] rather than via inhibition of NADPH oxidase. In addition, the recent report of Obeso et al. [75] shows that NADPH oxidase inhibition failed to prevent the hypoxia-induced release of catecholamine associated with carotid body activation in rat and rabbit carotid body chemoreceptor cells. NADPH OXIDASE AND ERYTHROPOIETIN GENE EXPRESSION
NADPH oxidase has also been suggested as an oxygen sensor in the control of erythropoietin (epo) production, which is increased under hypoxic conditions [76]. Mechanisms similar to those postulated to govern HPV (hypothesis 2) have been proposed. Fandrey et al. [77] suggested that since H2O2 was demonstrated to inhibit epo gene expression in human hepatoma cells, high H2O2 concentrations under conditions of normoxia may suppress, and lower levels under conditions of hypoxia may allow, epo gene expression. This is supported by the observation that agents that promote generation of H2O2 also lower human hepatic cell epo production [78]. H2O2 has also recently been shown to reduce epo synthesis in human hepatoma HepG2 cells [79], whereas decreased hydroxyl radical production stimulates epo synthesis [80, 81]. Evidence suggests that the reduced production of hydroxyl radicals may be acting as the second messenger of the oxygen-sensing pathway in these cells: 3D-scanning laser microscopy reveals significant changes in the levels of hydroxyl radicals in the perinuclear space of three HepG2cell lines exposed to conditions that mimic hypoxia [82]. An NADPH oxidase–like enzyme is proposed as a source of these ROS species since HepG2 cells contain a heme protein that possesses a 22 kDa subunit and exhibits an absorbance maxima of 559 nm, associated with the plasma membrane [82– 84]. Western blot analysis also shows the presence of p47phox in these cells [83]. Fur-
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thermore, cobalt has been demonstrated to reduce ROS production in HepG2 cells via interaction with cytochrome b558, resulting in oxidation of the central heme protein [81,84]. However, in contrast, ID has also been shown to inhibit the oxygen-sensing mechanism of epo production [85], suggesting that the process may in fact be upregulated by NADPH oxidase products.
predicted to contain an EF band [98]. The signal transduction pathways downstream of the NADPH oxidase in plant tissues are also coming to light. The addition of agents that promote H2O2 production in plant cells leads to alterations in the levels of expression of several genes, such as glutathione S-transferase. Such elicitors, and H2O2 itself, lead to programmed cell death in plants [99].
NADPH OXIDASE AND AIRWAY OXYGEN SENSING
CONCLUSION
Pulmonary neuroepithelial bodies (NEB) are structurally similar to carotid chemoreceptors and are widely distributed throughout the human airways. Evidence suggests that these structures may function as hypoxiasensitive airway receptors since hypoxia [86] and DPI [87] inhibit, and H2O2 [88] increases the outward potassium current of NEB. The presence of the gp91phox and p22phox subunits of NADPH oxidase have been identified in human and rabbit NEB [86 – 88], suggesting this enzyme as the source of ROS generation. Recent evidence also suggests that an NADPH oxidase may be responsible for the oxygen-sensing ability of guinea pig tracheal neuroepithelial endocrine cells (NEE) [89].
The existence of an enzyme similar to neutrophil NADPH oxidase in nonphagocytotic cells, and its ability to produce vasoactive and redox-active oxygen species in relation to oxygen concentrations, provide an extra dimension to the action of an enzyme that has long been recognized as a key structure in bacterial killing. Evidence supports a key role for this enzyme in the oxygensensing processes of pulmonary and systemic smooth muscle cells, which may have a profound influence on a number of common disease states. Evidence also supports the involvement of an NADPH oxidase in the oxygen-sensing process in the carotid body, airways, and in erythropoietin gene expression. An NADPH oxidase– like enzyme may also mediate oxygen-sensitive processes in plants. Because this enzyme exists in such a variety of cell types, it may constitute an evolutionaryconserved biochemical link; a ubiquitous oxygen-sensing mechanism.
NADPH OXIDASE AND OXYGEN SENSING IN PLANTS
Although most of the discussion in this review is centered on mammalian systems, plants also appear to show a sensitivity to oxygen, for example, in oxytropic responses [90]. Many plant tissues contain components of NADPH oxidase and are capable of generating ROS, either constitutively or in a stimulated manner. Therefore, is it possible that an NADPH oxidase type complex is involved in oxygen-sensing processes in plants too. Generation of ROS is one of the early responses seen when a plant is challenged with a micro-organism, the so-called hypersensitive response [91]. Both superoxide ions and H2O2 are produced by plant cells, and pharmacological evidence with iodonium compounds points to the involvement of an NADPH oxidase [92]. However recent studies have thrown into doubt the use of such compounds in plants, as they also inhibit peroxidases, the other putative source of ROS in these tissues [93]. Despite this, a role for an enzyme similar to neutrophil NADPH oxidase is substantiated by other observations; the presence of subunits of this enzyme complex have been reported in several plant species using mammalian NADPH oxidase antibodies [92,94], and homologues of the large flavocytochrome subunit gp91phox have been cloned [95–97]. Evidence exists of a direct role for intracellular calcium in NADPH oxidase activation in plants, since the N-terminal end of the cloned gp91phox polypeptide is
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ABBREVIATIONS
AOS—active oxygen species Ca2⫹— calcium ion Cd2⫹— cadmium ion CdSO4— cadmium sulphate CGD— chronic granulomatous disease cGMP— cyclic GMP DPI— diphenylene iodonium e⫺— electron GDP— guanosine diphosphate GTP— guanosine triphosphate H⫹—proton H2O2— hydrogen peroxide HPV— hypoxic pulmonary vasoconstriction ID—iodonium diphenyl mRNA—messenger ribonucleic acid NADH—nicotinamide adenine dinucleotide (reduced form) NADPH—nicotinamide adenine dinucleotide phosphate (reduced form) NEB—pulmonary neuroepithelial bodies NEE—neuroepithelial endocrine cells O2•⫺—superoxide radical OH•— hydroxyl radical PGF2␣—prostaglandin F2␣ PO2—partial pressure of oxygen (oxygen tension) ROS—reactive oxygen species Zn2⫹—zinc ion
