Key Points
-
The phagocyte NADPH oxidase (Phox) is a well-characterized enzyme that generates high levels of superoxide and secondary oxidants in phagocytes as part of the armoury of microbicidal mechanisms by which these cells function in host defence.
-
The catalytic moiety of Phox is the membrane-associated flavocytochrome gp91phox, which is inactive in resting phagocytes, but becomes activated in the phagosomal membrane by assembly with regulatory subunits such as p47phox, p67phox and RAC.
-
A new family of homologues of gp91phox, the NADPH oxidase (NOX)/dual oxidase (DUOX) family, which now contains seven members, has recently been described, and these are expressed in various cell types, including the epithelium, smooth-muscle cells and the endothelium. These enzymes 'deliberately' generate superoxide and secondarily produce other reactive oxygen species (ROS), including hydrogen peroxide (H2O2).
-
The NOX enzymes have been proposed to function to generate ROS as mediators of signal transduction related to growth, angiogenesis and apoptosis. In addition, circumstantial evidence indicates that NOX enzymes might in some cases function in innate immunity in barrier cells, such as the colon epithelium, in a manner that is analogous to Phox.
-
The DUOX enzymes are dual function enzymes, containing not only an ROS-generating domain that is homologous to gp91phox, but also a peroxidase domain that can use the H2O2 produced by the gp91phox-homology domain to carry out oxidations of other substrates. A DUOX enzyme in the thyroid has been shown to participate in thyroid-hormone biosynthesis.
-
A Duox enzyme in Caenorhabditis elegans catalyses the crosslinking of tyrosine residues in the cuticle — an exoskeletal structure in nematode worms. The structure and function of DUOX enzymes implies a general function in the oxidative modification of the extracellular matrix or other extracellular molecules.
-
The NOX/DUOX family is implicated in various pathological conditions, including atherosclerosis, hypertension, cancer and endocrine disorders. These enzymes provide an attractive target for new therapeutic agents.
-
Our understanding of the enzymology and subunit composition of the NOX/DUOX family of enzymes is evolving rapidly. However, definitive information regarding the biological roles of these enzymes is largely lacking and will require the development of animal model systems (Drosophila and knockout mice, for example).
Abstract
Professional phagocytes generate high levels of reactive oxygen species (ROS) using a superoxide-generating NADPH oxidase as part of their armoury of microbicidal mechanisms. The multicomponent phagocyte oxidase (Phox), which has been well characterized over the past three decades, includes the catalytic subunit gp91phox. Lower levels of ROS are seen in non-phagocytic cells, but are usually thought to be 'accidental' byproducts of aerobic metabolism. The discovery of a family of superoxide-generating homologues of gp91phox has led to the concept that ROS are 'intentionally' generated in these cells with distinctive cellular functions related to innate immunity, signal transduction and modification of the extracellular matrix.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 12 print issues and online access
$209.00 per year
only $17.42 per issue
Buy this article
- Purchase on Springer Link
- Instant access to full article PDF
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
McCord, J. M. & Fridovich, I. Superoxide dismutase. An enzymic function for erythrocuprein (hemocuprein). J. Biol. Chem. 244, 6049–6055 (1969). The authors describe the discovery of superoxide dismutase and provide an elegant example of the value of basic, rather than disease-oriented, investigations in making fundamentally important discoveries with widespread implications in normal biology and pathology.
Melov, S. et al. Lifespan extension and rescue of spongiform encephalopathy in superoxide dismutase 2 nullizygous mice treated with superoxide dismutase-catalase mimetics. J. Neurosci. 21, 8348–8353 (2001).
Suh, Y. -A. et al. Cell transformation by the superoxide-generating oxidase Mox1. Nature 401, 79–82 (1999). This paper describes the discovery of the first homologue of NADPH oxidase (NOX), NOX1 (originally known as MOX1), and its expression. The article, which proposed a growth-regulatory role for NOX1, has stimulated investigations into the role of NOX enzymes in cardiovascular disease and cancer.
Lambeth, J. D., Cheng, G., Arnold, R. S. & Edens, W. E. Novel homologs of gp91phox. Trends Biochem. Sci. 25, 459–461 (2000).
Babior, B. M., Lambeth, J. D. & Nauseef, W. The neutrophil NADPH oxidase. Arch. Biochem. Biophys. 397, 342–344 (2002).
Geiszt, M., Kopp, J. B., Varnai, P. & Leto, T. L. Identification of renox, an NAD(P)H oxidase in kidney. Proc. Natl Acad. Sci. USA 97, 8010–8014 (2000).
Shiose, A. et al. A novel superoxide-producing NAD(P)H oxidase in kidney. J. Biol. Chem. 276, 1417–1423 (2001).
Lee, S. -R., Kwon, K. -S., Kim, S. -R. & Rhee, S. G. Reversible inactivation of protein tyrosine phosphatase 1B in A431 cells stimulated with epidermal growth factor. J. Biol. Chem. 273, 15366–15372 (1998). This paper describes a mechanism by which reactive oxygen can act as a signal to regulate the function of enzymes such as protein tyrosine phosphatases, implying that reactive oxygen is involved in post-translational modification of proteins in a manner that is analogous to regulatory protein phosphorylations.
Lee, S. R. et al. Reversible inactivation of the tumor suppressor PTEN by H2O2 . J. Biol. Chem. 277, 20336–20342 (2002).
Meng, T. C., Fukada, T. & Tonks, N. K. Reversible oxidation and inactivation of protein tyrosine phosphatases in vivo. Mol. Cell 9, 387–399 (2002).
Banfi, B. et al. A Ca2+-activated NADPH oxidase in testis, spleen and lymph nodes. J. Biol. Chem. 276, 37594–37601 (2001).
Zeng, J. & Fenna, R. E. X-ray crystal structure of canine myeloperoxidase at 3 A resolution. J. Mol. Biol. 226, 185–207 (1992).
Edens, W. A. et al. Tyrosine crosslinking of extracellullar matrix is catalyzed by Duox, a multidomain oxidase/peroxidase with homology to the phagocyte oxidase subunit gp91phox. J. Cell Biol. 154, 879–891 (2001). The authors use biochemistry and reverse genetics in Caenorhabditis elegans to show the function of dual oxidase (Duox) in the oxidative modification of the worm cuticle, providing evidence for a general function of Duox in the modification of the extracellular matrix or other extracellular molecules.
Vignais, P. V. The superoxide-generating NADPH oxidase: structural aspects and activation mechanism. Cell. Mol. Life Sci. 59, 1428–1459 (2002).
Ago, T. et al. Phosphorylation of p47phox directs phox homology domain from SH3 domain toward phosphoinositides, leading to phagocyte NADPH oxidase activation. Proc. Natl Acad. Sci. USA 100, 4474–4479 (2003).
Groemping, Y., Lapouge, K., Smerdon, S. J. & Rittinger, K. Molecular basis of phosphorylation-induced activation of the NADPH oxidase. Cell 113, 343–355 (2003). This paper reports on the structure of p47phox domains and provides a structural rationale for the function of p47phox as a phosphorylation-regulated organizing protein that helps to orchestrate the assembly of the phagocyte NOX (Phox).
Hiroaki, H., Ago, T., Ito, T., Sumimoto, H. & Kohda, D. Solution structure of the PX domain, a target of the SH3 domain. Nature Struct. Biol. 8, 526–530 (2001).
Han, C. -H., Freeman, J. L. R., Lee, T., Motalebi, S. A. & Lambeth, J. D. Regulation of the neutrophil respiratory burst oxidase: identification of an activation domain in p67phox. J. Biol. Chem. 273, 16663–16668 (1998).
Nisimoto, Y., Motalebi, S., Han, C. -H. & Lambeth, J. D. The p67 phox activation domain regulates electron transfer flow from NADPH to flavin in flavocytochrome b558 . J. Biol. Chem. 274, 22999–23005 (1999).
Banfi, B., Clark, R. A., Steger, K. & Krause, K. H. Two novel proteins activate superoxide generation by the NADPH oxidase NOX1. J. Biol. Chem. 278, 3510–3513 (2003).
Geiszt, M., Lekstrom, K., Witta, J. & Leto, T. L. Proteins homologous to p47phox and p67phox support superoxide production by NAD(P)H oxidase 1 in colon epithelial cells. J. Biol. Chem. 278, 20006–20012 (2003).
Takeya, R. et al. Novel human homologues of p47phox and p67phox participate in activation of superoxide-producing NADPH oxidases. J. Biol. Chem. 278, 25234–25246 (2003).
Cheng, G. & Lambeth, J. D. NOXO1, regulation of lipid binding, localization and activation of Nox1 by the phox homology (PX) domain. J. Biol. Chem. 279, 4737–4742 (2004).
Dupuy, C. et al. Purification of a novel flavoprotein involved in the thyroid NADPH oxidase. J. Biol. Chem. 274, 37265–37269 (1999). A tour de force biochemical isolation and partial cloning of the hydrogen peroxide (H 2 O 2 )-generating enzyme in the thyroid gland, which subsequently was identified by molecular cloning to be DUOX.
Eklund, E. A., Jalava, A. & Kakar, R. PU. 1, interferon regulatory factor 1, and interferon consensus sequence-binding protein cooperate to increase gp91(phox) expression. J. Biol. Chem. 273, 13957–13965 (1998).
Lassegue, B. et al. Novel gp91(phox) homologues in vascular smooth muscle cells: Nox1 mediates angiotensin II-induced superoxide formation and redox-sensitive signaling pathways. Circ. Res. 88, 888–894 (2001).
Geiszt, M. et al. NAD(P)H oxidase 1, a product of differentiated colon epithelial cells, can partially replace glycoprotein 91(phox) in the regulated production of superoxide by phagocytes. J. Immunol. 171, 299–306 (2003).
Wingler, K. et al. Upregulation of the vascular NAD(P)H-oxidase isoforms Nox1 and Nox4 by the renin–angiotensin system in vitro and in vivo. Free Radic. Biol. Med. 31, 1456–1464 (2001).
Segal, A. W. The NADPH oxidase and chronic granulomatous disease. Mol. Med. Today 2, 129–135 (1996).
Roos, D. & Winterbourn, C. C. Immunology. Lethal weapons. Science 296, 669–671 (2002).
Reeves, E. P. et al. Killing activity of neutrophils is mediated through activation of proteases by K+ flux. Nature 416, 291–297 (2002). This article describes the activation of ion fluxes by activation of Phox, and hypothesizes that bacterial killing involves a change in the ionic environment of the phagosome, which in turn activates microbicidal proteases. Reactive oxygen species (ROS)-mediated oxidations might therefore be less important than NOX activation of ion channels in microbial killing by Phox.
Kawahara, T. et al. Type I Helicobacter pylori lipopolysaccharide stimulates toll-like receptor 4 and activates mitogen oxidase 1 in gastric pit cells. Infect. Immun. 69, 4382–4389 (2001).
Teshima, S., Rokutan, K., Nikawa, T. & Kishi, K. Guinea pig gastric mucosal cells produce abundant superoxide anion through an NADPH oxidase-like system. Gastroenterology 115, 1186–1196 (1998).
Thomas, E. L., Milligan, T. W., Joyner, R. E. & Jefferson, M. M. Antibacterial activity of hydrogen peroxide and the lactoperoxidase-hydrogen peroxide-thiocyanate system against oral streptococci. Infect. Immun. 62, 529–535 (1994).
Geiszt, M., Witta, J., Baffi, J., Lekstrom, K. & Leto, T. L. Dual oxidases represent novel hydrogen peroxide sources supporting mucosal surface host defense. FASEB J. 17, 1502–1504 (2003).
Burdon, R. Superoxide and hydrogen peroxide in relation to mammalian cell proliferation. Free Radic. Biol. Med. 18, 775–794 (1995).
Arnold, R. S. et al. Hydrogen peroxide mediates the cell growth and transformation caused by the mitogenic oxidase Nox1. Proc. Natl Acad. Sci. USA 98, 5550–5555 (2001).
Perner, A., Andresen, L., Pedersen, G. & Rask-Madsen, J. Superoxide production and expression of NAD(P)H oxidases by transformed and primary human colonic epithelial cells. Gut 52, 231–236 (2003).
Chamulitrat, W. et al. Association of gp91phox homolog Nox1 with anchorage-independent growth and MAP kinase-activation of transformed human keratinocytes. Oncogene 22, 6045–6053 (2003).
Lambeth, J. D. From SOD to Phox to Nox: a personal account of the evolving views of reactive oxygen species. Rec. Advances Res. Updates 4, 31–40 (2003).
Brar, S. S. et al. NOX5 NAD(P)H oxidase regulates growth and apoptosis in DU145 prostate cancer cells. Am. J. Physiol. Cell Physiol. 285, C353–C369 (2003).
Brar, S. S. et al. An NAD(P)H oxidase regulates growth and transcription in melanoma cells. Am. J. Physiol. Cell Physiol. 282, C1212–C1224 (2002).
Arbiser, J. L. et al. Reactive oxygen generated by Nox1 triggers the angiogenic switch. Proc. Natl Acad. Sci. USA 99, 715–720 (2002).
Katsuyama, M., Fan, C. & Yabe-Nishimura, C. NADPH oxidase is involved in prostaglandin F2α-induced hypertrophy of vascular smooth muscle cells: induction of NOX1 by PGF2α. J. Biol. Chem. 277, 13438–13442 (2002).
Sorescu, D. et al. Superoxide production and expression of nox family proteins in human atherosclerosis. Circulation 105, 1429–1435 (2002).
Szocs, K. et al. Upregulation of Nox-based NAD(P)H oxidases in restenosis after carotid injury. Arterioscler. Thromb. Vasc. Biol. 22, 21–27 (2002).
Brandes, R. P. Role of NADPH oxidases in the control of vascular gene expression. Antioxid. Redox Signal 5, 803–811 (2003).
Szatrowski, T. & Nathan, C. Production of large amounts of hydrogen peroxide by human tumor cells. Cancer Research 51, 794–798 (1991).
Teshima, S., Kutsumi, H., Kawahara, T., Kishi, K. & Rokutan, K. Regulation of growth and apoptosis of cultured guinea pig gastric mucosal cells by mitogenic oxidase 1. Am. J. Physiol. Gastrointest. Liver Physiol. 279, G1169–G1176 (2000).
Helenski, L. L., Clempus, R. E., Quinn, M. T., Lambeth, J. D. & Griendling, K. K. Distinct subcellular localizations of Nox1 and Nox4 in vascular smooth muscle cells. Arterioscler. Thromb. Vasc. Biol. (in the press).
Mahadev, K. et al. The NAD(P)H oxidase homolog Nox4 modulates insulin-stimulated generation of H2O2 and plays an integral role in insulin signal transduction. Mol. Cell Biol. (in the press).
Hwang, J. et al. Pulsatile versus oscillatory shear stress regulates NADPH oxidase subunit expression. Implication for native LDL oxidation. Circ. Res. 93, 1225–1232 (2003).
Moreno, J. C. et al. Inactivating mutations in the gene for thyroid oxidase 2 (THOX2) and congenital hypothyroidism. N. Engl. J. Med. 347, 95–102 (2002).
Inanami, O. et al. Activation of the leukocyte NADPH oxidase by phorbol ester requires the phosphorylation of p47phox on Serine 303 or 304. J. Biol. Chem. 273, 9539–9543 (1998).
Hoyal, C. R. et al. Modulation of p47PHOX activity by site-specific phosphorylation: Akt-dependent activation of the NADPH oxidase. Proc. Natl Acad. Sci. USA 100, 5130–5135 (2003).
Kanai, F. et al. The PX domains of p47phox and p40phox bind to lipid products of PI(3)K. Nature Cell Biol. 3, 675–678 (2001).
Brown, G. E., Stewart, M. Q., Liu, H., Ha, V. L. & Yaffe, M. B. A novel assay system implicates PtdIns(3,4)P(2), PtdIns(3)P, and PKC δ in intracellular production of reactive oxygen species by the NADPH oxidase. Mol. Cell 11, 35–47 (2003).
Zhan, Y., Virbasius, J. V., Song, X., Pomerleau, D. P. & Zhou, G. W. The p40phox and p47phox PX domains of NADPH oxidase target cell membranes via direct and indirect recruitment by phosphoinositides. J. Biol. Chem. 277, 4512–4518 (2002).
Yaffe, M. B. The p47phox PX domain: two heads are better than one! Structure (Camb) 10, 1288–1290 (2002).
Koga, H. et al. Tetratricopeptide repeat (TPR) motifs of p67phox participate in interaction with the small GTPase Rac and activation of the phagocyte NADPH oxidase. J. Biol. Chem. 274, 25051–25060 (1999).
Lapouge, K. et al. Structure of the TPR domain of p67phox in complex with Rac.GTP. Mol. Cell 6, 899–907 (2000).
Abo, A. et al. Activation of the NADPH oxidase involves the small GTP-binding protein p21rac1. Nature 353, 668–670 (1991).
Knaus, U. G., Heyworth, P. G., Evans, T., Curnutte, J. T. & Bokoch, G. M. Regulation of phagocyte oxygen radical production by the GTP-binding protein Rac 2. Science 254, 1512–1515 (1991).
Kanofsky, J. R. Singlet oxygen production by biological systems. Chem. Biol. Interact. 70, 1–28 (1989).
Wentworth, P., Jr. et al. Evidence for antibody-catalyzed ozone formation in bacterial killing and inflammation. Science 298, 2195–2199 (2002).
Nathan, C. Immunology. Catalytic antibody bridges innate and adaptive immunity. Science 298, 2143–2144 (2002).
Fu, X., Kassim, S. Y., Parks, W. C. & Heinecke, J. W. Hypochlorous acid generated by myeloperoxidase modifies adjacent tryptophan and glycine residues in the catalytic domain of matrix metalloproteinase-7 (matrilysin): an oxidative mechanism for restraining proteolytic activity during inflammation. J. Biol. Chem. 278, 28403–28409 (2003).
Cheng, G., Cao, Z., Xu, X., van Meir, E. G. & Lambeth, J. D. Homologs of gp91phox: cloning and tissue expression of Nox3, Nox4, and Nox5. Gene 269, 131–140 (2001).
De Deken, X. et al. Cloning of two human thyroid cDNAs encoding new members of the NADPH oxidase family. J. Biol. Chem. 275, 23227–23233 (2000).
Acknowledgements
I am grateful to D. Ritsick for suggesting important concepts regarding the biology and signalling of reactive oxygen species that have been incorporated into this article.
Author information
Authors and Affiliations
Ethics declarations
Competing interests
The author declares no competing financial interests.
Glossary
- SUPEROXIDE
-
The one-electron-reduced form of molecular oxygen.
- PEROXIDASES
-
Haeme-containing enzymes that metabolize hydrogen peroxide with the concomitant oxidation of a co-substrate, such as chloride in the case of myeloperoxidase.
- NADPH
-
The reduced form of nicotinamide adenine dinucleotide phosphate. This co-enzyme serves as an electron donor for various biochemical reactions.
- RESPIRATORY BURST
-
The large increase in oxygen consumption and reactive oxygen generation that accompanies exposure of neutrophils to microorganisms and/or inflammatory mediators.
- FLAVOPROTEIN
-
A protein or enzyme that contains a flavin co-enzyme such as flavin adenine dinucleotide (FAD).
- GASTRIC PIT CELLS
-
Specialized secretory epithelial cells found in the stomach.
- SENESCENCE
-
A cellular phenotype typically seen in primary cells that have undergone many cell divisions, and typified by altered morphology and an inability to continue to divide. Cell senescence is also seen in cells that have been exposed to oxidants and other stresses.
- NUDE MICE
-
A line of mice in which lymphocyte immune function is compromised due to the congenital absence of the thymus. These mice are frequently used for tumour studies, because there is no rejection of the tumour.
- THYROGLOBULIN
-
The protein that serves as a precursor to thyroid hormone. Tyrosine residues in thyroglobulin are first iodinated in a reaction catalysed by thyroid peroxidase, and are then crosslinked to form the precursor of thyroid hormone. Proteolysis of thyroglobulin then releases the thyroid hormone from the protein structure.
Rights and permissions
About this article
Cite this article
Lambeth, J. NOX enzymes and the biology of reactive oxygen. Nat Rev Immunol 4, 181–189 (2004). https://doi.org/10.1038/nri1312
Issue Date:
DOI: https://doi.org/10.1038/nri1312
This article is cited by
-
Pathological high intraocular pressure induces glial cell reactive proliferation contributing to neuroinflammation of the blood-retinal barrier via the NOX2/ET-1 axis-controlled ERK1/2 pathway
Journal of Neuroinflammation (2024)
-
Increase in antioxidant capacity associated with the successful subclone of hypervirulent carbapenem-resistant Klebsiella pneumoniae ST11-KL64
Nature Communications (2024)
-
Effects of fine particulate matter on bone marrow-conserved hematopoietic and mesenchymal stem cells: a systematic review
Experimental & Molecular Medicine (2024)
-
Deep Insight of Design, Mechanism, and Cancer Theranostic Strategy of Nanozymes
Nano-Micro Letters (2024)
-
Novel mutations in CYBB Gene Cause X-linked chronic Granulomatous Disease in Pakistani patients
Italian Journal of Pediatrics (2023)