Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:1.6.3.1 (NADPH oxidase)
 11,281 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A full-length cDNA clone was isolated for the 47-kilodalton (kDa) subunit of the NADPH oxidase system, whose absence is responsible for the most common form of autosomally inherited chronic granulomatous disease (CGD). It encodes a 44.7-kDa polypeptide, which contains two src homology (SH3) domains and several possible sites for phosphorylation by protein kinase C. We speculate that the SH3 domains may interact with the Rap1 protein associated with cytochrome b-245 (M.T. Quinn, C.A. Parkes, L. Walker, S. Orkin, M. Dinauer, and A. Jesaitis, Nature [London] 342:198-200, 1989). An antiserum raised to the predicted C terminus of the protein detects a polypeptide with an apparent molecular mass of 47 kDa in normal neutrophil granulocytes but not in those from patients with autosomal CGD. The antibody has been used to show that the protein associates with the vacuolar membrane and is phosphorylated in response to phorbol ester treatment. Analysis of a number of tissue types and cell lines shows that expression of the gene is confined to phagocytic cells and B lymphocytes. This observation suggests that patients with CGD may also have a defect in lymphocyte function. p47 protein and mRNA levels increase during retinoic acid-induced neutrophil differentiation of HL60 cells. Nuclear run-on transcription assays show that the gene for p47 is induced at the transcriptional level in a cycloheximide-insensitive manner. These data indicate that this gene is a primary target for regulation by retinoic acid.
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PMID:Characterization of the 47-kilodalton autosomal chronic granulomatous disease protein: tissue-specific expression and transcriptional control by retinoic acid. 239 96

Recombinant tumor necrosis factor (rTNF) and rIFN-gamma induce in the human leukemia cell lines HL-60, ML3, and U937 the accumulation of transcripts of the X chromosome-linked chronic granulomatous disease (X-CGD) gene, encoding the 91-kD heavy chain of cytochrome b-245, a component of the NADPH oxidase of phagocytic cells. The gene is induced within 6 h by either cytokine, and its accumulation is observed upon induction with rIFN-gamma up to 5 d. The combined effect of the two cytokines is more than additive. rIFN-gamma also induces accumulation of X-CGD mRNA in immature myeloid cells from peripheral blood of chronic myeloid leukemia (CML) patients, whereas rTNF has almost no effect. The cells from CML patients constitutively express TNF mRNA, suggesting that endogenously produced TNF may play a role in the effect of rIFN-gamma on these cells. rTNF induces X-CGD gene expression in the myeloid cell lines acting, at least in part, at the transcriptional level, as shown in nuclear run-on experiments. The gene encoding the 22-kD light chain of cytochrome b-245 is constitutively expressed in the human myeloid cell lines and the accumulation of its transcripts is affected by neither rTNF nor rIFN-gamma, rTNF and rIFN-gamma synergistically to induce the cell lines to express the cytochrome b-245 heterodimer (as evaluated by its visible spectrum), and to produce NADPH oxidase activity and H2O2 upon stimulation with phorbol diesters.
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PMID:Tumor necrosis factor and immune interferon synergistically induce cytochrome b-245 heavy-chain gene expression and nicotinamide-adenine dinucleotide phosphate hydrogenase oxidase in human leukemic myeloid cells. 249 43

Lipopolysaccharide (LPS) pretreatment "primes" neutrophils to release increased amounts of superoxide anion (O2-) when stimulated. We investigated the molecular basis of this enhanced activity. Comparison of kinetic parameters of the respiratory burst NADPH oxidase in unstimulated LPS-primed and control neutrophils disclosed a similar Km for NADPH and no difference was seen in the content of cytochrome b. Pertussis toxin, which inhibits some G proteins, did not prevent priming. Change in membrane potential (delta psi) was five-fold greater in LPS-primed cells and paralleled the increased O2- release. Cytofluorographic analysis indicated that the increased change in delta psi was due to the creation of a new population of active cells. Changes in the concentration of intracellular free Ca2+ ([Ca2+]i) are believed to antecede changes in delta psi. There was a consistent increment (67 +/- 8%, n = 12) in resting [Ca2+]i in cells preincubated with LPS compared with control. When stimulated, the peak [Ca2+]i was significantly higher in LPS-primed cells. Ca2+-dependent protein kinase C activity was unaltered in resting and FMLP-stimulated neutrophils preexposed to LPS. Addition to cells of the intracellular Ca2+ chelator MAPTAM before preincubation with LPS blocked the changes in [Ca2+]i and the enhanced respiratory burst that characterize LPS priming. The increased resting [Ca2+]i in LPS-primed cells may enhance stimulus-induced cellular activity by modifying a Ca2+-dependent step in signal transduction.
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PMID:Lipopolysaccharide priming of human neutrophils for an enhanced respiratory burst. Role of intracellular free calcium. 253 46

A superoxide-generating NADPH oxidase was solubilized from phorbol 12-myristate 13-acetate-activated human neutrophils with a mixture of sodium deoxycholate (0.125%, w/v) and Lubrol-PX (0.125%, v/v). The solubilized preparation contained FAD (577 pmol/mg of protein) and cytochrome b-245 (479 pmol/mg of protein) and produced 11.61 mol of O2-./s per mol of cytochrome b (340 nmol of O2-./min per mg of protein). On addition of NADPH, the cytochrome b-245 was reduced by 7.9% and the FAD by 38% in the aerobic steady state; NADH addition caused little steady-state reduction of cytochrome b and FAD. In this preparation, and several others, the measured rate of O2-. production correlated with the turnover of cytochrome b calculated from the extent of cytochrome b-245 reduction under aerobic conditions. Addition of diphenyleneiodonium abolished the reduction of both the FAD and cytochrome b-245 components and inhibited O2-. production. The haem ligand imidazole inhibited O2-. generation and cytochrome b reduction while permitting FAD reduction. These results support the suggestion that the human neutrophil NADPH oxidase has the electron-transport sequence: NADPH----FAD----cytochrome b-245----O2.
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PMID:Studies on the electron-transfer mechanism of the human neutrophil NADPH oxidase. 255 3

Studies were performed to examine the lateral organization of the NADPH oxidase system in the plasma membrane of human neutrophils. Analysis of the subcellular fractionation of human neutrophils by isopycnic sedimentation of cavitated cell lysates suggested that there may be more than one population of plasma membrane vesicles formed upon cell disruption. One population (30-32% sucrose) contained surface accessible wheat germ agglutinin binding sites, alkaline phosphatase activity, and cytochrome b. Another population (34-36% sucrose) contained membrane-bound flavin and, when the cells were prestimulated with phorbol myristate acetate (PMA), NADPH-dependent superoxide generating activity. Approximately 25% of the neutrophil cytochrome b cosedimented with the heavy population, confirming our previous hypothesis (Parkos et al. (1985) J. Biol. Chem. 260, 6541-6547) that only a fraction of the total cellular cytochrome b is involved in superoxide production. The heavy plasma membrane fraction was also enriched in membrane associated actin and fodrin as detected by Western blot analysis. After extraction of the plasma membrane vesicles with detergent cocktails, the majority of superoxide generating activity remained associated with the detergent insoluble pellet. Western blot analysis demonstrated that the pellets were also enriched in actin. Further analysis of these pellets using rate-zonal detergent-containing sucrose density gradients indicated that the superoxide generating complex had an approximate sedimentation coefficient of 80 S, suggesting that the neutrophil superoxide generating system may form a complex on the plasma membrane which is associated with or somehow organized by the membrane skeletal matrix. This organization may be of functional relevance not only to the actual production of superoxide, but also to the targeting of microbicidal oxidants.
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PMID:The lateral organization of components of the membrane skeleton and superoxide generation in the plasma membrane of stimulated human neutrophils. 255 84

We examined the potential of interferon gamma (IFN-gamma) to ameliorate the physiologic defect of chronic granulomatous disease (CGD) by studying its effects on CGD phagocyte superoxide generation, NADPH oxidase kinetics, cytochrome b559 content, and expression of X-CGD (the gene for the X-linked disease). Granulocytes and macrophages from three patients in two kindreds with "variant" X-linked CGD (i.e., with very low, but detectable, baseline superoxide-generating activity) responded to IFN-gamma with enhanced nitroblue tetrazolium reduction and two- to eightfold increases in superoxide generation. IFN-gamma did not augment the respiratory burst activity of phagocytes from patients with "classic" CGD (i.e., no detectable baseline superoxide generation) or autosomal variant CGD. Incubation of a responding patient's granulocytes with IFN-gamma nearly doubled the maximal velocity for the NADPH oxidase, but did not change its abnormal Michaelis constant. Although the interferon-treated CGD granulocytes produced superoxide at a rate 40% of normal, the cytochrome b spectrum remained undetectable. IFN-gamma treatment of cultured monocytes from an IFN-gamma-responsive CGD patient increased the steady state level of RNA transcripts from the X-CGD gene from barely detectable up to approximately 5% of normal.
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PMID:Recombinant interferon gamma augments phagocyte superoxide production and X-chronic granulomatous disease gene expression in X-linked variant chronic granulomatous disease. 282 Oct 69

Here we have investigated the ability of recombinant interferon-gamma (rIFN-gamma) to modulate human neutrophil (PMN) functions. PMN incubated in the presence of rIFN-gamma showed an enhanced hydrogen peroxide production in response to Con A, FMLP, PMA or immune complexes. The effect of rIFN-gamma was dose dependent, being half maximal at 2 U/ml, and required between 90 min and 240 min of incubation to reach optimal response. The enhancing effect of rIFN-gamma on the respiratory response of PMN was not blocked by polymixin B sulphate, but an anti-rIFN-gamma monoclonal antibody and cycloheximide and actinomycin D were effective inhibitors. The enhancement of the response to Con A was not accompanied by an enhanced binding of the lectin. Neither the kinetic properties of the Con A-stimulated NADPH oxidase nor the expression of cytochrome b-245 were altered in rIFN-gamma-treated PMN. rIFN-gamma also enhanced granule secretion in response to Con A, FMLP and PMA. Initial studies on the possible alterations of transmembrane signalling in rIFN-gamma-treated PMN showed that neither inositol phosphates formation nor cytoplasmic calcium transients were altered.
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PMID:Interferon-gamma activates human neutrophil oxygen metabolism and exocytosis. 283 15

The assignment of cytochrome b-558 as a component of the O2- (H2O2) -generating enzyme in guinea-pig alveolar macrophages was investigated. Guinea pig alveolar macrophages contained 76 pmol cytochrome b-558/mg protein, a value very similar to that of neutrophils. The rate of myristic acid-stimulated O2- generation by alveolar macrophages, calculated per cytochrome b-558, was only one-fourth that of neutrophils. An analysis of Percoll density gradient centrifugation profiles showed that the H2O2-generating activity of myristic acid-activated alveolar macrophages was concentrated in a single peak which was consistently associated with 5'-nucleotidase activity, a plasma membrane marker enzyme. A little H2O2-generating activity was seen with unactivated alveolar macrophages. Furthermore, the cytochrome b-558 of both myristic acid-activated and unactivated alveolar macrophages was also predominantly associated with 5'-nucleotidase activity and was found in trace amounts in a peak containing lysozyme activity, a marker of lysosome granules. Only about 6% of the cytochrome b-558 in plasma membranes from myristic acid-activated guinea-pig alveolar macrophages was anaerobically reduced by 0.5 mM NADPH, while under the same conditions about 30% of the heme protein of myristic acid-activated neutrophils was reduced. These results suggest two conclusions: firstly, that in both activated and unactivated alveolar macrophages, cytochrome b-558 is located in the plasma membrane, and the translocation of cytochrome b-558 does not occur during the activation of NADPH oxidase; and secondly, that a smaller part of cytochrome b-558 is associated with the activated NADPH oxidase of guinea pig alveolar macrophages compared with neutrophils.
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PMID:Presence of cytochrome b-558 in guinea-pig alveolar macrophages-subcellular localization and relationship with NADPH oxidase. 283 23

NADPH-dependent superoxide production by intact human neutrophils is inhibited by DPI (diphenyleneiodonium), when stimulated by either FMLP (N-formylmethionyl-leucyl-phenylalanine) or PMA (phorbol 12-myristate 13-acetate). Addition of 10 microM-DPI abolished the reduction of both the FAD and the cytochrome b components of the NADPH oxidase. DPI inhibition of the oxidase was associated with defective aerobic killing of staphylococci by human neutrophils. Anaerobic killing, phagocytosis, chemotaxis and motility were relatively unaffected by 10 microM-DPI. Degranulation of the azurophil and specific granules, induced by the soluble stimuli FMLP or PMA, and by particulate stimuli was decreased by the presence of DPI. The above effects of DPI on human neutrophils are similar to those found in chronic granulomatous disease.
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PMID:The effect of the NADPH oxidase inhibitor diphenyleneiodonium on aerobic and anaerobic microbicidal activities of human neutrophils. 284 66

Whether or not cytochrome b-559 is a necessary component of NADPH oxidase activity in neutrophils is still controversial. In highly purified plasma membranes isolated from resting neutrophils and lacking cytochrome b, addition of arachidonic acid induced an NADPH oxidase activity. This activity was similar to that of plasma membranes isolated from phorbol myristate acetate (PMA)-stimulated cells which possessed cytochrome b. Addition of arachidonic acid to the latter plasma membranes did not alter the oxidase activity. It can be concluded that plasma membranes isolated from resting neutrophils have, in the presence of arachidonic acid, an NADPH oxidase activity similar to that of PMA-stimulated cells, except that it is independent of cytochrome b-559.
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PMID:Role of cytochrome b-559 in arachidonic acid activation of resting human neutrophils. 284 62


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