Superoxide generation and tyrosine kinase

2000 ◽  
Vol 78 (1) ◽  
pp. 11-17 ◽  
Author(s):  
Su Yang ◽  
Martha Hardaway ◽  
George Sun ◽  
William L Ries ◽  
L Lyndon Key Jr
Keyword(s):  
Cell Surface ◽  
Nadph Oxidase ◽  
Tyrosine Kinase ◽  
B Lymphocytes ◽  
Specific Inhibitor ◽  
Superoxide Production ◽  
Regulatory Subunit ◽  
Superoxide Generation ◽  
Herbimycin A ◽  
Nadph Oxidase Complex

NADPH oxidase is a multi-subunit enzyme complex responsible for superoxide generation in many cells, for example, B-lymphocytes and osteoclasts. NADPH oxidase is localized on the cell surface and generates superoxide extracellularly. After synthesis, components of this oxidase are transported to the cell membrane where the functional NADPH oxidase complex is assembled. The mechanism by which the membrane-bound components are transported to the cell surface of osteoclasts remains unclear. In this study, we examined the role of tyrosine kinase activity in the transport of NADPH oxidase components. When B-lymphocytes and osteoclasts were treated with herbimycin A, a specific inhibitor of tyrosine kinase, superoxide production was significantly decreased. The amount of p91, the catalytic subunit of NADPH oxidase, was decreased in the cellular membrane of herbimycin A treated cells compared to untreated controls. Similar results were obtained for the movement of a regulatory subunit of the NADPH oxidase complex, p47, in B-lymphocytes. Thus, inhibition of tyrosine kinase decreases superoxide production by disrupting the translocation of the NADPH oxidase complex.

Diabetes induces pulmonary artery endothelial dysfunction by NADPH oxidase induction

2008 ◽  
Vol 295 (5) ◽  
pp. L727-L732 ◽  
Author(s):  
Jose G. Lopez-Lopez ◽  
Javier Moral-Sanz ◽  
Giovanna Frazziano ◽  
Maria J. Gomez-Villalobos ◽  
Jorge Flores-Hernandez ◽  
...  
Keyword(s):  
Endothelial Dysfunction ◽  
Nadph Oxidase ◽  
Pulmonary Arteries ◽  
Diabetic Rats ◽  
Oxidase Inhibitor ◽  
No Synthase ◽  
Superoxide Production ◽  
Regulatory Subunit ◽  
Sprague Dawley ◽  
Relaxant Response

Recent data suggest that diabetes is a risk factor for pulmonary hypertension. The aim of the present study was to analyze whether diabetes induces endothelial dysfunction in pulmonary arteries and the mechanisms involved. Male Sprague-Dawley rats were randomly divided into a control (saline) and a diabetic group (70 mg/kg−1 streptozotocin). After 6 wk, intrapulmonary arteries were mounted for isometric tension recording, and endothelial function was tested by the relaxant response to acetylcholine. Protein expression and localization were measured by Western blot and immunohistochemistry and superoxide production by dihydroethidium staining. Pulmonary arteries from diabetic rats showed impaired relaxant response to acetylcholine and reduced vasoconstrictor response to the nitric oxide (NO) synthase inhibitor l-NAME, whereas the response to nitroprusside and the expression of endothelial NO synthase remained unchanged. Endothelial dysfunction was reversed by addition of superoxide dismutase or the NADPH oxidase inhibitor apocynin. An increase in superoxide production and increased expression of the NADPH oxidase regulatory subunit p47phox were also found in pulmonary arteries from diabetic rats. In conclusion, the pulmonary circulation is a target for diabetes-induced endothelial dysfunction via enhanced NADPH oxidase-derived superoxide production.

scholarly journals Crumbs limits oxidase-dependent signaling to maintain epithelial integrity and prevent photoreceptor cell death

2012 ◽  
Vol 198 (6) ◽  
pp. 991-998 ◽  
Author(s):  
François J.-M. Chartier ◽  
Émilie J.-L. Hardy ◽  
Patrick Laprise
Keyword(s):  
Nadph Oxidase ◽  
Molecular Mechanisms ◽  
Light Exposure ◽  
Photoreceptor Cells ◽  
Protective Role ◽  
Superoxide Production ◽  
Epithelial Tissue ◽  
Complex Activity ◽  
Tissue Organization ◽  
Nadph Oxidase Complex

Drosophila melanogaster Crumbs (Crb) and its mammalian orthologues (CRB1–3) share evolutionarily conserved but poorly defined roles in regulating epithelial polarity and, in photoreceptor cells, morphogenesis and stability. Elucidating the molecular mechanisms of Crb function is vital, as mutations in the human CRB1 gene cause retinal dystrophies. Here, we report that Crb restricts Rac1–NADPH oxidase-dependent superoxide production in epithelia and photoreceptor cells. Reduction of superoxide levels rescued epithelial defects in crb mutant embryos, demonstrating that limitation of superoxide production is a crucial function of Crb and that NADPH oxidase and superoxide contribute to the molecular network regulating epithelial tissue organization. We further show that reduction of Rac1 or NADPH oxidase activity or quenching of reactive oxygen species prevented degeneration of Crb-deficient retinas. Thus, Crb fulfills a protective role during light exposure by limiting oxidative damage resulting from Rac1–NADPH oxidase complex activity. Collectively, our results elucidate an important mechanism by which Crb functions in epithelial organization and the prevention of retinal degeneration.

scholarly journals Occurrence of cytochrome b558 in B-cell lineage of human lymphocytes

Blood ◽  
1990 ◽  
Vol 75 (2) ◽  
pp. 458-461
Author(s):  
S Kobayashi ◽  
S Imajoh-Ohmi ◽  
M Nakamura ◽  
S Kanegasaki
Keyword(s):  
B Cells ◽  
Cell Surface ◽  
B Cell ◽  
B Lymphocytes ◽  
Plasma Cells ◽  
Human Lymphocytes ◽  
Cell Lineage ◽  
Superoxide Generation ◽  
Cell Stage ◽  
Cytochrome B558

Cytochrome b558, involved in superoxide generation in phagocytes, was found to be expressed on the cell surface of most normal peripheral B lymphocytes. The cytochrome was not found on the surface of peripheral T lymphocytes, natural killer cells, or peripheral lymphocytes derived from patients with X-linked chronic granulomatous disease. On stimulation, at least half of peripheral B lymphocytes could generate superoxide anion as detected by superoxide dismutase-sensitive nitroblue tetrazorium reduction. Cytochrome b558 was not present on the surface of pre-pre B cells or pre-B cells, but did appear at the early B-cell stage. It disappeared from the B-cell surface during terminal differentiation to plasma cells. The transient expression of the cytochrome in B-cell lineage may indicate that superoxide generation is important for the function of these cells at certain stages.

scholarly journals HBP1 Repression of the p47phox Gene: Cell Cycle Regulation via the NADPH Oxidase

2004 ◽  
Vol 24 (7) ◽  
pp. 3011-3024 ◽  
Author(s):  
Stephen P. Berasi ◽  
Mei Xiu ◽  
Amy S. Yee ◽  
K. Eric Paulson
Keyword(s):  
Cell Cycle ◽  
Nadph Oxidase ◽  
Cell Cycle Regulation ◽  
Dominant Negative ◽  
Cellular Growth ◽  
Superoxide Production ◽  
Regulatory Subunit ◽  
Dominant Negative Mutant ◽  
Ros Generation ◽  
Cycle Regulation

ABSTRACT Several studies have linked the production of reactive oxygen species (ROS) by the NADPH oxidase to cellular growth control. In many cases, activation of the NADPH oxidase and subsequent ROS generation is required for growth factor signaling and mitogenesis in nonimmune cells. In this study, we demonstrate that the transcriptional repressor HBP1 (HMG box-containing protein 1) regulates the gene for the p47phox regulatory subunit of the NADPH oxidase. HBP1 represses growth regulatory genes (e.g., N-Myc, c-Myc, and cyclin D1) and is an inhibitor of G1 progression. The promoter of the p47phox gene contains six tandem high-affinity HBP1 DNA-binding elements at positions −1243 to −1318 bp from the transcriptional start site which were required for repression. Furthermore, HBP1 repressed the expression of the endogenous p47phox gene through sequence-specific binding. With HBP1 expression and the subsequent reduction in p47phox gene expression, intracellular superoxide production was correspondingly reduced. Using both the wild type and a dominant-negative mutant of HBP1, we demonstrated that the repression of superoxide production through the NADPH oxidase contributed to the observed cell cycle inhibition by HBP1. Together, these results indicate that HBP1 may contribute to the regulation of NADPH oxidase-dependent superoxide production through transcriptional repression of the p47phox gene. This study defines a transcriptional mechanism for regulating intracellular ROS levels and has implications in cell cycle regulation.

scholarly journals Molecular Characterization of the NADPH Oxidase Complex in the Ergot Fungus Claviceps purpurea: CpNox2 and CpPls1 Are Important for a Balanced Host-Pathogen Interaction

2013 ◽  
Vol 26 (10) ◽  
pp. 1151-1164 ◽  
Author(s):  
Janine Schürmann ◽  
Dagmar Buttermann ◽  
Andrea Herrmann ◽  
Sabine Giesbert ◽  
Paul Tudzynski
Keyword(s):  
Nadph Oxidase ◽  
Pathogenic Fungi ◽  
Infection Process ◽  
Regulatory Subunit ◽  
Claviceps Purpurea ◽  
Oxygen Species ◽  
Disease Symptoms ◽  
Host Pathogen Interaction ◽  
Nadph Oxidase Complex

Reactive oxygen species producing NADPH oxidase (Nox) complexes are involved in defense reactions in animals and plants while they trigger infection-related processes in pathogenic fungi. Knowledge about the composition and localization of these complexes in fungi is limited; potential components identified thus far include two to three catalytical subunits, a regulatory subunit (NoxR), the GTPase Rac, the scaffold protein Bem1, and a tetraspanin-like membrane protein (Pls1). We showed that, in the biotrophic grass-pathogen Claviceps purpurea, the catalytical subunit CpNox1 is important for infection. Here, we present identification of major Nox complex partners and a functional analysis of CpNox2 and the tetraspanin CpPls1. We show that, as in other fungi, Nox complexes are important for formation of sclerotia; CpRac is, indeed, a complex partner because it interacts with CpNoxR, and CpNox1/2 and CpPls1 are associated with the endoplasmatic reticulum. However, unlike in all other fungi, Δcppls1 is more similar to Δcpnox1 than to Δcpnox2, and CpNox2 is not essential for infection. In contrast, Δcpnox2 shows even more pronounced disease symptoms, indicating that Cpnox2 controls the infection process and moderates damage to the host. These data confirm that fungal Nox complexes have acquired specific functions dependent of the lifestyle of the pathogen.

scholarly journals Leishmania pifanoi Amastigotes Avoid Macrophage Production of Superoxide by Inducing Heme Degradation

2005 ◽  
Vol 73 (12) ◽  
pp. 8322-8333 ◽  
Author(s):  
Nam-Kha Pham ◽  
Jennifer Mouriz ◽  
Peter E. Kima
Keyword(s):  
Nadph Oxidase ◽  
Subcellular Fractions ◽  
Superoxide Production ◽  
Heme Oxygenase 1 ◽  
Heme Degradation ◽  
Nadph Oxidase Complex ◽  
Immature Form ◽  
Oxidase Enzyme ◽  
Infected Cells ◽  
Rate Limiting

ABSTRACT Whereas infections of macrophages by promastigote forms of Leishmania mexicana pifanoi induce the production of superoxide, infections by amastigotes barely induce superoxide production. Several approaches were employed to gain insight into the mechanism by which amastigotes avoid eliciting superoxide production. First, in experiments with nitroblue tetrazolium, we found that 25% of parasitophorous vacuoles (PVs) that harbor promastigotes are positive for the NADPH oxidase complex, in contrast to only 2% of PVs that harbor amastigotes. Second, confocal microscope analyses of infected cells labeled with antibodies to gp91phox revealed that this enzyme subunit is found in PVs that harbor amastigotes. Third, in immunoblots of subcellular fractions enriched with PVs from amastigote-infected cells and probed with antibodies to gp91phox, only the 65-kDa premature form of gp91phox was found. In contrast, subcellular fractions from macrophages that ingested zymosan particles contained both the 91- and 65-kDa forms of gp91phox. This suggested that only the immature form of gp91phox is recruited to PVs that harbor amastigotes. Given that gp91phox maturation is dependent on the availability of heme, we found that infections by Leishmania parasites induce an increase in heme oxygenase 1 (HO-1), the rate-limiting enzyme in heme degradation. Infections by amastigotes performed in the presence of metalloporphyrins, which are inhibitors of HO-1, resulted in superoxide production by infected macrophages. Taken together, we propose that Leishmania amastigotes avoid superoxide production by inducing an increase in heme degradation, which results in blockage of the maturation of gp91phox, which prevents assembly of the NADPH oxidase enzyme complex.

Glutamine plays a role in superoxide production and the expression of p47phox, p22phox and gp91phox in rat neutrophils

2002 ◽  
Vol 103 (4) ◽  
pp. 403-408 ◽  
Author(s):  
Tania Cristina PITHON-CURI ◽  
Adriana C. LEVADA ◽  
Lúcia R. LOPES ◽  
Sonia Q. DOI ◽  
Rui CURI
Keyword(s):  
Amino Acid ◽  
Cytochrome C ◽  
Nadph Oxidase ◽  
Superoxide Anion ◽  
Scintillation Counting ◽  
Superoxide Production ◽  
Glutamine Metabolism ◽  
Nadph Oxidase Complex ◽  
O2 Production

The effect of glutamine on the activity of the NADPH oxidase complex from rat neutrophils was investigated. Superoxide anion (O2-) production was assessed: (1) by scintillation counting by using lucigenin, and (2) by reduction of cytochrome c over 10min. The effects of glutamine and PMA on the expression of the NADPH oxidase components p22phox, gp91phox and p47phox were also determined. Glutamine at 1 and 2mM increased O2- generation in the presence of PMA by 100% and 74% respectively, in neutrophils maintained previously for 3h in medium deprived of this amino acid. DON (6-diazo-5-oxo-l-norleucine), an inhibitor of phosphate-dependent glutaminase and thus of glutamine metabolism, caused a significant decrease in O2- production by neutrophils stimulated with PMA both in the absence (44%) and in the presence (66%) of glutamine. PMA markedly increased the expression of gp91phox, p22phox and p47phox mRNAs. Glutamine (2mM) increased the expression of these three proteins both in the absence and in the presence of PMA. We postulate that glutamine leads to O2- production in neutrophils, probably via the generation of ATP and regulation of the expression of components of NADPH oxidase.

scholarly journals Hypoxic induction of gene expression in chronic granulomatous disease- derived B-cell lines: oxygen sensing is independent of the cytochrome b558-containing nicotinamide adenine dinucleotide phosphate oxidase

Blood ◽  
1996 ◽  
Vol 87 (2) ◽  
pp. 756-761 ◽  
Author(s):  
RH Wenger ◽  
HH Marti ◽  
CC Schuerer-Maly ◽  
I Kvietikova ◽  
C Bauer ◽  
...  
Keyword(s):  
Nadph Oxidase ◽  
B Cell ◽  
Cell Lines ◽  
Nicotinamide Adenine Dinucleotide ◽  
Oxygen Sensing ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Wild Type ◽  
Superoxide Generation ◽  
Cytochrome B558 ◽  
Nadph Oxidase Complex

Reduced oxygenation of a variety of cells results in transcriptional upregulation of several genes, including the hematopoietic hormone erythropoietin, the angiogenic vascular endothelial growth factor (VEGF), and glycolytic enzymes such as aldolase. Recently, the heme protein cytochrome b558 of the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase complex has been proposed as a key component of the oxygen-sensing mechanism. Cytochrome b558 consists of the p22phox and gp91phox subunits and is essential for superoxide generation in phagocytes and B lymphocytes. Mutations in these subunits result in cytochrome b558-negative chronic granulomatous disease (cytb- CGD), an inherited disorder in humans characterized by reduced microbicidal activity due to deficient superoxide generation. To test whether NADPH oxidase is involved in oxygen sensing, we exposed wild- type B-cell lines as well as cytb- CGD-derived B cell lines, deficient in either p22phox or gp91phox, to hypoxia (1% oxygen) or CoCl2 (100 mumol/L) and compared the mRNA levels of VEGF and aldolase with the untreated controls. Northern blot analysis revealed unimpaired basal and inducible expression of VEGF and aldolase mRNA in all four cytb- CGD-derived B-cell lines compared with wild-type cells. Furthermore, reconstitution of cytochrome b558 expression in cytb- CGD-derived B cells by transfection with p22phox or gp91phox expression vectors did not modify VEGF and aldolase mRNA expression. Thus, cytochrome b558 of the NADPH oxidase complex appears not to be essential for hypoxia- activated gene expression and can be excluded as a candidate for the putative universal oxygen sensor.

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