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)

The intragastric alcohol infusion rat model (IAIRM) of alcoholic liver disease (ALD) has been utilized in various laboratories to study various aspects of ALD pathogenesis including oxidative stress, cytokine upregulation, hypoxic damage, apoptosis, ubiquitin-proteasome pathway and CYP2E1 induction. The basic value of the model is that it produces pathologic changes which resemble ALD including microvesicular and macrovesicular fat, megamitochondria, apoptosis, central lobular and pericellular fibrosis, portal fibrosis, bridging fibrosis, central necrosis, and mixed inflammatory infiltrate including PMNs and lymphocytes. The model is valuable because the diet and ethanol intake are totally under the control of the investigator. A steady state can be maintained with high or low blood alcohol levels for long periods. The cycling of the blood alcohol levels, when a constant infusion rate of alcohol is maintained, simulates binge drinking. Using this model the importance of dietary fat, especially the degree of saturation of the fatty acids on the induction of liver pathology, has been documented. The role of endotoxin, the Kupffer cell, TNFalpha, and NADPH oxidase have been demonstrated. The importance of 2E1 in oxidative stress induction has been shown using inhibitors of the isozyme. The importance of dietary iron in the pathogenesis of cirrhosis has been documented. Acetaldehyde has been shown to play a role in preventing liver pathology by preventing NFkappaB activation. Using the model, to maintain high blood alcohol levels is found to be necessary to demonstrate proteasomal peptidase inhibition. Ubiquitin synthesis is also inhibited at high blood alcohol levels in the IAIRM model. Oxidized proteins accumulate in the liver at high blood alcohol levels. Neoantigens derived from protein adducts formed with products of oxidation induce autoimmune mechanisms of liver injury. Thus, in many ways the model has revolutionized our understanding of the pathogenesis of ALD.
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PMID:Intragastric ethanol infusion model for cellular and molecular studies of alcoholic liver disease. 1117 72

Our previous studies indicated that an alternatively spliced variant mRNA of p40-phox, a cytosolic component of NADPH oxidase, is expressed but its protein is hardly detected in myeloid cells such as promyelocytic HL-60 cells and neutrophils. Here, we have examined the stability of p40-phox variant protein in undifferentiated HL-60 cells. When in vitro-translated proteins were incubated with subcellular fractions of HL-60 cells, p40-phox variant protein but not native p40-phox was degraded by the cytosol and granule fractions. The degradation of variant protein by the granule fraction was observed using sonicated but not intact granules, suggesting that the variant protein is unlikely to be degraded by the granules in intact cells. To identify the enzyme(s) involved, we examined the effects of various enzyme inhibitors on the degradation of variant protein by the cytosol fraction. Degradation was completely inhibited by proline-specific serine protease (prolyl endopeptidase) inhibitors but not by proteasome, calpain, and metalloprotease inhibitors. Furthermore, the variant protein was degraded by a purified prolyl endopeptidase, and the degradation was protected by treating HL-60 cells with a cell-permeable inhibitor (S17092-1) for prolyl endopeptidase. These observations suggest that a cytosolic prolyl endopeptidase is involved in the degradation of p40-phox variant protein in myeloid cells.
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PMID:Involvement of cytosolic prolyl endopeptidase in degradation of p40-phox splice variant protein in myeloid cells. 1140 83

Rac1 has been shown to activate a NADPH oxidase complex producing superoxide anions in a variety of mammalian cell types. We evaluated the impact of Rac1-induced reactive oxygen species production on the turnover of Rac1 itself in human aortic endothelial cells. The concentration of a constitutively active mutant of Rac1 (Rac1(V12)) was increased by treatment of the cells with diphenylene iodinium (DPI), an inhibitor of the NADPH oxidase. Such an effect was not observed for the dominant negative form of Rac1 (Rac1(N17)). We showed a decrease in proteolytic degradation of Rac1(V12) in the presence of DPI, and showed that short term treatment with H(2)O(2) reverses the effect of DPI. We found that proteasome inhibitors (lactacystin and MG132) increased Rac1(V12) protein level. In support of this finding, we have identified in the primary sequence of Rac1 a potential destruction box domain, which is known to be a signal for protein degradation mediated by the ubiquitin/proteasome system. We show that Rac1(V12) is ubiquitinated before degradation. By contrast Rac1(N17) induces an accumulation of the ubiquitinated form of Rac1. These results suggest that Rac1 activation of NADPH oxidase is necessary for the proteolytic degradation of Rac1 itself.
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PMID:Redox regulation of human Rac1 stability by the proteasome in human aortic endothelial cells. 1158 36

Adaptation to hypoxia is a topic of considerable clinical relevance, as it influences the pathophysiology of anaemia, polycythaemia, tissue ischaemia and cancer. A growing number of physiologically relevant genes are regulated in response to changes in intracellular oxygen tension. These include genes encoding erythropoietin, vascular endothelial growth factor and tyrosine hydroxylase. Studies on the regulation of the erythropoietin gene have provided insights into the common mechanism of oxygen sensing and signal transduction, leading to activation of the hypoxia-inducible transcription factor 1 (HIF-1). Activation of HIF-1 by hypoxia depends on rescue of its alpha-subunit from oxygen-dependent degradation in the proteasome, allowing it to form a heterodimer with HIF-1 beta. This then translocates to the nucleus. There, HIF-1 assembles with a highly conserved orphan nuclear receptor, HNF-4, and a critical transcriptional adaptor, p300. This complex binds to a 3' enhancer on the erythropoietin gene, enabling transcription of erythropoietin. HIF-1 also activates other genes, the cis-acting elements of which contain cognate hypoxia response elements. There is growing evidence that the oxygen sensor is a flavohaem protein and that the signal transduction pathway involves changes in the level of intracellular reactive oxygen intermediates. We have recently cloned a novel fusion protein called cytochrome b5/b5 reductase, which is a cyanide-insensitive NADPH oxidase and, therefore, a candidate to be the oxygen sensor. This flavohaem protein is widely expressed in cell lines and tissues, with localization in the perinuclear space. In the presence of oxygen and iron, it may induce oxidative modifications that target HIF-1 alpha for ubiquitination and degradation.
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PMID:Detecting and responding to hypoxia. 1181 5

The present study hypothesized that superoxide (O2(-)*) importantly contributes to the regulation of hypoxia-inducible factor (HIF)-1alpha expression at posttranscriptional levels in renal medullary interstitial cells (RMICs) of rats. By Western blot analysis, it was found that incubation of RMICs with O2(-)* generators xanthine/xanthine oxidase and menadione significantly inhibited the hypoxia- or CoCl(2)-induced increase in HIF-1alpha levels and completely blocked the increase in HIF-1alpha levels induced by ubiquitin-proteasome inhibition with CBZ-LLL in the nuclear extracts from these cells. Under normoxic conditions, a cell-permeable O2(-)* dismutase (SOD) mimetic, 4-hydroxyl-tetramethylpiperidin-oxyl (TEMPOL) and PEG-SOD, significantly increased HIF-1alpha levels in RMICs. Two mechanistically different inhibitors of NAD(P)H oxidase, diphenyleneiodonium and apocynin, were also found to increase HIF-1alpha levels in these renal cells. Moreover, introduction of an anti-sense oligodeoxynucleotide specific to NAD(P)H oxidase subunit, p22(phox), into RMICs markedly increased HIF-1alpha levels. In contrast, the OH* scavenger tetramethylthiourea had no effect on the accumulation of HIF-1alpha in these renal cells. By Northern blot analysis, scavenging or dismutation of O2(-)* by TEMPOL and PEG-SOD was found to increase the mRNA levels of an HIF-1alpha-targeted gene, heme oxygenase-1. These results indicate that increased intracellular O2(-)* levels induce HIF-1alpha degradation independently of H(2)O(2) and OH* radicals in RMICs. NAD(P)H oxidase activity may importantly contribute to this posttranscriptional regulation of HIF-1alpha in these cells under physiological conditions.
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PMID:Redox regulation of HIF-1alpha levels and HO-1 expression in renal medullary interstitial cells. 1259 75

Hypercholesterolemia (HC) and atherosclerosis often accompany and aggravate renal disease. Proteasome inhibitors (PSI) can decrease proliferation and inflammation, likely by reducing activation of the proinflammatory NF-kappaB. However, chronic proteasome inhibition has never been demonstrated in the HC kidney. Four groups of pigs (n = 7 each) were studied after a 12-wk normal (N) or 2% HC diet alone or supplemented (N+PSI and HC+PSI) with MLN-273 (0.08 mg/kg subcutaneously twice weekly). Renal hemodynamics and function were quantified in vivo using electron-beam computed tomography at baseline and after vasodilator challenge using acetylcholine. Renal tissue was studied ex vivo using immunoblotting, PCR, and immunohistochemistry. Serum cholesterol was similarly elevated in HC and HC+PSI. Basal renal blood flow was similar among the groups, whereas GFR was decreased in both N+PSI and HC+PSI. The blunted renovascular and functional responses to acetylcholine in HC were normalized in HC+PSI (suggesting renal endothelial function improvement), which was accompanied by decreased renal endothelin, NF-kappaB, and augmented endothelial nitric oxide synthase expression. In parallel, HC+PSI animals also showed elevated NAD(P)H oxidase expression and circulating oxidized LDL, suggesting a potential for increased oxidative stress. This study shows that chronic PSI intervention in HC improves renal endothelial functional responses to challenge, possibly by modulating nitric oxide availability and endothelin. Furthermore, PSI may decrease intrarenal inflammation through modulation of the NF-kappaB pathway but may potentially increase oxidative stress, which warrants further investigation. This study may support a role for the ubiquitin/proteasome system in the kidney in HC and early atherosclerosis.
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PMID:Effects of proteasome inhibition on the kidney in experimental hypercholesterolemia. 1571 31

Elicitation of defense reactions in tobacco by cryptogein, triggered a production of active oxygen species (AOS) via the NADPH oxidase, NtrbohD, and an accumulation of beta1din, a defense induced beta-type subunit of 20S proteasome. The proteasome inhibitor, MG132, stimulated this AOS production. Tobacco cells transformed with sense constructs of beta1din showed an inhibition of the AOS production following elicitin treatment, whereas the antisense transformed cells showed a strongly enhanced AOS production. In cells transformed with sense construct of beta1din, the NtrbohD transcripts failed to be induced by cryptogein as observed in control and antisense transformed cells. Conversely, in tobacco cells transformed with antisense constructs for NtrbohD, beta1din transcripts remained at a low level after elicitation. These results constitute the first demonstration of proteasome comprising beta1din acting as a negative regulator of NtrbohD and contributes to the regulation of AOS generation during plant defense reactions.
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PMID:Proteasome comprising a beta1 inducible subunit acts as a negative regulator of NADPH oxidase during elicitation of plant defense reactions. 1611 82

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a classical glycolytic enzyme that is involved in cellular energy production and has important housekeeping functions. We used the natural prooxidant and proatherogenic molecule oxidized low-density lipoprotein (OxLDL) to determine a potential link between OxLDL-promoted oxidative stress, GAPDH expression, and smooth muscle cell energy metabolism. OxLDL but not native LDL (nLDL) produced a 60% to 100% dose- and time-dependent reduction of GAPDH protein. OxLDL increased reactive oxygen species (ROS) formation, including rapid elevation of H2O2 levels. OxLDL decreased intracellular catalase expression, likely contributing to the increase in H2O2. Antioxidants, anti-CD36 receptor antibody, NADPH oxidase, or lipoxygenase blockers decreased OxLDL-specific ROS and prevented GAPDH downregulation. 12/15-Lipoxygenase or p47phox deficiency resulted in attenuation of GAPDH downregulation, but 5-lipoxygenase suppression had no effect. OxLDL or exogenous H2O2 oxidized GAPDH thiols, decreasing GAPDH protein half-life and increasing GAPDH sensitivity to proteasome-mediated protein degradation in vitro. OxLDL- or small interfering RNA-specific downregulation of GAPDH resulted in 65% reduction in glycolysis rate and 82% decrease in ATP levels. In conclusion, our data demonstrate that OxLDL downregulated GAPDH via a H2O2-dependent decrease in protein stability. GAPDH protein damage resulted in marked depletion of cellular ATP levels. Our data have important implications for understanding the metabolic effect of OxLDL on the vessel wall and mechanism of atherogenesis.
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PMID:Novel effect of oxidized low-density lipoprotein: cellular ATP depletion via downregulation of glyceraldehyde-3-phosphate dehydrogenase. 1677 34

Biallelic inactivation of the von Hippel-Lindau tumor suppressor gene (VHL) is linked to the development of hereditary and sporadic renal cell carcinoma (RCC). In the absence of VHL, the alpha subunits of heterodimeric hypoxia-inducible transcription factors (HIF-1alpha and HIF-2alpha) are stabilized. Reactive oxygen species, generated by NAD(P)H oxidases, are involved in signaling cascades of malignant growth. We show that in VHL-deficient cells p22phox, Nox4 protein levels and NADPH-dependent superoxide generation are increased. Reintroduction of VHL into the VHL-deficient cells down-regulates the expression of p22phox and NADPH-dependent superoxide generation. Inhibition of the 26 S proteasome in VHL-expressing cells increased p22phox protein levels, which correlated with an increase of NADPH-dependent superoxide generation. We also show that p22phox co-immunoprecipitates with VHL in vivo. Moreover, p22phox is a target of ubiquitination. Importantly, in VHL-deficient cells, diphenyleneiodonium chloride (DPI), an inhibitor of Nox oxidases, decreased the expression of HIF-2alpha. Down-regulation of Nox1, Nox4, and p22phox expression by small interfering RNA also decreased HIF-2alpha protein expression and inhibited Akt and 4E-BP1 phosphorylation, suggesting that a translational mechanism is involved in maintaining HIF-2alpha in VHL-deficient cells. Colony formation by RCC 786-O in soft agar was markedly inhibited by DPI. Moreover, DPI significantly inhibited RCC 786-O tumor formation in athymic mice. Collectively, the data demonstrate that VHL protein exerts its tumor suppressor action, at least partially, via inhibition of p22phox-based Nox4/Nox1 NADPH oxidase-dependent reactive oxygen species generation.
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PMID:NAD(P)H oxidases regulate HIF-2alpha protein expression. 1720 Jan 23

The antioxidants butylated hydroxytoluene (BHT, 1 mM) and D-alpha-tocopherol (10 microM) completely attenuated protein degradation in murine myotubes in response to both proteolysis-inducing factor (PIF) and angiotensin II (Ang II), suggesting that the formation of reactive oxygen species (ROS) plays an important role in this process. Both PIF and Ang II induced a rapid and transient increase in ROS formation in myotubes, which followed a parabolic dose-response curve, similar to that for total protein degradation. Antioxidant treatment attenuated the increase in expression and activity of the ubiquitin-proteasome proteolytic pathway by PIF and Ang II, by preventing the activation of the transcription factor nuclear factor-kappaB (NF-kappaB), through inhibition of phosphorylation of the NF-kappaB inhibitor protein (I-kappaB) and its subsequent degradation. ROS formation by both PIF and Ang II was attenuated by diphenyleneiodonium (10 microM), suggesting that it was mediated through the NADPH oxidase system. ROS formation was also attenuated by trifluoroacetyl arachidonic acid (10 microM), a specific inhibitor of cytosolic phospholipase A2, U-73122 (5 microM) and D609 (200 microM), inhibitors of phospholipase C and calphostin C (300 nM), a highly specific inhibitor of protein kinase C (PKC), all known activators of NADPH oxidase. Myotubes containing a dominant-negative mutant of PKC did not show an increase in ROS formation in response to either PIF or Ang II. The two Rac1 inhibitors W56 (200 microM) and NSC23766 (10 microM) also attenuated both ROS formation and protein degradation induced by both PIF and Ang II. Rac1 is known to mediate signalling between the phosphatidylinositol-3 kinase (PI-3K) product and NADPH oxidase, and treatment with LY24002 (10 microM), a highly selective inhibitor of PI-3K, completely attenuated ROS production in response to both PIF and Ang II, and inhibited total protein degradation, while the inactive analogue LY303511 (100 microM) had no effect. ROS formation appears to be important in muscle atrophy in cancer cachexia, since treatment of weight losing mice bearing the MAC16 tumour with D-alpha-tocopherol (1 mg kg(-1)) attenuated protein degradation and increased protein synthesis in skeletal muscle.
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PMID:Role of reactive oxygen species in protein degradation in murine myotubes induced by proteolysis-inducing factor and angiotensin II. 1753 11


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