Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.6.3.1 (NADPH oxidase)
11,281 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Human fat cells possess a multireceptor-linked H2O2-generating system that is activated by insulin. Previous studies revealed that manganese was the sole cofactor required for a hormonal regulation of NADPH-dependent H2O2 generation in vitro. In this report it is shown that the synergistic activation of NADPH-dependent H2O2 generation by Mn2+ and insulin was blocked by GDPbetaS (guanosine 5'-O-(2-thiodiphosphate)), pertussis toxin and COOH-terminal anti-Galphai1-2 or the corresponding peptide. Consistently, manganese could be replaced by micromolar concentrations of GTPgammaS (guanosine 5'-O-(3-thiotriphosphate)), which increased NADPH-dependent H2O2 generation by 20-40%. Insulin shifted the dose response curve for GTPgammaS to the left (>10-fold) and increased the maximal response. In the presence of 10 microM GTPgammaS, the hormone was active at picomolar concentrations, indicating that insulin acted via its cognate receptor. The insulin receptor and Gi were co-adsorbed on anti-Galphai and anti-insulin receptor beta-subunit (anti-IRbeta) affinity columns. Partially purified insulin receptor preparations contained Galphas, Galphai2, and Gbetagamma (but no Galphai1 or Galphai3). The functional nature of the insulin receptor-Gi2 complex was made evident by insulin's ability to modulate labeling of Gi by bacterial toxins. Insulin action was mimicked by activated Galphai, but not by Galphao or Gbetagamma, indicating that insulin's signal was transduced via Galphai2. Thus, NADPH oxidase is the first example of an effector system that is coupled to the insulin receptor via a heterotrimeric G protein.
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PMID:Insulin-induced activation of NADPH-dependent H2O2 generation in human adipocyte plasma membranes is mediated by Galphai2. 909 59

Cellular insulin stimulation generates a burst of H(2)O(2) that modulates protein-tyrosine phosphorylation in the insulin action pathway, in part by the inhibition of redox-sensitive protein-tyrosine phosphatases [J. Biol. Chem. 276 (2001) 21938]. Blocking the insulin-induced rise in H(2)O(2) with the NADPH oxidase inhibitor diphenyleneiodonium (DPI) strongly attenuated the activation of phosphatidylinositol 3' (PI 3')-kinase, Akt and GLUT4 translocation by insulin in 3T3-L1 adipocytes; however, under identical conditions, we observed a paradoxical increase in the activation of p42/p44 mitogen-activated protein (MAP) kinase. DPI inhibited the insulin-stimulated tyrosine phosphorylation of the insulin receptor and IRS-1/2, and also reduced the association of Grb2 with IRS-1, suggesting that the effect of DPI on MAP kinase activation occurred downstream of the IR and IRS proteins. DPI increased the insulin-stimulated phosphorylation of p42/p44 MAP kinase with no change in basal, and increased insulin-stimulated MAP kinase kinase (MEK) activity by a similar degree. DPI enhanced basal Grb2-Sos binding and reduced the effect of insulin to potentiate the dissociation of the Grb2-Sos complex, suggesting that the effect of DPI was mediated upstream of Raf-1. Cell treatment with dibutyryl cAMP significantly reduced the enhancement of MAP kinase activation in the presence of DPI. However, forskolin, acting in a PKA-independent manner, increased the insulin stimulation of MAP kinase and MEK, but fully abrogated the effect of DPI to enhance these insulin responses. PLCgamma inhibition with U73122 blocked the insulin stimulation of MAP kinase and MEK as well as the enhancing effect of DPI on these responses. PKC activation strongly stimulated MAP kinase and MEK activation, even in the presence of U73122, consistent with PKC acting downstream of PLCgamma. These data show that the insulin-stimulated oxidant signal differentially affects the two major downstream components of the insulin signaling pathway, PI 3'-kinase and MAP kinase, and cross-talk between insulin action, PLCgamma and, to a lesser extent, PKA modulates the net cellular effects of insulin-stimulated cellular H(2)O(2).
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PMID:Integration of multiple downstream signals determines the net effect of insulin on MAP kinase vs. PI 3'-kinase activation: potential role of insulin-stimulated H(2)O(2). 1468 62

Insulin stimulation of target cells elicits a burst of H(2)O(2) that enhances tyrosine phosphorylation of the insulin receptor and its cellular substrate proteins as well as distal signaling events in the insulin action cascade. The molecular mechanism coupling the insulin receptor with the cellular oxidant-generating apparatus has not been elucidated. Using reverse transcription-PCR and Northern blot analyses, we found that Nox4, a homolog of gp91phox, the phagocytic NAD(P)H oxidase catalytic subunit, is prominently expressed in insulin-sensitive adipose cells. Adenovirus-mediated expression of Nox4 deletion constructs lacking NAD(P)H or FAD/NAD(P)H cofactor binding domains acted in a dominant-negative fashion in differentiated 3T3-L1 adipocytes and attenuated insulin-stimulated H(2)O(2) generation, insulin receptor (IR) and IRS-1 tyrosine phosphorylation, activation of downstream serine kinases, and glucose uptake. Transfection of specific small interfering RNA oligonucleotides reduced Nox4 protein abundance and also inhibited the insulin signaling cascade. Overexpression of Nox4 also significantly reversed the inhibition of insulin-stimulated IR tyrosine phosphorylation induced by coexpression of PTP1B by inhibiting PTP1B catalytic activity. These data suggest that Nox4 provides a novel link between the IR and the generation of cellular reactive oxygen species that enhance insulin signal transduction, at least in part via the oxidative inhibition of cellular protein-tyrosine phosphatases (PTPases), including PTP1B, a PTPase that has been previously implicated in the regulation of insulin action.
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PMID:The NAD(P)H oxidase homolog Nox4 modulates insulin-stimulated generation of H2O2 and plays an integral role in insulin signal transduction. 1496 67

Previously, we have shown that the human insulin receptor (IR) interacts with G(i)2, independent of tyrosine kinase activity and stimulates NADPH oxidase via the Galpha subunit of G(i)2. We have now investigated the regulatory role of G(i)2-proteins in IR function. For the experiments, isolated IRs from plasma membranes of human fat cells were used. The activation of IR autophosphorylation by insulin was blocked by G-protein inactivation through GDPbetaS (guanosine 5'-[beta-thio]disphosphate). Consistently, activation of G-proteins by micromolar concentrations of GTPgammaS (guanosine 5'-[gamma-thio]triphosphate) induced receptor autophosphorylation 5-fold over baseline and increased insulin-induced autophosphorylation by 3-fold. In the presence of 10 microM GTPgammaS, insulin was active at picomolar concentrations, indicating that insulin acted via its cognate receptor. Pretreatment of the plasma membranes with pertussis toxin prevented insulin- and GTPgammaS-induced autophosphorylation, but did not disrupt the IR-G(i)2 complex. The functional nature of the IR-G(i)2 complex was made evident by insulin's ability to increase association of G(i)2 with the IR. This leads to an augmentation of maximal receptor autophosphorylation induced by insulin and GTPgammaS. The specificity of this mechanism was further demonstrated by the use of isolated preactivated G-proteins. Addition of G(i)2alpha and Gbetagamma mimicked maximal response of insulin, whereas Galphas or Galphao had no stimulatory effect. These results define a novel mechanism by which insulin signalling mediates tyrosine kinase activity and autophosphorylation of the IR through recruitment of G(i)-proteins.
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PMID:Ligand-dependent autophosphorylation of the insulin receptor is positively regulated by Gi-proteins. 1518 34

The role of endogenous kinins and their receptors in diabetes mellitus is being confirmed with the recent developments of molecular and genetic animal models. Compelling evidence suggests that the kinin B(2) receptor is organ-protective and partakes to the therapeutic effects of angiotensin 1-converting enzyme inhibitors (ACEI) and angiotensin AT(1) receptor antagonists. Benefits derive primarily from vasodilatory, antihypertensive, antiproliferative, antihypertrophic, antifibrotic, antithrombotic and antioxidant properties of kinin B(2) receptor activation. Mechanisms include the formation of nitric oxide and prostacyclin and the inhibition of NAD(P)H oxidase activity involving classical and novel signalling pathways. Kinin B(2) receptor also ameliorates insulin resistance by increasing glucose uptake and supply, and by inducing glucose transporter-4 translocation either directly or through phosphorylation of insulin receptor. The kinin B(1) receptor, which is induced by the cytokine network, growth factors and hyperglycaemia, mediates hyperalgesia, vascular hyperpermeability and leukocytes infiltration in diabetic animals. However, emerging data highlight reno- and cardio-protective effects mediated by kinin B(1) receptor under chronic ACEI therapy in diabetes mellitus. Thus, the Janus-faced of kinin receptors needs to be taken into account in future drug development. For instance, locally acting kinin B(1)/B(2) receptor agonists if used in a safe therapeutic window may represent a more rationale strategy in the prevention and management of diabetic complications. Because kinin B(2) receptor antagonists may further increase insulin resistance, the persisting dogma that restricts the development of kinin receptor analogues to antagonists (that is still relevant to abrogate pain and inflammation) needs to be revisited.
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PMID:Putative roles of kinin receptors in the therapeutic effects of angiotensin 1-converting enzyme inhibitors in diabetes mellitus. 1546 53

Propelled by the identification of a small family of NADPH oxidase (Nox) enzyme homologs that produce superoxide in response to cellular stimulation with various growth factors, renewed interest has been generated in characterizing the signaling effects of reactive oxygen species (ROS) in relation to insulin action. Two key observations made >30 years ago-that oxidants can facilitate or mimic insulin action and that H(2)O(2) is generated in response to insulin stimulation of its target cells-have led to the hypothesis that ROS may serve as second messengers in the insulin action cascade. Specific molecular targets of insulin-induced ROS include enzymes whose signaling activity is modified via oxidative biochemical reactions, leading to enhanced insulin signal transduction. These positive responses to cellular ROS may seem "paradoxical" because chronic exposure to relatively high levels of ROS have also been associated with functional beta-cell impairment and the chronic complications of diabetes. The best-characterized molecular targets of ROS are the protein-tyrosine phosphatases (PTPs) because these important signaling enzymes require a reduced form of a critical cysteine residue for catalytic activity. PTPs normally serve as negative regulators of insulin action via the dephosphorylation of the insulin receptor and its tyrosine-phosphorylated cellular substrates. However, ROS can rapidly oxidize the catalytic cysteine of target PTPs, effectively blocking their enzyme activity and reversing their inhibitory effect on insulin signaling. Among the cloned Nox homologs, we have recently provided evidence that Nox4 may mediate the insulin-stimulated generation of cellular ROS and is coupled to insulin action via the oxidative inhibition of PTP1B, a PTP known to be a major regulator of the insulin signaling cascade. Further characterization of the molecular components of this novel signaling cascade, including the mechanism of ROS generated by insulin and the identification of various oxidation-sensitive signaling targets in insulin-sensitive cells, may provide a novel means of facilitating insulin action in states of insulin resistance.
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PMID:Redox paradox: insulin action is facilitated by insulin-stimulated reactive oxygen species with multiple potential signaling targets. 1567 87

Hydrogen peroxide (H2O2) mimics many physiological responses of insulin, and increased H2O2 generation via the Nox-4 subunit of NAD(P)H oxidase was recently demonstrated to serve as a critical early step in the insulin signaling pathway. Exogenously added H2O2 has also been shown to activate several key components of the insulin signaling cascade. H2O2-induced signaling responses have been found to be associated with the activation of receptor and nonreceptor protein tyrosine kinases (PTK), including the insulin receptor (IR)-beta subunit. Therefore, in the present studies on Chinese hamster ovary cells overexpressing wild-type IR-PTK (CHO-IR) or a PTK-inactive form of IR (CHO-1018), we investigated whether IR-PTK plays a role in H2O2-induced signaling events. Treatment of CHO-IR cells with H2O2 increased the phosphorylation of extracellular signal-regulated kinases 1 and 2 (ERK1/2), protein kinase B (PKB), and glycogen synthase kinase-3beta while enhancing tyrosine phosphorylation of the IR-beta subunit and the p85 subunit of phosphatidylinositol 3-kinase (PI3K). Compared with CHO-IR cells, the stimulatory effect of H2O2 on ERK1/2 and PKB was partially reduced in CHO-1018 cells. However, pharmacological inhibition of Src family PTK by 4-amino-5-(4-chlorophenyl)-7-(tert-butyl)pyrazolo[3,4-d]pyrimidine (PP-2) almost completely blocked H2O2-stimulated phosphorylation of the p85 subunit of PI3K, ERK1/2, and PKB. Moreover, H2O2, but not insulin, induced Tyr-418 phosphorylation of Src, which was also suppressed by PP-2. Taken together, these data suggest that both IR-PTK and Src family PTKs contribute to H2O2-induced signaling in CHO-IR cells albeit IR-PTK has a less dominant role in this process.
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PMID:H2O2-induced phosphorylation of ERK1/2 and PKB requires tyrosine kinase activity of insulin receptor and c-Src. 1599 56

Leprechaunism features a clinical constellation characterized by extreme insulin resistance, growth retardation, and several distinct developmental abnormalities. One puzzling observation about leprechaunism is that mutations in the insulin receptor gene frequently associated with this syndrome cannot account for the aberrant responses of cultured cells to other growth factors. Here we report that the generation of reactive oxygen species (ROS) is impaired in cells from leprechaunism patients, thus shedding new light on this issue. Stimulation of patients' skin fibroblast cells with platelet-derived growth factor (PDGF) resulted in a lower-level tyrosine phosphorylation of cytosolic proteins compared with that seen in normal cells. In addition, consistent with the hypothesis that ROS mediate the level of tyrosine phosphorylation of cytosolic proteins through inactivation of protein tyrosine phosphatases (PTPases), patient fibroblast cells showed a significantly higher phosphatase activity than normal cells. We further showed that the lower-level tyrosine phosphorylation in response to growth factors results from the downregulation of an NADPH oxidase, Nox4, which in turn results in the reduction of ROS generation. Ectopic expression of Nox4 in the patient fibroblast cells consistently restored PDGF-induced ROS production and regulation of PTPase activities. Taken together, these data provide insight into the mechanisms through which growth retardation is associated with leprechaunism syndrome.
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PMID:Impaired generation of reactive oxygen species in leprechaunism through downregulation of Nox4. 1624 42

The renin-angiotensin system is a central component of the physiological and pathological responses of cardiovascular system. Its primary effector hormone, angiotensin II (ANG II), not only mediates immediate physiological effects of vasoconstriction and blood pressure regulation, but is also implicated in inflammation, endothelial dysfunction, atherosclerosis, hypertension, and congestive heart failure. The myriad effects of ANG II depend on time (acute vs. chronic) and on the cells/tissues upon which it acts. In addition to inducing G protein- and non-G protein-related signaling pathways, ANG II, via AT(1) receptors, carries out its functions via MAP kinases (ERK 1/2, JNK, p38MAPK), receptor tyrosine kinases [PDGF, EGFR, insulin receptor], and nonreceptor tyrosine kinases [Src, JAK/STAT, focal adhesion kinase (FAK)]. AT(1)R-mediated NAD(P)H oxidase activation leads to generation of reactive oxygen species, widely implicated in vascular inflammation and fibrosis. ANG II also promotes the association of scaffolding proteins, such as paxillin, talin, and p130Cas, leading to focal adhesion and extracellular matrix formation. These signaling cascades lead to contraction, smooth muscle cell growth, hypertrophy, and cell migration, events that contribute to normal vascular function, and to disease progression. This review focuses on the structure and function of AT(1) receptors and the major signaling mechanisms by which angiotensin influences cardiovascular physiology and pathology.
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PMID:Angiotensin II cell signaling: physiological and pathological effects in the cardiovascular system. 1687 Aug 27

The reduced capacity of insulin to stimulate glucose transport into skeletal muscle, termed insulin resistance, is a primary defect leading to the development of prediabetes and overt type 2 diabetes. Although the etiology of this skeletal muscle insulin resistance is multifactorial, there is accumulating evidence that one contributor is overactivity of the renin-angiotensin system (RAS). Angiotensin II (ANG II) produced from this system can act on ANG II type 1 receptors both in the vascular endothelium and in myocytes, with an enhancement of the intracellular production of reactive oxygen species (ROS). Evidence from animal model and cultured skeletal muscle cell line studies indicates ANG II can induce insulin resistance. Chronic ANG II infusion into an insulin-sensitive rat produces a markedly insulin-resistant state that is associated with a negative impact of ROS on the skeletal muscle glucose transport system. ANG II treatment of L6 myocytes causes impaired insulin receptor substrate (IRS)-1-dependent insulin signaling that is accompanied by augmentation of NADPH oxidase-mediated ROS production. Further critical evidence has been obtained from the TG(mREN2)27 rat, a model of RAS overactivity and insulin resistance. The TG(mREN2)27 rat displays whole body and skeletal muscle insulin resistance that is associated with local oxidative stress and a significant reduction in the functionality of the insulin receptor (IR)/IRS-1-dependent insulin signaling. Treatment with a selective ANG II type 1 receptor antagonist leads to improvements in whole body insulin sensitivity, enhanced insulin-stimulated glucose transport in muscle, and reduced local oxidative stress. In addition, exercise training of TG(mREN2)27 rats enhances whole body and skeletal muscle insulin action. However, these metabolic improvements elicited by antagonism of ANG II action or exercise training are independent of upregulation of IR/IRS-1-dependent signaling. Collectively, these findings support targeting the RAS in the design of interventions to improve metabolic and cardiovascular function in conditions of insulin resistance associated with prediabetes and type 2 diabetes.
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PMID:Improvement of insulin sensitivity by antagonism of the renin-angiotensin system. 1758 38


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