Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Co-regulation of constitutive nitric oxide synthases and NADPH oxidase by the small GTPase Rac.

ABSTRACT: Nitric oxide (NO), generated by NO synthases (NOSs), has multifarious roles in signal transduction. Reactive oxygen species (ROS), generated by ubiquitous NADPH oxidases (NOXs), also participate in cellular signaling. However, the coordination of signals conveyed by NO and ROS is poorly understood. We show that the small GTPase Rac, a component of some NOXs, also interacts with and regulates the constitutively-expressed NOSs. Cellular NO and O(2)(-) production increase or decrease together following activation or inhibition of Rac, and Rac inhibition reveals transduction mechanisms that depend upon NO (vasodilation), ROS (actin polymerization) or both (cytoskeletal organization). Thus, signaling by NO and ROS may be coordinated through a common control element.

SUBMITTER: Selvakumar B

PROVIDER: S-EPMC2519002 | biostudies-literature | 2008 Jun

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Co-regulation of constitutive nitric oxide synthases and NADPH oxidase by the small GTPase Rac.

Selvakumar Balakrishnan B Hess Douglas T DT Goldschmidt-Clermont Pascal J PJ Stamler Jonathan S JS

FEBS letters 20080522 15

Nitric oxide (NO), generated by NO synthases (NOSs), has multifarious roles in signal transduction. Reactive oxygen species (ROS), generated by ubiquitous NADPH oxidases (NOXs), also participate in cellular signaling. However, the coordination of signals conveyed by NO and ROS is poorly understood. We show that the small GTPase Rac, a component of some NOXs, also interacts with and regulates the constitutively-expressed NOSs. Cellular NO and O(2)(-) production increase or decrease together follo ...[more]

PMID: 18501711

Similar Datasets

Structures of human constitutive nitric oxide synthases.

Project description:Mammals produce three isoforms of nitric oxide synthase (NOS): neuronal NOS (nNOS), inducible NOS (iNOS) and endothelial NOS (eNOS). The overproduction of NO by nNOS is associated with a number of neurodegenerative disorders; therefore, a desirable therapeutic goal is the design of drugs that target nNOS but not the other isoforms. Crystallography, coupled with computational approaches and medicinal chemistry, has played a critical role in developing highly selective nNOS inhibitors that exhibit exceptional neuroprotective properties. For historic reasons, crystallography has focused on rat nNOS and bovine eNOS because these were available in high quality; thus, their structures have been used in structure-activity-relationship studies. Although these constitutive NOSs share more than 90% sequence identity across mammalian species for each NOS isoform, inhibitor-binding studies revealed that subtle differences near the heme active site in the same NOS isoform across species still impact enzyme-inhibitor interactions. Therefore, structures of the human constitutive NOSs are indispensible. Here, the first structure of human neuronal NOS at 2.03 Å resolution is reported and a different crystal form of human endothelial NOS is reported at 1.73 Å resolution.

| S-EPMC4188008 | biostudies-literature

Rational design of a fluorescent NADPH derivative imaging constitutive nitric-oxide synthases upon two-photon excitation.

Project description:We report the structure-based design and synthesis of a unique NOS inhibitor, called nanoshutter NS1, with two-photon absorption properties. NS1 targets the NADPH site of NOS by a nucleotide moiety mimicking NADPH linked to a conjugated push-pull chromophore with nonlinear absorption properties. Because NS1 could not provide reducing equivalents to the protein and competed with NADPH binding, it efficiently inhibited NOS catalysis. NS1 became fluorescent once bound to NOS with an excellent signal-to-noise ratio because of two-photon excitation avoiding interference from the flavin-autofluorescence and because free NS1 was not fluorescent in aqueous solutions. NS1 fluorescence enhancement was selective for constitutive NOS in vitro, in particular for endothelial NOS (eNOS). Molecular dynamics simulations suggested that two variable residues among NOS isoforms induced differences in binding of NS1 and in local solvation around NS1 nitro group, consistent with changes of NS1 fluorescence yield. NS1 colocalized with eNOS in living human umbilical vein endothelial cells. Thus, NS1 constitutes a unique class of eNOS probe with two-photon excitation in the 800-950-nm range, with great perspectives for eNOS imaging in living tissues.

| S-EPMC3411954 | biostudies-literature

Crystal structures of constitutive nitric oxide synthases in complex with de novo designed inhibitors.

Project description:New nitric oxide synthase (NOS) inhibitors were designed de novo with knowledge gathered from the studies on the nNOS-selective dipeptide inhibitors. Each of the new inhibitors consists of three fragments: an aminopyridine ring, a pyrrolidine, and a tail of various length and polarity. The in vitro inhibitory assays indicate good potency and isoform selectivity for some of the compounds. Crystal structures of these inhibitors bound to either wild type or mutant nNOS and eNOS have confirmed design expectations. The aminopyridine ring mimics the guanidinium group of L-arginine and functions as an anchor to place the compound in the NOS active site where it hydrogen bonds to a conserved Glu. The rigidity of the pyrrolidine ring places the pyrrolidine ring nitrogen between the same conserved Glu and the selective residue nNOS Asp597/eNOS Asn368, which results in similar interactions observed with the alpha-amino group of dipeptide inhibitors bound to nNOS. These structures provide additional information to help in the design of inhibitors with greater potency, physicochemical properties, and isoform selectivity.

| S-EPMC3103786 | biostudies-literature

Nitric oxide reduces NADPH oxidase 5 (Nox5) activity by reversible S-nitrosylation.

Project description:The NADPH oxidases (Noxs) are a family of transmembrane oxidoreductases that produce superoxide and other reactive oxygen species (ROS). Nox5 was the last of the conventional Nox isoforms to be identified and is a calcium-dependent enzyme that does not depend on accessory subunits for activation. Recently, Nox5 was shown to be expressed in human blood vessels and therefore the goal of this study was to determine whether nitric oxide (NO) can modulate Nox5 activity. Endogenously produced NO potently inhibited basal and stimulated Nox5 activity and this inhibition was reversible with chronic, but not acute, exposure to L-NAME. Nox5 activity was reduced by NO donors, iNOS, and eNOS and in endothelial cells and LPS-stimulated smooth muscle cells in a manner dependent on NO concentration. ROS production was diminished by NO in an isolated enzyme activity assay replete with surplus calcium and NADPH. There was no evidence for NO-dependent changes in tyrosine nitration, glutathiolation, or phosphorylation of Nox5. In contrast, there was evidence for the increased nitrosylation of Nox5 as determined by the biotin-switch assay and mass spectrometry. Four S-nitrosylation sites were identified and of these, mutation of C694 dramatically lowered Nox5 activity, NO sensitivity, and biotin labeling. Furthermore, coexpression of the denitrosylation enzymes thioredoxin 1 and GSNO reductase prevented NO-dependent inhibition of Nox5. The potency of NO against other Nox enzymes was in the order Nox1 ? Nox3 > Nox5 > Nox2, whereas Nox4 was refractory. Collectively, these results suggest that endogenously produced NO can directly S-nitrosylate and inhibit the activity of Nox5.

| S-EPMC3464050 | biostudies-literature

Nitric oxide and heme-NO stimulate superoxide production by NADPH oxidase 5.

Project description:Nitric oxide (NO) is a ubiquitous cell signaling molecule which mediates widespread and diverse processes in the cell. These NO dependent effects often involve activation (e.g. NO binding to the heme group of soluble guanylyl cyclase for cGMP production) or inactivation (e.g. S-nitrosation) of protein targets. We studied the effect of NO and heme-NO on the transmembrane signaling enzyme NADPH oxidase 5 (NOX5), a heme protein which produces superoxide in response to increases in intracellular calcium. We found that treatment with NO donors increases NOX5 activity through heme-dependent effects, and that this effect could be recapitulated by the addition of heme-NO. This work adds to our understanding of NOX5 regulation in the cell but also provides a framework for understanding how NO could cause widespread changes in hemeprotein activity based on different affinities for heme v. heme-NO, and helps explain the opposing roles NO plays in activation and inactivation of hemeprotein targets.

| S-EPMC8355125 | biostudies-literature

NADPH oxidase 4 promotes endothelial angiogenesis through endothelial nitric oxide synthase activation.

Project description:UnlabelledBACKGROUND- Reactive oxygen species serve signaling functions in the vasculature, and hypoxia has been associated with increased reactive oxygen species production. NADPH oxidase 4 (Nox4) is a reactive oxygen species-producing enzyme that is highly expressed in the endothelium, yet its specific role is unknown. We sought to determine the role of Nox4 in the endothelial response to hypoxia.Methods and resultsHypoxia induced Nox4 expression both in vitro and in vivo and overexpression of Nox4 was sufficient to promote endothelial proliferation, migration, and tube formation. To determine the in vivo relevance of our observations, we generated transgenic mice with endothelial-specific Nox4 overexpression using the vascular endothelial cadherin promoter (VECad-Nox4 mice). In vivo, the VECad-Nox4 mice had accelerated recovery from hindlimb ischemia and enhanced aortic capillary sprouting. Because endothelial nitric oxide synthase (eNOS) is involved in endothelial angiogenic responses and eNOS is activated by reactive oxygen species, we probed the effect of Nox4 on eNOS. In cultured endothelial cells overexpressing Nox4, we observed a significant increase in eNOS protein expression and activity. To causally address the link between eNOS and Nox4, we crossed our transgenic Nox4 mice with eNOS(-/-) mice. Aortas from these mice did not demonstrate enhanced aortic sprouting, and VECad-Nox4 mice on the eNOS(-/-) background did not demonstrate enhanced recovery from hindlimb ischemia.ConclusionsCollectively, we demonstrate that augmented endothelial Nox4 expression promotes angiogenesis and recovery from hypoxia in an eNOS-dependent manner.

| S-EPMC3589548 | biostudies-literature

Paradoxical activation of endothelial nitric oxide synthase by NADPH oxidase.

Project description:Increased formation of reactive oxygen species (ROS) has been identified as a causative factor in endothelial dysfunction by reducing NO bioavailability and uncoupling endothelial nitric oxide synthase (eNOS). However, the specific contribution of ROS to endothelial function is not well understood.A major source of intracellular ROS is the NADPH oxidase (Nox) family of enzymes. The goal of the current study was to directly assess the contribution of NADPH oxidase derived superoxide to eNOS function by expressing Nox5, a single gene product that constitutively produces superoxide within cells. Paradoxically, we found that instead of inhibiting eNOS, coexpression of Nox5 increased NO release from both bovine and human endothelial cells. To establish the functional significance of this observation in intact blood vessels, the endothelium of mouse aorta was transduced with Nox5 or control adenoviruses. Nox5 potently inhibited Ach-induced relaxation and potentiated contractile responses to phenylephrine. In precontracted aortae, acute exposure to superoxide dismutase induced significant vascular relaxation in vessels exposed to Nox5 versus control and unmasked the ability of Nox5 to activate eNOS in blood vessel endothelium.These findings suggest that ROS inhibit eNOS function via consumption of NO rather than direct inhibition of enzymatic activity.

| S-EPMC2771553 | biostudies-other

Periplasmic superoxide dismutase protects Salmonella from products of phagocyte NADPH-oxidase and nitric oxide synthase.

Project description:Superoxide dismutase (SOD) catalyzes the conversion of superoxide radical to hydrogen peroxide. Periplasmic localization of bacterial Cu,Zn-SOD has suggested a role of this enzyme in defense against extracellular phagocyte-derived reactive oxygen species. Sequence analysis of regions flanking the Salmonella typhimurium sodC gene encoding Cu,Zn-SOD demonstrates significant homology to lambda phage proteins, reflecting possible bacteriophage-mediated horizontal gene transfer of this determinant among pathogenic bacteria. Salmonella deficient in Cu,Zn-SOD has reduced survival in macrophages and attenuated virulence in mice, which can be restored by abrogation of either the phagocyte respiratory burst or inducible nitric oxide synthase. Moreover, a sodC mutant is extremely susceptible to the combination of superoxide and nitric oxide. These observations suggest that SOD protects periplasmic or inner membrane targets by diverting superoxide and limiting peroxynitrite formation, and they demonstrate the ability of the respiratory burst and nitric oxide synthase to synergistically kill microbial pathogens in vivo.

| S-EPMC28421 | biostudies-literature

Peroxynitrite generated by inducible nitric oxide synthase and NADPH oxidase mediates microglial toxicity to oligodendrocytes.

Project description:Reactive microglia in the CNS have been implicated in the pathogenesis of white matter disorders, such as periventricular leukomalacia and multiple sclerosis. However, the mechanism by which activated microglia kill oligodendrocytes (OLs) remains elusive. Here we show that lipopolysaccharide (LPS)-induced death of developing OLs is caused by microglia-derived peroxynitrite, the reaction product of nitric oxide (NO) and superoxide anion. Blocking peroxynitrite formation with nitric oxide synthase inhibitors, superoxide dismutase mimics, or a decomposition catalyst abrogated the cytotoxicity. Only microglia, but not OLs, expressed inducible NO synthase (iNOS) after LPS challenge; microglia from iNOS knockout mice were not cytotoxic upon activation. The molecular source for superoxide was identified as the superoxide-generating enzyme NADPH oxidase. The oxidase was activated upon LPS exposure, and its inhibition prevented microglial toxicity toward OLs. Furthermore, microglia isolated from mice deficient in the catalytic component of the oxidase, gp91(phox), failed to induce cell death. Our results reveal a role for NADPH oxidase in LPS-induced OL death and suggest that peroxynitrite produced by iNOS and NADPH oxidase in activated microglia may play an important role in the pathogenesis of white matter disorders.

| S-EPMC1174990 | biostudies-literature

Molecular architecture of mammalian nitric oxide synthases.

Project description:NOSs are homodimeric multidomain enzymes responsible for producing NO. In mammals, NO acts as an intercellular messenger in a variety of signaling reactions, as well as a cytotoxin in the innate immune response. Mammals possess three NOS isoforms--inducible, endothelial, and neuronal NOS--that are composed of an N-terminal oxidase domain and a C-terminal reductase domain. Calmodulin (CaM) activates NO synthesis by binding to the helical region connecting these two domains. Although crystal structures of isolated domains have been reported, no structure is available for full-length NOS. We used high-throughput single-particle EM to obtain the structures and higher-order domain organization of all three NOS holoenzymes. The structures of inducible, endothelial, and neuronal NOS with and without CaM bound are similar, consisting of a dimerized oxidase domain flanked by two separated reductase domains. NOS isoforms adopt many conformations enabled by three flexible linkers. These conformations represent snapshots of the continuous electron transfer pathway from the reductase domain to the oxidase domain, which reveal that only a single reductase domain participates in electron transfer at a time, and that CaM activates NOS by constraining rotational motions and by directly binding to the oxidase domain. Direct visualization of these large conformational changes induced during electron transfer provides significant insight into the molecular underpinnings governing NO formation.

| S-EPMC4156747 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data