Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.1 (protein kinase)
 81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The recognition of peptide antigens by T cells through their antigen receptors (T cell receptors, or TCR), together with the ligation of additional surface molecules called costimulatory receptors, rapidly induces interactive signaling pathways that lead to transcriptional initiation at genes such as that of the autocrine growth factor interleukin 2 (IL-2). Activation of the ERK and JNK subfamilies of MAPK mediates some of these signals. This unit presents procedures for a solid-phase kinase assay and immune-complex kinase assay to measure JNK and ERK activities, respectively, in T cells that have been appropriately stimulated. Also described is a procedure for preparing GSThyphen;cJun/GSH-Sepharose beads needed in the solid-phase JNK protein kinase activity assay.
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PMID:Analyzing mitogen-activated protein kinase (MAPK) activities in T cells. 1843 22

Although there has been considerable interest in the regulation of NFkappaB activation by glutathionylation, the possibility of IkappaB as a target for glutathionylation has not been investigated. We now report that Cys(189) of IkappaB alpha is a target for S-glutathionylation. This modification is reversed by thiols such as dithiothreitol and GSH. The glutathionylated IkappaB alpha appears to be significantly less susceptible than is native protein to phosphorylation by IkappaB kinase and casein kinase II, as well as to in vitro ubiquitination. This finding suggests that glutathionylation plays a regulatory role, presumably through structural alterations. HeLa cells treated with oxidant inducing GSH oxidation such as diamide showed the accumulation of glutathionylated IkappaB alpha. This mechanism suggests an alternative modification to the redox regulation of cysteine in IkappaB alpha and a possible mechanism in the regulation of NFkappaB activation.
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PMID:Glutathionylation regulates IkappaB. 1855 96

Estrogens are considered neurotrophic for dopamine neurons. Parkinson's disease is more frequent in males than in females, and more prevalent in females with short reproductive life. Estrogens are neuroprotective against neurotoxic agents for dopamine neurons in vivo and in vitro. Here, we have investigated the role of estrogens in wild-type (WT) and parkin null mice (PK-/-). WT mice present sexual dimorphisms in neuroprotective mechanisms (Bcl-2/Bax, chaperones, and GSH), but some of these inter-sex differences disappear in PK-/-. Tyrosine hydroxylase (TH) protein and TH+ cells decreased earlier and more severely in female than in male PK-/- mice. Neuronal cultures from midbrain of WT and PK-/- mice were treated with estradiol from 10 min to 48 h. Short-term treatments activated the mitogen-activated protein kinase pathway of WT and PK-/- neurons and the phosphatidylinositol 3'-kinase/AKT/glycogen synthase kinase-3 pathway of WT but not of PK-/- cultures. Long-term treatments with estradiol increased the number of TH+ neurons, the TH expression, and the extension of neurites, and decreased the level of apoptosis, the expression of glial fibrillary acidic protein, and the number of microglial cells in WT but not in PK-/- cultures. The levels of estrogen receptor-alpha were elevated in midbrain cultures and in the striatum of adult PK-/- male mice, suggesting that suppression of parkin changes the estrogen receptor-alpha turnover. From our data, it appears that parkin participates in the cellular estrogen response which could be of interest in the management of parkin-related Parkinson's disease patients.
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PMID:Gender differences and estrogen effects in parkin null mice. 1864 94

Oxidative stress is known to be a common feature of cholestatic syndrome. We have described the internalization of multidrug resistance-associated protein 2 (Mrp2), a biliary transporter involved in bile salt-independent bile flow, under acute oxidative stress, and a series of signaling pathways finally leading to the activation of novel protein kinase C were involved in this mechanism; however, it has been unclear whether the internalized Mrp2 localization was relocalized to the canalicular membrane when the intracellular redox status was recovered from oxidative stress. In this study, we demonstrated that decreased canalicular expression of Mrp2 induced by tertiary-butyl hydroperoxide (t-BHP) was recovered to the canalicular membrane by the replenishment of GSH by GSH-ethyl ester, a cell-permeable form of GSH. Moreover, pretreatment of isolated rat hepatocytes with colchicine and PKA inhibitor did not affect the t-BHP-induced Mrp2 internalization process but did prevent the Mrp2 recycling process induced by GSH replenishment. Moreover, intracellular cAMP concentration similarly changed with the change of intracellular GSH content. Taken together, our data clearly indicate that the redox-sensitive balance of PKA/PKC activation regulates the reversible Mrp2 localization in two different pathways, the microtubule-independent internalization pathway and -dependent recycling pathway of Mrp2.
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PMID:Canalicular Mrp2 localization is reversibly regulated by the intracellular redox status. 1933 16

Glyoxalase I (GLO1), together with glyoxalase II and the co-factor GSH, comprise the glyoxalase system, which is responsible for the detoxification of the cytotoxic glycolytic-derived metabolite methylglyoxal (MG). We, and others, have previously reported that GLO1 is subjected to several post-translational modifications, including a NO-mediated modification and phosphorylation. In this study, we demonstrate that GLO1 is a substrate for calcium, calmodulin-dependent protein kinase II (CaMKII). Site-directed mutagenesis of several serine and threonine residues revealed that CaMKII induced phosphorylation of GLO1 at a single site Thr-106. Mutagenesis of Thr-106 to Ala in GLO1 completely abolished the CaMKII-mediated phosphorylation. A phosphopeptide bracketing phosphothreonine-106 in GLO1 was used as an antigen to generate polyclonal antibodies against phosphothreonine-106. By using this phospho-specific antibody, we demonstrated that TNF induces phosphorylation of GLO1 on Thr-106. Furthermore, we investigated the role of NO-mediated modification and phosphorylation of GLO1 in the TNF-induced transcriptional activity of NF-kappaB. Overexpression of WT GLO1 suppressed TNF-induced NF-kappaB-dependent reporter gene expression. Suppression of the basal and TNF-induced NF-kappaB activity was significantly stronger upon expression of a GLO1 mutant that was either deficient for the NO-mediated modification or phosphorylation on Thr-106. However, upon overexpression of a GLO1 mutant that was deficient for both types of modification, the suppressive effect of GLO1 on TNF-induced NF-kappaB activity was completely abolished. These results suggest that NO-modification and phosphorylation of GLO1 contribute to the suppression of TNF-induced NF-kappaB-dependent reporter gene expression. In line with this, knock-down of GLO1 by siRNA significantly increased TNF-induced NF-kappaB-dependent reporter gene expression. These findings suggest that phosphorylation and NO-modification of glyoxalase I provides another control mechanism for modulating the basal and TNF-induced expression of NF-kappaB-responsive genes.
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PMID:Phosphorylation on Thr-106 and NO-modification of glyoxalase I suppress the TNF-induced transcriptional activity of NF-kappaB. 1919 7

Nitrite has recently emerged as an important bioactive molecule, capable of conferring cardioprotection and a variety of other benefits in the cardiovascular system and elsewhere. The mechanisms by which it accomplishes these functions remain largely unclear. To characterize the dose response and corresponding cardiac sequelae of transient systemic elevations of nitrite, we assessed the time course of oxidation/nitros(yl)ation, as well as the metabolomic, proteomic, and associated functional changes in rat hearts following acute exposure to nitrite in vivo. Transient systemic nitrite elevations resulted in: (1) rapid formation of nitroso and nitrosyl species; (2) moderate short-term changes in cardiac redox status; (3) a pronounced increase in selective manifestations of long-term oxidative stress as evidenced by cardiac ascorbate oxidation, persisting long after changes in nitrite-related metabolites had normalized; (4) lasting reductions in glutathione oxidation (GSSG/GSH) and remarkably concordant nitrite-induced cardioprotection, which both followed a complex dose-response profile; and (5) significant nitrite-induced protein modifications (including phosphorylation) revealed by mass spectrometry-based proteomic studies. Altered proteins included those involved in metabolism (eg, aldehyde dehydrogenase 2, ubiquinone biosynthesis protein CoQ9, lactate dehydrogenase B), redox regulation (eg, protein disulfide isomerase A3), contractile function (eg, filamin-C), and serine/threonine kinase signaling (eg, protein kinase A R1alpha, protein phosphatase 2A A R1-alpha). Thus, brief elevations in plasma nitrite trigger a concerted cardioprotective response characterized by persistent changes in cardiac metabolism, redox stress, and alterations in myocardial signaling. These findings help elucidate possible mechanisms of nitrite-induced cardioprotection and have implications for nitrite dosing in therapeutic regimens.
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PMID:Mechanistic insights into nitrite-induced cardioprotection using an integrated metabolomic/proteomic approach. 1922 60

In recent years, it has been accepted that oxidative stress is critically involved in the etiopathology of Parkinson's disease (PD) and as a result new therapeutic targets for reduction of oxidant injury and neuroprotection can be defined. Here we discuss the potential use of the transcription factor nuclear factor erythroid-2-related factor 2 (Nrf2), as a pharmacological target for neuroprotective therapy in PD. Data generated by various groups indicate that Nrf2 induces the expression of a group of cytoprotective, antixenobiotic and antioxidant enzymes that include heme oxygenase-1, NAD(P)H:quinone oxidoreductase and enzymes of glutathione (GSH) metabolism such as gamma-glutamyl cysteine ligase, GSH transferases and so on. Two strategies are known to increase Nrf2 transcriptional activity in PD: i) use of certain catechol-derived quinones for selective inhibition of the Nrf2 repressor Kelch-like ECH-associated protein to increase of Nrf2 protein levels; and ii) use of glycogen synthase kinase 3beta inhibitors to maintain high protein and activity levels of Nrf2 in the nucleus. This review provides a rationale for drug design of appropriate molecules that might endorse a neuroprotective strategy to PD on the basis of attenuation of oxidative stress.
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PMID:The transcription factor Nrf2 as a new therapeutic target in Parkinson's disease. 1923 54

Extracellular signal-regulated protein kinase (ERK), one of the mitogen-activated protein kinase, has been known to be involved in diverse cellular functions. In this work, we found that basically inhibition of this kinase in cultured cells markedly increased the gamma-glutamate-cysteine ligase (GCL; EC 6.3.2.2) activity, but without any considerable induction of the GCL genes. The increased GCL activity consequently elevated the cellular GSH level and eventually enhanced the cellular antioxidant capacity. Genetic inhibition of B-Raf, the upstream of ERK, also resulted in increased GCL activity and GSH level. Recent evidence also suggests that chronic pro-oxidant exposure results in the loss of ERK phosphorylation in vivo. Therefore, the findings in the present study suggest that inhibition of B-Raf/MEK/ERK pathway might be a promising physiological approach to up-regulate GCL activity and GSH. This study would also help us to understand the comprehensive role of the Raf/MEK/ERK pathway in overall physio/pathological conditions.
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PMID:Regulation of GCL activity and cellular glutathione through inhibition of ERK phosphorylation. 1927 32

The ubiquitous tripeptide glutathione (GSH) is an essential factor in many biological processes, thus its depletion has a major impact on cell function and survival. In this study, we examined regulation of GSH in cardiomyocytes under chronic oxidative stress elicited by myocardial infarction (MI). Cardiac dysfunction was induced in rats by coronary artery ligation, and experiments were conducted in myocytes isolated from non-infarcted left ventricle and septum after 6-8 weeks. Fluorescence microscopy studies using the probe monochlorobimane showed that [GSH] in myocytes from post-MI hearts was 42% less than in sham control hearts (P < 0.05). However, depleted GSH levels were normalized after 5-6 h by an insulin mimetic (bis-peroxovanadium-1,10-phenanthroline, bpV(phen); 10 micromol l(-1)) or by exogenous pyruvate (5 mmol l(-1)). The increase in [GSH] by bpV(phen) was partly inhibited by buthionine sulphoximine (BSO; 50 micromol l(-1)), a blocker of GSH synthesis, and by 1,3-bis-(2-chloroethyl)-1-nitrosourea (BCNU; 100 micromol l(-1)), an inhibitor of glutathione disulphide reductase. By comparison, the effect of pyruvate was not altered by BSO but was completely blocked by BCNU. Studies using inhibitors of signalling cascades indicated that upregulation of [GSH] by bpV(phen) in myocytes from post-MI hearts was mediated by mitogen activated protein kinase/extracellular signal-regulated kinase kinase 1/2 and p38 mitogen-activated protein kinase but not by phosphatidylinositol 3-kinase. The effect of pyruvate was not altered by any kinase inhibitor tested. In cells loaded with the probe TEMPO-9-AC to monitor superoxide anion, baseline fluorescence was 2.3-fold greater in post-MI myocytes than in sham control myocytes (P < 0.05) and was markedly decreased by diphenyleneiodonium (30 micromol l(-1)), an inhibitor of NADPH oxidase, exogenous GSH (10 mmol l(-1)) or bpV(phen). In parallel studies, [GSH] in post-MI myocytes was also normalized by diphenyleneiodonium or exogenous GSH. These data show that GSH is differentially regulated by receptor tyrosine kinase-dependent and -independent agonists that maintain functional GSH levels necessary to neutralize excess generation of reactive oxygen species in the failing heart.
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PMID:Glutathione homeostasis in ventricular myocytes from rat hearts with chronic myocardial infarction. 1939 62

Glutathione (GSH), the most abundant intracellular nonprotein thiol, is critical for many cellular functions. The rate-limiting step in GSH synthesis is catalyzed by glutamate cysteine ligase (GCL), a heterodimer composed of a catalytic (GCLC) and a modifier (GCLM) subunit. The tissue-specific regulation of GSH synthesis is poorly understood. We showed previously that gonadotropin hormones regulate ovarian GSH synthesis. In the present study, we sought to clarify the ovarian cell type-specific effects of follicle-stimulating hormone (FSH) and estradiol on GSH synthesis. Immature female rats were treated with estradiol to stimulate development of small antral follicles. Granulosa cells (GCs) from these follicles or whole follicles were cultured in serum-free media, with or without FSH and 17beta-estradiol. The GSH and GCLC protein and mRNA levels increased in GCs treated with FSH alone. The effects of FSH on GCLC and GCLM protein and mRNA levels, GCL enzymatic activity, and GSH concentrations in GCs were significantly enhanced by the addition of estradiol. Estradiol alone had no effects on GSH. Dibromo-cAMP mimicked and protein kinase A (PKA) inhibitors prevented FSH stimulation of GCL subunit protein levels. In cultured small antral follicles, FSH stimulated estradiol synthesis and robustly increased GCL subunit mRNA and protein levels and GSH concentrations. The GCL subunit mRNA expression increased in both the granulosa cells and theca cells of follicles with FSH stimulation. These data demonstrate that maximal stimulation of GSH synthesis by FSH in granulosa cells and follicles requires estradiol. Without estradiol, FSH causes lesser increases in GCL subunit expression via a PKA-dependent pathway.
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PMID:Follicle-stimulating hormone and estradiol interact to stimulate glutathione synthesis in rat ovarian follicles and granulosa cells. 1951 19


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