Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.13 (protein kinase C)
 49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Increased Na+/H+ exchanger (NHE) activity has been demonstrated in cells from patients with hypertension and diabetic nephropathy. Vascular myocytes from the spontaneously hypertensive rat (SHR) also exhibit increased NHE activity as compared with cells from the normotensive Wistar Kyoto rat (WKY). The interaction of increased glucose concentrations with NHE activity is unclear. The effect of glucose on NHE activity, NHE-1 (isoform 1) protein expression, and phosphorylation of cultured vascular myocytes from these rat strains was thus investigated. NHE activity was determined fluorometrically with 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF). A rabbit NHE-1-specific polyclonal antibody was used (1) to measure NHE-1 abundance in Western blots of cell extracts and (2) for immunoprecipitating 32P-labeled NHE-1. Cells from SHR exhibited increased NHE activity and NHE-1 phosphorylation as compared with cells from WKY, with similar NHE-1 protein content per cell. Incubation in 25 mmol.L-1 glucose for 24 hours led to increased NHE activity only in WKY cultures, with no change in NHE-1 protein but a concomitantly reduced NHE-1 phosphorylation. Changes in NHE activity in WKY cells were reversed by inhibition of protein kinase C. Incubation of SHR cells with 25 mmol.L-1 glucose did not enhance the increased NHE activity or NHE-1 phosphorylation present in these cells. Thus, high glucose levels have disparate effects on NHE activity and NHE-1 phosphorylation in cells from different rat strains. The glucose-induced increase in NHE-1 turnover number in WKY cells is not mediated by an increase in its direct phosphorylation, but is dependent on protein kinase C.
 ...
PMID:Glucose-induced changes in activity and phosphorylation of the Na+/H+ exchanger, NHE-1, in vascular myocytes from Wistar-Kyoto and spontaneously hypertensive rats. 854 67

Vascular myocytes from the spontaneously hypertensive rat (SHR) demonstrate elevated Na(+)-H(+) exchanger activity associated with increased cell proliferation and hyperresponsiveness to agonists such as phorbol esters. Since the Na(+)-H(+) exchanger isoform 1 (NHE-1) is stimulated by protein kinase C, we have investigated the effects of phorbol esters on NHE-1 activity and its phosphorylation in vascular myocytes of these rats. SHR cells demonstrated a larger alkalinization response to 12-O-tetradecanoylphorbol 13-acetate than Wistar-Kyoto rat (WKY) cells. Kinetic analyses indicated that whereas 12-O-tetradecanoylphorbol 13-acetate increased the maximal transport capacity of NHE-1 in both cell types, affinity for H+ was increased in WKY cells and cooperativity for H+ at the internal modifier site was reduced in SHR cells. In neither cell type was the subcellular distribution of NHE-1 altered by phorbol ester stimulation. NHE-1 phosphorylation was markedly reduced in WKY cells stimulated by the phorbol ester, an effect abolished by inhibition of protein kinase C. In contrast, NHE-1 phosphorylation in quiescent SHR cells was approximately double that of WKY cells and was reduced after phorbol ester treatment. Inhibition of protein kinase C in SHR cells led to a marked elevation of NHE-1 phosphorylation that was not associated with a change in the exchanger activity, but WKY cells exhibited a small, insignificant rise in NHE-1 phosphorylation. Thus, the kinetic responses of NHE-1 to phorbol esters in vascular myocytes of these rat strains are different, the changes in exchanger kinetics of SHR resembling those described in human hypertension. NHE-1 phosphorylation has an inverse relationship with protein kinase C activity. However, modulation of NHE-1 phosphorylation may not be associated with concurrent alterations in activity, indicating a role for non-phosphorylation-dependent mechanisms.
 ...
PMID:Phorbol ester activation of the rat vascular myocyte Na(+)-H(+) exchanger isoform 1. 861 61

Treatment of human platelets with phorbol 12-myristate 13-acetate (PMA) and arginine vasopressin (AVP) increase the phosphorylation and activation of mitogen-activated protein kinase (MAPK). Electrophoretic retardation of MAPK mobility on SDS-polyacrylamide gels was used for determination of MAPK phosphorylation. The activity of MAPK was tested in myelin basic protein (MBP)-containing polyacrylamide gels. In this study we compared the PMA and AVP signal transduction pathways leading to the activation of MAPKs and Na+/H+ exchanger (NHE). Both agonists stimulate MAPK and NHE activities in a similar time frame and concentration dependence. The MAPK and NHE activities induced by PMA were inhibited by staurosporine, a potent inhibitor for protein kinase C (PKC), and by MAPK kinase (MEK) inhibitor, PD98059, but were not affected by the tyrosine kinase inhibitor genistein. In contrast, both AVP-induced MAPK and NHE activities were inhibited by genistein and MEK inhibitor but were not affected by staurosporine. Immunoprecipitation studies demonstrate that PMA, but not AVP, enhances the basal phosphorylation of the NHE-1. In this study, MAPKs are suggested to be a part of converging signaling leading to NHE activation by PKC-dependent and AVP-tyrosine kinase-dependent pathways. We propose that the MAPK activation of the NHE-1 does not involve phosphorylation of this exchanger protein. On the other hand, PKC can lead to phosphorylation and to additional activation of the NHE-1 through a MAPK-independent pathway.
 ...
PMID:Stimulation of mitogen-activated protein kinase and Na+/H+ exchanger in human platelets. Differential effect of phorbol ester and vasopressin. 866

Two routes, associated with protein kinase C (PKC) and cyclic AMP (cAMP), to regulate the activation of Na(+)-H+ exchange (NHE) were investigated in the outer hair cell (OHC). The intracellular pH (pHi) of OHC loaded with the pHi indicator dye was measured by using fluorescence ratio imaging microscopy. The acid load was carried out by an "NH4Cl pre-pulse". The rate of acid extrusion after an acid load was increased as pHi became more acidic. The linear relationship between the rate of acid extrusion and pHi suggests the presence of an internal modifier site for the H+ ion on the NHE. Although addition of phorbol 12-myristate 13-acetate (TPA) had no effect on pHi in unstimulated conditions, the pHi recovery from an acid load was enhanced by exposure to TPA. This finding can be explained by the pHi dependence of the PKC action, namely, that the activating effect of PKC on the NHE is facilitated by internal acidification. In contrast to the PKC effect, addition of dibutyryl cAMP failed to change the rate of acid extrusion. These findings are consistent with the characteristic property of the NHE-1. Thus, intracellular signal transduction to regulate the activity of the NHE in the OHC involves phosphorylation by PKC.
 ...
PMID:Effects of protein kinase C on the Na(+)-H+ exchange in the cochlear outer hair cell. 897 15

The trout red blood cell Na+/H+ antiporter (beta NHE) plays two interesting properties: it is the only NHE own to be activated by cyclic AMP, and the activation process is followed by a desensitisation of the transport system itself. Cloning and expression of beta NHE have provided inificant information about Na+/H+ activation, in particular that activation by cyclic AMP is directly dependent upon the presence of two protein kinase A consensus sites in the cytoplasmic tail of the antiporter. Expression of beta NHE in fibroblasts demonstrates that the protein kinase A (PKA) and protein kinase C (PKC) activation pathways are independent and do not converge a common kinase. Moreover, the hydrophilic C-terminal fragment is essential to the mediation of the various hormonal responses. NHE1 (the human ubiquitous isoform) is not activated by cyclic AMP, but a "NHE1 transmembrane domain/beta NHE cytoplasmic domain' chimera is fully activated by cyclic AMP. In red cells, activation of beta NHE is the result of phosphorylation by PKA of at least two independent sites. Desensitisation, inhibited by the phosphatase inhibitor okadaic acid, may consist of the dephosphorylation of one of these two sites. Furthermore, Calyculin A (CIA), another specific protein phosphatase inhibitor, induces in unstimulated cells a Na+/H+ exchange activity whose exchange properties are very different from those of the adrenergically stimulated antiporter. It is suggested that CIA may be able to revive "sequestered' antiporters. We propose that the molecular events underlying beta NHE desensitisation could be similar to those involved in rhodopsin desensitisation. Antibodies were generated against trout red cell arrestin in order to analyse the binding of arrestin to the activated exchanger. Recombinant trout arrestin was produced in a protease-deficient strain of Escherichia coli and its functionality tested in a reconstituted rhodopsin assay.
 ...
PMID:Regulation of Na+/H+ antiporter in trout red blood cells. 905 Feb 44

We have cloned and sequenced the cDNA for a Na+/H+ exchanger (NHE) from Xenopus laevis oocytes. This cDNA contains an open reading frame encoding a protein of 782 amino acids with 12 putative transmembrane domains and a long cytoplasmic tail. The protein exhibits a strong homology at the amino acid level to the human NHE-1 as well as to the beta NHE from trout red blood cells: 69% and 58% respectively. Two potential N-linked glycosylation sites at Asn56 and Asn351 were identified. Three potential protein kinase C phosphorylation sites at the cytoplasmic tail were identified at Ser494, Thr726 and Ser747. RT-PCR revealed the expression of the X1-NHE in Xenopus heart, reticulocytes and skeletal muscle.
 ...
PMID:Cloning and sequencing of the cDNA encoding for a Na+/H+ exchanger from Xenopus laevis oocytes (X1-NHE). 910 79

In the red blood cell membrane, sodium-proton exchange (NHE-1) exchanges intracellular H(+), Li(+), and Na(+) with extracellular Na(+). In hypertensives (HT), the maximal velocity of translocation (V max)of Na(+)/H(+) and of Na(+)/Li(+) exchange modes are higher, while apparent affinity for external Na(+) of Na(+)/Li(+) exchange and Hill's coefficient for H(+) activation of Na(+)/H(+) exchange are lower than in normotensive subjects (NT). We have therefore examined the effects of protein kinase C (PKC) and insulin on red blood cell membrane phosphorylation and on the kinetic properties of cation heteroexchange. In red cell from NT, PMA-induced activation of PKC reduced K(m) for H(+) of NHE but it did not affect V(max) and K(m) for Na(+). In red cell from HT, PMA-induced a greater PKC stimulation and membrane phosphorylation of band 3,4.1,4.9 than in NT and it did not significantly reduced K(m) for H(i). On the contrary, in HT PKC activation significantly increased Hill's coefficient of NHE. The larger activation of PKC in HT could be due to downregulation secondary to higher membrane calpain activity. Incubation of red cells with insulin decreases K(m) for external Na(+) and increases V(max) of Na(+)/Li(+) exchange. Therefore, we have examined the relationships between Na(+)-activation kinetics of Na(+)/Li(+) exchange and fasting insulin levels. Na(+)-stimulated Li(+) efflux was studied by raising Na(+)up to 300 mM isoosmotically to measure K(m) for Na(+) and V (max). Li(+) efflux saturated at 150 mM external Na(+)in NT but not in HT because in HT it exhibited a two fold higher Na(+) Km. V(max) was higher in HT than in NT. In hyperinsulinemic (fasting insulin > 10 mu U/ml) HT, V(max) and Na(+) Km were higher than in normoinsulinemic HT. In NT, hyperinsulinemia was not associated to abnormal kinetic properties of Na(+)/Li(+)exchange. Stepwise multiple regression analysis confirmed that the main determinants of a high Km were blood pressure and insulin. Our results show that posttranslational effects of PKC and insulin affect the kinetic properties of NHE-1 in red blood cells and suggest that the differences observed between hypertensives and normotensive subjects can be accounted for by PKC activation and insulin exposure.
 ...
PMID:Posttranslational effects of protein kinase C and insulin on red cell membrane phosphorylation and cation heteroexchange in hypertension. 916 39

Increased activity of the Na(+)-H+ exchanger (NHE-1 isoform) has been observed in cells and tissues from hypertensive humans and animals, including the spontaneously hypertensive rat (SHR). No mutation in NHE-1 DNA sequence or alteration in NHE-1 mRNA and protein expression has been demonstrated in hypertension, indicating that alterations in proteins that regulate NHE-1 activity are responsible for increased activity. The recent finding that NHE-1 phosphorylation in SHR vascular smooth muscle cells (VSMCs) was greater than in Wistar-Kyoto rat (WKY) VSMCs suggested that NHE-1 kinases may represent an abnormal regulatory pathway present in hypertension. To define NHE-1 kinases altered in the hypertensive phenotype. We measured NHE-1 kinase activity by an in-gel-kinase assay using a recombinant glutathione S-transferase NHE-1 fusion protein as a substrate. At least 7 NHE-1 kinases (42 to 90 kD) were present in VSMCs. We studied a 90-kD kinase because it was the major NHE-1 kinase and exhibited differences between SHR and WKY. Comparison of 90-kD kinase activity revealed that SHR VSMCs had increased activity in growth-arrested cells and in cells stimulated by angiotensin II (100 nmol/L for 5 minutes). Activation of the 90-kD kinase by angiotensin II was Ca2+ dependent, PKC independent, and partially dependent on the mitogen-activated protein kinase pathway. These findings indicate that increased activity of a 90-kD NHE-1 kinase is a characteristic of SHR VSMCs in culture and suggest that alterations in the 90-kD NHE-1 kinase and/or proteins that regulate its activity may be a pathogenic component in hypertension in the SHR.
 ...
PMID:A 90-kD Na(+)-H+ exchanger kinase has increased activity in spontaneously hypertensive rat vascular smooth muscle cells. 918 Jun 27

Angiotensin II is a multifunctional agonist for vascular smooth muscle cells (VSMCs), stimulating increases in signal events, cell growth, and ion flux. We previously defined protein kinase C (PKC)-dependent and -independent mechanisms by which angiotensin II stimulated activity of the Na(+)-H+ exchanger isoform-1 (NHE-1) and identified a 90-kD kinase that exhibited increased activity in VSMCs isolated from genetically hypertensive rats. To determine whether this 90-kD kinase was p90rsk (RSK), VSMCs were stimulated with 100 nmol/L angiotensin II, and NHE-1 kinase activity was measured by phosphorylation of recombinant NHE-1 (a glutathione S-transferase fusion protein containing amino acids 516 to 815 of the cytoplasmic carboxyl tail) in vitro. NHE-1 kinase (90 kD) activity was markedly decreased by immunodepletion of RSK. Characterization of RSK activation by angiotensin II revealed many similarities to the 90-kD NHE-1 kinase, including time course and NHE-1 domain phosphorylation, as well as regulation by extracellular signal-regulated kinases (ERK1/2), intracellular Ca2+, and PKC. Specifically, angiotensin II stimulated a rapid and transient (peak, 5 minutes) increase in RSK activity. Analysis of several NHE-1 fusion proteins revealed that only proteins containing amino acids 670 to 714 were phosphorylated by RSK. Inhibiting ERK1/2 (30 mumol/L PD098059 for 30 minutes) or chelating intracellular Ca2+ prevented RSK activation. In contrast, downregulating PKC (1 mumol/L phorbol dibutyrate for 24 hours) had little effect. These findings establish RSK as a putative NHE-1 kinase and potential mediator of increased Na(+)-H+ exchange in hypertension.
 ...
PMID:Angiotensin II stimulates p90rsk in vascular smooth muscle cells. A potential Na(+)-H+ exchanger kinase. 924 88

Acid produces a dynamic effect on the cell phenotype of Barrett's esophagus (BE) ex vivo. An acid pulse induces hyperproliferation, whereas continuous acid exposure promotes differentiation. To examine the mechanism for acid pulse-induced hyperproliferation, we studied the Na+/H+ exchanger (NHE), which plays a role in the control of intracellular pH and cell proliferation. NHE was inhibited pharmacologically in endoscopic BE biopsies using amiloride analogs. Cell proliferation was assessed after pulsed or continuous acid exposure using tritiated thymidine incorporation assays and immunohistochemical analysis of proliferating cell nuclear antigen expression. The NHE-dependent intracellular pH response to an acid pulse was examined by pH-sensitive microfluorimetry using a Barrett's adenocarcinoma cell line TE7. NHE inhibition significantly reduced the hyperproliferative acid-pulse effect. Furthermore, the acid-pulse activation of NHE occurred via increased transporter activity (22Na uptake) without any change in NHE-1 protein levels. Inhibition of protein kinase C (PKC), an NHE activator, also reduced the hyperproliferative response. The response of TE7 cells to an acid pulse was similar to that of BE biopsies in terms of cell proliferation and NHE and PKC dependence. Acid-pulse exposure of TE7 cells resulted in intracellular acidification followed by reneutralization to an intracellular pH greater than preacidosis values. We conclude that NHE may mediate the hyperproliferative response of BE to an acid pulse via changes in intracellular pH.
 ...
PMID:Altered sodium-hydrogen exchange activity is a mechanism for acid-induced hyperproliferation in Barrett's esophagus. 965 83


<< Previous 1 2 3 4 5 6 7 Next >>