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:2.7.11.1 (protein kinase)
81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

This review focuses on the mechanisms whereby the cytosolic Ca2+ regulates the ubiquitous Na+/ H+ exchanger (NHE-1) and how these regulatory processes might modify the behavior of NHE-1 in essential hypertension. The pH setpoint for activation of the Na+/H+ exchanger is controlled by two interrelated and Ca(2+)-dependent pathways, namely, the protein kinase/ phosphatase cascade and Ca2+/calmodulin. The cytoplasmic domain of NHE-1 contains elements responsive to serine/theorine kinases and a high affinity binding site to Ca2+/calmodulin. Phosphorylation of NHE-1 or the binding of the Ca2+/calmodulin complex to its binding site promotes an alkaline shift in the pH setpoint for the exchanger. It is suggested that, in essential hypertension, an increased cellular Ca2+ load or an enhanced external Ca2+ entry stimulate the NHE-1 through protein kinase/phosphatase and Ca2+/calmodulin systems, thereby increasing its activity.
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PMID:The links between cellular Ca2+ and Na+/H+ exchange in the pathophysiology of essential hypertension. 880 85

The ubiquitously expressed Na+/H+ exchanger NHE1 is the target of multiple signaling pathways, including those activated by tyrosine kinase receptors, G protein-coupled receptors, and integrins. The intracellular pathways leading to activation of NHE1 are poorly understood. To gain more insight into these activation pathways, we examined the role of mitogen-activated protein kinases (MAPKs) as potential mediators of NHE1 activation by extracellular stimuli such as growth factors and hyperosmotic stress. Whereas p44 MAPK does not appear to phosphorylate NHE1 in vitro, we found that inhibition of the p42/p44 MAPK signaling by expression of a dominant negative form of p44 MAPK, by expression of the MAP kinase phosphatase MKP-1, or by inhibition of MAPK kinase 1 (MKK1) with the PD 98059 compound reduced by 50-60% NHE1 activation in response to growth factors. This inhibitory effect also was observed in C-terminal NHE1 deletion mutants in which the major phosphorylation sites have been deleted. Furthermore, the use of a CCL39-derived cell line expressing an estradiol-regulated form of oncogenic Raf-1 (CCL39-deltaRaf-1:ER) revealed that the exclusive activation of the Raf --> MKK1 --> p42/p44 MAPK cascade was capable of inducing NHE1 activation to the same extent as potent growth factors like thrombin. Together, our findings demonstrate that the p42/p44 MAPK cascade plays a predominant role in the regulation of NHE1 by growth factors, an action that is mediated via accessory proteins that remain to be identified. In contrast, we found no evidence in favor of the contribution of any MAPK, p42/p44, p38 MAPKs, and Jun kinase, in NHE1 activation by osmotic stress.
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PMID:The p42/p44 mitogen-activated protein kinase cascade is determinant in mediating activation of the Na+/H+ exchanger (NHE1 isoform) in response to growth factors. 899 58

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.
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PMID:Regulation of Na+/H+ antiporter in trout red blood cells. 905 Feb 44

To characterize and localize a K+/H+ antiport mechanism in the renal medullary thick ascending limb (MTAL), membrane vesicles were isolated from a rat MTAL homogenate. K+/H+ antiport (in > out H+ gradient-stimulated 86Rb+ uptake) was abolished by barium and verapamil (apparent Ki of 55 microM) but unaffected by other K+ channel blockers such as quinidine and high amiloride concentrations. SCH 28080, a H+/K+-ATPase blocker, did not affect K+/H+ antiport. K+/H+ antiport activity was correlated positively with the enrichment factor of the membranes in the apical marker enzyme alkaline phosphatase (r = 0.875, p < 0.01) and negatively correlated with the enrichment factor in basolateral Na+/K+-ATPase (r = -0.665, p < 0.05). Moreover, a functional interaction occurred with Na+/H+ exchange (NHE) consistent with colocation of K+/H+ antiport and apical NHE-3, not basolateral NHE-1. K+/H+ antiport was shown by intracellular pH measurements to be inhibited by arginine vasopressin and 8-bromo-cAMP through cAMP-dependent protein kinase (protein kinase A) activation. These results demonstrate the presence of a K+/H+ antiport mechanism, which is inhibited by arginine vasopressin via protein kinase A, in the apical membrane of the MTAL.
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PMID:Apical location and inhibition by arginine vasopressin of K+/H+ antiport of the medullary thick ascending limb of rat kidney. 932 90

The calmodulin (CaM)-binding domain reduces the affinity of the Na+/H+ exchanger NHE1 for intracellular H+ by exerting an autoinhibitory function in quiescent cells. We replaced this domain (aa 637-656) with homologous segments from other NHE isoforms (NHE2 and 4) or functionally similar regions from other sources (Na+/Ca2+ exchanger, CaM-dependent protein kinase II, plasma membrane Ca2+-pump, or CaM-binding peptide Trp3). The NHE-1-, NHE2-, and NHE4-segments bound CaM with Kds of 16, 130, and 27 nM, respectively. These chimeric molecules were expressed in the exchanger-deficient cell PS120. NHE1 with incorporated NHE2-segment was activated in response to Ca2+-mobilizing agents ionomycin and thrombin resulting in an alkaline shift of the intracellular pH (pHi)-dependence of 22Na+ uptake, as was the case with the intact rat NHE2. In contrast, incorporation of the NHE4-segment or other CaM-binding segments induced a constitutive alkaline shift of pHi-dependence with concomitant abolishment of Ca2+-dependent activation, indicating that these segments could not function as an autoinhibitory domain in NHE1. Detailed analyses revealed that Leu639, Lys651 and Tyr652, conserved in the NHE1- and NHE2-segments, but not in the NHE4-segment, are important for the autoinhibition. Furthermore, 125I-labeled CaM-binding peptide from NHE1 was efficiently crosslinked to the NHE1 protein, suggesting that the inhibitory domain physically interacts with part(s) of the molecule. Together, these findings support the notion that the reduction of H+ affinity in Na+/H+ exchange occurs through a mechanism involving a highly sequence-specific interaction of the inhibitory domain with its putative acceptor in NHE1.
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PMID:Calmodulin-binding autoinhibitory domain controls "pH-sensing" in the Na+/H+ exchanger NHE1 through sequence-specific interaction. 933 43

We examined factors important in regulation of expression of the Na+/H+ exchanger gene in NIH/3T3 cells. A stable fibroblast cell line was generated that contained a 1.1-kb proximal fragment of the mouse NHE1 promoter. The addition of serum to serum-starved cells resulted in an increase in activity of the NHE1 promoter. The mitogenic agonists insulin, thrombin, and epidermal growth factor also increased transcription from the NHE1 promoter. Phorbol esters also increased NHE1 promoter-directed transcription, whereas the serine/threonine protein kinase inhibitor 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine inhibited this stimulation. The protein kinase inhibitors GF-109203X, PD-98059, and genistein all stimulated promoter activity. Promoter deletion analysis and gel mobility shift assays showed that a region between 0.9 and 1.1 kb from the start site was involved in mediating the effect of mitogenic stimulation. The results show that a variety of mitogenic factors can activate the NHE1 promoter during cell growth and proliferation.
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PMID:Regulation of Na+/H+ exchanger gene expression: mitogenic stimulation increases NHE1 promoter activity. 953 Jan 16

The ubiquitously expressed Na-H exchanger, NHE1, acts downstream of RhoA in a pathway regulating focal adhesion and actin stress fiber formation. p160ROCK, a serine/threonine protein kinase, is a direct RhoA target mediating RhoA-induced assembly of focal adhesions and stress fibers. Here, stress fiber formation induced by p160ROCK was inhibited by the addition of a specific NHE1 inhibitor, ethylisopropylamiloride, in CCL39 fibroblasts, and was absent in PS120 mutant fibroblasts lacking NHE1. In CCL39 cells, NHE1 activity was stimulated by expression of mutationally active p160ROCK, but not by mutationally active protein kinase N, another RhoA target kinase. Expression of a dominant interfering p160ROCK inhibited RhoA-, but not Cdc42- or Rac-activation of NEH1. In addition, the p160ROCK-specific inhibitor Y-27632 inhibited increases in NHE1 activity in response to RhoA, and to lysophosphatidic acid (LPA), which stimulates RhoA, and it also inhibited LPA-increased phosphorylation of NHE1. A C-terminal truncation of NHE1 abolished both LPA-induced phosphorylation and activation of the exchanger. Furthermore, mutationally active p160ROCK phosphorylated an NHE1 C-terminal fusion protein in vitro, and this was inhibited in the presence of Y-27632. Phosphopeptide maps indicated that identical residues in NHE1 were phosphorylated by p160ROCK in vivo and in vitro. These findings identify p160ROCK as an upstream, possibly direct, activator of NHE1, and suggest that NHE1 activity and phosphorylation are necessary for actin stress fiber assembly induced by p160ROCK.
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PMID:p160ROCK mediates RhoA activation of Na-H exchange. 970 30

Replacement of external NaCl with LiCl induced cytoplasmic alkalinization in CCL-39 cells and rat L6 myoblasts expressing the endogenous Na+/H+ exchanger isoform NHE1. This Li+-induced alkalinization is due to activation of the Na+/H+ exchanger because it was completely inhibited by 100 microM ethylisopropylamiloride (apparent Kd=1 microM) and because it did not occur in exchanger-deficient PS120 cells. The effect of Li+ was not mimicked by Na+, K+, Cs+ and choline+. Li+ caused cytoplasmic alkalinization in PS120 cells expressing NHE1 or NHE2, but not NHE3, when Li+ was added to cells at a concentration high enough to saturate their external transport sites as predicted from Li+ affinities. Li+ did not induce phosphatidylinositol (PI) turnover or intracellular Ca2+ mobilization. Li+-induced alkalinization was not affected by protein kinase C down-regulation, loss of glycogen synthase kinase 3beta caused by antisense oligonucleotide treatment, or pretreatment with calphostin C, pertussis toxin, MEK inhibitor PD98059 and PI3-kinase inhibitor LY294002. However, it was markedly suppressed by the tyrosine kinase inhibitor genistein (10 microM). Thus, externally added Li+ activates NHE1 and NHE2 via a mechanism possibly involving a tyrosine kinase, causing an increase in cytoplasmic pH that could potentially affect various cell functions.
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PMID:Lithium activates mammalian Na+/H+ exchangers: isoform specificity and inhibition by genistein. 1067 42

Modulation of cell proliferation has often been thought to be connected to changes in the activity of pH-regulatory transporters and consequently intracellular pH (pH(i)). The influence of natriuretic peptides, diadenosine polyphosphates, adenosine and ATP as well as platelet-derived growth factor (PDGF) on pH(i) regulation of cultured rat mesangial cells was examined with the pH-sensitive dye 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein. The inhibitors of Na(+)/H(+) exchange, amiloride and HOE694, blocked pH(i) recovery completely in the absence of and by approximately 50% in the presence of HCO(3)(-)/CO(2). Natriuretic peptides (ANP, BNP, CNP, urodilatin) completely inhibited pH(i) recovery in the absence of and by approximately 40% in the presence of HCO(3)(-)/CO(2). These effects were abolished by the cGMP-dependent protein kinase inhibitor KT5823. Diadenosine polyphosphates (Ap3A-Ap6A), ATP and adenosine also inhibited pH(i) recovery completely in the absence of and partially (30-40%) in the presence of HCO(3)(-)/ CO(2). The effect of adenosine was abolished in the presence of the cAMP-dependent protein kinase inhibitor KT5720, and that of Ap5A by the protein kinase C inhibitor calphostin C. PDGF activated acid extrusion in these cells by approximately 40%. From the four cloned isoforms of the Na(+)/H(+) exchanger in the rat, only transcripts of NHE-1 were found in these mesangial cell cultures using RT-PCR analysis. These data suggest that in these rat mesangial cells the Na(+)/H(+) exchanger, specifically the NHE-1 isoform, accounts for around 50% of pH(i) recovery from an acid load under physiological conditions, and that Na(+)/H(+) exchange stimulated by acidification can be inhibited by activation of PKG, PKA, and PKC and stimulated by PDGF after acute exposition to these agonists.
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PMID:Natriuretic peptides and diadenosine polyphosphates modulate pH regulation of rat mesangial cells. 1074 97

The activity of the Na(+)/H(+) exchanger has been implicated as an important contributing factor in damage to the myocardium that occurs during ischemia and reperfusion. We examined regulation of the protein in ischemic and reperfused isolated hearts and isolated ventricular myocytes. In isolated myocytes, extracellular signal-regulated kinases were important in regulating activity of the exchanger after recovery from ischemia. Ischemia followed by reperfusion caused a strong inhibitory effect on NHE1 activity that abated with continued reperfusion. Four major protein kinases of size 90, 55, 44, and 40 kDa phosphorylated the Na(+)/H(+) exchanger. The Na(+)/H(+) exchanger-directed kinases demonstrated dramatic increases in activity of 2-10-fold that was induced by 3 different models of ischemia and reperfusion in intact hearts and isolated myocytes. p90(rsk) was identified as the 90-kDa protein kinase activated by ischemia and reperfusion while ERK1/2 was identified as accounting for some of the 44-kDa protein kinase phosphorylating the Na(+)/H(+) exchanger. The results demonstrate that MAPK-dependent pathways including p90(rsk) and ERK1/2 and are important in regulating the Na(+)/H(+) exchanger and show their dramatic increase in activity toward the Na(+)/H(+) exchanger during ischemia and reperfusion of the myocardium. They also show that ischemia followed by reperfusion have important inhibitory effects on Na(+)/H(+) exchanger activity.
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PMID:Activation of Na+/H+ exchanger-directed protein kinases in the ischemic and ischemic-reperfused rat myocardium. 1127 85


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