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:3.6.1.3 (ATPase)
65,361 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Increases in heart rate are accompanied by acceleration of relaxation. This effect is apparent at the single myocyte level and depends on sarcoplasmic reticulum (SR) Ca transport and Ca/calmodulin dependent protein kinase [CaMKII; see R. A. Bassani, A. Mattiazzi, and D. M. Bers. Am. J. Physiol. 268 (Heart Circ. Physiol. 37): H703-H712, 1995]. Because phosphorylation of phospholamban (PLB) by CaMKII can stimulate SR Ca transport, it is a plausible candidate mechanism. We examined this issue using ventricular myocytes isolated from wild-type (WT) mice and those in which the PLB gene was ablated by gene targeting (PLB-KO). During steady-state (SS) stimulation, twitch relaxation and intracellular Ca concentration ([Ca]i) decline were significantly faster than after a rest in both WT and PLB-KO myocytes. Furthermore, the CaMKII inhibitor KN-93 (1 microM) abolished the stimulation-dependent acceleration of twitch [Ca]i decline in PLB-KO. This indicates that neither PLB nor its phosphorylation are required for the CaMKII-dependent acceleration of the SS twitch [Ca]i decline and relaxation. Other quantitative aspects of Ca transport in WT and PLB-KO myocytes were also examined. As expected, the time constant (tau) of [Ca]i decline during the SS twitch is much faster in PLB-KO than in WT myocytes (112 +/- 6 vs. 188 +/- 14 ms, P < 0.0001). There was also an increase in SS SR Ca load, based on the change of [Ca]i during rapid caffeine-induced contractures (CafC) with Na/Ca exchange blocked (565 +/- 74 nM for WT, 1118 +/- 133 nM for PLB-KO, P < 0.01). Accounting for cytosolic Ca buffering, this implies a 37% increase in SR Ca content. The tau for [Ca]i decline of the cafC with Na present indicated slower extrusion by Na/Ca exchange in the PLB-KO mouse (2.2 +/- 0.2 s in WT vs. 3.2 +/- 0.2 in PLB-KO, P < 0.01), although exchanger protein expression was unchanged. Integrated Ca flux analysis in WT and PLB-KO myocytes, respectively, shows that 90 and 96% of Ca during twitch relaxation is removed by the SR Ca-ATPase, 9 and 3.4% by Na/Ca exchange, and 0.5 and 0.1% by slow mechanisms (mitochondria Ca uniporter and sarcolemmal Ca-ATPase). We conclude that the PLB-KO myocytes retain a CaMKII-dependent acceleration of SS twitch [Ca]i decline. The PLB-KO (vs. WT) myocytes also have higher SR Ca pump activity, higher SR Ca load, and reduced Na/Ca exchange activity.
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PMID:Cardiac myocyte calcium transport in phospholamban knockout mouse: relaxation and endogenous CaMKII effects. 957 39

Using SK-N-SH cells, we observe that muscarinic acetylcholine receptor activation by methacholine (MCh) rapidly and selectively diminishes l-NE transport capacity (Vmax) with little or no change in norepinephrine (NE) Km and without apparent effects on membrane potential monitored directly under current clamp. Over the same time frame, MCh exposure reduces the density of [3H]nisoxetine binding sites (Bmax) in intact cells but not in total membrane fractions, consistent with a loss of transport capacity mediated by sequestration of transporters rather than changes in intrinsic transport activity or protein degradation. Similar changes in NE transport and [3H]nisoxetine binding capacity are observed after phorbol ester (beta-PMA) treatment. Inhibition of PKC by antagonists and downregulation of PKC by chronic treatment with phorbol esters abolishes beta-PMA-mediated effects but produce only a partial blockade of MCh-induced effects. Neither muscarinic acetylcholine receptor nor PKC activation require extracellular Ca++ to diminish NET activity. In contrast, treatment of cells with the Ca++/ATPase antagonist, thapsigargin in Ca++-free medium, eliminates the staurosporine-insensitive component of MCh regulation. These findings were further corroborated by the ability of [1, 2-bis(o-amino-phenoxy)ethane-N,N,N',N'-tetraacetic acid tetra(acetoxymethyl)ester application in Ca++-free medium to abolish NET regulation by MCh. Although they may contribute to basal NET expression, we could not implicate CaMKII-, PKA- or nitric oxide-linked pathways in MCh regulation. Together, these findings 1) provide evidence in support of G-protein coupled receptor-mediated regulation of catecholamine transport, 2) reveal intracellular Ca++-sensitive, PKC-dependent and -independent pathways that serve to regulate NET expression and 3) indicate that the diminished capacity for NE transport evident after mAChR and PKC activation involves a redistribution of NET protein.
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PMID:Acute regulation of norepinephrine transport: I. protein kinase C-linked muscarinic receptors influence transport capacity and transporter density in SK-N-SH cells. 980 4

A smooth membrane system consisting of subsurface cisternae (SSC) underlies the lateral plasmalemma of auditory outer hair cells (OHCs). The SSC contain Ca-ATPase and are regarded as an intracellular Ca2+ reservoir like the sarcoplasmic reticulum of myocytes. Recently, it has been demonstrated that Ca-ATPase activity in sarcoplasmic reticulum is regulated by Ca2+/calmodulin-dependent protein kinases (CaM kinases). Here we investigated the presence of CaM kinases in OHCs and their possible association with the SSC. Inner ears collected from adult gerbils and from neonates at 2-day intervals between 0 and 20 days after birth were immunostained with antibodies specific for different CaM kinases. A polyclonal antiserum against CaM kinase IV yielded a strong immunostaining reaction along the lateral wall of OHCs. The staining appeared after the tenth postnatal day and continued into adulthood. No other site in the inner ear, including cochlear inner hair cells and vestibular hair cells, was reactive. The kinase's apparent association with the SSC strongly supports its involvement in intracellular Ca2+ homeostasis and suggests a role in regulating the OHCs' slow motile responses.
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PMID:Immunohistochemical localization of Ca2+/Calmodulin-dependent protein kinase IV in outer hair cells. 985 8

Calmodulin (CaM) and Ca(2+)/CaM-dependent protein kinase II (CaM kinase) are tightly associated with cardiac sarcoplasmic reticulum (SR) and are implicated in the regulation of transmembrane Ca(2+) cycling. In order to assess the importance of membrane-associated CaM in modulating the Ca(2+) pump (Ca(2+)-ATPase) function of SR, the present study investigated the effects of a synthetic, high affinity CaM-binding peptide (CaM BP; amino acid sequence, LKWKKLLKLLKKLLKLG) on the ATP-energized Ca(2+) uptake, Ca(2+)-stimulated ATP hydrolysis, and CaM kinase-mediated protein phosphorylation in rabbit cardiac SR vesicles. The results revealed a strong concentration-dependent inhibitory action of CaM BP on Ca(2+) uptake and Ca(2+)-ATPase activities of SR (50% inhibition at approximately 2-3 microM CaM BP). The inhibition, which followed the association of CaM BP with its SR target(s), was of rapid onset (manifested within 30 s) and was accompanied by a decrease in V(max) of Ca(2+) uptake, unaltered K(0.5) for Ca(2+) activation of Ca(2+) transport, and a 10-fold decrease in the apparent affinity of the Ca(2+)-ATPase for its substrate, ATP. Thus, the mechanism of inhibition involved alterations at the catalytic site but not the Ca(2+)-binding sites of the Ca(2+)-ATPase. Endogenous CaM kinase-mediated phosphorylation of Ca(2+)-ATPase, phospholamban, and ryanodine receptor-Ca(2+) release channel was also strongly inhibited by CaM BP. The inhibitory action of CaM BP on SR Ca(2+) pump function and protein phosphorylation was fully reversed by exogenous CaM (1-3 microM). A peptide inhibitor of CaM kinase markedly attenuated the ability of CaM to reverse CaM BP-mediated inhibition of Ca(2+) transport. These findings suggest a critical role for membrane-bound CaM in controlling the velocity of Ca(2+) pumping in native cardiac SR. Consistent with its ability to inhibit SR Ca(2+) pump function, CaM BP (1-2.5 microM) caused marked depression of contractility and diastolic dysfunction in isolated perfused, spontaneously beating rabbit heart preparations. Full or partial recovery of contractile function occurred gradually following withdrawal of CaM BP from the perfusate, presumably due to slow dissociation of CaM BP from its target sites promoted by endogenous cytosolic CaM.
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PMID:Reversible inhibition of the calcium-pumping ATPase in native cardiac sarcoplasmic reticulum by a calmodulin-binding peptide. Evidence for calmodulin-dependent regulation of the V(max) of calcium transport. 1066 Jun 12

The effects of ischemic preconditioning (IP) on changes in cardiac performance and sarcoplasmic reticulum (SR) function due to Ca(2+) paradox were investigated. Isolated perfused hearts were subjected to IP (three cycles of 3-min ischemia and 3-min reperfusion) followed by Ca(2+)-free perfusion and reperfusion (Ca(2+) paradox). Perfusion of hearts with Ca(2+)-free medium for 5 min followed by reperfusion with Ca(2+)-containing medium for 30 min resulted in a dramatic decrease in the left ventricular (LV) developed pressure and a marked increase in LV end-diastolic pressure. Alterations in cardiac contractile activity due to Ca(2+) paradox were associated with depressed SR Ca(2+)-uptake, Ca(2+)-pump ATPase, and Ca(2+)-release activities as well as decreased SR protein contents for Ca(2+)-pump and Ca(2+) channels. All these changes due to Ca(2+) paradox were significantly prevented in hearts subjected to IP. The protective effects of IP on Ca(2+) paradox changes in cardiac contractile activity as well as SR Ca(2+)-pump and Ca(2+)-release activities were lost when the hearts were treated with 8-(p-sulfophenyl)-theophylline, an adenosine receptor antagonist; KN-93, a specific Ca(2+)/calmodulin-dependent protein kinase II (CaMK II) inhibitor; or chelerythrine chloride, a protein kinase C (PKC) inhibitor. These results indicate that IP rendered cardioprotection by preventing a depression in SR function in Ca(2+) paradox hearts. Furthermore, these beneficial effects of IP may partly be mediated by adenosine receptors, PKC, and CaMK II.
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PMID:Mechanisms of ischemic preconditioning effects on Ca(2+) paradox-induced changes in heart. 1071 Mar 71

Although Ca(2+)/calmodulin-dependent protein kinase II (CaMK II) is known to modulate the function of cardiac sarcoplasmic reticulum (SR) under physiological conditions, the status of SR CaMK II in ischemic preconditioning (IP) of the heart is not known. IP was induced by subjecting the isolated perfused rat hearts to three cycles of brief ischemia-reperfusion (I/R; 5 min ischemia and 5 min reperfusion), whereas the control hearts were perfused for 30 min with oxygenated medium. Sustained I/R in control and IP groups was induced by 30 min of global ischemia followed by 30 min of reperfusion. The left ventricular developed pressure, rate of the left ventricular pressure, as well as SR Ca(2+)-uptake activity and SR Ca(2+)-pump ATPase activity were depressed in the control I/R hearts; these changes were prevented upon subjecting the hearts to IP. The beneficial effects of IP on the I/R-induced changes in contractile activity and SR Ca(2+) pump were lost upon treating the hearts with KN-93, a specific CaMK II inhibitor. IP also prevented the I/R-induced depression in Ca(2+)/calmodulin-dependent SR Ca(2+)-uptake activity and the I/R-induced decrease in the SR CaMK II activity; these effects of IP were blocked by KN-93. The results indicate that IP may prevent the I/R-induced alterations in SR Ca(2+) handling abilities by preserving the SR CaMK II activity, and it is suggested that CaMK II may play a role in mediating the beneficial effects of IP on heart function.
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PMID:Ischemic preconditioning prevents I/R-induced alterations in SR calcium-calmodulin protein kinase II. 1084 74

To decipher the mechanism(s) underlying glucocorticoid action on cardiac contractile function, this study investigated the effects of adrenalectomy and dexamethasone treatment on the contents of sarcoplasmic reticulum (SR) Ca(2+)-cycling proteins, their phosphorylation by endogenous Ca(2+)/calmodulin-dependent protein kinase II (CaM kinase II), and SR Ca(2+) sequestration in the rat myocardium. Cardiac SR vesicles from adrenalectomized rats displayed significantly diminished rates of ATP-energized Ca(2+) uptake in vitro compared with cardiac SR vesicles from control rats; in vivo administration of dexamethasone to adrenalectomized rats prevented the decline in SR function. Western immunoblotting analysis showed that the relative protein amounts of ryanodine receptor/Ca(2+)-release channel, Ca(2+)-ATPase, calsequestrin, and phospholamban were neither diminished significantly by adrenalectomy nor elevated by dexamethasone treatment. However, the relative amount of SR-associated CaM kinase II protein was increased 2.5- to 4-fold in dexamethasone-treated rats compared with control and adrenalectomized rats. Endogenous CaM kinase II activity, as judged from phosphorylation of ryanodine receptor, Ca(2+)-ATPase, and phospholamban protein, was also significantly higher (50--80% increase) in the dexamethasone-treated rats. The stimulatory effect of CaM kinase II activation on Ca(2+) uptake activity of SR was significantly depressed after adrenalectomy and greatly enhanced after dexamethasone treatment. These findings identify the SR as a major target for glucocorticoid actions in the heart and implicate modification of the SR CaM kinase II system as a component of the mechanisms by which dexamethasone influences SR Ca(2+)-cycling and myocardial contraction.
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PMID:Glucocorticoid modulation of protein phosphorylation and sarcoplasmic reticulum function in rat myocardium. 1140

The Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) is thought to be activated during the cholinergic stimulation of gastric acid secretion. The carbachol-induced acid production of cultured rabbit parietal cells was dose-dependently inhibited by the CaMKII inhibitor KN-62 as measured by accumulation of the weak base [(14)C]aminopyrine ([(14)C]-AP). Inhibition by KN-62 was most efficient at concentrations of carbachol >10(-6) M. After carbachol stimulation, we observed an activation of CaMKII activity, and its translocation to the apical membrane of gastric mucosal cells. We found a doubling of the abundance of CaMKII to the stimulus-associated apical membrane (SA vesicles) compared to the apical membrane from the resting state after carbachol induction. This was shown by both an anti-CaMKII serum and the 1.8-fold increase of the CaMKII phosphotransferase activity in vitro. The SA vesicles exhibited a strong increase of autoactivated CaMKII probed with an anti-autoactivated CaMKII antibody. Additionally, we observed a colocalization of both CaMKII and the H(+)-K(+)-ATPase of SA vesicles similar to the colocalization of both enzymes to the tubulovesicles suggesting them as at least one pool for the SA vesicular CaMKII. Our data indicate that the activation of CaMKII and the carbachol-dependent redistribution of CaMKII to the SA vesicles are distinct processes that occur in parallel to regulate the activity and localization of CaMKII. These findings contribute to the model implicating an involvement for CaMKII in the intracellular dynamics of the acid secretion.
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PMID:CaMKII is activated and translocated to the secretory apical membrane during cholinergically conveyed gastric acid secretion. 1178 Nov 41

Phospholamban (PLB) plays a primary role in regulating cardiac sarcoplasmic reticulum (SR) Ca(2+)-ATPase activity. Dephosphorylated PLB suppresses the SR Ca(2+) pump activity, whereas phosphorylation of PLB leads to deinhibition. A widely accepted sequential model of dual site PLB phosphorylation states that PKA-dependent phosphorylation of Ser(16) is obligatory to phosphorylation of Thr(17) by Ca(2+)/calmodulin-dependent kinase II, and mainly accounts for beta-adrenergic receptor mediated cardiac relaxation. However, emerging evidence supports independent phosphorylation of Ser(16) and Thr(17) and their independent contributions to cardiac relaxation. Furthermore, concurrent activation of PKA and CaMKII signaling pathways exhibits a robust synergistic effect on phosphorylation of Thr(17), but not of Ser(16). Thus, the synergistic interaction may masquerade as a sequential phosphorylation of Ser(16) and Thr(17) under certain circumstances. Further studies are required to determine the exact process of dual site PLB phosphorylation and its functional roles in healthy and diseased hearts.
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PMID:Dual site phospholamban phosphorylation and its physiological relevance in the heart. 1185 50

Cardiac ryanodine receptors (RyR2s) play a critical role in excitation-contraction coupling by providing a pathway for the release of Ca(2+) from the sarcoplasmic reticulum into the cytosol. RyR2s exist as macromolecular complexes that are regulated via binding of Ca(2+) and protein phosphorylation/dephosphorylation. The present study examined the association of endogenous CaMKII (calcium/calmodulin-dependent protein kinase II) with the RyR2 complex and whether this enzyme could modulate RyR2 function in isolated rabbit ventricular myocardium. Endogenous phosphorylation of RyR2 was verified using phosphorylation site-specific antibodies. Co-immunoprecipitation studies established that RyR2 was physically associated with CaMKIIdelta. Quantitative assessment of RyR2 protein was performed by [(3)H]ryanodine binding to RyR2 immunoprecipitates. Parallel kinase assays allowed the endogenous CaMKII activity associated with these immunoprecipitates to be expressed relative to the amount of RyR2. The activity of RyR2 in isolated cardiac myocytes was measured in two ways: (i) RyR2-mediated Ca(2+) release (Ca(2+) sparks) using confocal microscopy and (ii) Ca(2+)-sensitive [(3)H]ryanodine binding. These studies were performed in the presence and absence of AIP (autocamtide-2-related inhibitory peptide), a highly specific inhibitor of CaMKII. At 1 microM AIP Ca(2+) spark duration, frequency and width were decreased significantly. Similarly, 1 microM AIP decreased [(3)H]ryanodine binding. At 5 microM AIP, a more profound inhibition of Ca(2+) sparks and a decrease in [(3)H]ryanodine binding was observed. Separate measurements showed that AIP (1-5 microM) did not affect sarcoplasmic reticulum Ca(2+)-ATPase-mediated Ca(2+) uptake. These results suggest the existence of an endogenous CaMKIIdelta that associates directly with RyR2 and specifically modulates RyR2 activity.
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PMID:Calcium/calmodulin-dependent protein kinase IIdelta associates with the ryanodine receptor complex and regulates channel function in rabbit heart. 1455 49


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