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)

Our understanding of calcium homeostasis during the crustacean moulting cycle derives from research on intermoult animals that has been extrapolated to other stages. In terms of transepithelial Ca(2+) flux, the more interesting stages are those surrounding ecdysis since crustaceans experience a sizeable negative calcium balance in immediate premoult and a significant positive calcium balance in immediate postmoult. These stages are elusive in the sense that larger species such as lobsters are rarely captured at this time, and smaller species such as blue crabs and crayfish are seldom synchronized in their moulting cycle. The reductionist approaches employed in cellular physiology, such as vesicle techniques, employ pooling of fresh tissues from many organisms. Examination of the elusive moulting stages requires more sensitive approaches that can utilize tissue from an individual crustacean to characterize Ca(2+) pumps (Sarco/Endoplasmic Reticulum Ca(2+)-ATPase, SERCA; Plasma Membrane Ca(2+)-ATPase, PMCA) and the Na(+)/Ca(2+) eXchanger (NCX). An emerging subcellular approach described in this paper is to use flow cytometry as a technique to monitor Ca(2+) uptake into Fluo-3-loaded membrane vesicles. This paper illustrates the utility of this technique for measuring ATP-dependent Ca(2+) uptake into hepatopancreatic basolateral membrane vesicles. Obstacles to progress in molecular studies have not been limited by synchronization of moulting since tissue can be snap-frozen and collected from many animals over time. Here, the problem has been the lack of specific antibodies that hybridize with the Ca(2+) transporters of interest so that they can be localized within epithelia. In this paper, we introduce polyclonal antibodies raised in rabbits against crayfish SERCA, PMCA and NCX. Immunocytochemistry of SERCA in muscle, PMCA in antennal gland and NCX in heart confirms the specificity of the antibodies.
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PMID:Novel subcellular and molecular tools to study Ca(2+) transport mechanisms during the elusive moulting stages of crustaceans: flow cytometry and polyclonal antibodies. 1117 19

The fluorescent calcium probe, Fluo-3, AM was used to measure the intracellular calcium concentration in red blood cells (RBCs) of the teiid lizards Ameiva ameiva and Tupinambis merianae. The cytosolic [Ca2+] is maintained around 20 nM and the cells contain membrane-bound Ca2+ pools. One pool appears to be identifiable with the endoplasmic reticulum (ER) inasmuch as addition of the sarco-endoplasmic reticulum Ca2+ ATPase, SERCA, inhibitor thapsigargin induces an increase in cytosolic [Ca2+ both in the presence and in the absence of extracellular Ca2+. In addition to the ER, an acidic compartment appears to be involved in Ca2+ storage, as collapse of intracellular pHgradients by monensin, a Na+ -H+ exchanger, and nigericin, a K+ -H+ exchanger, induce the release of Ca2+ from internal pools. A vacuolar H+ pump, sensitive to NBD-Cl and bafilomycin appears to be necessary to load the acidic Ca2+ pools. Finally, the purinergic agonist ATP triggers a rapid and transient increase of [Ca2+]c in the cells from both lizard species, mostly by mobilization of the cation from internal stores.
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PMID:Signal transduction in red blood cells of the lizards Ameiva ameiva and Tupinambis merianae (Squamata, Teiidae). 1135 9

Altered cytosolic Ca2+ is implicated in the aetiology of many diseases including diabetes but there are few studies on the mechanism(s) of the altered Ca2+ regulation. Using human lymphocytes, we studied cytosolic calcium (Cai) and various Ca2+ transport mechanisms in subjects with Type 2 diabetes mellitus and control subjects. Ca2+-specific fluorescent probes (Fura-2 and Fluo-3) were used to monitor the Ca2+ signals. Thapsigargin, a potent and specific inhibitor of the sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA), was used to study Ca2+-store dependent Ca2+ fluxes. Significant (P<0.05) elevation of basal Ca, levels was observed in lymphocytes from diabetic subjects. Cai levels were positively correlated with fasting plasma glucose and HbA1c. There was also a significant (P<0.05) reduction in plasma membrane calcium (PMCA) ATPase activity in diabetic subjects compared to controls. Cells from Type 2 diabetics exhibited an increased Ca2+ influx (as measured both by Fluo-3 fluorescence and 45Ca assays) as a consequence of thapsigargin-mediated Ca2+ store depletion. Upon addition of Mn2+ (a surrogate of Ca2+), the fura-2 fluorescence decayed in an exponential fashion and the rate and extent of this decline was steeper and greater in cells from type 2 diabetic patients. There was also a significant (P<0.05) difference in the Na+/Ca2+ exchange activity in Type 2 diabetic patients, both under resting conditions and after challenging the cells with thapsigargin, when the internal store Ca2+ sequestration was circumvented. Pharmacological activation of protein kinase C (PKC) in cells from patients resulted in only partial inhibition of Ca2+ entry. We conclude that cellular Ca2+ accumulation in cells from Type 2 diabetes results from (a) reduction in PMCA ATPase activity, (b) modulation of Na+/Ca2+ exchange and (3) increased Ca2+ influx across the plasma membrane.
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PMID:Evidence for mechanistic alterations of Ca2+ homeostasis in Type 2 diabetes mellitus. 1146 18

Outer nuclear membrane is endowed with a SERCA type Ca(2+)-ATPase which pumps calcium into the nuclear envelope lumen and creates calcium stores. Variation in this calcium pool, among other things, regulates nuclear transport. The transport of Nuclear Localization Signal (NLS)-containing molecules into the nucleus is well established. Intermediate size molecules lacking an NLS translocate to the nucleus and its mechanism remains obscure. It is observed here that the treatment of HEK 293 cells in culture with dibutyryl cyclic AMP (db-cAMP) or forskolin (FK) triggered transport of Calcium Green 10 kDa dextran into the nucleus. Under similar conditions Fluo-3-AM accumulated around the nuclei. cAMP-dependent protein kinase phosphorylated 105 kDa nuclear Ca(2+)-ATPase (NCA) which served as a trigger for NLS-independent transport into the nucleus.
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PMID:In vivo nuclear Ca2+-ATPase phosphorylation triggers intermediate size molecular transport to the nucleus. 1268 66

In this study, we investigated whether nucleoplasmic free Ca2+ in aortic vascular smooth muscle cells (VSMCs) might be independently regulated from cytosolic free Ca2+. Understanding mechanisms and pathways responsible for this regulation is especially relevant given the role of a numerous intranuclear Ca2+-sensitive proteins in transcriptional regulation, apoptosis and cell division. The question of an independent regulatory mechanism remains largely unsettled because the previous use of intensitometric fluorophores (e.g., Fluo-3) has been criticized on technical grounds. To circumvent the potential problem of fluorescence artifact, we utilized confocal laser scanning microscopy to image intracellular Ca2+ movements with the ratiometric fluorophore Indo-1. In cultured rabbit VSMCs, we found sarcoplasmic reticulum (SR) Ca2+ ATPase (SERCA) pumps and ryanodine receptor (RyR) Ca2+ channel proteins to be discretely arranged within a perinuclear locus, as determined by fluorescent staining patterns of BODIPY FL thapsigargin and BODIPY FL-X Ry. When intracellular Ca2+ stores were mobilized by addition of thapsigargin (5 microM) and activatory concentrations of ryanodine (1 microM), Indo-1 ratiometric signals were largely restricted to the nucleoplasm. Cytosolic signals, by comparison, were relatively small and even then its spatial distribution was largely perinuclear rather homogeneous. These observations indicate perinuclear RyR and SERCA proteins are intimately involved in regulating VSMC nucleoplasmic Ca2+ concentrations. We also observed a similar pattern of largely nucleoplasmic Ca2+ mobilization upon exposure of cells to the immunosuppressant drug FK506 (tacrolimus), which binds to the RyR-associated immunophillin-binding proteins FKBP12 and FKBP12.6. However, initial FK506-induced nucleoplasmic Ca2+ mobilization was followed by marked reduction of Indo-1 signal intensity close to pretreatment levels. This suggested FK506 exerts both activatory and inhibitory effects upon RyR channels. The latter was reinforced by observed effects of FK506 to only reduce nucleoplasmic Indo-1 signal intensity when added following pretreatment with both activatory and inhibitory concentrations of ryanodine. These latter observations raise the possibility that VSMC nuclei represent an important sink of intracellular Ca2+ and may help explain vasodilatory actions of FK506 observed by others.
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PMID:Nucleoplasmic calcium regulation in rabbit aortic vascular smooth muscle cells. 1273 28

Altered intracellular Ca(2+) homeostasis and accumulated Ca(2+) deposition in arterial walls contribute to the natural arterial aging and aging-related vascular pathologies. To gain further insight into internal relationship between these two factors, a vitamin D(3)-induced vascular Ca(2+) overload rat model was employed. Mesenteric vascular smooth muscle cells (VSMCs) were isolated from both vitamin D(3) and Wistar control rats and were maintained in primary culture for 24 h. Cytosolic and nuclear Ca(2+) ([Ca(2+)](i), [Ca(2+)](n)) in VSMCs were compared between vitamin D(3) and Wistar groups using laser scanning confocal microscopy and Ca(2+)-sensitive-dye Fluo-3. Cytosolic and nuclear Ca(2+) were evaluated under both resting and agonist-stimulated conditions including the voltage-dependent Ca(2+) channel openers BayK8644 and KCl, the inositol-1,4,5-trisphosphate (IP(3))-sensitive Ca(2+) release channel activator IP(3), the ryanodine-sensitive Ca(2+) release channel activator trichloromethane, the sarcoplasmic reticulum Ca(2+) -ATPase inhibitor cyclopiazonic acid, and angiotensin II. Although the levels of [Ca(2+)](n) and [Ca(2+)](i) were comparable between vitamin D(3) and Wistar groups under the resting condition, the increase of [Ca(2+)](n) and [Ca(2+)](i) elicited by various agonists was significantly enhanced in VSMCs from the vitamin D(3) group compared with those from the Wistar group, suggesting abnormality of membrane Ca(2+) gating and intracellular Ca(2+) release under Ca(2+) overload condition. In conclusion, our study indicated that vitamin D(3)-induced vascular Ca(2+) overload may directly interrupt cytosolic and nuclear Ca(2+) homeostasis.
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PMID:Agonist-stimulated Ca(2+) transport in mesenteric vascular smooth muscle cells of vitamin D(3)-induced calcium overload rats. 1474 25

By using the fluorescent dye Rhod-2, we have investigated the ability of Plasmodium mitochondria to participate in cellular Ca2+ homeostasis. To this end, isolated parasites were simultaneously loaded with the mitochondrial Ca2+ probe Rhod-2 and the cytosolic Ca2+ dye Fluo-3 and their fluorescent intensities were monitored in the same cells by confocal microscopy. We here demonstrate that Ca2+ increases, as elicited by treatment of parasites with sarco-endoplasmic reticulum Ca2+ ATPase inhibitors or the hormone melatonin, induce rapid and reversible increases of the Ca2+ concentration in the mitochondria of both human and murine parasites. Pre-treatment of parasites with the mitochondrial uncoupler, FCCP, suppresses mitochondrial Ca2+ accumulation. Our data demonstrate that mitochondria of malaria parasites are able to reversibly accumulate part of the Ca2+ released in the cytoplasm by pharmacological and physiological agents and thus suggest that this organelle participate in the maintenance of Ca2+ homeostasis of Plasmodia.
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PMID:The malaria parasite mitochondrion senses cytosolic Ca2+ fluctuations. 1535 26

We investigated the aluminum (Al)-induced alterations in zeta potential, plasma membrane (PM) potential and intracellular calcium levels to elucidate their interaction with callose production induced by Al toxicity. A noninvasive confocal laser microscopy has been used to analyse the live tobacco (Nicotiana tabacum) cell events by means of fluorescent probes Fluo-3 acetoxymethyl ester (intracellular calcium) and DiBAC4 (PM potential) as well as to monitor callose accumulation. Log-phase cells showed no detectable changes in the PM potential during the first 30 min of Al treatment, but sustained large depolarization from 60 min onwards. Measurement of zeta potential confirmed the depolarization effect of Al, but the kinetics were different. The Al-treated cells showed a moderate increase in intracellular Ca2+ levels and callose production in 1 h, which coincided with the time course of PM depolarization. Compared with the Al treatment, cyclopiazonic acid, an inhibitor of endoplasmic reticulum Ca(2+)-ATPase, facilitated a higher increase in intracellular Ca2+ levels, but resulted in accumulation of only moderate levels of callose. Calcium channel modulators and Al induced similar levels of callose in the initial 1 h of treatment. Callose production induced by Al toxicity is dependent on both depolarization of the PM and an increase in intracellular Ca2+ levels.
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PMID:Early events responsible for aluminum toxicity symptoms in suspension-cultured tobacco cells. 1572 Jun 25

We have previously demonstrated that rat cerebellar Type-1 astrocytes express a very active genistein sensitive Na(+)/Ca(2+) exchanger, which accounts for most of the total plasma membrane Ca(2+) fluxes and for the clearance of loads induced by physiological agonists. In this work, we have explored the mechanism by which the reverse Na(+)/Ca(2+) exchange is involved in agonist-induced Ca(2+) signaling in rat cerebellar astrocytes. Microspectrofluorometric measurements of Cai(2+) with Fluo-3 demonstrate that the Cai(2+) signals associated long (> 20 s) periods of reverse operation of the Na(+)/Ca(2+) exchange are amplified by a mechanism compatible with calcium-calcium release, while those associated with short (< 20 s) pulses are not amplified. This was confirmed by pharmacological experiments using ryanodine receptors agonist (4-chloro-m-cresol) and the endoplasmic reticulum ATPase inhibitor (thapsigargin). Confocal microscopy demonstrates a high co-localization of immunofluorescent labeled Na(+)/Ca(2+) exchanger and RyRs. Low (< 50 micromol/L) or high (> 500 micromol/L) concentrations of L-glutamate (L-Glu) or L-aspartate causes a rise in which is completely blocked by the Na(+)/Ca(2+) exchange inhibitors KB-R7943 and SEA0400. The most important novel finding presented in this work is that L-Glu activates the reverse mode of the Na(+)/Ca(2+) exchange by inducing Na(+) entry through the electrogenic Na(+)-Glu-co-transporter and not through the ionophoric L-Glu receptors, as confirmed by pharmacological experiments with specific blockers of the ionophoric L-Glu receptors and the electrogenic Glu transporter.
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PMID:Na+ entry via glutamate transporter activates the reverse Na+/Ca2+ exchange and triggers Ca(i)2+-induced Ca2+ release in rat cerebellar Type-1 astrocytes. 1731 98

Vanadium, a trace element, as vanadate (VO4(3-)) is known to interfere with a wide variety of enzymes including Ca2+ ATPase and Na+/+ ATPase. VO4(3-) is excreted mainly via the kidney. In renal insufficiency, the impaired VO4(3-) excretion leads to VO4(3-) accumulation in blood.The present study explored the effect of VO4(3-) on eryptosis, the suicidal death of erythrocytes. Eryptosis is characterized by cell shrinkage and phosphatidylserine exposure at the erythrocyte surface. Eryptotic cells are phagocytosed and thus rapidly cleared from circulating blood. Stimulators of eryptosis include an increase of the cytosolic Ca2+ concentration. Erythrocyte Ca2+ activity was estimated from Fluo-3 fluorescence, phosphatidylserine exposure from annexin V-binding, and erythrocyte volume from forward scatter in FACS analysis. Exposure of erythrocytes to VO4(3-) increased cytosolic Ca2+ concentration, enhanced the percentage of annexin V-binding erythrocytes, decreased erythrocyte forward scatter, and lowered the intracellular ATP concentration. In conclusion, VO4(3-) induces eryptosis at least partially through increase of cytosolic Ca2+ concentration, an effect presumably contributing to the development of anemia in chronic renal failure.
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PMID:Vanadate-induced suicidal erythrocyte death. 1831 5


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