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1.
In an attempt to elucidate the relationship between phosphatidylinositol breakdown and alpha-adrenergic responses, effects of phosphatidic acid and phosphatidylinositol related metabolites on Ca2+ mobilization and glucose output in cultured hepatocytes were examined. Norepinephrine induced the net 45Ca2+ efflux from preloaded cells and stimulated glucose output via alpha-adrenergic receptor stimulation, whereas phosphatidic acid caused 45Ca2+ uptake to cells and did not stimulate glucose output. Myo-inositol-monophosphate, diglyceride and arachidonic acid, which are released by phosphatidylinositol breakdown, had no effect on 45Ca2+ efflux and glucose output in cells. These results suggest that phosphatidic acid and phosphatidylinositol related metabolites can not mimic the alpha-adrenergic actions in cultured hepatocytes. 相似文献
2.
S. S. Medvedev O. V. Tankelyun A. Yu. Batov O. V. Voronina J. Martinec I. Macháčková 《Russian Journal of Plant Physiology》2006,53(1):39-47
Effects of phosphatidic acid (PA), a product of phospholipase D activity, on Ca2+ and H+ transport were investigated in membrane vesicles obtained from roots and coleoptiles of maize (Zea mays L.). Calcium flows were measured with fluorescent probes indo-1 and chlorotetracycline loaded into the vesicles and added to the incubation medium, respectively. Phosphatidic acid (50–500 μM) was found to induce downhill flow of Ca2+ along the concentration gradient into the plasma membrane vesicles and endomembrane vesicles (tonoplast and endoplasmic reticulum). Protonophorous functions of PA were probed with acridine orange. First, the ionic H+ gradient was created on the tonoplast vesicles by means of H+-ATPase activation with Mg-ATP addition. Then, the vesicles were treated with 25–100 μM PA, which induced the release of protons from tonoplast vesicles and dissipation of the proton gradient. Thus, PA could function as an ionophore and was able to transfer Ca2+ and H+ across plant cell membranes along concentration gradients of these ions. The role of PA in mechanisms of intracellular signaling in plants is discussed. 相似文献
3.
Ca2+-induced transformation of phosphatidylcholine-phosphatidic acid vesicles to larger bilayer structures has been examined using nuclear magnetic resonance, electron microscopy, gel permeation and radioisotope tracer techniques. For concentrated vesicle preparations where phosphatidic acid content remains less than 50% of total lipid, transformation to larger well defined unilamellar structures can be induced. The size of the product formed is dependent on phosphatidic acid content and on Ca2+ content when Ca2+ levels are between 0.3 and 1.0 mol ratios with respect to phosphatidic acid. During transformation bilayer composition remains unchanged and internal contents are retained in the final structure. These properties are indicative of concerted two vesicle and multiple vesicle fusions. The controllable and concerted fusions make the phosphatidic acid system suitable for further mechanistic studies. 相似文献
4.
PA1b,a plant peptide,induces intracellular [Ca<Superscript>2+</Superscript>] increase via Ca<Superscript>2+</Superscript> influx through the L-type Ca<Superscript>2+</Superscript> channel and triggers secretion in pancreatic β cells
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Using alginic acid to adsorb polypeptides at pH 2.7, we isolated a peptide pea albumin 1b (PA1b) from pea seeds. The PA1b
is a single chain peptide consisting of 37 amino acid residues with 6 cysteines which constitutes the cystine-knot structure.
Using microfluorometry and patch clamp techniques, we found that PA1b significantly elevated the intracellular calcium level
([Ca2+]i) and elicited membrane capacitance increase in the primary rat pancreatic β cells. The PA1b effect on [Ca2+]i elevation was abolished in the absence of extracellular Ca2+ or in the presence of L-type Ca2+ channel blocker, nimodipine. Interestingly, we found that PA1b significantly depolarized membrane potential, which could
lead to the opening of voltage-dependent L-type Ca2+ channels and influx of extracellular Ca2+, and then evoke robust secretion. In this study we identified the plant peptide PA1b which is capable of affecting the excitability
and function of mammalian pancreatic β cell. 相似文献
5.
Huiyan Huang Michael A. Frohman 《Biochimica et Biophysica Acta (BBA)/Molecular and Cell Biology of Lipids》2009,1791(9):839-844
Regulated production and elimination of the signaling lipids phosphatidic acid (PA), diacylglycerol (DAG), and phosphatidylinositol 4,5-bisphosphate (PI4,5P2) creates a complex and interconnected signaling network that modulates a wide variety of eukaryotic cell biological events. PA production at the plasma membrane and on trafficking membrane organelles by classical Phospholipase D (PLD) through the hydrolysis of phosphatidylcholine (PC) has been studied widely. In this chapter, we review a newly identified, non-canonical member of the PLD superfamily, MitoPLD, which localizes to the mitochondrial surface and plays a role in mitochondrial fusion via the hydrolysis of cardiolipin (CL) to generate PA. The role of PA in facilitating the mitochondrial fusion event carried out by proteins known as Mitofusins is intriguing in light of the role classic PLD-generated PA plays in facilitating SNARE-mediated fusion of secretory membrane vesicles into the plasma membrane. In addition, however, PA on the mitochondrial surface may also trigger a signaling cascade that elevates DAG, leading to downstream events that affect mitochondrial fission and energy production. PA production on the mitochondrial surface may also stimulate local production of PI4,5P2 to facilitate mitochondrial fission and subcellular trafficking or facilitate Ca2+ influx. 相似文献
6.
Ca2+ was required for carbachol-induced decreases in phosphatidylinositol (PI) and increases in phosphatidic acid (PA) concentrations during incubation of rat submaxillary gland fragments, but was not required for increases in [32P]Pi incorporation into these phospholipids. Like carbachol, A23187 provoked a Ca2+-dependent decrease in PI mass. These results suggest concomitant operation of two separate mechanisms for stimulating PI hydrolysis and 32P labeling of PA and PI during carbachol action: one mechanism is not dependent on external Ca2+ and is manifested by rapid labeling in a relatively small PA-PI pool; the other mechanism is dependent on Ca2+ and involves a large PA-PI pool which appears to have a relatively slow renewal (labeling) rate. 相似文献
7.
Rafat A. Siddiqui Denis English 《Biochimica et Biophysica Acta (BBA)/Molecular and Cell Biology of Lipids》2000,1483(1):161-173
Phosphatidylinositol 3′-kinase (PI 3′-kinase) plays an important role in the migration of hepatocytes, endothelial cells and neoplastic cells to agonists which activate cellular tyrosine kinases. We examined the PI 3′-kinase-dependent chemotactic responses of neutrophilic leukocytes induced by phosphatidic acid (PA) in order to clarify mechanisms by which the enzyme potentially influences cellular migration. Western analysis of immunoprecipitates indicated that PA induced the tyrosine phosphorylation of three distinct proteins involved in functional activation which co-immunoprecipitated in PA-stimulated cells. These proteins were identified as lyn, syk and the 85 kDa regulatory subunit of PI 3′-kinase. Chemotactic responses to PA but not to several other neutrophil agonists were inhibited by the PI 3′-kinase inhibitors wortmannin and LY294002. Chemotactic inhibition resulted from upstream inhibition of calcium mobilization. Chelation of extracellular calcium by ethylene glycol-bis(β-aminoethyl ether) N,N,N′,N′-tetraacetic acid (EGTA) did not affect the PA-induced chemotaxis, whereas chelation of intracellular calcium by 1,2-bis(2-aminophenoxy)-ethane-N,N,N′,N′-tetraacetic acid (BAPTA) attenuated this response. Thus, changes in intracellular Ca2+ levels that can be effected by Ca2+ mobilized from intracellular stores in the absence of Ca2+ influx regulate PA-induced chemotaxis. Furthermore, PI 3′-kinase inhibition blunted the agonist-dependent generation of inositol 1,4,5-trisphosphate (IP3), suggesting that PI 3′-kinase exerted its effects on calcium mobilization from intracellular sources by mediating activation of phospholipase C (PLC) in PA-stimulated cells. Moreover, the PI 3′-kinase inhibitor LY294002 also inhibited phosphorylation of syk in PA-stimulated cells. We, therefore, propose that products of PI 3′-kinase confined to the inner leaflet of the plasma membrane play a role in activation of syk, calcium mobilization and induction of chemotactic migration. 相似文献
8.
Stimulation of washed rabbit platelets with AGEPC (1-O-alkyl-2-acetyl--glyceryl-3-phosphorylcholine) caused a 15–20% decrease in their phosphatidylinositol level within 15 seconds without affecting other major classes of phospholipids. In the same time frame the level of phosphatidic acid (PA) increased dramatically some four fold. LysoGEPC, which is inactive in stimulating rabbit platelets, did not cause any change in PI or PA. When [32Pi] was present during the stimulation of platelets by AGEPC, the incorporation of radiolabel into PI-4-phosphate (DPI), PI-4,5-bis phosphate (TPI) and PA was enhanced significantly within one minute while the incorporation into PI increased only after one minute. These results clearly established that AGEPC induced stimulation of rabbit platelets was associated with the metabolism of inositol phospholipids and phosphatidic acid. The relevance of these findings to the mode of action of AGEPC and Ca2+ mobilization is also discussed. 相似文献
9.
Atsushi Imai Hiroaki Hattori Masaru Takahashi Yoshinori Nozawa 《Biochemical and biophysical research communications》1983,112(2):693-700
The regulation of human platelet responses by cyclic AMP (cAMP) has been investigated by measuring thrombin-stimulated serotonin release, Ca2+ uptake and phospholipase activity. Thrombin-induced 1,2-diacylglycerol (DG) formation as a result of phospholipase C activation was inhibited by pretreatment with dibutyryl cAMP (dbcAMP) in a dose-dependent manner. Subsequent failure to produce phosphatidic acid (PA), which is converted from 1,2-DG by phosphorylation and would serve as intracellular Ca2+ ionophore, appeared to parallel the decrease in Ca2+ uptake activity. Phospholipase A2 activity, monitored by the production of [3H]lysophosphatidylcholine and [3H]lysophosphatidylethanolamine, was also suppressed by dbcAMP. These data indicate that the intracellular cAMP level may be closely associated with Ca2+ uptake and phospholipases activation. In addition, it is suggested that alteration of intracellular cAMP regulates phospholipase activation and consequently platelet responses, perhaps by controlling available Ca2+ content. 相似文献
10.
E. A. C. MacRobbie 《Planta》1989,178(2):231-241
The influx of 45Ca into isolated guard cells of Commelina communis L. has been measured, using short uptake times, and washing in ice-cold La3+-containing solutions to remove extracellular tracer after the loading period. Over 0.5–4 min the uptake was linear with time, through the origin. Over 20–200M external Ca2+ the influx measured with 10–20 mM external KCl was in the range 0.3–2.3 pmol·cm-2·s-1 (on the basis of estimated guard-cell area); with only 1 mM KCl externally the 45Ca influx was significantly reduced, in the range 0.3–1.1 pmol·cm-2·s-1 for external Ca2+ of 50–100 M. The results indicate that the Ca-channel is voltage-sensitive, opening with depolarisation. No consistent effect of the addition of abscisic acid could be found. In different experiments, on the addition of 0.1 mM abscisic acid the Ca2+ influx was sometimes stimulated by 28–79%, was sometimes unaffected, and was sometimes inhibited by 16–29%. The results rule out a long-lasting stimulation of 45Ca influx by ABA, but they do not rule out a transient stimulation followed by inhibition, perphaps as a consequence of down-regulation of Ca2+ influx by increasing cytoplasmic Ca2+. The hypothesis that ABA may act via an action on Ca2+ influx, increasing cytoplasmic Ca2+, with consequent effects on voltage-dependent and Ca2+-dependent ion channels in both plasmalemma and tonoplast, is neither proved nor disproved by these results.Abbreviations ABA
abscisic acid
- Cao, Ko
external Ca and K concentrations 相似文献
11.
Calcium- and calmodulin-regulated breakdown of phospholipid by microsomal membranes from bean cotyledons 总被引:11,自引:8,他引:3
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Evidence for the involvement of Ca2+ and calmodulin in the regulation of phospholipid breakdown by microsomal membranes from bean cotyledons has been obtained by following the formation of radiolabeled degradation products from [U-14C]phosphatidylcholine. Three membrane-associated enzymes were found to mediate the breakdown of [U-14C] phosphatidylcholine, viz. phospholipase D (EC 3.1.4.4), phosphatidic acid phosphatase (EC 3.1.3.4), and lipolytic acyl hydrolase. Phospholipase D and phosphatidic acid phosphatase were both stimulated by physiological levels of free Ca2+, whereas lipolytic acyl hydrolase proved to be insensitive to Ca2+. Phospholipase D was unaffected by calmodulin, but the activity of phosphatidic acid phosphatase was additionally stimulated by nanomolar levels of calmodulin in the presence of 15 micromolar free Ca2+. Calmidazolium, a calmodulin antagonist, inhibited phosphatidic acid phosphatase activity at IC50 values ranging from 10 to 15 micromolar. Thus the Ca2+-induced stimulation of phosphatidic acid phosphatase appears to be mediated through calmodulin, whereas the effect of Ca2+ on phospholipase D is independent of calmodulin. The role of Ca2+ as a second messenger in the initiation of membrane lipid degradation is discussed. 相似文献
12.
Alexey Agafonov Konstantin Belosludtsev Odile Gateau-Roesch Galina D. Mironova 《生物化学与生物物理学报:生物膜》2003,1609(2):153-160
Formation of palmitic acid/Ca2+ (PA/Ca2+) complexes was suggested to play a key role in the non-classical permeability transition in mitochondria (NCPT), which seems to be involved in the PA-induced apoptosis of cardiomyocytes. Our previous studies of complexation of free fatty acids (FFA) with Ca2+ showed that long-chain (C:16-C:22) saturated FFA had an affinity to Ca2+, which was much higher than that of other FFA and lipids. The formation of FFA/Ca2+ complexes in the black-lipid membrane (BLM) was demonstrated to induce a nonspecific ion permeability of the membrane. In the present work, we have found that binding of Ca2+ to PA incorporated into the membrane of sulforhodamine B (SRB)-loaded liposomes results in an instant release of a part of SRB, with the quantity of SRB released depending on the concentration of PA and Ca2+. The pH-optimum of this phenomenon, similar to that of PA/Ca2+ complexation, is in the alkaline range. The same picture of SRB release has been revealed for stearic, but not for linoleic acid. Along with Ca2+, some other bivalent cations (Ba2+, Sr2+, Mn2+, Ni2+, Co2+) also induce SRB release upon binding to PA-containing liposomes, while Mg2+ turns out to be relatively ineffective. As revealed by fluorescence correlation spectroscopy, the apparent size of liposomes does not alter after the addition of PA, Ca2+ or their combination. So it has been supposed that the cause of SRB release from liposomes is the formation of lipid pores. The effect of FFA/Ca2+-induced permeabilization of liposomal membranes has several analogies with NCPT, suggesting that both these phenomena are of similar nature. 相似文献
13.
Addition of the mitogenic lectin concanavalin A to rat spleen cells results in a small increase in the steady-state Ca2+ content of the cells. 45Ca2+ fluxes were measured under conditions where artifacts due to Ca2+ binding to concanavalin A could be excluded. Both 45Ca2+ influx into and efflux from these cells are significantly activated by the lectin. If 45Ca2+ is added 30 min after concanavalin A the rate of influx is further enhanced. The increase in 45Ca2+ influx correlates well with binding of concanavalin A to the cells. At low concentrations (optimal mitogenic) of the lectin (1 and 3 μg/ml) no significant increase in 45Ca2+ influx occurs but an increase in 45Ca2+ efflux is still observed. The results suggest that concanavalin A binding to the cell surface causes an increase in Ca2+ influx into the cells and that activation of Ca2+ efflux occurs as a response to an increase in the cytosolic Ca2+ activity. Thus, Ca2+ may well play a role in triggering lymphocyte activation. 相似文献
14.
15.
The characteristics of Ca2+ transport across the excitable membrane of Paramecium aurelia were studied by measuring 45Ca2+ influx and efflux. The intracellular concentration of free Ca2+ in resting P. aurelia was at least ten times less than the extracellular concentration. Ca2+ influx was easily measurable at 0°C, but not at 23°C. The influx of 45Ca2+ was stimulated by the same conditions which cause membrane depolarization and ciliary reversal. Addition of Na+ and K+ (which stimulate ciliary reversal) resulted in a 10-fold increase in the rate of Ca2+ influx. An externally applied, pulsed, electric field (1–2 mA/cm2 of electrode surface), caused the rate of Ca2+ influx to increase 3–5 times, with the extent of stimulation dependent on the current density and the pulse width Ca2+ influx had the characteristics of a passive transport system and was associated with the chemically or electrically triggered Ca2+ “gating” mechanism, which has been studied electrophysiologically. In contrast, Ca2+ efflux appeared to be catalyzed by an active transport system. With cells previously loaded at 0°C with 45Ca2+, Ca2+ efflux was rapid at 23°C, but did not occur at 0°C. This active Ca2+ efflux mechanism is probably responsible for maintaining the low internal Ca2+ levels in unstimulated cells. 相似文献
16.
Viswanathan Natarajan William M. Scribner Suryanarayana Vepa 《Chemistry and physics of lipids》1996,80(1-2)
Activation of phospholipase D (PLD) represents part of an important signalling pathway in mammalian cells, Phospholipase D catalyzed hydrolysis of phospholipids generates phosphatidic acid (PA) which is subsequently metabolized to lyso-PA (LPA) or diacylglycerol (DAG). While DAG is an endogenous activator of protein kinase C (PKC), PA and LPA have been recognized as second messengers as well, Activation of PLD in response to an external stimulus may involve PKC, Ca2+, G-proteins and/or tyrosine kinases. In this review, we will address the role of protein tyrosine phosphorylation in growth factor-, agonist- and oxidant-mediated activation of PLD. Furthermore, a possible link between PKC, Ca2+, G-proteins and tyrosine kinases is discussed to indicate the complexity involved in the regulation of PLD in mammalian cells. 相似文献
17.
The effects of Ca2+ on phosphatidic acid-phosphatidylcholine membranes have been studied using phospholipid spin labels. ESR spectra of spin-labeled phosphatidic acid-phosphatidylcholine membranes and phosphatidic acid-spin-labeled phosphatidylcholine membranes are exchange-broadened immediately upon addition of CaCl2. These changes directly and conclusively indicate Ca2+-induced clustering of spin-labeled phosphatidylcholine and aggregation of spin-labeled phosphatidic acid bridged by Ca2+-chelation in the binary phopholipid membranes. In the Ca2+-chelated aggregates, the motions of the alkyl chains of phosphatidic acid are greatly reduced and the lipid molecules are more closely packed. The clusters and aggregates are formed in patches and the sizes are dependent on the fractions. Ba2+ and Sr2+ induce the lateral phase separations to the same extent as Ca2+. Mg2+ is also effective but to a lesser extent. In acid solutions (pH 5.5), the Ca2+-induced lateral phase separations are of slightly lesser extent than in alkaline solution (pH 7.9). These results are compared with those for phosphatidylserine-phosphatidylcholine membranes reported previously and necessary conditions for the lateral phase separations are discussed. 相似文献
18.
Kai Y. Xu Weizhong Zhu Ling Chen Christopher DeFilippi Jin Zhang Rui-Ping Xiao 《Biochemical and biophysical research communications》2011,(2):200
(Na++K+)-ATPase (NKA) mediates positive inotropy in the heart. Extensive studies have demonstrated that the reverse-mode Na+/Ca2+-exchanger (NCX) plays a critical role in increasing intracellular Ca2+ concentration through the inhibition of NKA-induced positive inotropy by cardiac glycosides. Little is known about the nature of the NCX functional mode in the activation of NKA-induced positive inotropy. Here, we examined the effect of an NKA activator SSA412 antibody on 45Ca influx in isolated rat myocytes and found that KB-R7943, a NCX reverse-mode inhibitor, fails to inhibit the activation of NKA-induced 45Ca influx, suggesting that the Ca2+ influx via the reverse-mode NCX does not mediate this process. Nifedipine, an L-type Ca2+ channel (LTCC) inhibitor, completely blocks the activation of NKA-induced 45Ca influx, suggesting that the LTCC is responsible for the moderate increase in intracellular Ca2+. In contrast, the inhibition of NKA by ouabain induces 4.7-fold 45Ca influx compared with the condition of activation of NKA. Moreover, approximately 70% of ouabain-induced 45Ca influx was obstructed by KB-R7943 and only 30% was impeded by nifedipine, indicating that both the LTCC and the NCX contribute to the rise in intracellular Ca2+ and that the NCX reverse-mode is the major source for the 45Ca influx induced by the inhibition of NKA. This study provides direct evidence to demonstrate that the activation of NKA-induced Ca2+ increase is independent of the reverse-mode NCX and pinpoints a mechanistic distinction between the activation and inhibition of the NKA-mediated Ca2+ influx path ways in cardiomyocytes. 相似文献
19.
Addition of the mitogenic lectin concanavalin A to rat spleen cells results in a small increase in the steady-state Ca2+ content of the cells. 45Ca2+ fluxes were measured under conditions where artifacts due to Ca2+ binding to concanavalin A could be excluded. Both 45Ca2+ influx into and efflux from these cells are significantly activated by the lectin. If 45Ca2+ is added 30 min after concanavalin A the rate of influx is further enhanced. The increase in 45Ca2+ influx correlates well with binding of concanavalin A to the cells. At low concentrations (optimal mitogenic) of the lectin (1 and 3 μg/ml) no significant increase in 45Ca2+ influx occurs but an increase in 45Ca2+ efflux is still observed. The results suggest that concanavalin A binding to the cell surface causes an increase in Ca2+ influx into the cells and that activation of Ca2+ efflux occurs as a response to an increase in the cytosolic Ca2+ activity. Thus, Ca2+ may well play a role in triggering lymphocyte activation. 相似文献
20.
《生物化学与生物物理学报:生物膜》2004,1661(2):204-211
The basal 45Ca2+ influx in human red blood cells (RBC) into intact RBC was measured. 45Ca2+ was equilibrated with cells with t1/2=15-20 s and the influx reached the steady state value in about 90-100 s and the steady state level was 1.5±0.2 μmol/lpacked cells (n=6) at 37 °C. The average value of the Ca2+ influx rate was 43.2±8.9 μmol/lpacked cells hour. The rate of the basal influx was pH-dependent with a pH optimum at pH 7.0 and on the temperature with the temperature optimum at 25 °C. The basal Ca2+ influx was saturable with Ca2+ up to 5 mmol/l but at higher extracellular Ca2+ concentrations caused further increase of basal Ca2+ influx. The 45Ca2+ influx was stimulated by addition of submicromolar concentrations of phorbol esters (phorbol 12-myristate-13-acetate (PMA) and phorbol-12,13-dibutyrate (PDBu)) and forskolin. Uncoupler (3,3′,4′,5-tetrachloro-salicylanilide (TCS) 10−6-10−5 mol/l) inhibited in part the Ca2+ influx. The results show that the basal Ca2+ influx is mediated by a carrier and is under control of intracellular regulatory circuits. The effect of uncoupler shows that the Ca2+ influx is in part driven by the proton-motive force and indicates that the influx and efflux of Ca2+ are coupled via the RBC H+ homeostasis. 相似文献