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1.
Acholeplasma laidlawii was grown with different fatty acids for membrane lipid synthesis (saturated straight- and branched-chain acids and mono- and di-unsaturated acids). The ability of 12 different sterols to affect cell growth, lipid head group composition, the order parameter of the acyl chains, and the phase equilibria of in vivo lipid mixtures was studied. The following two effects were observed with respect to cell growth: with a given acyl chain composition of the membrane lipids, growth was stimulated, unaffected, reduced, or completely inhibited (lysis), depending on the sterol structure; and the effect of a certain sterol depended on the acyl chain composition (most striking for epicoprostanol, cholest-4-en-3-one, and cholest-5-en-3-one, which stimulated growth with saturated acyl chains but caused lysis with unsaturated chains). The three lytic sterols were the only sterols that caused a marked decrease in the ratio between the major lipids monoglucosyldiglyceride and diglucosyldiglyceride and hence a decrease in bilayer stability when the membranes were enriched in saturated (palmitoyl) chains. With these chains correlations were found for several sterols between the glucolipid ratio and the order parameter of the acyl chains, as well as the lamellar-reversed hexagonal phase transition, in model systems. A shaft experiment revealed a marked decrease in the ratio of monoglucosyldiglyceride to diglucosyldiglyceride with the lytic sterols in unsaturated (oleoyl) membranes. The two cholestenes induced nonlamellar phases in in vivo mixtures of oleoyl A. laidlawii lipids. The order parameters of the oleoyl chains were almost unaffected by the sterols. Generally, the observed effects cannot be explained by an influence of the sterols on the gel-to-liquid crystalline phase transition.  相似文献   

2.
In membranes of Acholeplasma laidlawii, lipid composition is regulated as a function of several stimuli affecting the volume and length of the hydrocarbon chains and the hydrocarbon-water interfacial area. This regulation is vizualized as changes in the relative amounts of the major polar lipids monoglucosyl diglyceride and diglucosyl diglyceride. These lipids form reversed hexagonal and lamellar phases with water, respectively. However, mixtures of the two lipids, in the molar proportions found in the A. laidlawii membrane, form a lamellar phase. By adjustment of the glycolipid ratio as a response to environmental stimuli, a certain stability of the lamellar membrane is maintained. In growing cells with oleoyl membrane lipids, a transmembrane electrical potential of approximately -50 mV (inside negative), but no transmembrane pH difference, was found. Addition of the K+ ionophore valinomycin caused a rapid and dose-dependent hyperpolarization remaining for at least 7 h. Simultaneously, a rapid and lasting metabolic decrease in the ratio monoglucosyl diglyceride/diglucosyl diglyceride occurred. The increase in potential and the decrease in the lipid ratio were both reversed in a dose-dependent manner by extracellular KCl. Likewise, the lipophilic cation tetraphenylphosphonium caused a dose-dependent decrease in membrane potential and an increase in the monoglucosyl diglyceride/diglucosyl diglyceride ratio, respectively. The ionophores monensin and particularly nigericin had similar but less pronounced effects on the potential and lipid ratios as valinomycin. The uncoupler carbonyl cyanide m-chlorophenylhydrazone had no effect on cell growth, membrane potential, or lipid regulation at 10 microM. These dissimilar structures and the low concentrations used make a direct disturbance of drug molecules on lipid packing in membranes less likely.(ABSTRACT TRUNCATED AT 250 WORDS)  相似文献   

3.
In the perfused rat liver, administration of glucagon causes a hyperpolarization of the liver cell membrane and increases gluconeogenesis. Insulin, a hormone which is known to antagonize the effect of glucagon on gluconeogenesis also blocks the hyperpolarizing effect of glucagon. Because of this inhibitory effect of insulin of the glucagon-evoked hyperpolarization, a systematic study of possible correlation between changes in membrane potential and gluconeogenesis was undertaken. The membrane potential was changed by valinomycin, tetracaine, or by varying the ionic composition of the perfusate. A highly significant correlation between changes in membrane potential and the rate of gluconeogenesis was noticed. The possibility was raised that changes in membrane potential might exert an influence on metabolic process by a yet unknown mechanism.  相似文献   

4.
The membrane potential of Ehrlich ascites tumor cells and the effects of valinomycin and ouabain upon it have been determined. The membrane potential in control cells was 12.0 mV, inside negative. Neither valinomycin nor ouabain alone affected this value. However, valinomycin and ouabain in combination resulted in a slight hyperpolarization of the membrane. Concomitant determinations of cellular Na+, K+ and Cl- showed that valinomycin induced net losses of K+ and Cl- and a net gain in Na+ when compared to ouabain-inhibited cells. K+ permeability was increased by approximately 30% in the presence of valinomycin. In addition, valinomycin caused a rapid depletion of cellular ATP. Inhibition of Na/K transport by ouabain was without sparing effect on the rate of ATP depletion. Possible mechanisms for the electroneutral increase in K+ permeability induced by valinomycin are discussed.  相似文献   

5.
In cells of Leuconostoc oenos, the fermentation of L-malic acid generates both a transmembrane pH gradient, inside alkaline, and an electrical potential gradient, inside negative. In resting cells, the proton motive force ranged from -170 mV to -88 mV between pH 3.1 and 5.6 in the presence Of L-malate. Membrane potentials were calculated by using a model for probe binding that accounted for the different binding constants at the different pH values at the two faces of the membrane. The delta psi generated by the transport of monovalent malate, H-malate-, controlled the rate of fermentation. The fermentation rate significantly increased under conditions of decreased delta psi, i.e., upon addition of the ionophore valinomycin in the presence of KCl, whereas in a buffer depleted of potassium, the addition of valinomycin resulted in a hyperpolarization of the cell membrane and a reduction of the rate of fermentation. At the steady state, the chemical gradient for H-malate- was of the same magnitude as delta psi. Synthesis of ATP was observed in cells performing malolactic fermentation.  相似文献   

6.
It has been suggested that the insulin-induced hyperpolarization might be a mediator of the stimulatory action of insulin on glucose transport. The purpose of the present study was to investigate the relationship between the insulin-induced hyperpolarization and the stimulatory action of insulin on glucose transport in skeletal muscle. Satorius muscles dissected from bullfrogs (Rana catesbeiana) were used. Insulin induced a hyperpolarization of the membrane and an increase in the 3-O-Methyl-D-glucose (3-O-MG) uptake and extrusion. In the presence of valinomycin, insulin had no significant effect on the membrane potential. Insulin still had the stimulatory action on both the 3-O-MG uptake and extrusion even in the presence of valinomycin, under whose condition insulin had no significant effect on the membrane potential. The magnitude of the stimulatory action of insulin on the 3-O-MG uptake in the presence of valinomycin was smaller than that in the absence of valinomycin. The magnitude of the stimulatory action of insulin on the 3-O-MG extrusion was, on the contrary, larger than that in the absence of valinomycin. The abolishment of the insulin-induced hyperpolarization decreased the 3-O-MG uptake and increased the 3-O-MG extrusion. The observation in the present study concludes that insulin has two different actions on glucose transport. One of them is developed through the insulin-induced hyperpolarization, which increases the 3-O-MG uptake and decreases the 3-O-MG extrusion. The other action is irrelevant of the insulin-induced hyperpolarization and stimulates both the 3-O-MG uptake and extrusion.  相似文献   

7.
Since several anti-cancer drugs interact with cell membrane lipids, the effects of anti-cancer dietary factors on liposomal membranes with different lipid composition were comparatively studied by measuring fluorescence polarization. Fluidity was imparted on both hydrophobic and hydrophilic regions of lipid bilayers by decreasing cholesterol and increasing unsaturated phosphatidylcholine in membranes. At 0.625-10 microM, (-)-epigallocatechin gallate, genistein, apigenin, resveratrol and a reference anti-cancer drug, doxorubicin, rigidified the tumor cell model membranes consisting of 20 mol% cholesterol and 80 mol% phosphatidylcholine with the acyl chain 18:1/16:0 ratio of 1.0, but not daidzein. They were more effective on the membrane core than the membrane surface. Quercetin showed a biphasic effect on the hydrophobic regions of membrane lipid bilayers to rigidify above 5 microM and fluidize below 2.5 microM. In contrast, anti-cancer dietary factors and doxorubicin were not or much less effective in rigidifying the normal cell model membranes consisting of 40 mol% cholesterol and 60 mol% phosphatidylcholine with the acyl chain 18:1/16:0 ratio of 0.5. The membrane-rigidifying effects were greater depending on a decrease of the cholesterol/phosphatidylcholine ratio and an increase of the phosphatidylcholine unsaturation degree. Membrane-active dietary factors and doxorubicin inhibited the growth of mouse myeloma cells at 10-100 microM, while the growth inhibition by membrane-inactive daidzein was relatively weak. Anti-cancer dietary factors appear to act on more fluid membranes like tumor cells as well as doxorubicin to induce rigidification, especially in the hydrocarbon core of membrane lipids, which is determined by the composition of cholesterol and unsaturated phospholipids.  相似文献   

8.
Disorders of the carnitine cycle and of the beta oxidation spiral impair the ability to obtain energy from fats at time of fasting and stress. This can result in hypoketotic hypoglycemia, cardiomyopathy, cardiac arrhythmia and other chronic medical problems. The in vitro study of fibroblasts from patients with these conditions is impaired by their limited oxidative capacity. Here we evaluate the capacity of valinomycin, a potassium ionophore that increases mitochondrial respiration, to increase the oxidation of fatty acids in cells from patients with inherited fatty acid oxidation defects. The addition of valinomycin to fibroblasts decreased the accumulation of the lipophilic cation tetraphenylphosphonium (TPP+) at low concentrations due to the dissipation of the mitochondrial membrane potential. At higher doses, valinomycin increased TPP+ accumulation due to the increased potassium permeability of the plasma membrane and subsequent cellular hyperpolarization. The incubation of normal fibroblasts with valinomycin increased [14C]-palmitate oxidation (measured as [14C]O2 release) in a dose-dependent manner. By contrast, valinomycin failed to increase palmitate oxidation in fibroblasts from patients with very long chain acyl CoA dehydrogenase (VLCAD) deficiency. This was not observed in fibroblasts from patients heterozygous for this condition. These results indicate that valinomycin can increase fatty acid oxidation in normal fibroblasts and could be useful to differentiate heterozygotes from patients affected with VLCAD deficiency.  相似文献   

9.
《BBA》2022,1863(7):148587
Cardiolipin is the signature phospholipid of the mitochondrial inner membrane. It participates in shaping the inner membrane as well as in modulating the activity of many membrane-bound proteins. The acyl chain composition of cardiolipin is finely tuned post-biosynthesis depending on the surrounding phospholipids to produce mature or unsaturated cardiolipin. However, experimental evidence showing that immature and mature cardiolipin are functionally equivalents for mitochondria poses doubts on the relevance of cardiolipin remodeling. In this work, we studied the role of cardiolipin acyl chain composition in mitochondrial bioenergetics, including a detailed bioenergetic profile of yeast mitochondria. Cardiolipin acyl chains were modified by genetic and nutritional manipulation. We found that both the bioenergetic efficiency and osmotic stability of mitochondria are dependent on the unsaturation level of cardiolipin acyl chains. It is proposed that cardiolipin remodeling and, consequently, mature cardiolipins play an important role in mitochondrial inner membrane integrity and functionality.  相似文献   

10.
Proximal mechanisms describing the evolution of high levels of basal metabolic rate (BMR) in endotherms are one of the most intriguing problems of evolutionary physiology. Because BMR mostly reflects metabolic activity of internal organs, evolutionary increase in BMR could have been realized by an increase in relative organ size and/or mass-specific cellular metabolic rate. According to the "membrane pacemaker" theory of metabolism, the latter is mediated by an increase in the average number of double bonds (unsaturation index) in cell membrane fatty acids. To test this, we investigated the effect of divergent artificial selection for body-mass-corrected BMR on the mass of internal organs and the fatty acid composition of cell membranes in laboratory mice (Mus musculus). Mice from the high-BMR line had considerably larger liver, kidneys, heart, and intestines. In contrast, the unsaturation index of liver cell membranes was significantly higher in low-BMR mice, mainly because of the significantly higher content of highly polyunsaturated 22 : 6 docosahexanoic fatty acid. Thus, divergent selection for BMR did not affect fatty acyl composition of liver and kidney phospholipids in the direction predicted by the membrane pacemaker theory. We conclude that an intraspecific increase in BMR may rapidly evolve mainly as a result of the changes in size of internal organs, without simultaneous increase of the unsaturation index in cell membrane lipids.  相似文献   

11.
The relationship between alterations in transmembrane potential, cell volume, and phospholipid fatty acid turnover has been examined in human erythrocytes by treating the cells with the monovalent cation ionophore valinomycin. Valinomycin increases the cellular uptake of tetra[3H]phenylphosphonium ion by erythrocytes, indicating membrane hyperpolarization, and causes net loss of potassium chloride and water from the cells leading to a decrease in cell volume. Treatment of erythrocytes with valinomycin also enhances incorporation of [9, 10-(3)H]oleic acid into phospholipids, primarily diacylphosphatidylethanolamine. After replacing intracellular chloride with sulfate and treating cells with the anion transport inhibitor 4,4'-diisothiocyanostilbene-2,2'-disulfonate, exposure to valinomycin results in uptake of tetra[3H]phenylphosphonium ion and stimulation of [9, 10-(3)H]oleic acid incorporation, but, because anion efflux is prevented, no decrease in cell volume occurs. When tetra[3H]phenylphosphonium ion uptake is also prevented by suspending these cells in 125 mM KCl to dissipate the transmembrane potassium gradient, valinomycin still enhances [9, 10-(3)H] oleic acid incorporation into phospholipid. These results suggest that the presence of valinomycin in the membrane directly alters phospholipid fatty acid turnover and that some of the effects of this ionophore on cellular function previously attributed to alterations in transmembrane potential or cellular potassium content may instead be due to altered phospholipid turnover. Since it is possible that valinomycin may directly perturb phospholipid fatty acid turnover in other cells, the possibility that valinomycin-induced alterations in cellular function are due to altered phospholipid turnover rather than membrane hyperpolarization or altered potassium content should be considered in the interpretation of studies employing this ionophore.  相似文献   

12.
Using indirect methods based on uptake of [3H]tetraphenylphosphonium cation and [14C]benzoic acid by cells of the fungus Trichoderma viride we found that the illumination-induced transient hyperpolarization of the plasma membrane is followed immediately by a rapid temporary decrease in intracellular pH. Hyperpolarization and intracellular acidification were completely suppressed by 150 mM-KCl and by the K(+)-ionophore valinomycin. The light-induced acidification of the cytoplasm was not observed in the presence of the cytochrome respiratory chain inhibitors antimycin A and mucidin. Based on these results, we hypothesize that the hyperpolarization of the cells is the consequence of an efflux of K+ through a light-activated K(+)-channel in the plasma membrane. The loss of positive charge in the cytoplasm caused by this efflux of cations is counterbalanced by H+ originating from the light-activated mitochondrial respiratory chain.  相似文献   

13.
Glycosphingolipids (GSLs) play major roles in cellular growth and development. Mammalian glycolipid transfer proteins (GLTPs) are potential regulators of cell processes mediated by GSLs and display a unique architecture among lipid binding/transfer proteins. The GLTP fold represents a novel membrane targeting/interaction domain among peripheral proteins. Here we report crystal structures of human GLTP bound to GSLs of diverse acyl chain length, unsaturation, and sugar composition. Structural comparisons show a highly conserved anchoring of galactosyl- and lactosyl-amide headgroups by the GLTP recognition center. By contrast, acyl chain chemical structure and occupancy of the hydrophobic tunnel dictate partitioning between sphingosine-in and newly-observed sphingosine-out ligand-binding modes. The structural insights, combined with computed interaction propensity distributions, suggest a concerted sequence of events mediated by GLTP conformational changes during GSL transfer to and/or from membranes, as well as during GSL presentation and/or transfer to other proteins.  相似文献   

14.
It has been recently shown that mitochondrial creatine kinase (mtCK) organizes mitochondrial model membrane by modulating the state and fluidity of lipids and by promoting the formation of protein-cardiolipin clusters. This report shows, using Brewster angle microscopy, that such clustering is largely dependent on the acyl chain composition of phospholipids. Indeed, mtCK-cardiolipin domains were observed not only with unsaturated cardiolipins, but also with the cardiolipin precursor phosphatidylglycerol. On the other hand, in the case of saturated dimyristoylphosphatidylglycerol and tetramyristoylcardiolipin, mtCK was homogeneously distributed underneath the monolayer. However, an overall decrease in membrane fluidity was indicated by infrared spectroscopy as well as by extrinsic fluorescence spectroscopy using Laurdan as a fluorescent probe, both for tetramyristoylcardiolipin and bovine heart cardiolipin containing liposomes. The binding mechanism implicated the insertion of protein segments into monolayers, as evidenced from alternative current polarography, regardless of the chain unsaturation for the phosphatidylglycerols and cardiolipins tested.  相似文献   

15.
Isolated frog skin, mounted in a Ussing apparatus, was investigated electrophysiologically. Application of amiloride, an inhibitor of sodium ion transport, and bumetanide, known to block the transport of chloride ions, revealed the effect of these ions on PD, both under control conditions and following mechanical stimulation. Under control conditions, mechanical stimulation of the skin caused hyperpolarization, i.e. a transient increase in the electrical potential difference. Preincubation in the presence of amiloride, or amiloride plus bumetanide, brought about both a decrease in electrical potential and an inhibition of the reaction upon stimulation. On the other hand, incubation with bumetanide resulted in a decrease in electrical potential, but did not affect the skin reaction after mechanical stimulation. The above results indicate that hyperpolarization of the frog skin following mechanical stimulation is caused by enhanced transepithelial transport of sodium ions which, in turn, is induced by stimulation of sensory receptors.  相似文献   

16.
Goldfish were acclimated to 5, 15, and 30°C, and the acyl group composition of choline phosphoglycerides (CPG) and ethanolamine phosphoglycerides (EPG) from whole brain and brain subcellular particles was examined. With the exception of synaptosomal CPG, the acyl group composition of CPG from whole brain and subcellular particles, including myelin, from cold-acclimated fish showed little response to the change in environmental temperature. Those changes that did occur were consistent with the expected trend toward a higher degree of unsaturation of the CPG acyl groups in fish acclimated to 5°C. The acyl group composition of CPG from synaptosomes of the cold-acclimated fish did, however, differ markedly in having a reduced unsaturation index (U.I.) and unsaturated: saturated fatty acid ratio (UFA:SFA) which was caused mainly by the decrease in 226n-3 content. In contrast, changes in the acyl group composition of EPG on cold acclimation were greater than those observed in any CPG fraction. The generally expected trend toward greater unsaturation was observed only in mitochondrial and myelin EPG. Moreover, in all fractions the amount of 226n-3 in EPG was lower at decreased environmental temperatures. In the synaptosomal and microsomal EPG, the reduction in 226n-3 was such that a markedly reduced U.I. was obtained. It is suggested that two compensatory mechanisms maintain the necessary degree of membrane permeability and fluidity in order to prevent transition to a crystalline state at lower temperatures.  相似文献   

17.
Data concerning the acyl composition of tissue phospholipids from mammal species, ranging in size from the shrew (7 g) to cattle (370 kg), has been collated from the literature and analysed allometrically. Phospholipids from heart, skeletal muscle, liver and kidney exhibited similar allometric trends whereby phospholipids had a significant decrease in unsaturation index (number of double bonds per 100 acyl chains) as species body size increased whilst there was no change in the percent of unsaturated acyl chains. Whilst total polyunsaturate content did not change with body mass, both heart and skeletal muscle phospholipids showed a significant allometric decrease in the omega-3 polyunsaturate content. The content of the highly polyunsaturated docosahexaenoic acid (22:6 n-3) in phospholipids showed significant and substantial allometric decline with increasing body mass in all four tissues (exponents ranged from -0.19 in liver to -0.40 in skeletal muscle). Brain phospholipids showed no allometric trends in acyl composition and were highly polyunsaturated in all species. These trends are discussed in light of the hypothesis that the relative content of polyunsaturated acyl chains in membranes, and especially docosahexaenoate (22:6 n-3), can act as a membrane pacemaker for metabolic activity.  相似文献   

18.
In the ascidian Ciona intestinalis (and C. savignyi), sperm-activating and -attracting factor (SAAF) is released from the egg at fertilization and stimulates both Ca(2+) influx and a transient increase in cAMP level of the sperm, leading to the activation of sperm motility (M. Yoshida et al., 1994, Dev. Growth Differ. 36, 589-595). In this paper we show in C. intestinalis that valinomycin, a potassium-selective ionophore, as well as SAAF, activated sperm motility, and this activation was suppressed by extracellular high K(+). Membrane potential measurements showed that both SAAF and valinomycin increase K(+) permeability of sperm and induce membrane hyperpolarization, the amplitude of which depends on the external K(+) concentration. The membrane potential and intracellular K(+) concentration of Ciona sperm without SAAF were estimated to be about -50 mV and 560 +/- 40 mM, respectively. After treatment with SAAF or valinomycin the membrane potential became almost equal to the equilibrium potential of K(+) (-100 mV), and the cAMP level increased in artificial seawater. A potent voltage-dependent K(+) channel blocker, MCD peptide, at the concentration of 10 microM blocked SAAF-induced hyperpolarization of the cells, increase in cAMP, and sperm motility. These results suggest that membrane hyperpolarization produced by the opening of K(+) channels elevates cAMP synthesis and leads to the activation of sperm motility in Ciona.  相似文献   

19.
The membrane potentials of mycoplasmas were investigated by using potential-sensitive cyanine dyes. The fluorescence response results from a potential-dependent partition of the dyes between the cells and the extracellular medium. Cell hyperpolarization (inside more negative), e.g., by the addition of valinomycin, results in uptake of the dyes into the cells and, by formation of dye aggregates, in quenching of the fluorescence intensity. The magnitude of the fluorescence change upon addition of valinomycin depended on the external K+ concentration. At a defined external K+ concentration, no change in fluorescence occurred. The intracellular K+ concentration was determined by atomic absorption spectroscopy. Mycoplasma membrane potentials were calculated according to the Nernst equation. The membrane potential of bothMycoplasma mycoides subsp.capri andMycoplasma gallisepticum was −48 mV±10%; the membrane potential ofAcholeplasma laidlawii was −28 mV±20%.  相似文献   

20.
A fluorescent dye sensitive to membrane potential was used to follow the plasma-membrane potential in the unicellular halo-tolerant alga Dunaliella salina. The signal observed during dissipation of the plasma membrane potential by the addition of excess K+ and valinomycin, or a protonophore, was taken as a measure of the preexisting potential. A resting potential of −85 to −100 millivolts (negative inside) was calculated. Following a hypertonic shock, the plasma membrane was rapidly hyperpolarized. This hyperpolarization was transient, and the algae resumed their resting potential about 30 minutes after the shock. The resting plasma membrane potential was decreased by vanadate and is concluded to be generated mostly by the plasma membrane ATPase of Dunaliella. The transient hyperpolarization following a hypertonic shock indicates, therefore, a transient activation of the ATPase. This is further corroborated by a rapid transient decrease in the intracellular ATP following a hypertonic shock and its inhibition by vanadate. It is suggested that activation of the plasma membrane ATPase may be the trigger for osmoregulation in Dunaliella.  相似文献   

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