Acid Composition Of Membrane Lipids Research Articles

Chilling sensitivity of Arabidopsis thaliana with genetically engineered membrane lipids.

Upon transfer of a genetically engineered Escherichia coli gene for glycerol-3-phosphate acyltransferase (plsB) to Arabidopsis thaliana (L.) Heynh., the gene is transcribed and translated into an enzymatically active polypeptide. This leads to an alteration in fatty acid composition of membrane lipids. From these alterations it is evident that the enzyme is located mainly inside the plastids. The amount of saturated fatty acids in plastidial membrane lipids increased. In particular, the fraction of high-temperature melting species of phosphatidylglycerol is elevated. These molecules are thought to play a crucial role in determining chilling sensitivity of plants. An increase in sensitivity could be observed in the transgenic plants during recultivation after chilling treatment. Implications for the hypothesis of phosphatidylglycerol-determined chilling sensitivity are discussed.

Low temperature-induced fatty acid desaturation in Brassica napus: thermal deactivation and reactivation of the process

When Brassica napus plants are grown at low temperatures (e.g., 5°C) the rate of desaturation in leaves of newly formed fatty acids in both chloroplastic (MGDG) and cytosolic (PC) diacylglycerols is higher or more rapid than in plants grown at higher temperatures (e.g., 30°C). This low temperature-induced increase in the rate of desaturation is lost within hours if plants are transferred to higher temperatures. However, if plants are then returned to low temperatures they regain the ability to rapidly desaturate fatty acids. This process is restored relatively slowly (over days) in contrast to the more rapid loss at high temperatures. This has important physiological consequences on the level of unsaturated fatty acids in plant membranes and the process of temperature control of the fatty acid composition of membrane lipids.

Effect of dietary fats on some membrane-bound enzyme activities, membrane lipid composition and fatty acid profiles of rat heart sarcolemma.

The effect of various dietary fats on membrane lipid composition, fatty acid profiles and membrane-bound enzyme activities of rat cardiac sarcolemma was assessed. Four groups of male weanling Charles Foster Young rats were fed diets containing 20% of groundnut, coconut, safflower or mustard oil for 16 weeks. Cardiac sarcolemma was prepared from each group and the activities of Na+, K(+)-ATPase, 5'-nucleotidase, Ca(2+)-ATPase and acetylcholinesterase were examined. ATPase activities were similar in all groups except the one fed coconut oil, which had the highest activities. Acetylcholinesterase activity was also similar in all the groups, however, it was significantly higher in the group fed mustard oil. No significant changes were observed among the groups in 5'-nucleotidase activity, in the cholesterol-to-phospholipid molar ratio and in sialic acid content. The coconut, safflower and mustard oil diets significantly increased cholesterol and phospholipid contents and the lipid-to-protein ratio of cardiac sarcolemma as compared to feeding the groundnut oil diet. The fatty acid composition of membrane lipids was quite different among the various groups, reflecting the type of dietary fat given. The total unsaturated-to-saturated fatty acid ratio was not different among the various groups; however, the levels of some major fatty acids such as palmitic (16:0), oleic (18:1) and linoleic (18:2) acids were significantly different. Cardiac sarcolemma of the group fed safflower oil had the highest polyunsaturated fatty acid content. The results suggest that dietary fats induce changes not only in the fatty acid composition of the component lipids but also in the activities of sarcolemmal enzymes involved in the regulation of cardiac function.

Functional Consequences of Abnormal Fatty Acid Profiles in Cultured Airway Epithelial Cells

Maintenance of serum-free conditions for the culture of TE or other airway epithelial cells provides a defined environment in which to explore the regulation of cellular functions. Yet TE cells appear to be dependent on the medium for essential, if not all, polyunsaturated fatty acids. At present, some laboratories routinely use serum to support the growth of airway epithelial cells, presumably in part through recognition that cells of mammalian origin require an exogenous source of lipids. While 5% FBS can increase the linoleic and arachidonic acid content of cultured rabbit and human TE cells, it does not fully restore the fatty acid composition of cultured TE cells to that of freshly isolated cells, particularly in the case of human TE cells. Equally good, if not better, repair of membrane fatty acid composition can be achieved by addition of a defined, commercial non-serum source of lipids (Excyte III) plus exogenous arachidonic acid. Cultured TE cells maintained in serum-free medium have been shown to be deficient in prostaglandin and HETE production, both at baseline and in response to physiological stimuli compared to TE cells with greater endogenous content of arachidonic acid. Differences between lipid supplemented and unsupplemented cultured TE cells in cAMP response to PGE2 and in susceptibility to hyperoxic injury have been observed. Other cellular functions regulated by the fatty acid composition of membrane lipids may also be impaired in lipid unsupplemented cells. It is evident that the maintenance of as normal as possible membrane fatty acid content is essential to the use of cultured TE cells as experimental models of airway epithelium.

Effects of estrogen-induced hyperlipidemia on the erythrocyte membrane in chicks.

The effects of estrogen-induced hyperlipidemia on plasma lipid peroxidation, fatty acid composition and osmotic fragility of erythrocytes in chickens were studied. Young male chickens implanted with estrogen for three wk developed a marked hyperlipidemia. Plasma levels of triglyceride, cholesterol and phospholipid were elevated 68-, four- and 24-fold, respectively, over controls. There was also a two-fold increase in plasma lipid peroxidation measured by the thiobarbituric acid test. Vitamin E supplement (1,000 IU/kg diet) reduced the plasma lipid peroxidation to the control level, but had no effect on the plasma lipid content. Estrogen-induced hyperlipidemia resulted in changes in the fatty acid composition of membrane lipids of erythrocytes. The major changes were an increase in oleic acid from 10.0% to 14.2% and a decrease in linoleic acid from 31.3% to 26.0%. The erythrocytes with an altered membrane fatty acid composition were found to have an increased osmotic fragility. It was apparent that there was a direct correlation between the oleic acid content and the osmotic fragility of erythrocytes.

Lipids and platelet function in runners.

The effect of distance running on the fatty acid composition of platelet and erythrocyte membranes has been investigated, together with platelet aggregation and levels of high density lipoprotein-cholesterol (HDLc), low density lipoprotein cholesterol (LDLc), total cholesterol and triglycerides in runners (n = 11) and healthy age-matched non-running controls (n = 12). Platelet aggregation and fatty acid composition of membrane lipids in both platelets and erythrocytes are similar in both groups with the following exceptions: in platelets docosahexaenoic acid (C22:6 omega 3) is significantly higher in runners; in erythrocytes the fatty acids C20:3 omega 6/C22:1 omega 9 and C22:5 omega 3 are significantly higher in runners. There were no significant differences between the levels of HDLc, LDLc and total cholesterol in runners and controls although triglycerides were significantly lower in runners. Possible beneficial effects of running are probably mediated through effects on serum lipid and lipoprotein concentrations, and probably not due to any antithrombotic effect of platelets.

Lipid interconversions in aging Mycoplasma capricolum cultures.

During the progression of Mycoplasma capricolum cultures from the early exponential to the stationary phase of growth, a decrease in the phospholipid-to-protein ratio and increases in both the unsaturated-to-saturated fatty acid ratio and the diphosphatidylglycerol (DPG)-to-phosphatidylglycerol (PG) ratio were found. The freedom of motion of spin-labeled fatty acids incorporated into the membrane remained unchanged throughout the growth cycle. The increase in DPG was almost stoichiometric with the decrease in PG. Furthermore, exogenous PG added to the medium was incorporated by the cells and partially converted to DPG. The DPG that was accumulated upon aging was always more unsaturated than the PG. This accumulation was enhanced in palmitic acid-poor media, but was inhibited even in aged cells when the cells were grown in palmitic acid-rich media, suggesting that the accumulation of DPG upon aging was associated with changes in the fatty acid composition of membrane lipids rather than with the transition of the cells from the exponential- to stationary-growth phase.

Effect of Essential Fatty Acid Deficiency on Acyl Group Composition, Membrane Fluidity and Adenylate Cyclase Activity in Plasma Membranes of Rat Submandibular Salivary Glands

The effects of a marginal and severe deficiency of essential fatty acids (EFA) on the fatty acid composition, fluorescence polarization of diphenyl-hexatriene (DPH) and adenylate cyclase activity were studied in plasma membranes of rat submandibular salivary glands (SMSG). Three groups of male weanling Sprague-Dawley rats were fed ad libitum EFA-deficient (EFAD), marginally EFA-deficient (MEFAD) or the control diets. Rats were killed after 8–17 wk of feeding and plasma membranes were prepared from the SMSG and the assayed for basal and fluoride-stimulated adenylate cyclase activity. In membranes prepared from EFAD and MEFAD rats, the fluoride-stimulated enzyme activity was 30–35% higher than that of the control group. Lower double-bond index of phospholipids in plasma membranes of the SMSG from EFAD and MEFAD rats was observed, suggesting a lower fluidity compared with the control group. Higher fluorescence polarization of DPH in membranes of EFAD and MEFAD rats was more evidence of lower fluidity. It is suggested that higher adenylate cyclase activity in SMSG of rats fed EFAD or MEFAD diets may be related to lower membrane fluidity. Fluoride-stimulated enzyme activity and fatty acid composition of membrane lipids were restored to normal values after feeding the control diets for 6 wk to the EFAD and MEFAD rats.

低温ショックによるウメ果実の低温障害抑制効果と膜透過性及び膜脂質構成脂肪酸の変化との関係

It has been reported that chilling injury in mume (Japanese apricot) fruits usually occurs much sooner at 5-8°C (moderate low temperature) than at 0-1°C, and that a cold-shock treatment by hydrocooling in ice water immediately after harvest is effective in retarding the development of the chilling injury. This paper deals with the mechanism of the cold-shock treatment.1. As an index of membrane permeability, the rate of K+ion leakage from tissue slices into water was measured at various temperatures ranging from 1°C to 20°C. While the leakage rate of chilling-insensitive potato tuber decreased as the temperature was lowered, that of chilling-sensitive cucumber fruits showed no decrease at low temperatures, remaining at an almost constant level below 10°C. However leakage rate of mume fruits, showed an unusual increase at moderate low temperatures (5-8°C). Using 4 cultivars of mume fruits, ‘Yosei’‘Nanko’‘Gyokuei’and ‘Gojiro’it was found that the temperature at which the unusual increase of K+ion leakage occurred on harvest day was almost the same as that at which severe chilling injury occurred during storage.2. When‘Oshuku’mume fruits, a very chilling-sensitive cultivar, were given a cold-shock in ice water for 2 hours and then stored at 6°C, chilling injury was retarded. Except for a temporary rise soon after the treatment, the rate of K+ion leakage remained at a constant low level. However, in untreated fruits, the rate of K+ion leakage increaesed markedly during storage at 6°C.Fatty acid composition of membrane lipids was analyzed, and C16:0 was found to be most abundant, followed by C18:2, C18:3, C18:1, C16:1, C14:0, C18:0 in that order. The ratio of unsaturated fatty acids to total fatty acids was 52%, a relatively low value compared with other commodities, suggesting a lower tolerance to chilling temperature. Cold-shock treatment increased the above ratio.Total phenol content gradually increased both in the untreated and treated fruits during 6°C storage, the values being slightly lower in the treated fruits. Activities of polyphenol oxidase and peroxidase increased during 6°C storage. No relation tocold-shock effect was found. NADH-cytochrome C reductase activity, an index of activity of the enzymatic system involved in unsaturation of fatty acids, increased rapidly during the cold-shock treatment and remained at a higher level than in untreated fruits throughout the 6°C storage period.3. From these results, it was concluded that the effect of the cold-shock treatment was to accelerate the desaturation of fatty acids in membrane lipids.

Studies on thermal adaptation of Tetrahymena lipids : Alteration in fatty acid composition and its mechanism in the growth temperature shift-up

A rise in the growth temperature of Tetrahymena pyriformis strain NT-1 from 15 to 34°C resulted in changes in the fatty acid composition of membrane lipids toward higher saturation after a shift-up. The proportion of palmitate (16:0) increased and γ-linolenate (18:3 (6,9,12)) decreased. Alteration in phospholipid composition was completed by a marked increase in phosphatidylethanolamine, with a compensatory decrease in 2-aminoethylphosphonolipid. However, the level of phosphatidylcholine remained unchanged. The rate of incorporation of [1- 14C]acetate into saturated fatty acids (myristate, 14:0; palmitate; stearate, 18:0) was maximum at 1 h after a shift, while formation of 14C-labeled unsaturated fatty acids (palmitoleate, 16:1 (9); linoleate, 18:2 (9,12); γ-linolenate) decreased by about 50%. These findings could be explained by the reduced desaturase activities which were due principally to the decrease in the terminal component, cyanide-sensitive factor (CSF), of the microsomal desaturase system. Furthermore, changes induced by temperature shift-up in activities of microsomal reductases and the content of cytochrome b 5 were observed to behave comparably with changes in activities of desaturases.

Phase separations in membranes of Anacystis nidulans grown at different temperatures

Freeze fracture electron microscopy studies were performed on samples of Anacystis nidulans quenched from different temperatures. Membrane lipid phase separations were observed to take place over the ranges 15–30°C, 5–25°C and –5–15°C for cultures grown at 38, 28 and 18°C, respectively. Differential scanning calorimetry heating curves showed endotherms which coincided with these temperature ranges. Variations of phase separation temperatures with growth temperature, and hysteresis effects in the calorimetric measurements, were related to changes in the fatty acid composition of membrane lipids.

Is the vertical disposition of mycoplasma membrane proteins affected by membrane fluidity?

The influence of the physical state of the membrane lipid matrix on the vertical disposition of membrane proteins was studied with Acholeplasma laidlawii. Changes in membrane fluidity were brought about by altering the fatty acid composition of membrane lipids, by changing the growth temperature, by aging of cultures and by inducing changes in the membrane lipid-to-protein ratio through treatment with chloramphenicol. The lactoperoxidase-mediated iodination technique was used to label membrane proteins exposed to the aqueous surroundings. The degree of exposure of the iodine-binding sites of membrane proteins on the external surface of intact cells was found to undergo significant changes on varying growth conditions, but the changes could not be consistently correlated with changes in membrane fluidity, nor were they discernible on iodination of isolated membranes.

Cholesterol in mytoplasma membranes. II Components of Acholeplasma laidlawii cell membranes responsible for cholesterol binding

1. 1. Cholesterol uptake by Acholeplasma laidlawii membranes was unaffected by the removal of 75% of the membrane protein by pronase digestion. Lipid depletion of the membranes by aqueous acetone extraction greatly reduced the amount of cholesterol taken up, indicating that the major portion of the cholesterol bound to the mycoplasma membrane is incorporated into its lipid domain. 2. 2. The influence of the fatty acid composition of membrane lipids on cholesterol uptake was tested using for this purpose oleate- and palmitate-enriched A. laidlawii membranes at various temperatures. The activation energy for cholesterol uptake was about the same for all the membranes tested, but the rate constant for uptake was significantly higher for the oleate-enriched membranes. The amount of cholesterol in the membrane at equilibrium was unaffected by the fatty acid composition. 3. 3. The isolated membrane preparations took up about twice as much cholesterol as membranes of intact, non-growing A. laidlawii cells and about six times as much cholesterol as membranes of growing cells. 4. 4. It is proposed that growing cells utilize some mechanism for excluding cholesterol from the membrane during growth.

Cholesterol in mycoplasm membranes. Correlation of enzymic and transport activities with physical state of lipids in membranes of Mycoplasma mycoides var. capri adapted to grow with low cholesterol concentrations

1. 1. Membranes of a Mycoplasma mycoides var. capri strain adapted to grow with very low concentrations of cholesterol undergo a reversible phase transition detectable by differential-scanning calorimetry and fluorescence measurements. No phase transition could be detected in membranes of the cholesterol-containing native strain. 2. 2. EPR spectrometry, as well as fluorescence measurements, demonstrated that at temperatures above phase transition the membranes of the adapted strain were more fluid than membranes of the native strain, although the former contained a higher percentage of saturated fatty acids. 3. 3. Arrhenius plots of the ATPase activity of the adapted strain membranes showed breaks at temperatures corresponding to those of the phase transition of membrane lipids. The temperature of the break depended on the fatty acid composition of membrane lipids and on the age of the culture. No break could be detected in Arrhenius plots of the ATPase activity of the native strain. 4. 4. No break could be demonstrated in the Arrhenius plot of α-methylgucoside uptake by the adapted strain. Yet the activation energy of the uptake process by the adapted strain was much higher than that of the native strain. On the other hand, activation energies of α-methylglucoside phosphorylation were the same for membranes of both strains. However, Arrhenius plots of α-methylglucoside efflux from cells of the adapted strain showed breaks at temperatures corresponding to those of the lipid phase transition. 5. 5. It is concluded that cholesterol, by preventing the crystallization of membrane lipids maintains them in a state of fluidity essential for the optimal manifestation of several key activities of the membrane.

Cholesterol in mycoplasma membranes. Composition, ultrastructure and biological properties of membranes from Mycoplasma mycoides var. capri cells adapted to grow with low cholesterol concentrations

1. 1. Serial passages of the sterol-requiring Mycoplasma mycoides var. capri in a serum-free medium supplemented with decreasing concentrations of cholesterol resulted in the adaptation of the organisms to grow with no added cholesterol. The cells of the adapted strain were osmotically more fragile than cells of the native strain and were more permeable to erythritol. 2. 2. The cell membrane of the adapted strain contained low amounts of cholesterol (up to 3% of the total membrane lipid as against 22–25% in the native strain) but its polar lipids were more saturated than those of the native strain, as reflected by the preferential incorporation of [ 14C]palmitic acid when added to the growth medium together with [ 14C]oleic acid. 3. 3. The native strain was capable of growing at temperatures as low as 25°C, whereas the adapted strain could not. The lowering of the growth temperature of the native strain resulted in a decrease in the cholesterol content of the membrane (from 24–15% of the total lipid) but had no effect on the fatty acid composition of the membrane polar lipids. On the other hand, aging of the culture increased the ratio of saturated to unsaturated fatty acids in both adapted and native strains, and decreased the cholesterol content of the native strain. 4. 4. Freeze-etching of cells of the native and adapted strains, kept at 37°C prior to freezing, showed a random distribution of particles on the fracture faces of the cell membranes. Keeping the cells at 4°C prior to freezing caused the aggregation of particles on the fracture faces of the adapted strain but had no effect on the distribution of particles in the native strain. 5. 5. Our data support the thesis that cholesterol functions as a regulator of membrane fluidity and that changes in the fatty acid composition of membrane lipids may act to compensate for the lack of cholesterol.

Are mycoplasma membrane proteins affected by variations in membrane fatty acid composition?

1. 1. The influence of the fatty acid composition of membrane lipids on the quantitative distribution of membrane protein of Mycoplasma laidlawii has been investigated. 2. 2. Cells were grown in media supplemented with either a saturated fatty acid (palmitic acid or stearic acid) or an unsaturated fatty acid (palmitoleic acid or oleic acid). Lipids isolated from membranes of these cells were markedly enriched in the supplemented fatty acid. 3. 3. The proteins of the different types of membranes were analyzed by sodium dodecyl sulfate-acrylamide-gel electrophoresis of membranes from cells grown in the presence of radioactively labelled amino acids. In general, protein composition and relative abundance were unaffected by fatty acid variation. One high molecular weight protein fraction was present in higher concentration in membranes rich in saturated fatty acids than in those rich in saturated fatty acids. 4. 4. The membranes rich in saturated fatty acids were also found to have a slightly higher density as determined by isopycnic centrifugation.

Lipid composition of the isolated rat intestinal microvillus membrane.

1. Rat intestinal microvillus plasma membranes were prepared from previously isolated brush borders and the lipid composition was analysed. 2. The molar ratio of cholesterol to phospholipid was greatest in the membranes and closely resembled that reported for myelin. 3. Unesterified cholesterol was the major neutral lipid. However, 30% of the neutral lipid fraction was accounted for by glycerides and fatty acid. 4. Five phospholipid components were identified and measured, including phosphatidylethanolamine, phosphatidylcholine, phosphatidylserine, sphingomyelin and lysophosphatidylcholine. Though phosphatidylethanolamine was the chief phospholipid, no plasmalogen was detected. 5. In contrast with other plasma membranes in the rat, the polar lipids of the microvillus membrane were rich in glycolipid. The cholesterol:polar lipid (phospholipid+glycolipid) ratio was about 1:3 for the microvillus membrane. Published data suggest that this ratio resembles that of the liver plasma membrane more closely than myelin or the erythrocyte membrane. 6. The fatty acid composition of membrane lipids was altered markedly by a single feeding of safflower oil. Membrane polar lipids did not contain significantly more saturated fatty acids than cellular polar lipids. Differences in the proportion of some fatty acids in membrane and cellular glycerides were noted. These differences may reflect the presence of specific membrane glycerides.