The effect of polyene antibiotics on fractions of DNP both bound and not bound to dog kidney nuclear membranes]

Effects of amphotericine B and nistatine on nuclear membrane-bound DNP (DNPm) and free DNP (DNPf) from dog kidney are studied. Intravenous injection of amphotericine B resulted in the increase of binding of DNP particles with nuclear membrane: the content of DNA in DNPm fraction was 50-fold increased. The injection of nistatine did not affect DNP binding with nuclear membrane. Amphotericine B alone increased the protein-DNA ratio and decreased the RNA/DNA ratio in DNPf fraction. Both amphotericine B and nistatine sharply increased the protein/DNA ratio and practically did not change the RNA/DNA ratio in DNPm fraction. Amphotericine B produced considerable changes in temperature denaturation of DNA in DNPo, while nistatine produced no effect. Both antibiotics considerably changed the composition of acid soluble proteins in DNPm and DNPf, non-histone proteins in DNPf, and also they caused the changed and quantitative redistribution of separate lipid components in DNPm lipids. Polyene antibiotics are suggested to effect on animal cell nuclear structures.     

Effects of polyene antibiotics on the membranes of dog kidney isolated nuclei]

The effects of amphotericin B and nistatin on the membranes of dog kidney isolated nuclei after their incubation with the antibiotics in question, have been studied. It is found that the polyene antibiotics, though they are superficially-active compounds, have no solubilizing effect on nuclear membranes and do not change their chemical composition. Electrophoretic study has revealed that nuclear membrane proteins, besides high- and low-molecular protein components, also contain a large amount of histones. The incubation of the nuclei with the polyene antibiotics results in marked changes in the fractional composition of nuclear membrane proteins, the most significant changes being induced by amphotericin B. It is assumed that polyene antibiotics induce proteolytic degradation of nuclear membrane proteins.     

Effect of amphoterecin B on the plasma membranes of a tissue culture of puppy kidney]

The effect of amphotoericin B on the chemical composition of the plasmic membranes of the puppy kidney tissue was studied. It was shown that amphotericin B in a concentration of 10 Units/ml induced changes in the quantitative content of proteins, lipids and RNA in the chemical composition of the plasmic membranes. The effect of amphotericin B on the composition of proteins and lipids in the membranes was also studied. It was found with the method of electrophoresis in polyacrylamide gel that after exposure of the culture cells to the antibiotic significant shifts in the composition of the membrane proteins took place. According to the findings of thin-layer chromatography amphotericin B markedly changed the quantitative content of the fractions in the composition of lipids in the plasmic membranes. Significant deviations under the effect of the antibiotic were observed in the quantitative ratio of separate fatty acids contained in the lipids of the membrane preparations studied. On the basis of the results obtained it was supposed that the mode of action of amphotericin B on the cell was associated with impairement of the plasmic membrane structure by the antibiotic.     

The change of template activity of dog kidney chromtin by polyene antibiotics in vivo and in vitro]

Effect of amphotericin B and nistatin on template activity of nuclear membrane-bound (DNPm) and free (DNPo) dog kidney chromatin after intravenous injections of antibiotics and after the incubation of isolated kidney cell nuclei with the antibiotics is studied. It is found that injections of amphotericin B and nistatin resulted in the increase of DNPo template activity in RNA polymerase system, the stimulating effect of nistatin being higher than that of amphotericin B. Injections of nistatine stimulated also template activity of DNPm, while amphotericin B produced no effect on DNPm. When studing the effect of polyene antibiotics on template activity of DNPo and DNPm in vitro, it is found that the intensity of RNA synthesis after incubation of isolated nuclei with antibiotics is considerably increased, and stimulating effect of nistatin is higher than of amphotericin B. Both antibiotics produced no effect on template activity of DNP in vitro. Thus, comparative analysis of changes in template activity of dog kidney chromatin under the effect of polyene antibiotics in vivo and in vitro has revealed the similarity of these drugs and draws to the conclusion that nistatin and amphotericin B produce a direct effect on template activity of chromatin.     

Changes in ATPase activity of dog kidney nuclear membrane under the effect of polyenic antibiotics]

Effect of amphotericin B and nistatin on ATPase activity of dog kidney nuclear membranes is studied in vivo and in vitro. Long-term intravenous injections of the antibiotics do not change the ATPase activity of kidney nuclear membranes. However, short-term injections of polyenic antibiotics have some effect on ATPase activity on nuclear membranes: amphotericin B, considerably activates the enzyme . In vitro incubation of isolated dog kidney nuclei with amphotericin and nistatin at concentrations of 1 and 10 mcg/ml does not affect the activity of nuclear membrane ATPase, while increased concentrations of polyenic antibiotics, (up to 200 mcg/ml) results in a slight inhibition of the enzyme activity. The role of the data obtained for solving molecular basis of the toxic effect of polyenic antibiotics.     

The evaluation of the age-related characteristics of the structural status of the plasma membrane lipid phase in rat fatty tissue by using the method of inductive-resonance energy transfer

Using the method of inductance-resonance energy transfer from tryptophanyl residues to fluorescent pyrene probe the structural state of plasmatic membranes from adipose tissue of different age rats has been studied. The structural heterogeneity of membrane lipid phase has been revealed. The differences in physical properties of annular and bilayer lipids don't depend on age. During aging the membrane lipid viscosity including lipids of near protein area decreases, the conformation of membrane protein components alters during aging as well. The data on various effectiveness of energy transfer from tryptophanyls to pyrene probe in young and aged animals with stable polypeptide composition of membrane proteins indicates that. The structure of membrane lipid phase is suggested to be the main factor affecting the conformational state and functional activity of membrane-bound proteins during aging.     

The interactions of bilirubin with model and biological membranes

The partitioning of bilirubin between albumin and model and biological membranes and the differential partitioning of bilirubin between membranes with different lipid and protein compositions were measured. Partition coefficients were independent of the concentration of bilirubin in membranes up to at least 7 mol of bilirubin/mol of phospholipid. The avidity of albumin for bilirubin was greater than that of membranes, but the avidity of the latter for bilirubin depended on the composition of the membrane. Bilirubin partitioned preferentially into model membranes comprised of microsomal lipids greater than dioleoylphosphatidylcholine = plasma membrane lipids much greater than egg phosphatidylcholine = dimyristoylphosphatidylcholine. Partitioning into membranes was increased if these contained proteins, but the effect of proteins could not be attributed to specific binding to sites on proteins, as reflected by the temperature independence of partition coefficients. Differential partitioning of bilirubin into different membranes of pure lipids also was independent of temperature. Differences in the bulk phase fluidity of membranes does not appear to account for the preferential partitioning of bilirubin into some membranes. It appears that bilirubin partitions into elements of free volume of differing sizes in membranes with variable lipid compositions and that the size of these elements can be increased by adding proteins to membranes.     

Thermotropic lipid phase transition and the behavior of hydrolytic enzymes in the kidney cortex brush border membrane

Functional interactions of lipids and proteins were examined in brush-border membranes isolated from the kidney cortex by studying the temperature dependence of the hydrolytic enzyme activities. A close relationship was observed for the membrane proteins and the thermotropic lipid phase transitions. Three lines of evidences were provided for such dependence: a) Arrhenius relationship of the membrane-bound enzyme activities, and the effect of temperature in native and partially delipidated membranes, b) differential scanning calorimetric study of the membrane lipid phase transitions in the native and delipidated membranes, multilamellar vesicles prepared from the membrane extracted lipids, and in vesicles from dimyristoyl phosphatidylcholine, and c) the excimer (dimer)-formation studies of the membrane extrinsic fluorescent probe, pyrene, and the resultant membrane microviscosity. The brush-border membranes were partially delipidated with BuOH and 2,2,2-trifluoroethanol. The functional interactions of the delipidated membranes, which were greatly lost on lipid removal, were largely restored by the addition of exogenous lipids in the reconstitution process, which indicate the critical dependence of the membrane integral proteins on the neighboring lipid molecules in the bulk lipid phase.     

Ultrastructure and functions of the cells of proximal and distal segments of the nephron of anadromous and freshwater fishes]

In the carp kidney, the proximal tubule cells have no infoldings of the basilar plasmatic membrane. In the Salmonidae nephrons the numerous parallel membranes of proximal tubule cell are formed by foldings of the basilar plasmatic membrane with numerous mitochondria in between. In O. nerka and S. malma experiments with MgCl2 injection showed that their kidneys secrete magnesium in contrast to the carp kidney. The numerous invaginations of proximal tubule cells membrane are suggested to be a morphological equivalent of the Na/Mg exchange mechanism.     

Lipid Sorting and Multivesicular Endosome Biogenesis

Intracellular organelles, including endosomes, show differences not only in protein but also in lipid composition. It is becoming clear from the work of many laboratories that the mechanisms necessary to achieve such lipid segregation can operate at very different levels, including the membrane biophysical properties, the interactions with other lipids and proteins, and the turnover rates or distribution of metabolic enzymes. In turn, lipids can directly influence the organelle membrane properties by changing biophysical parameters and by recruiting partner effector proteins involved in protein sorting and membrane dynamics. In this review, we will discuss how lipids are sorted in endosomal membranes and how they impact on endosome functions.It is now well established that membranes along the endocytic and secretory pathway show differences not only in protein but also in lipid composition. For example, lipid gradients exist along the biosynthetic pathway with increasing density of cholesterol and sphingolipids from the endoplasmic reticulum (ER) to the plasma membrane (Maxfield and van Meer 2010). Also, phosphoinositides show distributions restricted to relatively well-characterized membrane territories (Di Paolo and De Camilli 2006). Given the facts that lipids are small and contain little structural information when compared with proteins, that they can diffuse rapidly within membranes, and that membranes are connected by membrane flow during transport, it is not always obvious how different lipids are segregated from each other.In this article, we will evoke different mechanisms that may contribute to the heterogeneous lipid composition of endocytic membranes, including physicochemical properties of the membrane, interactions with other proteins or lipids, and synthesis or degradation. In addition, it has also become apparent that peripheral membrane proteins often interact with membranes via diverse lipid-binding motifs, and thus that lipids directly contribute to the distribution of many peripheral membrane proteins. For example, phosphatidylinositol 3-phosphate (PI(3)P) is detected predominantly on early endosomes, where most characterized PI(3)P-binding proteins encoded by the human genome are found as well (Raiborg et al. 2013). We will also discuss how some lipids may regulate protein sorting and membrane transport within the endosomal system.     

Randomly organized lipids and marginally stable proteins: A coupling of weak interactions to optimize membrane signaling

Eukaryotic lipids in a bilayer are dominated by weak cooperative interactions. These interactions impart highly dynamic and pliable properties to the membrane. C2 domain-containing proteins in the membrane also interact weakly and cooperatively giving rise to a high degree of conformational plasticity. We propose that this feature of weak energetics and plasticity shared by lipids and C2 domain-containing proteins enhance a cell's ability to transduce information across the membrane. We explored this hypothesis using information theory to assess the information storage capacity of model and mast cell membranes, as well as differential scanning calorimetry, carboxyfluorescein release assays, and tryptophan fluorescence to assess protein and membrane stability. The distribution of lipids in mast cell membranes encoded 5.6–5.8 bits of information. More information resided in the acyl chains than the head groups and in the inner leaflet of the plasma membrane than the outer leaflet. When the lipid composition and information content of model membranes were varied, the associated C2 domains underwent large changes in stability and denaturation profile. The C2 domain-containing proteins are therefore acutely sensitive to the composition and information content of their associated lipids. Together, these findings suggest that the maximum flow of signaling information through the membrane and into the cell is optimized by the cooperation of near-random distributions of membrane lipids and proteins. This article is part of a Special Issue entitled: Interfacially Active Peptides and Proteins. Guest Editors: William C. Wimley and Kalina Hristova.     

Features of lung tissue cell membrane lipid composition under acute emotional stress in rats

The lipid composition of the lung plasmatic membrane in rats which have been under the acute emotional pain stress action is studied. These results are compared with the control group of animals. It is shown that at acute stress the changes of lipid composition of the lung plasmatic membranes are manifested in decrease the phospholipids and increase of cholesterol levels. The correlation of phospholipids/cholesterol in plasmic membranes in the lungs decreases at stress. At the same time the decrease of triglyceroles and diglyceroles contents is observed as well as the increase of fat acids' number. The changes that take place in the lipid contents of the lung plasmatic membranes at acute stress can play an essential role in the mechanism of cell damage development.     

Identification of P-glycoprotein in renal brush border membranes

A monoclonal antibody (C219) that recognizes the P-glycoprotein (Mr = 170,000) in plasma membranes of multidrug-resistant Chinese hamster ovary (CHO) cell lines was used to assay renal brush border membrane (BBM) and basolateral membrane (BLM) fractions for the presence of a cross-reactive polypeptide. The C219 antibody bound to a 155,000 dalton protein in immunoblots of rat BBM but not BLM proteins resolved by sodium dodecyl sulfate gel electrophoresis. The corresponding human kidney BBM and dog kidney BBM proteins had molecular weights of 170,000 and 160,000 respectively. The glycoprotein nature of the renal protein was shown by its sensitivity to N-glycanase treatment which reduced the apparent molecular weight of the dog protein to 120,000. In addition, dog P-glycoprotein could be bound to and eluted from immobilized wheat germ agglutinin. The molecular weight, antibody crossreactivity, glycosidase sensitivity and lectin binding show that this protein is a normal kidney analogue of the P-glycoprotein induced in multidrug resistant cell lines.     

Modulation of entry of enveloped viruses by cholesterol and sphingolipids (Review)

Enveloped animal viruses infect host cells by fusion of viral and target membranes. This crucial fusion event occurs either with the plasma membrane of the host cells at the physiological pH or with the endosomal membranes at low pH and is triggered by specific glycoproteins in the virus envelope. Both lipids and proteins play critical and co-operative roles in the fusion process. Interactions of viral proteins with their receptors direct which membranes fuse and viral fusion proteins then drive the process. These fusion proteins operate on lipid assemblies, whose physical and mechanical properties are equally important to the proper functioning of the process. Lipids contribute to the viral fusion process by virtue of their distinct chemical structure, composition and/or their preferred partitioning into specific microdomains in the plasma membrane called 'rafts'. An involvement of lipid rafts in viral entry and membrane fusion has been examined recently. However, the mechanism(s) by which lipids as dynamic raft components control viral envelope-glycoprotein-triggered fusion is not clear. This paper will review literature findings on the contribution of the two raft-associated lipids, cholesterol and sphingolipids in viral entry.     

Modulation of entry of enveloped viruses by cholesterol and sphingolipids (Review)

Enveloped animal viruses infect host cells by fusion of viral and target membranes. This crucial fusion event occurs either with the plasma membrane of the host cells at the physiological pH or with the endosomal membranes at low pH and is triggered by specific glycoproteins in the virus envelope. Both lipids and proteins play critical and co-operative roles in the fusion process. Interactions of viral proteins with their receptors direct which membranes fuse and viral fusion proteins then drive the process. These fusion proteins operate on lipid assemblies, whose physical and mechanical properties are equally important to the proper functioning of the process. Lipids contribute to the viral fusion process by virtue of their distinct chemical structure, composition and/or their preferred partitioning into specific microdomains in the plasma membrane called 'rafts'. An involvement of lipid rafts in viral entry and membrane fusion has been examined recently. However, the mechanism(s) by which lipids as dynamic raft components control viral envelope-glycoprotein-triggered fusion is not clear. This paper will review literature findings on the contribution of the two raft-associated lipids, cholesterol and sphingolipids in viral entry.     

Effects of 2-hydroxyoleic acid on the structural properties of biological and model plasma membranes

Genetic hypertension is associated with alterations in lipid metabolism, membrane lipid composition and membrane-protein function. 2-Hydroxyoleic acid (2OHOA) is a new antihypertensive molecule that regulates the structure of model membranes and their interaction with certain peripheral signalling proteins in vitro. While the effect of 2OHOA on elevated blood pressure is thought to arise through its influence on signalling proteins, its effects on membrane lipid composition remain to be assessed. 2OHOA administration altered the lipid membrane composition of hypertensive and normotensive rat plasma membranes, and increased the fluidity of reconstituted liver membranes from hypertensive rats. In spontaneously hypertensive rats (SHR), treatment with 2OHOA increased the cholesterol and sphingomyelin content while decreasing that of phosphatidylserine-phosphatidylinositol lipids. In addition, monounsaturated fatty acid levels increased as well as the propensity of reconstituted membranes to form HII-phases. These data suggest that 2OHOA regulates lipid metabolism that is altered in hypertensive animals, and that it affects the structural properties of liver plasma membranes in SHR. These changes in the structural properties of the plasma membrane may modulate the activity of signalling proteins that associate with the cell membrane such as the Galphaq/11 protein and hence, signal transduction.     

Influence of lipid chain unsaturation on membrane-bound melittin: a fluorescence approach

Melittin, a cationic hemolytic peptide, is intrinsically fluorescent due to the presence of a single functionally important tryptophan residue. The organization of membrane-bound melittin is dependent on the physical state and composition of membranes. In particular, polyunsaturated lipids have been shown to modulate the membrane-disruptive action of melittin. Phospholipids with polyunsaturated acyl chains are known to modulate a number of physical properties of membranes and play an important role in regulating structure and function of membrane proteins. In this study, we have used melittin to address the influence of unsaturated lipids in modulating lipid-protein interactions. Our results show that fluorescence parameters such as intensity, emission maximum, polarization, lifetime and acrylamide quenching of melittin incorporated in membranes are dependent on the degree of unsaturation of lipids in membranes. Importantly, melittin in membranes composed of various unsaturated lipids shows red edge excitation shift (REES) implying that melittin is localized in a motionally restricted region in membranes. The extent of REES was found to increase drastically in membranes with increasing unsaturation, especially when the lipids contained more than two double bonds. In addition, increasing unsaturation in membranes causes a considerable change in the secondary structure of membrane-bound melittin. Taken together, our results assume significance in the overall context of the role of unsaturated lipids in membranes in the organization and function of membrane proteins and membrane-active peptides.     

Sec14 related proteins in yeast

Lipid transport between membranes of eukaryotic organisms represents an essential aspect of organelle biogenesis. This transport must be strictly selective and directional to assure specific lipid composition of individual membranes. Despite the intensive research effort in the last few years, our understanding of how lipids are sorted and moved within cells is still rather limited. Evidence indicates that at least some of the mechanisms generating and maintaining non-random distribution of lipids in cells are linked to the action of phosphatidylinositol transfer proteins (PITPs). The major PITP in yeast Saccharomyces cerevisiae, Sec14p, is essential in promoting Golgi secretory function by modulating of its membrane lipid composition. This review focuses on a group of five yeast proteins that share significant sequence homology with Sec14p. Based on this sequence identity, they were termed Sfh (Sec fourteen homologue) proteins. It is a diverse group of proteins with distinct subcellular localizations and varied physiological functions related to lipid metabolism, phosphoinositide mediated signaling and membrane trafficking.     

Effects of 2-hydroxyoleic acid on the structural properties of biological and model plasma membranes

Genetic hypertension is associated with alterations in lipid metabolism, membrane lipid composition and membrane-protein function. 2-Hydroxyoleic acid (2OHOA) is a new antihypertensive molecule that regulates the structure of model membranes and their interaction with certain peripheral signalling proteins in vitro. While the effect of 2OHOA on elevated blood pressure is thought to arise through its influence on signalling proteins, its effects on membrane lipid composition remain to be assessed. 2OHOA administration altered the lipid membrane composition of hypertensive and normotensive rat plasma membranes, and increased the fluidity of reconstituted liver membranes from hypertensive rats. In spontaneously hypertensive rats (SHR), treatment with 2OHOA increased the cholesterol and sphingomyelin content while decreasing that of phosphatidylserine-phosphatidylinositol lipids. In addition, monounsaturated fatty acid levels increased as well as the propensity of reconstituted membranes to form H_II-phases. These data suggest that 2OHOA regulates lipid metabolism that is altered in hypertensive animals, and that it affects the structural properties of liver plasma membranes in SHR. These changes in the structural properties of the plasma membrane may modulate the activity of signalling proteins that associate with the cell membrane such as the Gαq/11 protein and hence, signal transduction.     

Membrane fusion: a structural perspective on the interplay of lipids and proteins

The fusion of biological membranes is governed by the carefully orchestrated interplay of membrane proteins and lipids. Recently determined structures of fusion proteins, individual domains of fusion proteins and their complexes with regulatory proteins and membrane lipids have yielded much suggestive insight into how viral and intracellular membrane fusion might proceed. These structures may be combined with new knowledge on the fusion of pure lipid bilayer membranes in an attempt to begin to piece together the complex puzzle of how biological membrane fusion machines operate on membranes.