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1.
This review will discuss the use of small-angle X-ray diffraction approaches to study the organization of lipids in plasma membranes derived from two distinct mammalian cell types: arterial smooth muscle cells and ocular lens fiber cells. These studies indicate that cholesterol at an elevated concentration can self-associate and form immiscible domains in the plasma membrane, a phenomenon that contributes to both physiologic and pathologic cellular processes, depending on tissue source. In plasma membrane samples isolated from atherosclerotic smooth muscle cells, the formation of sterol-rich domains is associated with loss of normal cell function, including ion transport activity and control of cell replication. Analysis of meridional diffraction patterns from intact and reconstituted plasma membrane samples indicates the presence of an immiscible cholesterol domain with a unit cell periodicity of 34 A, consistent with a cholesterol monohydrate tail-to-tail bilayer, under disease conditions. These cholesterol domains were observed in smooth muscle cells enriched with cholesterol in vitro as well as from cells obtained ex vivo from an animal model of atherosclerosis. By contrast, well-defined cholesterol domains appear to be essential to the normal physiology of fiber cell plasma membranes of the human ocular lens. The organization of cholesterol into separate domains underlies the role of lens fiber cell plasma membranes in maintaining lens transparency. These domains may also interfere with cataractogenic aggregation of soluble lens proteins at the membrane surface. Taken together, these analyses provide examples of both physiologic and pathologic roles that sterol-rich domains may have in mammalian plasma membranes. These findings support a model of the membrane in which cholesterol aggregates into structurally distinct regions that regulate the function of the cell membrane. 相似文献
2.
Direct evidence for immiscible cholesterol domains in human ocular lens fiber cell plasma membranes.
The molecular structure of human ocular lens fiber cell plasma membranes was examined directly using small angle x-ray diffraction approaches. A distinct biochemical feature of these membranes is their high relative levels of free cholesterol; the mole ratio of cholesterol to phospholipid (C/P) measured in these membranes ranges from 1 to 4. The organization of cholesterol in this membrane system is not well understood, however. In this study, the structure of plasma membrane samples isolated from nuclear (3.3 C/P) and cortical (2.4 C/P) regions of human lenses was evaluated with x-ray diffraction approaches. Meridional diffraction patterns obtained from the oriented membrane samples demonstrated the presence of an immiscible cholesterol domain with a unit cell periodicity of 34.0 A, consistent with a cholesterol monohydrate bilayer. The dimensions of the sterol-rich domains remained constant over a broad range of temperatures (5-20 degrees C) and relative humidity levels (31-97%). In contrast, dimensions of the surrounding sterol-poor phase were significantly affected by experimental conditions. Similar structural features were observed in membranes reconstituted from fiber cell plasma membrane lipid extracts. The results of this study indicate that the lens fiber cell plasma membrane is a complex structure consisting of separate sterol-rich and -poor domains. Maintenance of these separate domains may be required for the normal function of lens fiber cell plasma membrane and may interfere with the cataractogenic aggregation of soluble lens proteins at the membrane surface. 相似文献
3.
Previous studies in our laboratory have provided direct evidence for the existence of distinct cholesterol domains within the plasma membranes of human ocular lens fiber cells. The fiber cell plasma membrane is unique in that it contains unusually high concentrations of cholesterol, with cholesterol to phospholipid (C/P) mole ratios ranging from 1 to 4. Since membrane cholesterol content is disturbed in the development of cataracts, it was hypothesized that perturbation of cholesterol domain structure occurs in cataracts. In this study, fiber cell plasma membranes were isolated from both normal (control) and cataractous lenses and assayed for cholesterol and phospholipid. Control and cataractous whole lens membranes had C/P mole ratios of 3.1 and 1.7, respectively. Small angle x-ray diffraction approaches were used to directly examine the structural organization of the cataractous lens plasma membrane versus control. Both normal and cataractous oriented membranes yielded meridional diffraction peaks corresponding to a unit cell periodicity of 34.0 A, consistent with the presence of immiscible cholesterol domains. However, comparison of diffraction patterns indicated that cataractous lens membranes contained more pronounced and better defined cholesterol domains than controls, over a broad range of temperature (5-40 degrees C) and relative humidity (52-92%) levels. In addition, diffraction analyses of the sterol-poor regions of cataractous membranes indicated increased membrane rigidity as compared with control membranes. Modification of the membrane lipid environment, such as by oxidative insult, is believed to be one potential mechanism for the formation of highly resolved cholesterol domains despite significantly reduced cholesterol content. The results of this x-ray diffraction study provide evidence for fundamental changes in the lens fiber cell plasma membrane structure in cataracts, including the presence of more prominent and highly ordered, immiscible cholesterol domains. 相似文献
4.
Specialized membrane domains containing lipid rafts are thought to be important for membrane processes such as signaling and trafficking. An unconventional type I myosin has been shown to reside in lipid rafts and function to target a disaccharidase to rafts in brush borders of intestinal mammalian cells. In the fission yeast Schizosaccharomyces pombe, distinct sterol-rich membrane domains are formed at the cell division site and sites of polarized cell growth at cell tips. Here, we show that the sole S. pombe myosin I, myo1p, is required for proper organization of these membrane domains. myo1 mutants lacking the TH1 domain exhibit a uniform distribution of sterol-rich membranes all over the plasma membrane throughout the cell cycle. These effects are independent of endocytosis because myo1 mutants exhibit no endocytic defects. Conversely, overexpression of myo1p induces ectopic sterol-rich membrane domains. Myo1p localizes to nonmotile foci that cluster in sterol-rich plasma membrane domains and fractionates with detergent-resistant membranes. Because the myo1p TH1 domain may bind directly to acidic phospholipids, these findings suggest a model for how type I myosin contributes to the organization of specialized membrane domains. 相似文献
5.
Little is known about the lipid environment of lens fiber junctions, the plasma membrane structure proposed to be responsible for passage of low molecular weight metabolites between adjacent lens fiber cells. Plasma membranes of the ocular lens are especially rich in fiber junctions. The resistance of junctional domains to disruption by detergent or alkali treatment provides the opportunity to isolate a lens plasma membrane fraction enriched in fiber junctions. When examined by electron microscopy, the fiber junction fraction prepared from bovine lenses was enriched with junctional structures by about twofold when compared to total plasma membrane. We compared the protein, phospholipid, and cholesterol concentration of total plasma membrane with fiber junctional membrane from rat and cow lens and from aged normal cataractous human lenses. The principal finding was that junctional membrane contained 20-40% more total lipid than that of the total plasma membrane. This was due to a proportionate increase in the relative content (mg/mg protein) of both phospholipid and cholesterol. Exclusive of one exception (nucleus of bovine lens), the cholesterol/phospholipid molar ratios of the two fractions were similar. In the bovine nucleus, the cholesterol/phospholipid molar ratio was substantially higher in the fiber junctional-enriched membrane fraction than in the total plasma membrane, suggesting a special association of cholesterol with bovine nuclear fiber junctions. The relative lipid compositions of the plasma membrane and fiber junction-enriched fractions from human normal and cataractous lenses were similar, suggesting that human senile cataractogenesis involves changes in the lens plasma membrane more subtle than would be reflected by gross changes in the membrane lipid composition. 相似文献
6.
Jessica F. Frisz Haley A. Klitzing Kaiyan Lou Ian D. Hutcheon Peter K. Weber Joshua Zimmerberg Mary L. Kraft 《The Journal of biological chemistry》2013,288(23):16855-16861
The plasma membranes of mammalian cells are widely expected to contain domains that are enriched with cholesterol and sphingolipids. In this work, we have used high-resolution secondary ion mass spectrometry to directly map the distributions of isotope-labeled cholesterol and sphingolipids in the plasma membranes of intact fibroblast cells. Although acute cholesterol depletion reduced sphingolipid domain abundance, cholesterol was evenly distributed throughout the plasma membrane and was not enriched within the sphingolipid domains. Thus, we rule out favorable cholesterol-sphingolipid interactions as dictating plasma membrane organization in fibroblast cells. Because the sphingolipid domains are disrupted by drugs that depolymerize the cells actin cytoskeleton, cholesterol must instead affect the sphingolipid organization via an indirect mechanism that involves the cytoskeleton. 相似文献
7.
Petri Sane Susanne K. Wiedmer Ilpo Vattulainen Juha M. Holopainen 《生物化学与生物物理学报:生物膜》2010,1798(5):958-965
The function of mammalian ocular lens is to provide a sharp image to the retina. Accordingly, the lens needs to be transparent and minimize light scattering. To do so the lens fiber cells first loose intracellular organelles, organize the cytoplasm and arrange the fiber cell membranes. Because the fiber cells are metabolically inactive, the plasma membrane becomes the only cellular organelle and consequently, the phase behavior of these membranes determines the physiological state of the lens. Previous studies have shown that lipids extracted from the nuclear and cortical region of human lens show a temperature-induced phase transition close to the body temperature. Yet, the physiological function of this phase transition is not known, and even the presence of the phase transition in intact lenses is unknown. Positron annihilation lifetime spectroscopy (PALS) was used to characterize the sub-nanometer-sized local structure of intact porcine lens and these studies were complemented with differential scanning calorimeter and mass spectrometric analysis in extracted porcine lens lipids. Using PALS, we present evidence for the presence of a temperature-dependent structural transition centered at 35.5 °C in-situ in clear extracted porcine lenses. Further studies employing extracted lens lipids and purified egg-yolk sphingomyelin and cholesterol mixtures suggest that the nano-scale transition emerges from the phase behavior of lens lipids. Based on our results, PALS seems to be a viable method for gaining additional information on biological tissues, especially since it enables non-destructive studies on intact tissues. 相似文献
8.
Transparency of the ocular lens depends on symmetric packing and membrane organization of highly elongated hexagonal fiber cells. These cells possess an extensive, well-ordered cortical cytoskeleton to maintain cell shape and to anchor membrane components. Periaxin (Prx), a PDZ domain protein involved in myelin sheath stabilization, is also a component of adhaerens plaques in lens fiber cells. Here we show that Prx is expressed in lens fibers and exhibits maturation dependent redistribution, clustering discretely at the tricellular junctions in mature fiber cells. Prx exists in a macromolecular complex with proteins involved in membrane organization including ankyrin-B, spectrin, NrCAM, filensin, ezrin and desmoyokin. Importantly, Prx knockout mouse lenses were found to be softer and more easily deformed than normal lenses, revealing disruptions in fiber cell hexagonal packing, membrane skeleton and membrane stability. These observations suggest a key role for Prx in maturation, packing, and membrane organization of lens fiber cells. Hence, there may be functional parallels between the roles of Prx in membrane stabilization of the myelin sheath and the lens fiber cell. 相似文献
9.
Swine vascular smooth muscle cells were exposed to homologous low-density or high-density lipoprotein fractions for 24 h. Total cell membranes were isolated from the post-nuclear supernatant of the cell homogenates, fractionated by sucrose density gradient centrifugation and characterized by enzyme assays. The membrane fraction with the lowest density was enriched in plasma membrane marker enzymes. Cholesterol analysis showed that cells exposed to low-density lipoprotein had higher cholesterol-to-protein ratios in total cells, total cell membranes and individual membrane fractions than had the cells exposed to high-density lipoproteins. Cholesterol-to-phospholipid ratios of the plasma membrane-enriched fraction from cells exposed to low-density lipoprotein were higher than the same membrane fraction of cells exposed to high-density lipoprotein. Studies with iodinated lipoproteins showed that these compositional changes could not be due to lipoprotein contamination. Membrane microviscosity was determined by fluorescence depolarization with diphenylhexatriene and the microviscosity of the plasma membrane-enriched fraction was different in the cells exposed to the two different lipoprotein fractions. This difference in membrane microviscosity was significant only when the medium cholesterol content was 40 μg per ml or greater; cells exposed to low-density lipoprotein gave membranes with higher microviscosity.These results demonstrate that the properties of vascular smooth muscle cell membranes are influenced by exposure of the cells to homologous lipoprotein fractions. 相似文献
10.
C Wolf P Quinn K Koumanov C Chachaty B Tenchov 《Journal de la Société de Biologie》1999,193(2):117-123
Detection of immiscible lipid domains in biological membranes offers an alternative support to protein sorting. Liquid ordered domains ("rafts") comprising cholesterol and saturated sphingolipids incorporate saturated glycosyl-phosphatidylinositol (GPI)-anchored or acylated (palmitoyl- and myristoyl-) proteins or particular transmembrane protein sequences. These lipid domains can be isolated in the form of Detergent resistant membranes (DRM) from biological plasma membrane preparations. Caveolae appear to be a differentiated fraction of plasma membranes comprising such numerous cross-linked microdomains associated with caveolin in different cell types. While the biological relevance of such membrane domains is evidenced in vivo by co-patching of proteins sharing the identical affinity for sphingolipids and by the disruption of co-patching following cell cholesterol depletion, only a few physical studies confort the principle of membrane heterogeneity. Results are now presented where cholesterol addition in a tertiary lipid mixture forces outphase-separation, as a realistic model where the lipid segregation can promote protein sorting to the segregated Lo phase. A lipid mixture comprising phosphatidylserine, phosphatidylethanolamine and sphingomyelin of natural origin in the ratio (1/4/3: mole/mole) has been rendered neatly heterogeneous after the addition of cholesterol (27 mole%). Xray diffraction (Small angle Xray scattering) showed the splitting of two neatly resolved lamellar diffractions in the presence of cholesterol. Above 37 degrees C the heterogeneity was traceable by a broadened diffraction spot up to the complete get-to-liquid transition of sphingomyelin at temperatures > 40 degrees C where the spot became again symmetrical and narrow. The large temperature range where the immiscible lamellar phases are detected, the specific requirement for cholesterol association with sphingomyelin, the positive influence of calcium and the reversibility of domain formation support the occurrence for such domains at the inner side of the plasma membrane whereon lipids-bound proteins concentrate. 相似文献
11.
In various eukaryotes, sterol-rich membrane domains have been proposed to play an important role in polarization and compartmentalization of the plasma membrane. Several studies have reported the cellular distribution of sterols in genetically tractable yeast species and the identification of molecules that might regulate the localization of sterol-rich membrane domains. Here, we attempt to synthesize our understanding of the function and organization of these domains from the study of fungi and identify some outstanding issues. 相似文献
12.
Characterization of lipid rafts from Medicago truncatula root plasma membranes: a proteomic study reveals the presence of a raft-associated redox system 总被引:1,自引:0,他引:1
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Lefebvre B Furt F Hartmann MA Michaelson LV Carde JP Sargueil-Boiron F Rossignol M Napier JA Cullimore J Bessoule JJ Mongrand S 《Plant physiology》2007,144(1):402-418
Several studies have provided new insights into the role of sphingolipid/sterol-rich domains so-called lipid rafts of the plasma membrane (PM) from mammalian cells, and more recently from leaves, cell cultures, and seedlings of higher plants. Here we show that lipid raft domains, defined as Triton X-100-insoluble membranes, can also be prepared from Medicago truncatula root PMs. These domains have been extensively characterized by ultrastructural studies as well as by analysis of their content in lipids and proteins. M. truncatula lipid domains are shown to be enriched in sphingolipids and Delta(7)-sterols, with spinasterol as the major compound, but also in steryl glycosides and acyl-steryl glycosides. A large number of proteins (i.e. 270) have been identified. Among them, receptor kinases and proteins related to signaling, cellular trafficking, and cell wall functioning were well represented whereas those involved in transport and metabolism were poorly represented. Evidence is also given for the presence of a complete PM redox system in the lipid rafts. 相似文献
13.
Analysis of detergent-resistant membranes in Arabidopsis. Evidence for plasma membrane lipid rafts 总被引:1,自引:0,他引:1
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Borner GH Sherrier DJ Weimar T Michaelson LV Hawkins ND Macaskill A Napier JA Beale MH Lilley KS Dupree P 《Plant physiology》2005,137(1):104-116
The trafficking and function of cell surface proteins in eukaryotic cells may require association with detergent-resistant sphingolipid- and sterol-rich membrane domains. The aim of this work was to obtain evidence for lipid domain phenomena in plant membranes. A protocol to prepare Triton X-100 detergent-resistant membranes (DRMs) was developed using Arabidopsis (Arabidopsis thaliana) callus membranes. A comparative proteomics approach using two-dimensional difference gel electrophoresis and liquid chromatography-tandem mass spectrometry revealed that the DRMs were highly enriched in specific proteins. They included eight glycosylphosphatidylinositol-anchored proteins, several plasma membrane (PM) ATPases, multidrug resistance proteins, and proteins of the stomatin/prohibitin/hypersensitive response family, suggesting that the DRMs originated from PM domains. We also identified a plant homolog of flotillin, a major mammalian DRM protein, suggesting a conserved role for this protein in lipid domain phenomena in eukaryotic cells. Lipid analysis by gas chromatography-mass spectrometry showed that the DRMs had a 4-fold higher sterol-to-protein content than the average for Arabidopsis membranes. The DRMs were also 5-fold increased in sphingolipid-to-protein ratio. Our results indicate that the preparation of DRMs can yield a very specific set of membrane proteins and suggest that the PM contains phytosterol and sphingolipid-rich lipid domains with a specialized protein composition. Our results also suggest a conserved role of lipid modification in targeting proteins to both the intracellular and extracellular leaflet of these domains. The proteins associated with these domains provide important new experimental avenues into understanding plant cell polarity and cell surface processes. 相似文献
14.
Subczynski WK Raguz M Widomska J Mainali L Konovalov A 《The Journal of membrane biology》2012,245(1):51-68
The most unique feature of the eye lens fiber-cell plasma membrane is its extremely high cholesterol content. Cholesterol
saturates the bulk phospholipid bilayer and induces formation of immiscible cholesterol bilayer domains (CBDs) within the
membrane. Our results (based on EPR spin-labeling experiments with lens-lipid membranes), along with a literature search,
have allowed us to identify the significant functions of cholesterol specific to the fiber-cell plasma membrane, which are
manifest through cholesterol–membrane interactions. The crucial role is played by the CBD. The presence of the CBD ensures
that the surrounding phospholipid bilayer is saturated with cholesterol. The saturating cholesterol content in fiber-cell
membranes keeps the bulk physical properties of lens-lipid membranes consistent and independent of changes in phospholipid
composition. Thus, the CBD helps to maintain lens-membrane homeostasis when the membrane phospholipid composition changes
significantly. The CBD raises the barrier for oxygen transport across the fiber-cell membrane, which should help to maintain
a low oxygen concentration in the lens interior. It is hypothesized that the appearance of the CBD in the fiber-cell membrane
is controlled by the phospholipid composition of the membrane. Saturation with cholesterol smoothes the phospholipid-bilayer
surface, which should decrease light scattering and help to maintain lens transparency. Other functions of cholesterol include
formation of hydrophobic and rigidity barriers across the bulk phospholipid-cholesterol domain and formation of hydrophobic
channels in the central region of the membrane for transport of small, nonpolar molecules parallel to the membrane surface.
In this review, we provide data supporting these hypotheses. 相似文献
15.
Membrane cholesterol dynamics: cholesterol domains and kinetic pools 总被引:10,自引:0,他引:10
F Schroeder J R Jefferson A B Kier J Knittel T J Scallen W G Wood I Hapala 《Proceedings of the Society for Experimental Biology and Medicine. Society for Experimental Biology and Medicine (New York, N.Y.)》1991,196(3):235-252
Nonreceptor mediated cholesterol uptake and reverse cholesterol transport in cells occur through cellular membranes. Thus, elucidation of cholesterol dynamics in membranes is essential to understanding cellular cholesterol accumulation and loss. To this end, it has become increasingly evident that cholesterol is not randomly distributed in either model or biologic membranes. Instead, membrane cholesterol appears to be organized into structural and kinetic domains or pools. Cholesterol-rich and poor domains can even be observed histochemically and physically isolated from epithelial cell surface membranes. The physiologic importance of these domains is 2-fold: (i) Select membrane proteins (receptors, transporters, etc.) are localized in either cholesterol-rich or cholesterol-poor domains. Consequently, the structure and properties of the domains rather than of the bulk lipid may selectively affect the function of proteins residing therein. (ii) Kinetic evidence suggests that cholesterol transport through and between membranes may occur through specific domains or pools. Regulation of the size and properties of such domains may be controlling factors of cholesterol transport or accumulation in cells. Recent technologic advances in the use of fluorescent sterols have allowed examination of cholesterol domain structure in model and biologic membranes. These techniques have been applied to examine the role of high-density lipoprotein, cholesterol lowering drugs, and intracellular lipid transfer proteins in membrane sterol domain structure and sterol movement between membranes. 相似文献
16.
N-Acyl phosphatidylethanolamines are negatively charged phospholipids, which are naturally occurring albeit at low abundance. In this study, we have examined how the amide-linked acyl chain affected the membrane behavior of the N-acyl-1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylethanolamine (N-acyl-POPE) or N-acyl-dipalmitoyl-sn-glycero-3-phosphatidylethanolamine (N-acyl-DPPE), and how the molecules interacted with cholesterol. The gel-->liquid crystalline transition temperature of sonicated N-acyl phosphatidylethanolamine vesicles in water correlated positively with the number of palmitic acyl chains in the molecules. Based on diphenylhexatriene steady state anisotropy measurements, the presence of 33 mol% cholesterol in the membranes removed the phase transition from N-oleoyl-POPE bilayers, but failed to completely remove it from N-palmitoyl-DPPE and N-palmitoyl-POPE bilayers, suggesting rather weak interaction of cholesterol with the N-saturated NAPEs. The rate of cholesterol desorption from mixed monolayers containing N-palmitoyl-DPPE and cholesterol (1:1 molar ratio) was much higher compared to cholesterol/DPPE binary monolayers, suggesting a weak cholesterol interaction with N-palmitoyl-DPPE also in monolayers. In bilayer membranes, both N-palmitoyl-POPE and N-palmitoyl-DPPE failed to form sterol-rich domains, and in fact appeared to displace sterol from sterol/N-palmitoyl-sphingomyelin domains. The present data provide new information about the effects of saturated NAPEs on the lateral distribution of cholesterol in NAPE-containing membranes. These findings may be of relevance to neural cells which accumulate NAPEs during stress and cell injury. 相似文献
17.
Where examined, cholesterol is synthesized in the endoplasmic reticulum; however, its precursor, zymosterol, is found mostly in the plasma membrane. The novel implication of these disparate findings is that zymosterol circulates within the cell. In tracing its movements, we have now established the following: (a) in human fibroblasts, zymosterol is converted to cholesterol solely in the rough ER. (b) Little or no zymosterol or cholesterol accumulates in the rough ER in vivo. (c) Newly synthesized zymosterol moves to the plasma membrane without a detectable lag and with a half-time of 9 min, about twice as fast as cholesterol. (d) The pool of radiolabeled zymosterol in the plasma membrane turns over rapidly, faster than does intracellular cholesterol. Thus, plasma membrane zymosterol is not stagnant. (e) [3H]Zymosterol pulsed into intact cells is initially found in the plasma membrane. It is rapidly internalized and is then converted to [3H] cholesterol. Half of the [3H]cholesterol produced returns to the plasma membrane within 30 min of the initial [3H]zymosterol pulse. (f) Nascent zymosterol accumulates in a buoyant sterol-rich intracellular membrane before it reaches the plasma membrane. This membrane also acquires nascent cholesterol, exogenous [3H]zymosterol pulsed into intact cells, and [3H]cholesterol synthesized from the exogenous [3H] zymosterol. These results suggest that at least one sterol moves rapidly and in both directions among the rough endoplasmic reticulum, a sterol-rich intracellular membrane bearing nascent cholesterol, and the plasma membrane. 相似文献
18.
The spectrin-based membrane skeleton plays an important role in determining the distributions and densities of receptors, ion channels, and pumps, thus influencing cell shape and deformability, cell polarity, and adhesion. In the paradigmatic human erythrocyte, short tropomodulin-capped actin filaments are cross-linked by spectrin into a hexagonal network, yet the extent to which this type of actin filament organization is utilized in the membrane skeletons of nonerythroid cells is not known. Here, we show that associations of tropomodulin and spectrin with actin in bovine lens fiber cells are distinct from that of the erythrocyte and imply a very different molecular organization. Mechanical disruption of the lens fiber cell membrane skeleton releases tropomodulin and actin-containing oligomeric complexes that can be isolated by gel filtration column chromatography, sucrose gradient centrifugation and immunoadsorption. These tropomodulin-actin complexes do not contain spectrin. Instead, spectrin is associated with actin in different complexes that do not contain tropomodulin. Immunofluorescence staining of isolated fiber cells further demonstrates that tropomodulin does not precisely colocalize with spectrin along the lateral membranes of lens fiber cells. Taken together, our data suggest that tropomodulin-capped actin filaments and spectrin-cross-linked actin filaments are assembled in distinct structures in the lens fiber cell membrane skeleton, indicating that it is organized quite differently from that of the erythrocyte membrane skeleton. 相似文献
19.
Cholesterol does not induce segregation of liquid-ordered domains in bilayers modeling the inner leaflet of the plasma membrane
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A fluorescence-quenching method has been used to assess the potential formation of segregated liquid-ordered domains in lipid bilayers combining cholesterol with mixtures of amino and choline phospholipids like those found in the cytoplasmic leaflet of the mammalian cell plasma membrane. When present in proportions >20-30 mol %, different saturated phospholipids show a strong proclivity to form segregated domains when combined with unsaturated phospholipids and cholesterol, in a manner that is only weakly affected by the nature of the phospholipid headgroups. By contrast, mixtures containing purely unsaturated phospholipids and cholesterol do not exhibit detectable segregation of domains, even in systems whose components differ in headgroup structure, mono- versus polyunsaturation and/or acyl chain heterogeneity. These results indicate that mixtures of phospholipids resembling those found in the inner leaflet of the plasma membrane do not spontaneously form segregated liquid-ordered domains. Instead, our findings suggest that factors extrinsic to the inner-monolayer lipids themselves (e.g., transbilayer penetration of long sphingolipid acyl chains) would be essential to confer a distinctive, more highly ordered organization to the cytoplasmic leaflet of "lipid raft" structures in animal cell membranes. 相似文献
20.
In this study, we used cholestatrienol (CTL) as a fluorescent reporter molecule to study sterol-rich L(o) domains in complex lipid bilayers. CTL is a fluorescent cholesterol analog that mimics the behavior of cholesterol well. The ability of 12SLPC to quench the fluorescence of cholestatrienol gives a measure of the amount of sterol included in L(o) domains in mixed lipid membranes. The stability of sterol-rich domains formed in complex lipid mixtures containing saturated sphingomyelins, phosphatidylcholines, or galactosylceramide as potential domain-forming lipids were studied. The amount of sterol associated with sterol-rich domains seemed to always increase with increasing temperature. The quenching efficiency was highly dependent on the domain-forming lipid present in complex lipid mixtures. Sphingomyelins formed stable sterol-enriched domains and were able to shield CTL from quenching better than the other lipids included in this study. The saturated phosphatidylcholines also formed sterol-rich domains, but the quenching efficiency in membranes with these was higher than with sphingomyelins and the domains melted at lower temperatures. PGalCer was not able to form sterol-enriched domains. However, we found that PGalCer stabilized sterol-rich domains formed in PSM-containing bilayers. Using a fluorescent ceramide analog, we also demonstrated that N-palmitoyl-ceramide displaced the sterol from sphingolipid-rich domains in mixed bilayer membranes. 相似文献