Membrane rafts, domains

Recently, due to increased interest in membrane raft domains, extensive attention has been paid to the cholesterol-dependent liquid-ordered phase in the membrane (Subczynski and Kusumi 2003). The pulse EPR spin-labeling DOT method detected two coexisting phases in the DMPC/cholesterol membranes the liquid-ordered and the liquid-disordered domains above the phase-transition temperature (Subczynski et al. 2007b). However, using the same method for DMPC/lutein (zeaxanthin) membranes, only the liquid-ordered-like phase was detected above the phase-transition temperature (Widomska, Wisniewska, and Subczynski, unpublished data). No significant differences were found in the effects of lutein and zeaxanthin on the lateral organization of lipid bilayer membranes. We can conclude that lutein and zeaxanthin—macular xanthophylls that parallel cholesterol in its function as a regulator of both membrane fluidity and hydrophobicity—cannot parallel the ability of cholesterol to induce liquid-ordered-disordered phase separation. 

FIGURE 10.12 The mole ratio of carotenoid/phospholipid and carotenoid/total lipid (phospholipid + cholesterol) in raft domain (detergent-resistant membrane, DRM) and bulk domain (detergent-soluble membrane, DSM) isolated from membranes made of raft-forming mixture (equimolar ternary mixture of dioleoyl-PC (DOPC)/sphingomyelin/cholesterol) with 1 mol% lutein (LUT), zeaxanthin (ZEA), P-cryptoxanthin (P-CXT), or P-carotene (P-CAR). 

FIGU RE 10.13 Schematic drawing of the distribution of xanthophyll molecules between raft domain (DRM) and bulk domain (DSM) in lipid bilayer membranes. For this illustration, the xanthophyll partition coefficient between domains is the same as obtained experimentally for raft-forming mixture. However, to better visualize the observed effect in the drawing, the number of lipid molecules was decreased and the total number of xanthophyll molecules was increased about 10 times. (From Wisniewska, A. and Subczynski, W.K., Free Radio. Biol. Med., 40, 1820, 2006. With permission.)... 

Despite the weakness and short-range nature of protein-lipid and lipid-lipid interactions, cells have nevertheless evolved means of laterally assembling into membrane-mi-crodomains. Sphingolipid-cholesterol rafts serve to recmit a specific set of membrane proteins and exclude others [24]. Caveolae are deeply invaginated raft domains that are stabilized by caveolin protein oligomers (binding cholesterol) [25]. 

Targeting of proteins to specialized domains of a membrane are less well understood. These include caveolae and lipid rafts, domains that are high in cholesterol and sphingolipids and which function in endocytosis and in cell signaling. A recent proposal is that proteins with hydrophobic surfaces needed in these domains become coated with a lipid "shell" before entering the membrane.6173... 

Lipid raft domains of plasma membranes are enriched in cholesterol and sphingolipids. As a consequence, compounds that extract or sequester cholesterol, such as fS-cyclodextrins, nystatin, and filipin, can block selectively endocytosis of cholera toxin, GPI-linked proteins, and other receptors that associate with lipid rafts and caveolae. However, cholesterol is also critical for CME, secretion of proteins, and the actin network. Therefore, conditions designed to affect selectively raft-mediated endocytosis by perturbing cholesterol levels must be carefully controlled to avoid disrupting other mechanisms of endocytosis (40). 

Apart of forming the bilayer, membrane lipids exhibit dynamic structures within the lamellas, forming microdomains with specific functionalities. The so called membrane rafts are sphingolipid-cholesterol domains that contribute to signal transduction, as well as to lipid and protein sorting and transport [18]. 

In order to understand the complex behavior of cellular membranes and their response to external perturbations Uke electric flelds, one has to elucidate the basic mechanical properties of the lipid bilayer. The signiflcant expansion in recent years of the field of membrane raft-hke domain formation [11, 132, 133] imposes the compelling need for understanding the effect of hpid bilayer composition on membrane properties. Cholesterol, a ubiquitous species in eukaryotic membranes, is an important component in raft-hke domains in cells and in vesicles, which mohvates studies aimed at understanding its influence on the mechanical properties and stability of membranes. 

Mendez AI, Lin G, Wade DP, et al. (2001) Membrane lipid domains distinct from cholesterol/sphingomyelin-rich rafts are involved in the ABCAl-mediated lipid secretory pathway. J Biol Chem 276 3158-3166... 

Recent studies suggest that detergent-resistant subdomains of the plasma membrane ( rafts , discussed previously) may be involved in formation of PrP . Consistent with this idea, both PrP and PrP are found in raft domains isolated biochemically (Gorodinsky and Harris, 1995 Taraboulos et al, 1995 Naslavsky et al., 1996 Vey et al., 1996 Naslavsky et al., 1999). In addition, pharmacological depletion of cellular cholesterol, which is known to disrupt rafts, inhibits PrP formation (Taraboulos et al, 1995), whereas sphingolipid depletion, which does not alter the raft localization of PrP, actually enhances PrP production (Naslavsky et al, 1999). Finally, artificially constructed transmembrane forms of PrP, which are excluded from rafts, are poor substrates for conversion to PrP (Kaneko et al., 1997). 

Our kinetic studies of mutant PrPs synthesized in CHO cells suggest that individual steps in formation of PrP may take place in at least two different cellular locations (Fig. 5). Because mutant PrPs become PIPLC-resistant within minutes of synthesis in pulse-labeling experiments, this early step must take place in the ER. Consistent with this conclusion, acquisition of PIPLC resistance is not affected by treatment of cells with brefeldin A or by incubation at 18°C, manipulations that block exit of proteins beyond the Golgi (Daude et al, 1997). In contrast, detergent insolubility and protease resistance, which do not develop until later times of chase, and are reduced by brefeldin A and 18°C incubation, are likely to be acquired after arrival of the protein at the cell surface, either on the plasma membrane itself or in endocytic compartments. Raft domains may be involved in these changes (unpublished data). 

The model reproduces the most prominent phase transitions of phospholipid monolayers [78] and bilayers [80]. In particular, it reproduces a main transitirm from a fluid membrane phase (L to a tilted gel phase Lpi) with an intermediate ripple phase Pp ), in agreement with experiments. The elastic parameters have been studied in the fluid phase and are in reasonable agreement with those of saturated DPPC (dipalmitoyl-phosphatidylcholine) bilayers. Recently, the Lenz model has been supplemented with a simple cholesterol model [81]. Cholesterol molecules are taken to be shorter and stiffer than lipids, and they have a slight affinity to lipids. Mixtures of lipids and cholesterol were found to develop nanoscale raft domains... 

Early descriptions of lipid rafts noted their enrichment in cholesterol and glycosph-ingolipids and focused on their ability to resist extraction by nonionic detergents [32]. Later experiments showed that lipid rafts were a heterogeneous collection of domains that differ in protein and lipid composition as well as in temporal stability [33, 34]. The distinctive lipid composition of membrane rafts as demonstrated by lipidomics makes a clearer picture of membrane rafts. 

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