Ceramide-Domain Formation and Collapse in Lipid Rafts: Membrane Reorganization by an Apoptotic Lipid

Abstract

The effect of physiologically relevant ceramide concentrations (< or = 4 mol %) in raft model membranes with a lipid composition resembling that of cell membranes, i.e., composed of different molar ratios of an unsaturated glycerophospholipid, sphingomyelin, and cholesterol (Chol) along a liquid-disordered-liquid-ordered tie line was explored. The application of a fluorescence multiprobe and multiparameter approach, together with multiple fluorescence resonance energy transfer (FRET) pairs, in the well-characterized palmitoyl-oleoyl-phosphocholine (POPC)/palmitoyl-sphingomyelin (PSM)/Chol ternary mixture, revealed that low palmitoyl-ceramide (PCer) concentrations strongly changed both the biophysical properties and lipid lateral organization of the ternary mixtures in the low-to-intermediate Chol/PSM-, small raft size range (<25 mol % Chol). For these mixtures, PCer recruited up to three PSM molecules for the formation of very small ( approximately 4 nm) and highly ordered gel domains, which became surrounded by rafts (liquid-ordered phase) when Chol/PSM content increased. However, the size of these rafts did not change, showing that PCer did not induce the formation of large platforms or the coalescence of small rafts. In the high Chol/PSM-, large raft domains range (>33 mol % Chol), Chol completely abolished the effect of PCer by competing for PSM association. Lipid rafts govern the biophysical properties and lateral organization in these last mixtures.

Enzymatic generation of ceramide induces membrane restructuring: Correlated AFM and fluorescence imaging of supported bilayers

“…These conclusions are consistent with reports that ceramide incorporation leads to displacement of cholesterol from ordered domains in both model membranes and cells (Ali et al, 2006;Chiantia et al, 2006Chiantia et al, , 2007Megha et al, 2006;Megha and London, 2004;Yu et al, 2005b) and with the reported ability of ceramide to substitute for cholesterol in liquid-ordered phases (Carrer et al, 2006;Silva et al, 2007). However, the correlated imaging results suggest that the membrane restructuring initiated by SMase may be more complex than simple loss of cholesterol from a ceramide-rich domain.…”

“…The importance of ceramide in modulating membrane physical properties and biological function has prompted a number of studies of ceramide incorporation in model membranes. Several recent investigations have examined the effects of ceramide incorporation on coexisting ordered and fluid domains in phase separated binary and ternary lipid mixtures that are used to model membrane rafts Fidorra et al, 2006;Silva et al, 2007;Sot et al, 2006Sot et al, , 2008. This work has demonstrated that ceramide leads to increased membrane heterogeneity and that its effects depend on the membrane composition.…”

“…These studies in monolayers and vesicles indicate that enzymatic generation of ceramide leads to more complex changes in membrane morphology than does direct ceramide incorporation in the same lipid membranes. Of particular relevance to the influence of ceramide on membrane raft domains are several recent studies that have compared direct and enzymatic generation of ceramide in vesicles and supported bilayers prepared from ternary lipid mixtures with coexisting ordered and fluid phases (Chiantia et al, 2006Johnston, 2006, 2008;Lopez-Mentero et al, 2007;Silva et al, 2007;Trajkovic et al, 2008). Atomic force microscopy (AFM) and fluorescence microscopy have shown that both direct and enzymatic incorporation of C 16 or C 18 ceramide causes formation of a new ceramide-rich ordered phase for various ternary lipid mixtures.…”

Sifuvirtide Screens Rigid Membrane Surfaces. Establishment of a Correlation between Efficacy and Membrane Domain Selectivity among HIV Fusion Inhibitor Peptides

“…Nevertheless, studies of sphingomyelin/cholesterol/ganglioside (or ceramide) ternary mixtures revealed the existence of gel-phase domains, rich in sphingomyelin and ganglioside (or ceramide), distinct and separated from the liquid-ordered sphingomyelin and cholesterol-rich domains. [54][55][56] The presence of GSL-binding domains on the V3 domain of HIV envelope protein gp120 is an evidence of the importance of GSL microdomains for the infection of HIV into the immune cells. 52,53 The affinity of the fusion inhibitor peptide T-1249 to the liquid-ordered rafts explains its improved efficacy relative to enfuvirtide.…”

Cholesterol homeostasis and the escape tendency (activity) of plasma membrane cholesterol

“…Indeed, ceramides have a higher affinity than cholesterol for ordered phases of 1-palmitoyl-2-oleoyl-phosphatidylcholine [95]. It also appears that ceramides can form condensed complexes with phospholipids like sphingomyelin [96,97]. Thus, the more general premise has been advanced that ceramides can not only form gel-like domains with raftforming phospholipids in vivo, but also elaborate them into platforms upon which signaling proteins can carry out informational functions [98,99].…”