On the neural networks of empathy: A principal component analysis of an fMRI study

© 2008 Nomi et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution Licens

Phosphatidylethanolamine critically supports internalization of cell-penetrating protein C inhibitor

Although their contribution remains unclear, lipids may facilitate noncanonical routes of protein internalization into cells such as those used by cell-penetrating proteins. We show that protein C inhibitor (PCI), a serine protease inhibitor (serpin), rapidly transverses the plasma membrane, which persists at low temperatures and enables its nuclear targeting in vitro and in vivo. Cell membrane translocation of PCI necessarily requires phosphatidylethanolamine (PE). In parallel, PCI acts as a lipid transferase for PE. The internalized serpin promotes phagocytosis of bacteria, thus suggesting a function in host defense. Membrane insertion of PCI depends on the conical shape of PE and is associated with the formation of restricted aqueous compartments within the membrane. Gain- and loss-of-function mutations indicate that the transmembrane passage of PCI requires a branched cavity between its helices H and D, which, according to docking studies, precisely accommodates PE. Our findings show that its specific shape enables cell surface PE to drive plasma membrane translocation of cell-penetrating PCI

Lipid Dynamics and Domain Formation in Model Membranes Composed of Ternary Mixtures of Unsaturated and Saturated Phosphatidylcholines and Cholesterol

In recent years, the implication of sphingomyelin in lipid raft formation has intensified the long sustained interest in this membrane lipid. Accumulating evidences show that cholesterol preferentially interacts with sphingomyelin, conferring specific physicochemical properties to the bilayer membrane. The molecular packing created by cholesterol and sphingomyelin, which presumably is one of the driving forces for lipid raft formation, is known in general to differ from that of cholesterol and phosphatidylcholine membranes. However, in many studies, saturated phosphatidylcholines are still considered as a model for sphingolipids. Here, we investigate the effect of cholesterol on mixtures of dioleoyl-phosphatidylcholine (DOPC) and dipalmitoyl-phosphatidylcholine (DPPC) or distearoyl-phosphatidylcholine (DSPC) and compare it to that on mixtures of DOPC and sphingomyelin analyzed in previous studies. Giant unilamellar vesicles prepared from ternary mixtures of various lipid compositions were imaged by confocal fluorescence microscopy and, within a certain range of sterol content, domain formation was observed. The assignment of distinct lipid phases and the molecular mobility in the membrane bilayer was investigated by fluorescence correlation spectroscopy. Cholesterol was shown to affect lipid dynamics in a similar way for DPPC and DSPC when the two phospholipids were combined with cholesterol in binary mixtures. However, the corresponding ternary mixtures exhibited different spatial lipid organization and dynamics. Finally, evidences of a weaker interaction of cholesterol with saturated phosphatidylcholines than with sphingomyelin (with matched chain length) are discussed

Probing Lipid Mobility of Raft-exhibiting Model Membranes by Fluorescence Correlation Spectroscopy

Confocal fluorescence microscopy and fluorescence correlation spectroscopy (FCS) have been employed to investigate the lipid spatial and dynamic organization in giant unilamellar vesicles (GUVs) prepared from ternary mixtures of dioleoyl-phosphatidylcholine/sphingomyelin/cholesterol. For a certain range of cholesterol concentration, formation of domains with raft-like properties was observed. Strikingly, the lipophilic probe 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI-C18) was excluded from sphingomyelin-enriched regions, where the raft marker ganglioside GM1 was localized. Cholesterol was shown to promote lipid segregation in dioleoyl-phosphatidylcholine-enriched, liquid-disordered, and sphingomyelin-enriched, liquid-ordered phases. Most importantly, the lipid mobility in sphingomyelin-enriched regions significantly increased by increasing the cholesterol concentration. These results pinpoint the key role, played by cholesterol in tuning lipid dynamics in membranes. At cholesterol concentrations >50 mol%, domains vanished and the lipid diffusion slowed down upon further addition of cholesterol. By taking the molecular diffusion coefficients as a fingerprint of membrane phase compositions, FCS is proven to evaluate domain lipid compositions. Moreover, FCS data from ternary and binary mixtures have been used to build a ternary phase diagram, which shows areas of phase coexistence, transition points, and, importantly, how lipid dynamics varies between and within phase regions.

Fluorescence Correlation Spectroscopy Relates Rafts in Model and Native Membranes

The lipid raft model has evoked a new perspective on membrane biology. Understanding the structure and dynamics of lipid domains could be a key to many crucial membrane-associated processes in cells. However, one shortcoming in the field is the lack of routinely applicable techniques to measure raft association without perturbation by detergents. We show that both in cell and in domain-exhibiting model membranes, fluorescence correlation spectroscopy (FCS) can easily distinguish a raft marker (cholera toxin B subunit bound to ganglioside (GM1) and a nonraft marker (dialkylcarbocyanine dye diI)) by their decidedly different diffusional mobilities. In contrast, these markers exhibit only slightly different mobilities in a homogeneous artificial membrane. Performing cholesterol depletion with methyl-β-cyclodextrin, which disrupts raft organization, we find an analogous effect of reduced mobility for the nonraft marker in domain-exhibiting artificial membranes and in cell membranes. In contrast, cholesterol depletion has differential effects on the raft marker, cholera toxin B subunit-GM1, rendering it more mobile in artificial domain-exhibiting membranes but leaving it immobile in cell membranes, where cytoskeleton disruption is required to achieve higher mobility. Thus, fluorescence correlation spectroscopy promises to be a valuable tool to elucidate lipid raft associations in native cells and to gain deeper insight into the correspondence between model and natural membranes