Characterization of the Lateral Distribution of Fluorescent Lipid in Binary-Constituent Lipid Monolayers by Principal Component Analysis

Lipid lateral organization in binary-constituent monolayers consisting of fluorescent and nonfluorescent lipids has been investigated by acquiring multiple emission spectra during measurement of each force-area isotherm. The emission spectra reflect BODIPY-labeled lipid surface concentration and lateral mixing with different nonfluorescent lipid species. Using principal component analysis (PCA) each spectrum could be approximated as the linear combination of only two principal vectors. One point on a plane could be associated with each spectrum, where the coordinates of the point are the coefficients of the linear combination. Points belonging to the same lipid constituents and experimental conditions form a curve on the plane, where each point belongs to a different mole fraction. The location and shape of the curve reflects the lateral organization of the fluorescent lipid mixed with a specific nonfluorescent lipid. The method provides massive data compression that preserves and emphasizes key information pertaining to lipid distribution in different lipid monolayer phases. Collectively, the capacity of PCA for handling large spectral data sets, the nanoscale resolution afforded by the fluorescence signal, and the inherent versatility of monolayers for characterization of lipid lateral interactions enable significantly enhanced resolution of lipid lateral organizational changes induced by different lipid compositions

\u3ci\u3eArabidopsis\u3c/i\u3e Accelerated Cell Death 11, ACD11, Is a Ceramide-1-Phosphate Transfer Protein and Intermediary Regulator of Phytoceramide Levels

The accelerated cell death 11 (acd11) mutant of Arabidopsis provides a genetic model for studying immune response activation and localized cellular suicide that halt pathogen spread during infection in plants. Here, we elucidate ACD11 structure and function and show that acd11 disruption dramatically alters the in vivo balance of sphingolipid mediators that regulate eukaryotic-programmed cell death. In acd11 mutants, normally low ceramide-1- phosphate (C1P) levels become elevated, but the relatively abundant cell death inducer phytoceramide rises acutely. ACD11 exhibits selective intermembrane transfer of C1P and phyto-C1P. Crystal structures establish C1P binding via a surface-localized, phosphate headgroup recognition center connected to an interior hydrophobic pocket that adaptively ensheaths lipid chains via a cleft-like gating mechanism. Point mutation mapping confirms functional involvement of binding site residues. A π helix (π bulge) near the lipid binding cleft distinguishes apo-ACD11 from other GLTP folds. The global two-layer, α-helically dominated, ‘‘sandwich’’ topology displaying C1P-selective binding identifies ACD11 as the plant prototype of a GLTP fold subfamily

Phosphatidylserine stimulates ceramide 1-phosphate (C1P) intermembrane transfer by C1P transfer proteins

Genetic models for studying localized cell suicide that halt the spread of pathogen infection and immune response activation in plants include Arabidopsis accelerated-cell-death 11 mutant (acd11). In this mutant, sphingolipid homeostasis is disrupted via depletion of ACD11, a lipid transfer protein that is specific for ceramide 1-phosphate (C1P) and phyto-C1P. The C1P binding site in ACD11 and in human ceramide-1-phosphate transfer protein (CPTP) is surrounded by cationic residues. Here, we investigated the functional regulation of ACD11 and CPTP by anionic phosphoglycerides and found that 1-palmitoyl-2-oleoyl-phosphatidic acid or 1-palmitoyl-2-oleoyl-phosphatidylglycerol (≤15 mol %) in C1P source vesicles depressed C1P intermembrane transfer. By contrast, replacement with 1-palmitoyl-2-oleoyl-phosphatidylserine stimulated C1P transfer by ACD11 and CPTP. Notably, “soluble” phosphatidylserine (dihexanoyl-phosphatidylserine) failed to stimulate C1P transfer. Also, none of the anionic phosphoglycerides affected transfer action by human glycolipid lipid transfer protein (GLTP), which is glycolipid-specific and has few cationic residues near its glycolipid binding site. These findings provide the first evidence for a potential phosphoglyceride headgroup-specific regulatory interaction site(s) existing on the surface of any GLTP-fold and delineate new differences between GLTP superfamily members that are specific for C1P versus glycolipid

Diol lipids are activators of protein kinase C

AbstractDiol lipids (dioleoyl- and dioctanoylethylene glycol) at relatively low concentrations (∼ 10 μM) were found to activate significantly protein kinase C in the presence of phosphatidylserine or phosphatidylinositol. Since diol lipids are widespread minor lipid constituents of many cells [(1974) Chem. Ind., 597–604], it has been suggested that they may be involved in the maintaining of basal protein kinase C activity in the absence of external stimuli