Changes in molecular species profiles of glycosylphosphatidylinositol anchor precursors in early stages of biosynthesis.

Glycosylphosphatidylinositol (GPI) anchor is a major lipidation in posttranslational modification. GPI anchor precursors are biosynthesized from endogenous phosphatidylinositols (PIs) and attached to proteins in the endoplasmic reticulum. Endogenous PIs are characterized by domination of diacyl species and the presence of polyunsaturated fatty acyl chain, such as 18:0-20:4, at the sn-2 position.

Fatty acid remodeling of GPI-anchored proteins is required for their raft association.

Whereas most of the cellular phosphatidylinositol (PI) contain unsaturated fatty chains and are excluded from rafts, GPI-anchored proteins (APs) unusually contain two saturated fatty chains in their PI moiety, and they are typically found within lipid rafts. However, the origin of the saturated chains and whether they are essential for raft association are unclear. Here, we report that GPI-APs, with two saturated fatty chains, are generated from those bearing an unsaturated chain by fatty acid remodeling that occurs most likely in the Golgi and requires post-GPI-attachment to proteins (PGAP)2 and PGAP3.

Overexpression of caveolin-1 in a human melanoma cell line results in dispersion of ganglioside GD3 from lipid rafts and alteration of leading edges, leading to attenuation of malignant properties.

Caveolin-1 is a component of lipid rafts, and is considered to be a tumor suppressor molecule. However, the mechanisms by which caveolin-1 functions in cancer cells are not well understood. We generated caveolin-1 transfectant cells (Cav-1(+) cells) using a human melanoma cell line (SK-MEL-28) and investigated the effects of caveolin-1 overexpression on the GD3-mediated malignant properties of melanomas.

Focused lipidomics by tandem mass spectrometry.

In this paper we performed focused analyses of phospholipids by using the data of precursor ion scanning and neutral loss scanning of their polar head groups and fatty acyl moieties for the specific search of categorical phospholipids. By using precursor ion scanning or neutral loss scanning of polar head groups in the positive ion mode, more sensitive identification were obtained than that in the negative ion mode. Precursor ion scanning of carbonic anions in the negative ion mode was also effective to identify molecular species of phospholipids having specified fatty acyl moieties.

A shotgun tandem mass spectrometric analysis of phospholipids with normal-phase and/or reverse-phase liquid chromatography/electrospray ionization mass spectrometry.

Electrospray ionization mass spectrometry is used in lipidomics studies. The present research established a top-down liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) shotgun analysis method for phospholipids (PLs) using a normal-phase column or a C30 reverse-phase column with the data-dependent MS/MS scanning mode. A normal-phase column can separate most of the major different classes of PLs.

Rapid and selective identification of molecular species in phosphatidylcholine and sphingomyelin by conditional neutral loss scanning and MS3.

Analyses of molecular species of phospholipids containing choline (Ch), such as phosphatidylcholine (PC) and sphingomyelin (SM), are reported. Neutral loss scanning was applied for the selective detection of these lipids using a quadrupole-linear ion trap mass spectrometer. By using ammonium formate as an elution buffer, both PC and SM were detected as [M+HCOO]- ions in the negative ion mode.

High-resolution analysis by nano-electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry for the identification of molecular species of phospholipids and their oxidized metabolites.

Nano-electrospray ionization (ESI) Fourier transform ion cyclotron resonance mass spectrometry (FTICRMS) was applied to identify the molecular species of phosphatidylethanolamine of Caenorhabditis elegans, which has a high concentration of phospholipids with a fatty acyl chain having an odd number of carbon atoms. The molecular species of diacyl phosphatidylethanolamine with one fatty acyl chain having an odd number of carbon atoms and one fatty acyl chain having an even number of carbon atoms was identified separately from alkyl-acyl phosphatidylethanolamine with an alkyl chain having an even number of carbon atoms and a fatty acyl chain having an even number of carbon atoms. Furthermore, nano-ESI-FTICRMS was applied to the direct identification of oxidized phosphatidylcholine from soybean.

Human group IVC phospholipase A2 (cPLA2gamma). Roles in the membrane remodeling and activation induced by oxidative stress.

To create the unique properties of a certain cellular membrane, both the composition and the metabolism of membrane phospholipids are key factors. Phospholipase A(2) (PLA(2)), with hydrolytic enzyme activities at the sn-2 position in glycerophospholipids, plays critical roles in maintaining the phospholipid composition as well as producing bioactive lipid mediators. In this study we examined the contribution of a Ca(2+)-independent group IVC PLA(2) isozyme (cPLA(2)gamma), a paralogue of cytosolic PLA(2)alpha (cPLA(2)alpha), to phospholipid remodeling.

The surface of lipid droplets is a phospholipid monolayer with a unique Fatty Acid composition.

We found that caveolin-2 is targeted to the surface of lipid droplets (Fujimoto, T., Kogo, H., Ishiguro, K.