title: Glycosylinositol phosphorylceramides from Rosa cell cultures are boron-bridged in the plasma membrane and form complexes with rhamnogalacturonan II.
abstract: Boron (B) is essential for plant cell-wall structure and membrane functions. Compared with its role in cross-linking the pectic domain rhamnogalacturonan II (RG-II), little information is known about the biological role of B in membranes. Here, we investigated the involvement of glycosylinositol phosphorylceramides (GIPCs), major components of lipid rafts, in the membrane requirement for B. Using thin-layer chromatography and mass spectrometry, we first characterized GIPCs from Rosa cell culture. The major GIPC has one hexose residue, one hexuronic acid residue, inositol phosphate, and a ceramide moiety with a C18 trihydroxylated mono-unsaturated long-chain base and a C24 monohydroxylated saturated fatty acid. Disrupting B bridging (by B starvation in vivo or by treatment with cold dilute HCl or with excess borate in vitro) enhanced the GIPCs' extractability. As RG-II is the main B-binding site in plants, we investigated whether it could form a B-centred complex with GIPCs. Using high-voltage paper electrophoresis, we showed that addition of GIPCs decreased the electrophoretic mobility of radiolabelled RG-II, suggesting formation of a GIPC-B-RG-II complex. Last, using polyacrylamide gel electrophoresis, we showed that added GIPCs facilitate RG-II dimerization in vitro. We conclude that B plays a structural role in the plasma membrane. The disruption of membrane components by high borate may account for the phytotoxicity of excess B. Moreover, the in-vitro formation of a GIPC-B-RG-II complex gives the first molecular explanation of the wall-membrane attachment sites observed in vivo. Finally, our results suggest a role for GIPCs in the RG-II dimerization process.
authors.affiliation: The Edinburgh Cell Wall Group, Institute of Molecular Plant Sciences, The University of Edinburgh, Edinburgh, EH9 3JH, UK.
authors.forename: Aline
authors.initials: A
authors.lastname: Voxeur
authors.forename: Stephen C
authors.initials: SC
authors.lastname: Fry
mesh_terms: D001881:Borates / Q000378:metabolism; D001895:Boron / Q000378:metabolism*; D002478:Cells, Cultured; D002855:Chromatography, Thin Layer; D006028:Glycosphingolipids / Q000302:isolation & purification / Q000378:metabolism*; D017261:Glycosylphosphatidylinositols / Q000302:isolation & purification / Q000378:metabolism*; D021962:Membrane Microdomains / Q000378:metabolism*; D010368:Pectins / Q000378:metabolism*; D027826:Rosa / Q000378:metabolism*; D021241:Spectrometry, Mass, Electrospray Ionization
chemical_list: D001881:Borates; D006028:Glycosphingolipids; D017261:Glycosylphosphatidylinositols; C025524:inositolphosphoceramides; C042492:rhamnogalacturonan II; D010368:Pectins; D001895:Boron
keywords: Rosa sp; boron; cell wall; cross-linking; glycosylinositol phosphoceramides; glycosylinositol phosphorylceramides; lipid rafts; plasma membrane; rhamnogalacturonan II