Differential solubilization of inner plasma membrane leaflet components by Lubrol WX and Triton X-100.
A commonly-used method for analysing raft membrane domains is based on their resistance to extraction by non-ionic detergents at 4 degrees C. However, the selectivity of different detergents in defining raft membrane domains has been questioned. We have compared the lipid composition of detergent-resistant membranes (DRMs) obtained after Triton X-100 or Lubrol WX extraction in MDCK cells in order to understand the differential effect of these detergents on membranes and their selectivity in solubilizing or not proteins. Both Lubrol and Triton DRMs were enriched with cholesterol over the lysate, thus exhibiting characteristics consistent with the properties of membrane rafts. However, the two DRM fractions differed considerably in the ratio between lipids of the inner and outer membrane leaflets. Lubrol DRMs were especially enriched with phosphatidylethanolamine, including polyunsaturated species with long fatty acyl chains. Lubrol and Triton DRMs also differed in the amount of raft transmembrane proteins and raft proteins anchored to the cytoplasmic leaflet. Our results suggest that the inner side of rafts is enriched with phosphatidylethanolamine and cholesterol, and is more solubilized by Triton X-100 than by Lubrol WX.
Delaunay
Jean-Louis
JL
Institut National de la Santé et de la Recherche Médicale UMR S 538, Université Pierre et Marie Curie, Paris, France. delaunay@st-antoine.inserm.fr
Breton
Michelyne
M
Trugnan
Germain
G
Maurice
Michèle
M
D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects* / Q000378:metabolism*; D004285:Dogs; D005227:Fatty Acids / Q000032:analysis; D005732:Gangliosides / Q000032:analysis; D021962:Membrane Microdomains / Q000187:drug effects / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D017830:Octoxynol / Q000494:pharmacology*; D010713:Phosphatidylcholines / Q000032:analysis; D010714:Phosphatidylethanolamines / Q000032:analysis; D011092:Polyethylene Glycols / Q000494:pharmacology*; D012995:Solubility / Q000187:drug effects
D015415:Biomarkers; D005227:Fatty Acids; D005732:Gangliosides; D008565:Membrane Proteins; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; C017474:Lubrol WX; C483858:phosphatidylethanolamine; D011092:Polyethylene Glycols; D017830:Octoxynol