High-density-lipoprotein subfraction 3 interaction with glycosylphosphatidylinositol-anchored proteins.
To elucidate further the binding of high-density-lipoprotein subfraction 3 (HDL3) to cells, the involvement of glycosylphosphatidylinositol-anchored proteins (GPI-proteins) was studied. Treatment of cultured cells, such as fibroblasts or SK-MES-1 cells, with a phosphatidylinositol-specific phospholipase C (PI-PLC) significantly decreases specific HDL3 binding. Moreover, PI-PLC treatment of cultured cells or cellular plasma membrane fractions results in releasing proteins. These proteins have a soluble form and can also bind HDL3, as revealed by ligand blotting experiments with HDL3. In order to obtain enriched GPI-proteins, we used a detergent-free purification method to prepare a caveolar membrane fraction. In the caveolar fraction, we obtained, by ligand blotting experiments, the enrichment of two HDL3-binding proteins with molecular masses of 120 and 80 kDa. These proteins were also revealed in a plasma membrane preparation with two other proteins, with molecular masses of 150 and 104 kDa, and were sensitive to PI-PLC treatment. Electron microscopy also showed the binding of Au-labelled HDL3 inside the caveolar membrane invaginations. In SK-MES-1 cells, HDL3 are internalized into a particular structure, resulting in the accumulation and concentration of such specific membrane domains. To sum up, a demonstration has been made of the implication of GPI-proteins as well as caveolae in the binding of HDL3 to cells.
Nion
S
S
Unité INSERM 325, Institut Pasteur, 1 rue du Professeur Calmette, BP 245, 59019 Lille Cedex, France.
Briand
O
O
Lestavel
S
S
Torpier
G
G
Nazih
F
F
Delbart
C
C
Fruchart
J C
JC
Clavey
V
V
D001692:Biological Transport; D002462:Cell Membrane / Q000378:metabolism*; D002478:Cells, Cultured; D002784:Cholesterol / Q000378:metabolism*; D004705:Endocytosis; D005347:Fibroblasts / Q000378:metabolism; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D006801:Humans; D008075:Lipoproteins, HDL / Q000378:metabolism*; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054801:Phosphoinositide Phospholipase C; D011485:Protein Binding / Q000187:drug effects; D012867:Skin / Q000166:cytology / Q000378:metabolism; D010738:Type C Phospholipases / Q000494:pharmacology
D017261:Glycosylphosphatidylinositols; D008075:Lipoproteins, HDL; D002784:Cholesterol; D010738:Type C Phospholipases; D054801:Phosphoinositide Phospholipase C; D043265:Phosphatidylinositol Diacylglycerol-Lyase