title: Glucosylceramidase mass and subcellular localization are modulated by cholesterol in Niemann-Pick disease type C.
abstract: Niemann-Pick disease type C (NPC) is characterized by the accumulation of cholesterol and sphingolipids in the late endosomal/lysosomal compartment. The mechanism by which the concentration of sphingolipids such as glucosylceramide is increased in this disease is poorly understood. We have found that, in NPC fibroblasts, the cholesterol storage affects the stability of glucosylceramidase (GCase), decreasing its mass and activity; a reduction of cholesterol raises the level of GCase to nearly normal values. GCase is activated and stabilized by saposin C (Sap C) and anionic phospholipids. Here we show by immunofluorescence microscopy that in normal fibroblasts, GCase, Sap C, and lysobisphosphatidic acid (LBPA), the most abundant anionic phospholipid in the endolysosomal system, reside in the same intracellular vesicular structures. In contrast, the colocalization of GCase, Sap C, and LBPA is markedly impaired in NPC fibroblasts but can be re-established by cholesterol depletion. These data show for the first time that the level of cholesterol modulates the interaction of GCase with its protein and lipid activators, namely Sap C and LBPA, regulating the GCase activity and stability.
authors.affiliation: Department of Hematology, Istituto Superiore Sanita', 00161 Roma, Italy.
authors.forename: Rosa
authors.initials: R
authors.lastname: Salvioli
authors.forename: Susanna
authors.initials: S
authors.lastname: Scarpa
authors.forename: Fiorella
authors.initials: F
authors.lastname: Ciaffoni
authors.forename: Massimo
authors.initials: M
authors.lastname: Tatti
authors.forename: Carlo
authors.initials: C
authors.lastname: Ramoni
authors.forename: Marie T
authors.initials: MT
authors.lastname: Vanier
authors.forename: Anna Maria
authors.initials: AM
authors.lastname: Vaccaro
mesh_terms: D015153:Blotting, Western; D002460:Cell Line; D002784:Cholesterol / Q000378:metabolism*; D011992:Endosomes / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D005962:Glucosylceramidase / Q000096:biosynthesis* / Q000737:chemistry*; D006023:Glycoproteins / Q000737:chemistry; D006801:Humans; D050356:Lipid Metabolism; D008246:Lysophospholipids / Q000737:chemistry; D008247:Lysosomes / Q000378:metabolism; D008856:Microscopy, Fluorescence; D050178:Monoglycerides; D009154:Mutation; D009542:Niemann-Pick Diseases / Q000378:metabolism*; D010743:Phospholipids / Q000378:metabolism; D011233:Precipitin Tests; D049231:Saposins; D013997:Time Factors
chemical_list: D006023:Glycoproteins; D008246:Lysophospholipids; D050178:Monoglycerides; C483345:PSAP protein, human; D010743:Phospholipids; D049231:Saposins; C012786:bis(monoacylglyceryl)phosphate; D002784:Cholesterol; D005962:Glucosylceramidase