Golgi localization and phosphorylation of oxysterol binding protein in Niemann-Pick C and U18666A-treated cells

Oxysterol binding protein (OSBP) translocation between Golgi and vesicular/cytoplasmic compartments is affected by conditions that alter cholesterol and sphingomyelin homeostasis, indicating a role in lipid and sterol regulation in this organelle. In this study, we show that OSBP dissociation from t...

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Published in:Journal of lipid research 2001-07, Vol.42 (7), p.1062-1071
Main Authors: Mohammadi, A, Perry, R J, Storey, M K, Cook, H W, Byers, D M, Ridgway, N D
Format: Article
Language:English
Subjects:
Androstenes - pharmacology
Animals
CHO Cells
cholesterol
Cholesterol - biosynthesis
Cholesterol, LDL - metabolism
Cholinergic Antagonists - pharmacology
Cricetinae
Fibroblasts - drug effects
Fibroblasts - metabolism
Golgi apparatus
Golgi Apparatus - metabolism
Golgi Apparatus - ultrastructure
Humans
Mutagenesis, Site-Directed - physiology
Niemann-Pick Diseases - metabolism
Niemann-Pick type C
Phosphorylation
Protein Transport - physiology
Receptors, Steroid - metabolism
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Summary:Oxysterol binding protein (OSBP) translocation between Golgi and vesicular/cytoplasmic compartments is affected by conditions that alter cholesterol and sphingomyelin homeostasis, indicating a role in lipid and sterol regulation in this organelle. In this study, we show that OSBP dissociation from the Golgi apparatus was inhibited when LDL cholesterol efflux from lysosomes was blocked in Niemann-Pick C (NPC) or U18666A [3-beta-[2-(diethylamino)ethoxy]androst-5-en-17-one]-treated fibroblasts. Dissociation of OSBP from the Golgi apparatus in response to LDL was independent of de novo cholesterol biosynthesis. OSBP did not localize with filipin-stained lysosomal cholesterol, and the NPC defect did not alter OSBP expression or phosphorylation. However, OSBP in the Golgi apparatus was progressively dephosphorylated (as assessed by a molecular mass shift on SDS-PAGE) in U18666A-treated fibroblasts or Chinese hamster ovary cells as a result of combined inhibition of LDL cholesterol transport and de novo cholesterol synthesis. In vivo phosphopeptide mapping and mutagenesis of OSBP was used to identify the cholesterol-sensitive phosphorylation sites at serines 381, 384, and 387 that were responsible for the altered mobility on SDS-PAGE. NPC-1 protein-mediated release of LDL-derived cholesterol and de novo biosynthesis regulates OSBP localization and phosphorylation. This indicates that OSBP responds to or senses altered cellular sterol content and transport.
ISSN:0022-2275
DOI:10.1016/s0022-2275(20)31595-9