title: Sterol liganding of OSBP-related proteins (ORPs) regulates the subcellular distribution of ORP-VAPA complexes and their impacts on organelle structure.
abstract: Oxysterol-binding protein (OSBP) and its homologues (ORPs) are lipid-binding/transfer proteins with affinity for oxysterols, cholesterol and glycerophospholipids. In addition to a ligand-binding domain, a majority of the ORPs carry a pleckstrin homology domain that targets organelle membranes via phosphoinositides, and a motif targeting the endoplasmic reticulum (ER) via VAMP-associated proteins (VAPs). We employed here Bimolecular Fluorescence Complementation (BiFC) to systematically assess the effects of sterol manipulation of HuH7 cells on complexes of established sterol-binding ORPs with their ER receptor, VAMP-associated protein A (VAPA). Depletion of cellular cholesterol with lipoprotein-deficient medium and Mevastatin caused concentration of OSBP-VAPA complexes and Golgi complex markers at a juxtanuclear position, an effect reversed by low-density lipoprotein treatment. A similar redistribution of OSBP-VAPA but not of sterol-binding deficient mutant OSBP(ΔELSK)-VAPA, occurred upon treatment with the high-affinity ligand, 25-hydroxycholesterol (25OHC), which reduced total and free cholesterol. ORP2-VAPA complexes, which localize in untreated cells at blob-like ER structures with associated lipid droplets, were redistributed upon treatment with the ORP2 ligand 22(R)OHC to a diffuse cytoplasmic/ER pattern and the plasma membrane. Analogously, distribution of ORP4L-VAPA complexes between the plasma membrane and vimentin intermediate filament associated compartments was modified by statin or 25OHC treatment. The treatments resulted in loss of vimentin co-localization, and sterol-binding deficient ORP4L(ΔELSR)-VAPA localized predominantly to the plasma membrane. In conclusion, treatment with statin or oxysterol ligands modify the subcellular targeting of ORP-VAPA complexes, consistent with the notion that this machinery controls lipid homeostasis and signaling at organelle interfaces.
authors: Minerva Foundation Institute for Medical Research, Biomedicum 2U, FI-00290 Helsinki, Finland.
authors: Henriikka
authors: H
authors: Kentala
authors: Institute of Biomedicine, Anatomy, FI-00014 University of Helsinki, Finland.
authors: Simon G
authors: SG
authors: Pfisterer
authors: Minerva Foundation Institute for Medical Research, Biomedicum 2U, FI-00290 Helsinki, Finland; Institute of Biomedicine, Anatomy, FI-00014 University of Helsinki, Finland.
authors: Vesa M
authors: VM
authors: Olkkonen
authors: Minerva Foundation Institute for Medical Research, Biomedicum 2U, FI-00290 Helsinki, Finland. Electronic address: marion.weber@helsinki.fi.
authors: Marion
authors: M
authors: Weber-Boyvat
mesh_terms: D045744:Cell Line, Tumor; D005453:Fluorescence; D006056:Golgi Apparatus / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D006888:Hydroxycholesterols / Q000494:pharmacology; D042541:Intracellular Space / Q000378:metabolism; D008024:Ligands; D046912:Multiprotein Complexes / Q000378:metabolism*; D015388:Organelles / Q000187:drug effects / Q000378:metabolism*; D021381:Protein Transport / Q000187:drug effects; D011987:Receptors, Steroid / Q000378:metabolism*; D013261:Sterols / Q000378:metabolism*; D013347:Subcellular Fractions / Q000378:metabolism; D014280:Triglycerides / Q000378:metabolism; D033921:Vesicular Transport Proteins / Q000378:metabolism*; D014746:Vimentin / Q000378:metabolism; D000098825:Oxysterol Binding Proteins
chemical_list: D006888:Hydroxycholesterols; D008024:Ligands; D046912:Multiprotein Complexes; D011987:Receptors, Steroid; D013261:Sterols; D014280:Triglycerides; C114051:VAPA protein, human; D033921:Vesicular Transport Proteins; D014746:Vimentin; D000098825:Oxysterol Binding Proteins; C007997:25-hydroxycholesterol
keywords: Cholesterol; Lipid signaling; Lipid transport; Oxysterol; Oxysterol-binding protein; VAMP-associated protein