Translocation of oxysterol binding protein to Golgi apparatus triggered by ligand binding.
A cDNA encoding a cytoplasmic oxysterol binding protein was expressed at high levels by transfection in animal cells. This protein binds oxysterols such as 25-hydroxycholesterol that regulate sterol metabolism by transcriptional and posttranscriptional effects. In the transfected cells, some of the oxysterol binding protein (OSBP) was distributed diffusely in the cytoplasm, and some was bound to small vesicles near the nucleus, as revealed by indirect immunofluorescence. Upon addition of 25-hydroxycholesterol, most of the OSBP became concentrated in large perinuclear structures that stained with lentil lectin, a protein that stains the Golgi apparatus. The structures that contained OSBP were disrupted by brefeldin A, confirming their identification as Golgi. A mutant OSBP lacking the COOH-terminal oxysterol binding domain localized to the Golgi spontaneously, suggesting that this domain normally occludes the domain that binds to the Golgi and that sterols relieve this occlusion. The previously noted potential leucine zipper sequence in OSBP was not required for Golgi localization, nor was it essential for homodimer formation. We conclude that OSBP is triggered to bind extrinsically to Golgi membranes when it binds oxysterols and speculate that this translocation may play a role in the transport, metabolism, or regulatory actions of oxysterols.
Ridgway
N D
ND
Department of Molecular Genetics, University of Texas Southwestern Medical Center, Dallas 75235.
Dawson
P A
PA
Ho
Y K
YK
Brown
M S
MS
Goldstein
J L
JL
D000818:Animals; D001483:Base Sequence; D001692:Biological Transport; D020126:Brefeldin A; D016466:CHO Cells; D006224:Cricetinae; D003432:Cross-Linking Reagents / Q000737:chemistry; D003517:Cyclopentanes / Q000494:pharmacology; D004252:DNA Mutational Analysis; D005455:Fluorescent Antibody Technique; D005976:Glutaral / Q000737:chemistry; D006056:Golgi Apparatus / Q000378:metabolism*; D006888:Hydroxycholesterols / Q000378:metabolism; D066298:In Vitro Techniques; D016350:Leucine Zippers; D008969:Molecular Sequence Data; D011987:Receptors, Steroid / Q000378:metabolism*; D011994:Recombinant Proteins; D013329:Structure-Activity Relationship; D000098825:Oxysterol Binding Proteins
D003432:Cross-Linking Reagents; D003517:Cyclopentanes; D006888:Hydroxycholesterols; D011987:Receptors, Steroid; D011994:Recombinant Proteins; D000098825:Oxysterol Binding Proteins; D020126:Brefeldin A; C007997:25-hydroxycholesterol; D005976:Glutaral