A Caveolin Dominant Negative Mutant Associates with Lipid Bodies and Induces Intracellular Cholesterol Imbalance

doi:10.1083/jcb.152.5.1057

Published online 5 March 2001. doi:10.1083/jcb.152.5.1057

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© The Rockefeller University Press, 0021-9525/2001//1057 $5.00
The Journal of Cell Biology, Volume 152, Number 5, , 2001 1057-1070

Original Article

A Caveolin Dominant Negative Mutant Associates with Lipid Bodies and Induces Intracellular Cholesterol Imbalance

Albert Pol^a, Robert Luetterforst^a, Margaret Lindsay^a, Sanna Heino^b, Elina Ikonen^b, and Robert G. Parton^a

^a Institute for Molecular Bioscience, Centre for Microscopy and Microanalysis and Department of Physiology and Pharmacology, University of Queensland, Queensland 4072, Australia
^b Department of Biochemistry, National Public Health Institute, 00300 Helsinki, Finland
University Of Queensland, Queensland 4072, Brisbane, Australia.(61) 7 3365 4422(61) 7 3365 6468

r.parton{at}mailbox.uq.edu.au

Recent studies have indicated a role for caveolin in regulatingcholesterol-dependent signaling events. In the present studywe have analyzed the role of caveolins in intracellular cholesterolcycling using a dominant negative caveolin mutant. The mutantcaveolin protein, cav-3^DGV, specifically associates with themembrane surrounding large lipid droplets. These structurescontain neutral lipids, and are accessed by caveolin 1–3upon overexpression. Fluorescence, electron, and video microscopyobservations are consistent with formation of the membrane-enclosedlipid rich structures by maturation of subdomains of the ER.The caveolin mutant causes the intracellular accumulation offree cholesterol (FC) in late endosomes, a decrease in surfacecholesterol and a decrease in cholesterol efflux and synthesis.The amphiphile U18666A acts synergistically with cav^DGV to increaseintracellular accumulation of FC. Incubation of cells with oleicacid induces a significant accumulation of full-length caveolinsin the enlarged lipid droplets. We conclude that caveolin canassociate with the membrane surrounding lipid droplets and isa key component involved in intracellular cholesterol balanceand lipid transport in fibroblasts.

Key Words: caveolin • caveolae • cholesterol • lipid droplets • endoplasmic reticulum

The online version of this article contains supplemental material.

Abbreviations used in this paper: BFA, brefeldin A; CDV, cav^DGVenriched vesicles; FC, free cholesterol; GFP, green fluorescentprotein; NPC, Niemann-Pick disease C; PDI, phosphodisulphideisomerase; PM, plasma membrane; SCAP, SREBP-cleavage activatingprotein; SREBP, sterol-regulatory element-binding protein; YFP,yellow fluorescent protein.

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