Characterization of a Cytosolic Heat-shock Protein-Caveolin Chaperone Complex
INVOLVEMENT IN CHOLESTEROL TRAFFICKING*
- From the ‡University of Kentucky Medical School, Department of Physiology, MS 508, Lexington, Kentucky 40536 and the§University of Texas Southwestern Medical Center, Department of Cell Biology and Neuroscience, Dallas, Texas 75235
Abstract
Caveolin is a 22-kDa protein that appears to play a critical role in regulating the cholesterol concentration of caveolae. Even though caveolin is thought to be a membrane protein, several reports suggest that this peculiar protein can traffic independently of membrane vesicles. We now present evidence that a cytosolic pool of caveolin is part of a heat-shock protein-immunophilin chaperone complex consisting of caveolin, heat-shock protein 56, cyclophilin 40, cyclophilin A, and cholesterol. Treatment of NIH 3T3 cells with 1 μm cyclosporin A or 100 nmrapamycin disrupted the putative transport complex and prevented rapid (10–20 min) transport of cholesterol to caveolae. The lymphoid cell line, L1210-JF, does not express caveolin, does not form an immunophilin-caveolin complex, and does not transport newly synthesized cholesterol to caveolae. Transfection of caveolin cDNA into L1210-JF cells allowed the assembly of a transport complex identical to that found in NIH 3T3 cells. In addition, newly synthesized cholesterol in transfected cells was rapidly (10–20 min) and specifically transported to caveolae. These data strongly suggest that a caveolin-chaperone complex is a mechanism by which newly synthesized cholesterol is transported from the endoplasmic reticulum through the cytoplasm to caveolae.
Footnotes
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↵* This work was supported by Grant IRG-1631 from the American Cancer Society, NHLBI, National Institutes of Health, Grant R29HL58475-01, and Grant-in-Aid KY-97-GS-2 from the American Heart Association, Kentucky Affiliate.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵¶ To whom correspondence should be addressed: University of Kentucky, Dept. of Physiology, 800 Rose St., MS 508 C, Lexington, KY 40536. E-mail: ejsmart{at}pop.uky.edu.
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↵1 The abbreviations used are: ER, endoplasmic reticulum; HSP, heat-shock protein; DMEM, Dulbecco’s modified Eagle’s medium; Tricine,N-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycine; MES, 4-morpholineethanesulfonic acid; BPM, bulk plasma membranes; PVDF, polyvinylidene difluoride; PAGE, polyacrylamide gel electrophoresis.
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- Received October 17, 1997.
- Revision received December 13, 1997.
- The American Society for Biochemistry and Molecular Biology, Inc.
