Sterol Carrier Protein-2 Is Involved in Cholesterol Transfer from the Endoplasmic Reticulum to the Plasma Membrane in Human Fibroblasts

Puglielli, Luigi; Rigotti, Attilio; Greco, Aldo V.; Santos, Manuel J.; Nervi, Flavio

doi:10.1074/jbc.270.32.18723

Cited by 142 publications

(92 citation statements)

References 25 publications

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“…The physiological impact of cholesterol binding proteins such as SCP-2 and L-FABP on cholesterol uptake, esterification, and excretion into bile was confirmed by studies with SCP-2 transgenic (i.e. overexpressing or antisense cDNA treated) mice and rats (177-179) Studies with radiolabeled cholesterol were consistent with many of the findings shown with the fluorescent DHE and other sterols in cultured cells (43,45,172,(190)(191)(192)(193)(194)(195)(196)(197). In one study, it was shown that overexpression of L-FABP in L-cell fibroblasts enhanced the transfer or radiolabled cholesterol from the plasma membrane to the endoplasmic reticulum for esterification (194).…”

Section: Physiological Significance Of In Vitro Studies With Dhe and mentioning

confidence: 78%

Fluorescence Techniques Using Dehydroergosterol to Study Cholesterol Trafficking

McIntosh

Atshaves

Huang

et al. 2008

Lipids

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Cholesterol itself has very few structural/chemical features suitable for real-time imaging in living cells. Thus, the advent of dehydroergosterol [ergosta-5,7,9(11),22-tetraen-3β-ol, DHE] the fluorescent sterol most structurally and functionally similar to cholesterol to date, has proven to be a major asset for real-time probing/elucidating the sterol environment and intracellular sterol trafficking in living organisms. DHE is a naturally-occurring, fluorescent sterol analog that faithfully mimics many of the properties of cholesterol. Because these properties are very sensitive to sterol structure and degradation, such studies require the use of extremely pure (>98%) quantities of fluorescent sterol. DHE is readily bound by cholesterol-binding proteins, is incorporated into lipoproteins (from the diet of animals or by exchange in vitro), and for real-time imaging studies is easily incorporated into cultured cells where it co-distributes with endogenous sterol. Incorporation from an ethanolic stock solution to cell culture media is effective, but this process forms an aqueous dispersion of dehydroergosterol crystals which can result in endocytic cellular uptake and distribution into lysosomes which is problematic in imaging DHE at the plasma membrane of living cells. In contrast, monomeric DHE can be incorporated from unilamellar vesicles by exchange/fusion with the plasma membrane or from DHE-methyl-β-cyclodextrin (DHE-MβCD) complexes by exchange with the plasma membrane. Both of the latter techniques can deliver large quantities of monomeric dehydroergosterol with significant distribution into the plasma membrane. The properties and behavior of DHE in protein-binding, lipoproteins, model membranes, biological membranes, lipid rafts/caveolae, and real-time imaging in living cells indicate that this naturally-occurring fluorescent sterol is a useful mimic for probing the properties of cholesterol in these systems. Keywordsdehydroergosterol; cholesterol; ergosterol; fluorescent sterols; microscopy Historical PerspectiveIn order to resolve the dynamics and factors governing cholesterol trafficking within cells, it is important to recognize that the cholesterol molecule has very few polar constituents (Fig. 1A). Consequently, cholesterol is poorly soluble in aqueous environments such as cytosol as evidenced by critical micellar concentrations of cholesterol and other sterols being very low, *To whom correspondence should be addressed: Department of Physiology and Pharmacology, Texas A&M University, TVMC, College Station, TX 862-1433, FAX: (979) NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript near 20-30 nM (1-5). The physiological impact of cholesterol's low aqueous solubility is that spontaneous cholesterol desorption from the cytofacial leaflet of the plasma membrane or lysosomal membrane into the cytosol for transfer to intracellular sites for esterification, oxidation, or secretion is extremely slow (t 1/2 3 hours-days) (6-9). Therefore, it is important to resolve factors...

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Section: Physiological Significance Of In Vitro Studies With Dhe and mentioning

confidence: 78%

Fluorescence Techniques Using Dehydroergosterol to Study Cholesterol Trafficking

McIntosh

Atshaves

Huang

et al. 2008

Lipids

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“…Transport of de novo-synthesized cholesterol to the plasma membrane is energy-and temperature-dependent and thought to be mediated via lipid-rich transport intermediates (11,(30)(31)(32). Inhibitors of glycoprotein export, such as BFA, result in only modest reductions in cholesterol export, suggesting that the majority of cholesterol transported to the plasma membrane bypasses the Golgi apparatus in BFA-treated cells (28).…”

Section: Discussionmentioning

confidence: 99%

Golgi-dependent transport of cholesterol to the Chlamydia trachomatis inclusion

Carabeo

Mead

Hackstadt

2003

Proc. Natl. Acad. Sci. U.S.A.

251

279

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Cholesterol, a lipid not normally found in prokaryotes, was identified in purified Chlamydia trachomatis elementary bodies and in the chlamydial parasitophorous vacuole (inclusion) membrane of infected HeLa cells. Chlamydiae obtained eukaryotic host cell cholesterol both from de novo synthesis or low-density lipoprotein. Acquisition of either de novo-synthesized cholesterol or low-density lipoprotein-derived cholesterol was microtubule-dependent and brefeldin A-sensitive, indicating a requirement for the Golgi apparatus. Transport also required chlamydial protein synthesis, indicative of a pathogen-directed process. The cholesterol trafficking pathway appears to coincide with a previously characterized delivery of sphingomyelin to the inclusion in that similar pharmacological treatments inhibited transport of both sphingomyelin and cholesterol. These results support the hypothesis that sphingomyelin and cholesterol may be cotransported via a Golgidependent pathway and that the chlamydial inclusion receives cholesterol preferentially from a brefeldin A-sensitive pathway of cholesterol trafficking from the Golgi apparatus to the plasma membrane.C hlamydia trachomatis is the etiologic agent of the most common form of sexually transmitted disease in the United States and preventable blindness worldwide (1). Chlamydiae are obligate intracellular bacteria with a biphasic life cycle consisting of an extracellular infectious form and an intracellular replicative form. Infection is initiated by the metabolically inert elementary body (EB), which develops into the noninfectious but metabolically active reticulate body (RB). RBs differentiate back to EBs before release from the infected cell (2). Chlamydiae replicate within a specialized parasitophorous vacuole, termed an inclusion, that is neither acidified nor fusogenic with lysosomes (3, 4). Rather than interacting with the endocytic pathway, the chlamydial inclusion is fusogenic with exocytic vesicles containing sphingomyelin en route from the Golgi apparatus to the plasma membrane (5, 6). Acquisition of sphingomyelin requires chlamydial de novo transcription and translation of proteins that are thought to modify the inclusion membrane (7).Because sphingomyelin is delivered to the chlamydial inclusion and EBs contain both cholesterol and sphingomyelin (5, 8, 9), we asked whether cholesterol may be similarly regulated and transported to the inclusion. Cholesterol is an essential component of eukaryotic membranes and is enriched in the plasma membrane. Although synthesis occurs in the endoplasmic reticulum (ER), the role of the Golgi apparatus in the transport of cholesterol to the plasma membrane has been controversial. Dissection of intracellular cholesterol transport is complicated by the fact that there are a number of possible sources of this lipid. Eukaryotic cells acquire cholesterol from either de novo synthesis or from the extracellular media via the low-density lipoprotein (LDL) pathway (10-12). We find that C. trachomatis has the ability to obtain host cholest...

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“…Another protein, sterol carrier protein x/2 (SCPx/2), has also been implicated in the initial rapid transport of cellular cholesterol to the plasma membrane. 50 The impaired HDL-and apoA-I-mediated lipid efflux in FHD fibroblasts does not appear to be related to a defective HDL interaction with cell surface binding proteins. We found that binding of HDL 3 at 4°C to cholesterol-loaded FHD fibroblasts was not significantly different from HDL 3 binding to normal cells.…”

Section: Discussionmentioning

confidence: 99%

Cellular Cholesterol Transport and Efflux in Fibroblasts Are Abnormal in Subjects With Familial HDL Deficiency

Marcil

Krimbou

et al. 1999

ATVB

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Abstract-Familial high density lipoprotein (HDL) deficiency (FHD) is a genetic lipoprotein disorder characterized by a severe decrease in the plasma HDL cholesterol (-C) level (less than the fifth percentile). Unlike Tangier disease, FHD is transmitted as an autosomal dominant trait. FHD subjects have none of the clinical manifestations of Tangier disease (lymphoid tissue infiltration with cholesteryl esters and/or neurological manifestations). Plasmas from FHD subjects contain pre-␤-migrating HDLs but are deficient in ␣-migrating HDLs. We hypothesized that a reduced HDL-C level in FHD is due to abnormal transport of cellular cholesterol to the plasma membrane, resulting in reduced cholesterol efflux onto nascent HDL particles, leading to lipid-depleted HDL particles that are rapidly catabolized. Cellular cholesterol metabolism was investigated in skin fibroblasts from FHD and control subjects. HDL 3 -and apolipoprotein (apo) A-I-mediated cellular cholesterol and phosphatidylcholine efflux was examined by labeling cells with

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Sterol Carrier Protein-2 Is Involved in Cholesterol Transfer from the Endoplasmic Reticulum to the Plasma Membrane in Human Fibroblasts

Cited by 142 publications

References 25 publications

Fluorescence Techniques Using Dehydroergosterol to Study Cholesterol Trafficking

Fluorescence Techniques Using Dehydroergosterol to Study Cholesterol Trafficking

Golgi-dependent transport of cholesterol to the Chlamydia trachomatis inclusion

Cellular Cholesterol Transport and Efflux in Fibroblasts Are Abnormal in Subjects With Familial HDL Deficiency

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