Membrane Cholesterol Dynamics: Cholesterol Domains and Kinetic Pools

Schroeder, Friedhelm; Jefferson, John R.; Kier, Ann B.; Knittel, James J.; Scallen, Terrence J.; Wood, W. Gibson; Hapala, Ivan

doi:10.3181/00379727-196-43185

Cited by 251 publications

(218 citation statements)

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“…The lifetime studies resolved for the first time at least two DHE domains in model membranes, one less sensitive to the aqueous than the other, and that these domains were dependent on the lipid composition, temperature, and other properties of the membrane (54,119). This possibility was further supported by model membrane studies of DHE exchange which provided kinetic evidence for the existence of multiple sterol containing domains in membranes (118,(121)(122)(123)(124).…”

Section: Model Membranesmentioning

confidence: 68%

“…in (54,55). The exchange kinetics of DHE and radiolabeled cholesterol between model membranes was indistinguishable (117).…”

Section: Dhe Architecture In Purified Biological Membranesmentioning

confidence: 94%

“…Because of the tremendous interest of biologists and membraneologists in cholesterol-rich microdomains (also called lipid rafts or caveolae) over the past 15 years (rev. in (54)(55)(56)(57)(58)(59)(60)(61)(62)(63), DHE has emerged as a popular, potentially less perturbing fluorescent sterol probe since it does not contain additional bulky fluorescent groups added to the sterol structure. Although this ultraviolet (UV) light absorbing and fluorescent sterol (i.e.…”

Section: Advent Of Fluorescent Sterolsmentioning

confidence: 99%

“…It is important to note that not only the quantity, but also the chemical structure of the sterol and the presence of cholesterol oxidation products are known to significantly affect the structure and/or function of model membranes (rev. in (54,55,70,72,73), biological membranes (13,(74)(75)(76)(77)(78)(79)(80), lipid raft/caveolae (58,(81)(82)(83)(84)(85)(86)(87)(88), and cholesterol-requiring plasma membrane receptors (66,(89)(90)(91). Because of this sensitivity it is essential that fluorescent sterols such as DHE not contain other sterols (e.g.…”

Section: Effect Of Sterol Structure and Purity On Sterol Architecturementioning

confidence: 99%

“…DHE has been used to probe the structural (polarization, limiting anisotropy, order), dynamic (lifetime, rotational rate), and kinetic (exchange) environment of cholesterol in plasma membranes (6,7,54,55,135,(138)(139)(140)(141), microsomes (6,7), mitochondria (6,7,142,143), and lysosomes (8,129). DHE has revealed several important structural and dynamic aspects of the cholesterol microenvironment in biological membranes.…”

Section: Dhe Architecture In Purified Biological Membranesmentioning

confidence: 99%

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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: Model Membranesmentioning

confidence: 68%

“…in (54,55). The exchange kinetics of DHE and radiolabeled cholesterol between model membranes was indistinguishable (117).…”

Section: Dhe Architecture In Purified Biological Membranesmentioning

confidence: 94%

Section: Advent Of Fluorescent Sterolsmentioning

confidence: 99%

Section: Effect Of Sterol Structure and Purity On Sterol Architecturementioning

confidence: 99%

Section: Dhe Architecture In Purified Biological Membranesmentioning

confidence: 99%

See 3 more Smart Citations

Fluorescence Techniques Using Dehydroergosterol to Study Cholesterol Trafficking

McIntosh

Atshaves

Huang

et al. 2008

Lipids

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Modulation of ?5?1 integrin functions by the phospholipid and cholesterol contents of cell membranes

Gopalakrishna¹,

Chaubey²,

Manogaran³

et al. 2000

J. Cell. Biochem.

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Several modifications of the alpha5beta1 integrin, which alter its intracellular and extracellular interaction with fibronectin and other proteins, have been reported. However, the significance of the lateral mobility of integrin molecules in the plasma membrane, as a regulator of their distribution and function, is poorly understood. We examined this problem by increasing the cholesterol content of plasma membranes, and consequently modifying the fluidity of membrane phospholipids, in rat fibroblasts. Under these conditions, the clustering of alpha5beta1 integrin molecules in focal adhesions, their adhesion to the cell-binding domain of fibronectin, and their association with the cytoskeletal protein talin were significantly enhanced as compared to control cells. However, the activation of MAP-kinase pathways by the association of fibronectin with alpha5beta1 integrin, and its association with integrin-linked kinase (ilk), were suppressed. The treated cells also showed distinct changes in shape, and their actin stress fiber network was more dense and thick as compared to control cells. The changes in fluidity of phospholipids occurred differentially and fluidity of phosphatidyl-ethanolamine increased, while that of phosphatidyl-choline was reduced. Our results suggest that proteins in focal adhesions could be partitioned in specific lipid domains, which regulate specific aspects of alpha5beta1 integrin functions.

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Regulatory role of adenosine deaminase complexing protein (dipeptidyl peptidase IV = CD26) on the malignancy marker adenosine deaminase: Effect of membrane cholesterol and phase-transition

et al. 2000

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Membrane Cholesterol Dynamics: Cholesterol Domains and Kinetic Pools

Cited by 251 publications

References 0 publications

Fluorescence Techniques Using Dehydroergosterol to Study Cholesterol Trafficking

Fluorescence Techniques Using Dehydroergosterol to Study Cholesterol Trafficking

Modulation of ?5?1 integrin functions by the phospholipid and cholesterol contents of cell membranes

Regulatory role of adenosine deaminase complexing protein (dipeptidyl peptidase IV = CD26) on the malignancy marker adenosine deaminase: Effect of membrane cholesterol and phase-transition

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