A Role for Lipid Shells in Targeting Proteins to Caveolae, Rafts, and Other Lipid Domains

Anderson, Richard G. W.; Jacobson, Ken

doi:10.1126/science.1068886

Cited by 1,083 publications

(884 citation statements)

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“…Larger, even more stable domains can be formed, but these require proteinprotein interactions and/or crosslinking -as with the creation of T-cell-signalling domains 31 . Note that this model is different from the previously advanced lipid-shell hypothesis 32 , which suggests that lipid shells that are formed around lipid anchors operate as targeting motifs that allow interaction of the shelled protein with pre-existing lipid rafts or caveolae 32 .…”

Section: Lipid Rafts In the Intact Plasma Membrane?mentioning

confidence: 66%

Lipid rafts: contentious only from simplistic standpoints

Hancock

2006

Nat Rev Mol Cell Biol

751

716

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The hypothesis that lipid rafts exist in plasma membranes and have crucial biological functions remains controversial. The lateral heterogeneity of proteins in the plasma membrane is undisputed, but the contribution of cholesterol-dependent lipid assemblies to this complex, non-random organization promotes vigorous debate. In the light of recent studies with model membranes, computational modelling and innovative cell biology, I propose an updated model of lipid rafts that readily accommodates diverse views on plasma-membrane micro-organization.A widely accepted hypothesis in contemporary cell biology is that freely diffusing, stable, lateral assemblies of sphingolipids and cholesterol, which are termed lipid rafts 1-3 , constitute an important organizing principle for the plasma membrane. The basic concept is that lipid rafts can facilitate selective protein-protein interactions by selectively excluding or including proteins. This lipid-based sorting mechanism has been widely implicated in the assembly of transient signalling platforms and more permanent structures such as the immunological synapse, as well as in the sorting of proteins for entry into specific exocytic and endocytic trafficking pathways [1][2][3] . Despite the undoubted theoretical utility of lipid rafts to many cell biological processes, the basic hypothesis that stable lipid rafts exist at all in biological membranes is under intense scrutiny 4,5 . This is partly because lipid rafts, if they exist in resting cell membranes, are too small to be resolved by fluorescent microscopy and have no defined ultrastructure; therefore, proving their existence is problematic. This article will consider the biophysical properties of model membranes that underpin the lipid raft hypothesis, and the limitations of the biochemical approaches that have been used to study rafts in biological membranes that largely account for the current debate on the hypothesis mentioned above. After examining the challenges that are involved in correlating observations, which have been made in model and cellular membranes, I focus on synthesizing recent data on the size of lipid domains in model membranes with observations that have been obtained by imaging intact plasma membranes. These imaging approaches include single particle tracking (SPT), single fluorophore video tracking (SFVT), fluorescence resonance energy transfer (FRET), homo-FRET and electron microscopy (EM). On the one hand, these studies challenge the simplistic null hypothesis that lipid-based assemblies, such as lipid rafts, do not exist in biological membranes. On the other hand, it is timely to reconsider the raft hypothesis in the light of these new data because the consensus model that emerges is more complex than a simplistic notion of stable, freely diffusing lipid rafts. Some intriguing new questions aboutCompeting interests statement The author declares no competing financial interests. DATABASES

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Section: Lipid Rafts In the Intact Plasma Membrane?mentioning

confidence: 66%

Lipid rafts: contentious only from simplistic standpoints

Hancock

2006

Nat Rev Mol Cell Biol

751

716

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“…In this interpretation, each membrane protein is embedded in a unique lattice consisting of tens of specific lipid molecules [1]. This allows for a high variety of different membrane domains potentially formed.…”

Section: Plasma Membrane Domains In Yeastmentioning

confidence: 99%

The lateral compartmentation of the yeast plasma membrane

Malínský

Opekarová

Tanner

2010

Yeast

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The plasma membrane of Saccharomyces cerevisiae contains large microdomains enriched in ergosterol, which house at least nine integral proteins, including proton symporters. The domains adopt a characteristic structure of furrow-like invaginations typically seen in freeze-fracture pictures of fungal cells. Being stable for the time comparable with the cell cycle duration, they might be considered as fixed islands (rafts) in an otherwise fluid yeast plasma membrane. Rapidly moving endocytic marker proteins avoid the microdomains; the domain-accumulated proton symporters consequently show a reduced rate of substrate-induced endocytosis and turnover.

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“…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%

“…Successful application of the fluorescent DHE to investigation of the function and organization of sterols in membranes, especially lipid raft/ caveolae microdomains of living cells, requires the use of highly purified DHE. This is due to the fact that lipid rafts/caveolae are highly sensitive not only to cholesterol content (58,60,82,84,85,87,88), but also to sterol structure (89), and sterol oxidation. The present review yielded new insights into this problem and potential applications of DHE imaging in living cells.…”

mentioning

confidence: 99%

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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A Role for Lipid Shells in Targeting Proteins to Caveolae, Rafts, and Other Lipid Domains

Cited by 1,083 publications

References 78 publications

Lipid rafts: contentious only from simplistic standpoints

Lipid rafts: contentious only from simplistic standpoints

The lateral compartmentation of the yeast plasma membrane

Fluorescence Techniques Using Dehydroergosterol to Study Cholesterol Trafficking

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