Lipid Domain Structure of the Plasma Membrane Revealed by Patching of Membrane Components

Harder, Thomas; Scheiffele, Peter; Verkade, Paul; Simons, Kai

doi:10.1083/jcb.141.4.929

Cited by 1,132 publications

(1,114 citation statements)

References 69 publications

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“…The observed FRET between Thy-1.1 and Thy-1.2 could thus reflect small changes in topography of the target molecules induced by their dimerization and movement into the same clusters, a situation that has been observed in another system [17]. In order to determine whether Ab-induced dimerization does indeed contribute to FRET, we labeled Thy-1.1 with either monovalent OX7(Fab)-FITC or dimerizing OX7(IgG 1 )-FITC probes.…”

Section: Resultsmentioning

confidence: 65%

Topography of plasma membrane microdomains and its consequences for mast cell signaling

et al. 2006

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Thy-1 (CD90) is a glycoprotein bound to the plasma membrane by a GPI anchor. Aggregation of Thy-1 in mast cells and basophils induces activation events independent of the expression of Fce receptor I (FceRI). Although we and others have previously suggested that plasma membrane microdomains called lipid rafts are implicated in both Thy-1 and FceRI signaling, properties of these microdomains are still poorly understood. In this study we used rat basophilic leukemia cells and their transfectants expressing both endogenous Thy-1.1 and exogenous Thy-1.2 genes and analyzed topography of the Thy-1 isoforms and Thy-1-induced signaling events. Light microscopy showed that both Thy-1 isoforms were in the plasma membrane distributed randomly and independently. Electron microscopy on isolated membrane sheets and fluorescence resonance energy transfer analysis indicated cross-talk between Thy-1 isoforms and between Thy-1 and FceRI. This cross-talk was dependent on actin filaments. Thy-1 aggregates colocalized with two transmembrane adaptor proteins, non-T cell activation linker (NTAL) and linker for activation of T cells (LAT), which had been shown to inhabit different membrane microdomains. Thy-1 aggregation led to tyrosine phosphorylation of these two adaptors. The combined data indicate that aggregated GPI-anchored proteins can attract different membrane proteins in different clusters and thus can trigger different signaling pathways.

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Section: Resultsmentioning

confidence: 65%

Topography of plasma membrane microdomains and its consequences for mast cell signaling

et al. 2006

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“…The size of these domains could be increased in response to protein oligomerization (Harder et al, 1998). On the basis of these presumptions, dIgA-mediated dimerization of pIgR (Singer and Mostov, 1998) could have increased interactions Figure 7.…”

Section: Discussionmentioning

confidence: 99%

Cholesterol-sensitive Modulation of Transcytosis

Leyt

Melamed-Book

Vaerman

et al. 2007

MBoC

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Cholesterol-rich membrane domains (e.g., lipid rafts) are thought to act as molecular sorting machines, capable of coordinating the organization of signal transduction pathways within limited regions of the plasma membrane and organelles. The significance of these domains in polarized postendocytic sorting is currently not understood. We show that dimeric IgA stimulates the incorporation of its receptor into cholesterol-sensitive detergent-resistant membranes confined to the basolateral surface/basolateral endosomes. A fraction of human transferrin receptor was also found in basolateral detergent-resistant membranes. Disrupting these membrane domains by cholesterol depletion (using methyl-␤-cyclodextrin) before ligand-receptor internalization caused depolarization of traffic from endosomes, suggesting that cholesterol in basolateral lipid rafts plays a role in polarized sorting after endocytosis. In contrast, cholesterol depletion performed after ligand internalization stimulated cargo transcytosis. It also stimulated caveolin-1 phosphorylation on tyrosine 14 and the appearance of the activated protein in dimeric IgA-containing apical organelles. We propose that cholesterol depletion stimulates the coupling of transcytotic and caveolin-1 signaling pathways, consequently prompting the membranes to shuttle from endosomes to the plasma membrane. This process may represent a unique compensatory mechanism required to maintain cholesterol balance on the cell surface of polarized epithelia. INTRODUCTIONIn humans, the major antibody that mediates immunological defense against mucosal infections is dimeric IgA (dIgA). This antibody is produced by plasma cells in the lamina propria located underneath the mucosal surface (reviewed in Lamm et al., 1995;Rojas and Apodaca, 2002) and is carried from the blood to mucosal secretions by the polymeric immunoglobulin receptor (pIgR). The itinerary of pIgR and its ligand was intensively studied in the polarized MadinDarby canine kidney (MDCK) cell line that heterologously expresses rabbit pIgR (for a recent review, see Rojas and Apodaca, 2002). The receptor is initially delivered from the trans-Golgi network (TGN) to the basolateral plasma membrane, where it encounters and binds dIgA. The complex is subsequently internalized and transcytosed to the apical plasma membrane of the cell. At that surface, the ectodomain of pIgR is proteolytically cleaved off to yield the secretory component (SC), which is released together with dIgA into mucosal secretions. In the course of transcytosis, pIgRdIgA complexes traverse various endosomal compartments, among which is an endocytic compartment located immediately underneath the apical surface (Gibson et al., 1998). This apical compartment, defined as apical recycling endosomes (AREs), is enriched in dIgA and is relatively deficient in recycling transferrin (Apodaca et al., 1994;Barroso and Sztul, 1994;Brown et al., 2000). AREs are thought to play a unique role in transcytosis, possibly by exploiting apical recycling mechanisms to shuttle transcy...

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“…Such domains containing clustered GPI-anchored proteins can sometimes be detected by conventional light or electron microscopy (Matsuura et al 1984;Latker et al 1987;Kobayashi and Robinson, 1991;van den Berg et al, 1995) (reviewed in Anderson [1998]). In other cases clusters of GPI-anchored proteins or other lipid raft components are only apparent when they have been crosslinked with secondary antibodies (Howell et al 1987;Rothberg et al, 1990;Fra et al, 1994;Mayor et al, 1994;Parton et al, 1994;Fujimoto, 1996;Harder et al, 1998). This suggests that in vivo, lipid rafts may be small or dynamic structures that are aggregated and stabilized by detergent extraction or by antibody-induced cross-linking (Edidin, 1997;Harder and Simons, 1997;Simons and Ikonen, 1997;Weimbs et al, 1997;Brown and London, 1998;Jacobson and Dietrich, 1999).…”

Section: Introductionmentioning

confidence: 99%

High-Resolution FRET Microscopy of Cholera Toxin B-Subunit and GPI-anchored Proteins in Cell Plasma Membranes

Kenworthy

Petranova

Edidin

2000

MBoC

406

311

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“Lipid rafts” enriched in glycosphingolipids (GSL), GPI-anchored proteins, and cholesterol have been proposed as functional microdomains in cell membranes. However, evidence supporting their existence has been indirect and controversial. In the past year, two studies used fluorescence resonance energy transfer (FRET) microscopy to probe for the presence of lipid rafts; rafts here would be defined as membrane domains containing clustered GPI-anchored proteins at the cell surface. The results of these studies, each based on a single protein, gave conflicting views of rafts. To address the source of this discrepancy, we have now used FRET to study three different GPI-anchored proteins and a GSL endogenous to several different cell types. FRET was detected between molecules of the GSL GM1 labeled with cholera toxin B-subunit and between antibody-labeled GPI-anchored proteins, showing these raft markers are in submicrometer proximity in the plasma membrane. However, in most cases FRET correlated with the surface density of the lipid raft marker, a result inconsistent with significant clustering in microdomains. We conclude that in the plasma membrane, lipid rafts either exist only as transiently stabilized structures or, if stable, comprise at most a minor fraction of the cell surface.

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Lipid Domain Structure of the Plasma Membrane Revealed by Patching of Membrane Components

Cited by 1,132 publications

References 69 publications

Topography of plasma membrane microdomains and its consequences for mast cell signaling

Topography of plasma membrane microdomains and its consequences for mast cell signaling

Cholesterol-sensitive Modulation of Transcytosis

High-Resolution FRET Microscopy of Cholera Toxin B-Subunit and GPI-anchored Proteins in Cell Plasma Membranes

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