Dynamin-mediated Internalization of Caveolae

Henley, John R.; Krueger, Eugene W.; Oswald, Barbara; McNiven, Mark A.

doi:10.1083/jcb.141.1.85

Cited by 703 publications

(570 citation statements)

References 97 publications

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“…Hudy-1 antibody was generously supplied by S. Schmid (The Scripps Research Institute) (18), and ␣-dyn (D25520) was purchased from Transduction Laboratories. Peptide antibodies MC13 and MC63 were kindly provided by M. McNiven (28). All other chemicals were reagent grade.…”

Section: Methodsmentioning

confidence: 99%

Dynamin Assembles into Spirals under Physiological Salt Conditions upon the Addition of GDP and γ-Phosphate Analogues

Carr

Hinshaw

1997

Journal of Biological Chemistry

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Dynamin is a 100-kDa GTPase that is believed to be involved in the constriction of clathrin-coated pits and the fission of clathrin-coated vesicles during receptormediated endocytosis and during membrane retrieval in nerve termini. It has been shown that purified dynamin incubated under low salt conditions forms rings and spirals that, in dimension and appearance, resemble the dense material occasionally observed at the necks of coated pits. In this report we show that purified dynamin forms spirals under physiological salt conditions when incubated with GDP and ␥-phosphate analogues (beryllium and aluminum fluoride) or when dialyzed into guanosine 5 -3-O-(thio)triphosphate. Moreover, spirals still form when dynamin is proteolyzed to either a predominant ϳ90-kDa species, lacking the C terminus, or to two smaller fragments, a ϳ55-kDa species originating from the N-terminal half of the protein and a ϳ30-kDa species lacking both the N and C termini. This work indicates that the addition of GDP and ␥-phosphate analogues arrests dynamin in a GTP or transition state that markedly stabilizes the spiral conformation under physiological ionic strength conditions and thereby suggests that dynamin in the absence of a receptor is capable of assembly into spirals at the necks of coated pits prior to vesicle fission.

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Section: Methodsmentioning

confidence: 99%

Dynamin Assembles into Spirals under Physiological Salt Conditions upon the Addition of GDP and γ-Phosphate Analogues

Carr

Hinshaw

1997

Journal of Biological Chemistry

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“…CTXB is enriched in biochemical lipid raft fractions and in caveolae (Fra et al, 1994;Parton et al, 1994;Parton, 1996;Smart et al, 1995;Schnitzer et al, 1996;Harder and Simons, 1997;Stauffer and Meyer, 1997;Henley et al, 1998;Orlandi and Fishman, 1998;Wolf et al, 1998;Lencer et al, 1999). Three GPI-anchored proteins, which are major components of detergentinsoluble raft fractions (Skibbens et al, 1989;Stefanova et al, 1991;Brown and Rose, 1992;Fiedler et al, 1993;Sargiacomo et al, 1993), were used as protein markers for lipid rafts.…”

Section: Plasma Membrane Distribution Of Lipid Raft Markers Detected mentioning

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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“…The better characterized pathways are mediated by clathrin-and caveolae-coated endocytic invaginations and require dynamin activity to be severed from the plasma membrane (Damke et al, 1994;Hinshaw and Schmid, 1995;Henley et al, 1998;Oh et al, 1998). There also seem to be one or more clathrin-independent pathways where dynamin is required, for example, the pathway for internalization of the interleukin (IL)-2 receptor subunits (Lamaze et al, 2001).…”

Section: Introductionmentioning

confidence: 99%

Arf6-independent GPI-anchored Protein-enriched Early Endosomal Compartments Fuse with Sorting Endosomes via a Rab5/Phosphatidylinositol-3′-Kinase–dependent Machinery

Kalia

Kumari

Chadda

et al. 2006

MBoC

107

117

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In the process of internalization of molecules from the extracellular milieu, a cell uses multiple endocytic pathways, consequently generating different endocytic vesicles. These primary endocytic vesicles are targeted to specific destinations inside the cell. Here, we show that GPI-anchored proteins are internalized by an Arf6-independent mechanism into GPI-anchored protein-enriched early endosomal compartments (GEECs). Internalized GPI-anchored proteins and the fluid phase are first visualized in GEECs that are acidic, primary endocytic structures, negative for early endosomal markers, Rab4, Rab5, and early endosome antigen (EEA)1. They subsequently acquire Rab5 and EEA1 before homotypic fusion with other GEECs, and heterotypic fusion with endosomes containing cargo from the clathrin-dependent endocytic pathway. Although, the formation of GEECs is unaffected by inhibition of Rab5 GTPase and phosphatidylinositol-3 -kinase (PI3K) activity, their fusion with sorting endosomes is dependent on both activities. Overexpression of Rab5 reverts PI3K inhibition of fusion, providing evidence that Rab5 effectors play important roles in heterotypic fusion between the dynamin-independent GEECs and clathrin-and dynamin-dependent sorting endosomes. INTRODUCTIONA cell uses multiple endocytic pathways for the uptake of molecules from the extracellular milieu (Mukherjee et al., 1997;Conner and Schmid, 2003). The better characterized pathways are mediated by clathrin-and caveolae-coated endocytic invaginations and require dynamin activity to be severed from the plasma membrane (Damke et al., 1994;Hinshaw and Schmid, 1995;Henley et al., 1998;Oh et al., 1998). There also seem to be one or more clathrin-independent pathways where dynamin is required, for example, the pathway for internalization of the interleukin (IL)-2 receptor subunits (Lamaze et al., 2001). In addition, there are a growing number of dynamin-independent endocytic processes that serve as internalization routes for a number of cell surface proteins, including lipid-linked proteins, such as GPI-anchored proteins (Lamaze and Schmid, 1995;Conner and Schmid, 2003;Nichols, 2003;Mayor and Riezman, 2004;Naslavsky et al., 2004;.GPI-anchored proteins are internalized via a dynamin, caveolin, and clathrin-independent pathway that is also responsible for the entry of a sizable fraction of the fluid phase in a number of cell types (Fivaz et al., 2002;Sabharanjak et al., 2002;Guha et al., 2003;. GPI-anchored proteins and the fluid phase enter the cell via tubular invaginations at the cell surface, into compartments termed GPI-anchored protein-enriched early endosomal compartments (GEECs) . At early times, these structures are largely devoid of cargo from the clathrin-dependent pathway. Besides a requirement for the small GTPase cdc42, very little is known about the molecular players that govern the formation and trafficking of GEECs. Much less is known about the detailed endocytic itinerary of these pathways (Lamaze and Schmid, 1995;Mayor and Riezman, 2004).After internalization...

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Dynamin-mediated Internalization of Caveolae

Cited by 703 publications

References 97 publications

Dynamin Assembles into Spirals under Physiological Salt Conditions upon the Addition of GDP and γ-Phosphate Analogues

Dynamin Assembles into Spirals under Physiological Salt Conditions upon the Addition of GDP and γ-Phosphate Analogues

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

Arf6-independent GPI-anchored Protein-enriched Early Endosomal Compartments Fuse with Sorting Endosomes via a Rab5/Phosphatidylinositol-3′-Kinase–dependent Machinery

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