Detergent-resistant membrane isolation

2009 ◽  
Author(s):  
Abdur Rub
Keyword(s):  
Membrane Isolation ◽  
Detergent Resistant Membrane

Comparative lipid analysis and structure of detergent-resistant membrane raft fractions isolated from human and ruminant erythrocytes

2005 ◽  
Vol 434 (1) ◽  
pp. 150-158 ◽  
Author(s):  
Kamen S. Koumanov ◽  
Cedric Tessier ◽  
Albena B. Momchilova ◽  
Dominique Rainteau ◽  
Claude Wolf ◽  
...  
Keyword(s):  
Lipid Analysis ◽  
Membrane Raft ◽  
Detergent Resistant Membrane

scholarly journals The ER–Golgi intermediate compartment is a key membrane source for the LC3 lipidation step of autophagosome biogenesis

eLife ◽  
2013 ◽  
Vol 2 ◽  
Author(s):  
Liang Ge ◽  
David Melville ◽  
Min Zhang ◽  
Randy Schekman
Keyword(s):  
Autophagosome Formation ◽  
Functional Studies ◽  
A Cell ◽  
Intermediate Compartment ◽  
Membrane Isolation ◽  
Necessary And Sufficient ◽  
Organelle Membrane ◽  
Catabolic Process

Autophagy is a catabolic process for bulk degradation of cytosolic materials mediated by double-membraned autophagosomes. The membrane determinant to initiate the formation of autophagosomes remains elusive. Here, we establish a cell-free assay based on LC3 lipidation to define the organelle membrane supporting early autophagosome formation. In vitro LC3 lipidation requires energy and is subject to regulation by the pathways modulating autophagy in vivo. We developed a systematic membrane isolation scheme to identify the endoplasmic reticulum–Golgi intermediate compartment (ERGIC) as a primary membrane source both necessary and sufficient to trigger LC3 lipidation in vitro. Functional studies demonstrate that the ERGIC is required for autophagosome biogenesis in vivo. Moreover, we find that the ERGIC acts by recruiting the early autophagosome marker ATG14, a critical step for the generation of preautophagosomal membranes.

Fatty acid synthase drives the synthesis of phospholipids partitioning into detergent-resistant membrane microdomains

2003 ◽  
Vol 302 (4) ◽  
pp. 898-903 ◽  
Author(s):  
Johannes V Swinnen ◽  
Paul P Van Veldhoven ◽  
Leen Timmermans ◽  
Ellen De Schrijver ◽  
Koen Brusselmans ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Fatty Acid Synthase ◽  
Membrane Microdomains ◽  
Detergent Resistant Membrane

scholarly journals Membrane isolation on polylysine-coated beads. Plasma membrane from HeLa cells.

1977 ◽  
Vol 75 (1) ◽  
pp. 119-134 ◽  
Author(s):  
C M Cohen ◽  
D I Kalish ◽  
B S Jacobson ◽  
D Branton
Keyword(s):  
Adenosine Triphosphatase ◽  
Wheat Germ ◽  
Plasma Membranes ◽  
Cell Attachment ◽  
Isolation Procedure ◽  
Chemical Analyses ◽  
Isolation Techniques ◽  
Cell Plasma ◽  
Membrane Bound ◽  
Membrane Isolation

HeLa cell plasma membranes have been purified after binding cells to polylysine-coated polyacrylamide beads. Cell attachment to beads and membrane recovery were maximal in a sucrose-acetate buffer, pH 5.0, at 25 degrees C. Measurements of ouabain-sensitive NaK-adenosine triphosphatase, membrane-bound 125I-wheat germ agglutinin, and chemical analyses showed that membranes on beads were of comparable or greater purity than membranes isolated by conventional methods. Because the isolation procedure is rapid (approximately 2.5 h), and produces membranes whose protoplasmic surfaces are fully exposed, it should be a useful supplement to standard isolation techniques.

scholarly journals Detergent-Resistant Membrane Microdomains Facilitate Ib Oligomer Formation and Biological Activity of Clostridium perfringens Iota-Toxin

2004 ◽  
Vol 72 (4) ◽  
pp. 2186-2193 ◽  
Author(s):  
Martha L. Hale ◽  
Jean-Christophe Marvaud ◽  
Michel R. Popoff ◽  
Bradley G. Stiles
Keyword(s):  
Clostridium Perfringens ◽  
Cell Types ◽  
Cell Receptor ◽  
Vero Cells ◽  
Membrane Microdomains ◽  
Oligomer Formation ◽  
A Cell ◽  
Clostridium Perfringens Iota Toxin ◽  
Detergent Resistant Membrane ◽  
Soluble Fractions

ABSTRACT Clostridium perfringens iota-toxin consists of two separate proteins identified as a cell binding protein, iota b (Ib), which forms high-molecular-weight complexes on cells generating Na+/K+-permeable pores through which iota a (Ia), an ADP-ribosyltransferase, presumably enters the cytosol. Identity of the cell receptor and membrane domains involved in Ib binding, oligomer formation, and internalization is currently unknown. In this study, Vero (toxin-sensitive) and MRC-5 (toxin-resistant) cells were incubated with Ib, after which detergent-resistant membrane microdomains (DRMs) were extracted with cold Triton X-100. Western blotting revealed that Ib oligomers localized in DRMs extracted from Vero, but not MRC-5, cells while monomeric Ib was detected in the detergent-soluble fractions of both cell types. The Ib protoxin, previously shown to bind Vero cells but not form oligomers or induce cytotoxicity, was detected only in the soluble fractions. Vero cells pretreated with phosphatidylinositol-specific phospholipase C before addition of Ib indicated that glycosylphosphatidyl inositol-anchored proteins were minimally involved in Ib binding or oligomer formation. While pretreatment of Vero cells with filipin (which sequesters cholesterol) had no effect, methyl-β-cyclodextrin (which extracts cholesterol) reduced Ib binding and oligomer formation and delayed iota-toxin cytotoxicity. These studies showed that iota-toxin exploits DRMs for oligomer formation to intoxicate cells.

Sign in / Sign up

Export Citation Format

Share Document