Fusion of Liposomes with the Plasma Membrane of Epithelial Cells: Fate of Incorporated Lipids as Followed by Freeze Fracture and Autoradiography of Plastic Sections

• Published in: The Journal of cell biology Vol. 107; no. 6; pp. 2511 - 2521
• Main Authors: Knoll, Gerd, Koert N. J. Burger, Bron, Romke, van Meer, Gerrit, Verkleij, Arie J.
• Format: Journal Article
• Language: English
• Published: New York, NY Rockefeller University Press 01.12.1988; The Rockefeller University Press
• Subjects: Animals; Autoradiography; Biological and medical sciences; Cell Line; Cell Membrane - metabolism; Cell membranes; Cell physiology; Cells; Cold Temperature; Endocytosis; Epithelial Cells; epithelium; Epithelium - metabolism; Freeze Fracturing; Freezing; Fundamental and applied biological sciences. Psychology; Hydrogen-Ion Concentration; kidney; Lipids; Lipids - analysis; Liposomes; Liposomes - metabolism; Membrane Fusion; Microscopy, Electron; Molecular and cellular biology; Orthomyxoviridae; P branes; Phospholipids; Plasma diffusion; Fissipedia; Cell culture; Membrane fusion; Carnivora; Incorporation; Radiolabelling; Liposome; Lipids; Electron microscopy; Kidney; Vertebrata; Endocytosis; Optical microscopy; Mammalia; Plasma membrane; Epithelial cell; Freeze etching; Dog
• ISSN: 0021-9525; 1540-8140
• DOI: 10.1083/jcb.107.6.2511

The fusion of liposomes with the plasma membrane of influenza virus-infected monolayers of an epithelial cell line, Madin-Darby canine kidney cells, has been analyzed by morphological techniques. The distribution of liposomal lipids over the apical and basolateral plasma membrane domains after fusion was assessed by autoradiography of liposomal [3H]dipalmitoylphosphatidylcholine after rapid freezing or chemical fixation and further processing by freeze substitution and low temperature embedding. Before fusion, radioactivity was solely detected on the apical cell surface, indicating the absence of redistribution artifacts and demonstrating the reliability of lipid autoradiography on both a light and electron microscopical level. After induction of fusion by a low pH treatment, the basolateral plasma membrane domain became progressively labeled, indicative of rapid lateral diffusion of [3H]dipalmitoylphosphatidylcholine in the plasma membrane. Analysis of individual fusion events by freeze fracture after rapid freezing confirmed the rapid diffusion of the liposomal lipids into the plasma membrane, as intramembrane particle-free lipid patches were never observed. After the induction of liposome-cell fusion, well-defined intramembrane particles were present on the otherwise smooth liposomal fracture faces and on the fracture faces of the plasma membrane. Morphological evidence thus was obtained in favor of a local point fusion mechanism with an intramembrane particle as a specific structural fusion intermediate.