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© The Rockefeller University Press, 0021-9525/1998//929 $5.00
The Journal of Cell Biology, Volume 141, Number 4, , 1998 929-942

Articles

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



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

European Molecular Biology Laboratory, Cell Biology Programme, 69117 Heidelberg, Germany

Lateral assemblies of glycolipids and cholesterol, "rafts," have been implicated to play a role in cellular processes like membrane sorting, signal transduction, and cell adhesion. We studied the structure of raft domains in the plasma membrane of non-polarized cells. Overexpressed plasma membrane markers were evenly distributed in the plasma membrane. We compared the patching behavior of pairs of raft markers (defined by insolubility in Triton X-100) with pairs of raft/non-raft markers. For this purpose we cross-linked glycosyl-phosphatidylinositol (GPI)-anchored proteins placental alkaline phosphatase (PLAP), Thy-1, influenza virus hemagglutinin (HA), and the raft lipid ganglioside GM1 using antibodies and/or cholera toxin. The patches of these raft markers overlapped extensively in BHK cells as well as in Jurkat T–lymphoma cells. Importantly, patches of GPI-anchored PLAP accumulated src-like protein tyrosine kinase fyn, which is thought to be anchored in the cytoplasmic leaflet of raft domains. In contrast patched raft components and patches of transferrin receptor as a non-raft marker were sharply separated. Taken together, our data strongly suggest that coalescence of cross-linked raft elements is mediated by their common lipid environments, whereas separation of raft and non-raft patches is caused by the immiscibility of different lipid phases. This view is supported by the finding that cholesterol depletion abrogated segregation. Our results are consistent with the view that raft domains in the plasma membrane of non-polarized cells are normally small and highly dispersed but that raft size can be modulated by oligomerization of raft components.

Abbreviations used in this paper: CTx, choleratoxin B subunit; DIGs, detergent-insoluble glycolipid-enriched complexes; GM1, ganglioside GM1; GPI, glycosyl-phosphatidylinositol; HA, hemagglutinin; hTfR, human transferrin receptor; LDL-R, low density lipoprotein receptor; PC, phosphatidylcholine; PLAP, placental alkaline phosphatase; VSV-G, vesicular stomatitis virus glycoprotein.

T. Harder is supported by a Deutsche Forschurgsgemeinschaft research fellowship.

T. Harder and P. Scheiffele contributed equally to this work.


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