Binding of cross-linked glycosylphosphatidylinositol-anchored proteins to discrete actin-associated sites and cholesterol-dependent domains

Kenichi Suzuki, Michael Sheetz

Research output: Contribution to journalArticle

51 Citations (Scopus)

Abstract

The mechanism by which cross-linked glycosylphosphatidylinositol (GPI)-anchored proteins are immobilized has been a mystery because both the binding to a transmembrane protein and attachment to a rigid cytoskeleton are needed. Using laser tweezers surface scanning resistance (SSR) technology, we obtained physical evidence for cross-linked GPI-anchored protein, Qa-2, binding to a transmembrane protein and for diffusion to discrete cytoskeleton attachment sites. At low levels of cross-linking of Qa-2 molecules, the resistance to lateral movement was that expected of monomeric lipid-anchored proteins, and no specific binding to cytoskeleton-attached structures was observed. When aggregates of the GPI-anchored protein, Qa-2, were scanned across plasma membranes, the background resistance was much higher than expected for a GPI-anchored protein alone and submicron domains of even higher resistance were observed (designated as elastic or non-elastic barriers) at a density of 82 (61 elastic and 21 small non-elastic barriers) per 100 μm2. Elastic barriers involved weak but specific bonds to the actin cytoskeleton (broken by forces of 2 or 4 pN and were removed by cytochalasin D). Small non-elastic barriers (50-100 nm) depended upon membrane cholesterol and were closely correlated with caveolae density. Thus, cross-linked GPI-anchored proteins can diffuse through the membrane in complex with a transmembrane protein and bind weakly to discrete cytoskeleton attachment sites either associated with flexible actin networks or sphingolipid-cholesterol rich microdomains in live cell membranes. Our SSR measurements provide the first description of the physical characteristics of the interactions between rafts and stable membrane structures.

Original languageEnglish (US)
Pages (from-to)2181-2189
Number of pages9
JournalBiophysical Journal
Volume81
Issue number4
DOIs
StatePublished - Jan 1 2001
Externally publishedYes

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GPI-Linked Proteins
Actins
Cytoskeleton
Cholesterol
Glycosylphosphatidylinositols
Proteins
Membranes
Lipid-Linked Proteins
Cell Membrane
Optical Tweezers
Cytochalasin D
Caveolae
Sphingolipids
Actin Cytoskeleton
Technology

ASJC Scopus subject areas

  • Biophysics

Cite this

Binding of cross-linked glycosylphosphatidylinositol-anchored proteins to discrete actin-associated sites and cholesterol-dependent domains. / Suzuki, Kenichi; Sheetz, Michael.

In: Biophysical Journal, Vol. 81, No. 4, 01.01.2001, p. 2181-2189.

Research output: Contribution to journalArticle

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