Assembly of glycoprotein-80 adhesion complexes in Dictyostelium. Receptor compartmentalization and oligomerization in membrane rafts.
The phospholipid-anchored membrane glycoprotein (gp)-80 mediates cell-cell adhesion through a homophilic trans-interaction mechanism during Dictyostelium development and is enriched in a Triton X-100-insoluble floating fraction. To elucidate how gp80 adhesion complexes assemble in the plasma membrane, gp80-gp80 and gp80-raft interactions were investigated. A low density raft-like membrane fraction was isolated using a detergent-free method. It was enriched in sterols, the phospholipid-anchored proteins gp80, gp138, and ponticulin, as well as DdCD36 and actin, corresponding to components found in the Triton X-100-insoluble floating fraction. Chemical cross-linking revealed that gp80 oligomers were enriched in the raft-like membrane fraction, implicating stable oligomer-raft interactions. However, gp80 oligomers resisted sterol sequestration and were partially dissociated with Triton X-100, suggesting that compartmentalization in rafts was not solely responsible for their formation. The trans-dimer known to mediate adhesion was identified, but cis-oligomerization predominated and displayed greater accumulation during development. In fact, oligomerization was dependent on the level of gp80 expression and occurred among isolated gp80 extracellular domains, indicating that it was mediated by direct gp80-gp80 interactions. Rafts existed in gp80-null cells and such pre-existent membrane domains may provide optimal microenvironments for gp80 cis-oligomerization and the assembly of adhesion complexes.
Harris
T J
TJ
Banting and Best Department of Medical Research, University of Toronto, Toronto, Ontario M5G 1L6, Canada.
Ravandi
A
A
Siu
C H
CH
D000818:Animals; D002448:Cell Adhesion / Q000502:physiology*; D002458:Cell Fractionation; D003432:Cross-Linking Reagents / Q000494:pharmacology; D003902:Detergents / Q000737:chemistry; D004023:Dictyostelium / Q000166:cytology / Q000254:growth & development / Q000378:metabolism*; D050356:Lipid Metabolism; D008055:Lipids / Q000737:chemistry; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D021962:Membrane Microdomains / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D008954:Models, Biological; D017830:Octoxynol / Q000737:chemistry; D011108:Polymers / Q000737:chemistry / Q000378:metabolism; D013261:Sterols / Q000378:metabolism
D003432:Cross-Linking Reagents; D003902:Detergents; D008055:Lipids; D008562:Membrane Glycoproteins; D011108:Polymers; D013261:Sterols; C057189:membrane glycoprotein 80; D017830:Octoxynol