title: Membrane cholesterol regulates LFA-1 function and lipid raft heterogeneity.
abstract: Many surface receptors and signaling molecules are thought to associate with unique membrane microdomains termed lipid rafts. We examined the involvement of lipid rafts in the activation of leukocyte function-associated antigen-1 (LFA-1). Depletion or sequestration of cholesterol with methyl-beta-cyclodextrin (MCD) or filipin, respectively, strongly inhibited LFA-1-mediated adhesion of T-cell lines and primary T cells. This inhibition was reversed by cholesterol reconstitution. LFA-1 on T-cell lines was detected in cold Triton X-100-insoluble lipid rafts, which were disrupted by MCD or filipin treatment. However, no LFA-1 on primary T cells was detected in lipid rafts isolated by the same procedures, and these rafts were resistant to cholesterol depletion or sequestration. Association of LFA-1 with lipid rafts of primary T cells could be detected only when they were isolated with another nonionic detergent, Brij 35. Upon treatment with MCD, LFA-1 in Brij 35-insoluble lipid rafts partially shifted to nonraft fractions. T-cell lines were found to have a high level of cholesterol and a low level of ganglioside GM1, a common marker for lipid rafts, whereas primary T cells have a much lower level of cholesterol and a very high amount of GM1. Cross-linking of LFA-1 on primary T cells induced cocapping of cholesterol but not GM1. These results suggest that lipid rafts of T cells are heterogenous, and LFA-1 associates with a subset of lipid rafts containing a high level of cholesterol. This association seems to regulate LFA-1 functions, possibly by facilitating LFA-1 clustering.
authors.affiliation: Terry Fox Laboratory, British Columbia Cancer Agency, 601 W 10th Ave, Vancouver, BC, V5Z 1L3, Canada.
authors.forename: Muhammad Reza
authors.initials: MR
authors.lastname: Marwali
authors.forename: Jose
authors.initials: J
authors.lastname: Rey-Ladino
authors.forename: Lisa
authors.initials: L
authors.lastname: Dreolini
authors.forename: Douglas
authors.initials: D
authors.lastname: Shaw
authors.forename: Fumio
authors.initials: F
authors.lastname: Takei
mesh_terms: D000818:Animals; D002448:Cell Adhesion; D002784:Cholesterol / Q000032:analysis / Q000502:physiology*; D003505:Cyclodextrins / Q000494:pharmacology; D005372:Filipin / Q000494:pharmacology; D005677:G(M1) Ganglioside / Q000032:analysis; D007152:Immunologic Capping; D016169:Lymphocyte Function-Associated Antigen-1 / Q000032:analysis / Q000276:immunology / Q000502:physiology*; D021962:Membrane Microdomains / Q000737:chemistry* / Q000187:drug effects / Q000502:physiology; D051379:Mice; D017830:Octoxynol; D000077423:Polidocanol; D011092:Polyethylene Glycols; D013601:T-Lymphocytes / Q000166:cytology / Q000378:metabolism / Q000648:ultrastructure*; D014407:Tumor Cells, Cultured; D047392:beta-Cyclodextrins
chemical_list: D003505:Cyclodextrins; D016169:Lymphocyte Function-Associated Antigen-1; D047392:beta-Cyclodextrins; C108732:methyl-beta-cyclodextrin; D000077423:Polidocanol; D005677:G(M1) Ganglioside; D011092:Polyethylene Glycols; D005372:Filipin; D017830:Octoxynol; D002784:Cholesterol