title: Interdependence of laminin-mediated clustering of lipid rafts and the dystrophin complex in astrocytes.
abstract: Astrocyte endfeet surrounding blood vessels are active domains involved in water and potassium ion transport crucial to the maintenance of water and potassium ion homeostasis in brain. A growing body of evidence points to a role for dystroglycan and its interaction with perivascular laminin in the targeting of the dystrophin complex and the water-permeable channel, aquaporin 4 (AQP4), at astrocyte endfeet. However, the mechanisms underlying such compartmentalization remain poorly understood. In the present study we found that AQP4 resided in Triton X-100-insoluble fraction, whereas dystroglycan was recovered in the soluble fraction in astrocytes. Cholesterol depletion resulted in the translocation of a pool of AQP4 to the soluble fraction indicating that its distribution is indeed associated with cholesterol-rich membrane domains. Upon laminin treatment AQP4 and the dystrophin complex, including dystroglycan, reorganized into laminin-associated clusters enriched for the lipid raft markers GM1 and flotillin-1 but not caveolin-1. Reduced diffusion rates of GM1 in the laminin-induced clusters were indicative of the reorganization of raft components in these domains. In addition, both cholesterol depletion and dystroglycan silencing reduced the number and area of laminin-induced clusters of GM1, AQP4, and dystroglycan. These findings demonstrate the interdependence between laminin binding to dystroglycan and GM1-containing lipid raft reorganization and provide novel insight into the dystrophin complex regulation of AQP4 polarization in astrocytes.
authors.affiliation: Department of Cellular and Physiological Sciences, University of British Columbia, Vancouver V6T 1Z3, Canada.
authors.forename: Geoffroy
authors.initials: G
authors.lastname: Noël
authors.forename: Daniel Kai Long
authors.initials: DK
authors.lastname: Tham
authors.forename: Hakima
authors.initials: H
authors.lastname: Moukhles
mesh_terms: D000818:Animals; D051401:Aquaporin 4 / Q000235:genetics / Q000378:metabolism; D001253:Astrocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D015153:Blotting, Western; D002478:Cells, Cultured; D002784:Cholesterol / Q000378:metabolism; D003902:Detergents / Q000737:chemistry / Q000494:pharmacology; D049030:Dystroglycans / Q000235:genetics / Q000378:metabolism; D016189:Dystrophin / Q000378:metabolism*; D016650:Fluorescein-5-isothiocyanate / Q000737:chemistry; D036681:Fluorescence Recovery After Photobleaching; D005455:Fluorescent Antibody Technique; D005677:G(M1) Ganglioside / Q000378:metabolism; D007797:Laminin / Q000378:metabolism* / Q000494:pharmacology; D021962:Membrane Microdomains / Q000187:drug effects / Q000378:metabolism*; D011485:Protein Binding; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D014162:Transfection
chemical_list: C496032:Aqp4 protein, rat; D051401:Aquaporin 4; D003902:Detergents; D016189:Dystrophin; D007797:Laminin; D034741:RNA, Small Interfering; C121346:laminin 1; D049030:Dystroglycans; D005677:G(M1) Ganglioside; D002784:Cholesterol; D016650:Fluorescein-5-isothiocyanate