title: Gangliosides and beta1-integrin are required for caveolae and membrane domains.
abstract: Caveolae are plasma membrane domains involved in the uptake of certain pathogens and toxins. Internalization of some cell surface integrins occurs via caveolae suggesting caveolae may play a crucial role in modulating integrin-mediated adhesion and cell migration. Here we demonstrate a critical role for gangliosides (sialo-glycosphingolipids) in regulating caveolar endocytosis in human skin fibroblasts. Pretreatment of cells with endoglycoceramidase (cleaves glycosphingolipids) or sialidase (modifies cell surface gangliosides and glycoproteins) selectively inhibited caveolar endocytosis by >70%, inhibited the formation of plasma membrane domains enriched in sphingolipids and cholesterol ('lipid rafts'), reduced caveolae and caveolin-1 at the plasma membrane by approximately 80%, and blunted activation of beta1-integrin, a protein required for caveolar endocytosis in these cells. These effects could be reversed by a brief incubation with gangliosides (but not with asialo-gangliosides or other sphingolipids) at 10 degrees C, suggesting that sialo-lipids are critical in supporting caveolar endocytosis. Endoglycoceramidase treatment also caused a redistribution of focal adhesion kinase, paxillin, talin, and PIP Kinase Igamma away from focal adhesions. The effects of sialidase or endoglycoceramidase on membrane domains and the distribution of caveolin-1 could be recapitulated by beta1-integrin knockdown. These results suggest that both gangliosides and beta1-integrin are required for maintenance of caveolae and plasma membrane domains.
authors.affiliation: Departments of Medicine, Biochemistry and Molecular Biology, Mayo Clinic College of Medicine, Rochester, MN 55905, USA.
authors.forename: Raman Deep
authors.initials: RD
authors.lastname: Singh
authors.forename: David L
authors.initials: DL
authors.lastname: Marks
authors.forename: Eileen L
authors.initials: EL
authors.lastname: Holicky
authors.forename: Christine L
authors.initials: CL
authors.lastname: Wheatley
authors.forename: Tatiana
authors.initials: T
authors.lastname: Kaptzan
authors.forename: Satoshi B
authors.initials: SB
authors.lastname: Sato
authors.forename: Toshihide
authors.initials: T
authors.lastname: Kobayashi
authors.forename: Kun
authors.initials: K
authors.lastname: Ling
authors.forename: Richard E
authors.initials: RE
authors.lastname: Pagano
mesh_terms: D021941:Caveolae / Q000378:metabolism*; D051242:Caveolin 1 / Q000378:metabolism; D004705:Endocytosis; D005347:Fibroblasts / Q000378:metabolism*; D051416:Focal Adhesion Protein-Tyrosine Kinases / Q000378:metabolism; D005732:Gangliosides / Q000378:metabolism*; D006026:Glycoside Hydrolases / Q000494:pharmacology; D006801:Humans; D019012:Integrin beta1 / Q000378:metabolism*; D021962:Membrane Microdomains / Q000378:metabolism; D009439:Neuraminidase / Q000494:pharmacology; D051419:Paxillin / Q000378:metabolism; D012867:Skin / Q000378:metabolism*; D016608:Talin / Q000378:metabolism
chemical_list: D051242:Caveolin 1; D005732:Gangliosides; D019012:Integrin beta1; D051419:Paxillin; D016608:Talin; D051416:Focal Adhesion Protein-Tyrosine Kinases; D006026:Glycoside Hydrolases; C050427:endoglycoceramidase; D009439:Neuraminidase