Gangliosides and beta1-integrin are required for caveolae and membrane domains.
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.
Singh
Raman Deep
RD
Departments of Medicine, Biochemistry and Molecular Biology, Mayo Clinic College of Medicine, Rochester, MN 55905, USA.
Marks
David L
DL
Holicky
Eileen L
EL
Wheatley
Christine L
CL
Kaptzan
Tatiana
T
Sato
Satoshi B
SB
Kobayashi
Toshihide
T
Ling
Kun
K
Pagano
Richard E
RE
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
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