Factor XII binding to endothelial cells depends on caveolae.
It is now generally accepted that factor XII (FXII) binds to cellular surfaces in the vascular system. One of the suggested receptors of this binding is the glycosylphosphatidylinositol-anchored urokinase-like plasminogen activator (u-PAR) harbored in caveolae/lipid rafts. However, binding of FXII to human umbilical vein endothelial cells (HUVEC) has never been shown to be localized to these specialized membrane structures. Using microscopical techniques, we here report that FXII binds to specific patches of the HUVEC plasma membrane with a high density of caveolae. Further investigations of FXII binding to caveolae were performed by sucrose density-gradient centrifugations. This showed that the majority of FXII, chemically cross-linked to HUVEC, could be identified in the same fractions of the gradient as caveolin-1, a marker of caveolae, while the majority of u-PAR was identified in noncaveolae lipid rafts. Accordingly, cholesterol-depleted cells were found to bind significantly reduced amounts of FXII. These observations, combined with the presence of a minority of u-PAR in caveolae concomitant with FXII binding, indicate that FXII binding to u-PAR may be secondary and depends upon the structural elements within caveolae. Thus, FXII binding to HUVEC depends on intact caveolae on the cellular surface.
Schousboe
Inger
I
Department of Medical Biochemistry & Genetics, The Panum Institute, University of Copenhagen, Denmark. schousboe@imbg.ku.dk
Thomsen
Peter
P
van Deurs
Bo
B
D000818:Animals; D021941:Caveolae / Q000378:metabolism*; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D002478:Cells, Cultured; D002784:Cholesterol / Q000378:metabolism; D016895:Culture Media, Serum-Free; D003505:Cyclodextrins / Q000378:metabolism; D042783:Endothelial Cells / Q000378:metabolism* / Q000648:ultrastructure; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism; D005174:Factor XII / Q000378:metabolism*; D006801:Humans; D011485:Protein Binding; D013347:Subcellular Fractions / Q000737:chemistry / Q000378:metabolism; D047392:beta-Cyclodextrins
D016895:Culture Media, Serum-Free; D003505:Cyclodextrins; D047392:beta-Cyclodextrins; C108732:methyl-beta-cyclodextrin; D005174:Factor XII; D002784:Cholesterol