Interaction of fibronectin with cultured human endothelial cells: characterization of the specific receptor.
In this study we provide a characterization of the fibronectin (FN) binding to endothelial cells (EC), and we identify the FN binding site on these cells. 125I-FN binding to EC in suspension was time dependent and reached a plateau at 4 h. Cold FN inhibited this interaction in a concentration-dependent way, but vitronectin, fibrinogen, and IgG were poorly effective. About 80% of the total FN associated to EC at the equilibrium was specifically bound; of this, 60% was reversibly bound, while 20% appeared to be internalized. The FN binding was saturable and an apparent dissociation constant of about 0.23 x 10(-6) mol/L and a maximal number of binding sites of about 9.8 x 10(5) was estimated from binding isotherms. Autoradiography data showed that EC-associated 125I-FN was all in high mol wt form that did not enter the gel. We then characterize the FN receptor (FNR) in EC. An antiserum to the FNR isolated from human placenta inhibited FN binding to EC by 89%, and using the immunoblotting technique, it recognized two bands in the EC detergent extract of mol wt 125/160 Kd. This antiserum also recognized the EC membrane protein complex eluted from the FN affinity column by an arg-gly-asp (RGD) peptide. When this complex was included into liposomes, it poorly bound to FN. However, the binding was strikingly increased by addition of Mn in the buffer and was specific for FN in respect to other substrata. These data define the FN binding site in EC and indicate that it is functionally and structurally related to that isolated from human placenta.
Conforti
G
G
Istituto di Ricerche Farmacologiche "Mario Negri," Milano, Italy.
Zanetti
A
A
Colella
S
S
Abbadini
M
M
Marchisio
P C
PC
Pytela
R
R
Giancotti
F
F
Tarone
G
G
Languino
L R
LR
Dejana
E
E
D000954:Antigens, Surface; D015815:Cell Adhesion Molecules; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000502:physiology*; D005109:Extracellular Matrix / Q000378:metabolism; D005340:Fibrinogen / Q000378:metabolism; D005353:Fibronectins / Q000502:physiology*; D006023:Glycoproteins / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D008081:Liposomes; D008970:Molecular Weight; D017469:Receptors, Fibronectin; D011971:Receptors, Immunologic / Q000502:physiology*; D019096:Vitronectin
D000954:Antigens, Surface; D015815:Cell Adhesion Molecules; D005353:Fibronectins; D006023:Glycoproteins; D008081:Liposomes; D017469:Receptors, Fibronectin; D011971:Receptors, Immunologic; D019096:Vitronectin; D005340:Fibrinogen