Vitronectin regulates the synthesis and localization of urokinase-type plasminogen activator in HT-1080 cells.
The effect of extracellular matrix composition on the location, amount, and activity of cell-associated urokinase-type plasminogen activator was tested using HT-1080 cells adherent to either fibronectin or vitronectin. Specific immunoprecipitation of newly synthesized urokinase indicated that cells adherent to fibronectin synthesized 2-3-fold more urokinase than cells adherent to vitronectin. Complexes of urokinase and plasminogen activator inhibitor type 1 (PAI-1) were detected in cell layers of vitronectin-adherent but not fibronectin-adherent cells. Inhibition of PAI-1 using a neutralizing monoclonal antibody resulted in a 3-fold increase in urokinase enzymatic activity on vitronectin adherent cells. Urokinase activity on fibronectin adherent cells was only slightly increased following PAI-1 neutralization. Examination of both HT-1080 and normal human fibroblast cells by immunofluorescent microscopy localized urokinase-type plasminogen activator to discrete, focal areas underneath cells adherent to vitronectin. Urokinase was not detectable by immunofluorescence on cells adherent to fibronectin. The addition of exogenous prourokinase to locate urokinase receptors on adherent HT-1080 cells indicated that the focal localization of cell-surface urokinase resulted from the clustering of urokinase receptors following adhesion to vitronectin but not fibronectin-coated substrates. These results suggest that vitronectin can contribute to the control of cell-surface plasmin activity by regulating the synthesis of urokinase and directing the localization of urokinase receptors.
Ciambrone
G J
GJ
Department of Physiology and Cell Biology, Albany Medical College, New York 12208.
McKeown-Longo
P J
PJ
D000911:Antibodies, Monoclonal; D002448:Cell Adhesion; D005353:Fibronectins / Q000302:isolation & purification; D005455:Fluorescent Antibody Technique; D006023:Glycoproteins / Q000302:isolation & purification / Q000494:pharmacology*; D006801:Humans; D010958:Plasminogen / Q000302:isolation & purification; D011233:Precipitin Tests; D011956:Receptors, Cell Surface / Q000378:metabolism; D055293:Receptors, Urokinase Plasminogen Activator; D013379:Substrate Specificity; D014407:Tumor Cells, Cultured; D014568:Urokinase-Type Plasminogen Activator / Q000096:biosynthesis* / Q000378:metabolism; D019096:Vitronectin
D000911:Antibodies, Monoclonal; D005353:Fibronectins; D006023:Glycoproteins; C524898:PLAUR protein, human; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D019096:Vitronectin; D010958:Plasminogen; D014568:Urokinase-Type Plasminogen Activator