title: Effects of urokinase receptor occupancy on plasmin generation and proteolysis of basement membrane by human tumor cells.
abstract: The goal of the present study was to assess the relative importance of receptor-bound and secreted plasminogen activator urokinase (u-PA) in generating cell-surface plasmin and fostering destruction of normal tissue by tumor cells. We first showed that active site-inhibited u-PA could displace endogenous u-PA from the surface of the human colon adenocarcinoma cell line HCT 116. We then prepared expression vectors for u-PA and for a mutant molecule in which the codon for the active site serine residue was changed to encode alanine. Expression of non-functional mutant u-PA decreased the level of cell-bound active u-PA by more than 95% via a mechanism that involved competition for receptor sites. Decreased cell-surface u-PA activity was associated with a decrease in cell-bound plasmin activity to undetectable levels, suggesting that receptor-bound u-PA plays an important role in the generation of plasmin on the cell surface. Transfectants that secreted eightfold to 20-fold elevated levels of active wild-type u-PA showed approximately 50% increases in cell-associated u-PA and only twofold to fourfold increases in cell-associated plasmin, suggesting that the role of secreted u-PA in generating cell-surface plasmin activity was relatively minor. In parent cells and both types of transfectants there was a good correlation between the amount of plasmin bound to the tumor cell surface and the extent to which a basement membrane substrate was degraded. These studies show that receptor-bound u-PA provides an efficient mechanism for plasmin generation on the surface of tumor cells, which, in turn, contributes significantly to their degradative potential.
authors.affiliation: Cancer Research Institute, University of California-San Francisco 94143.
authors.forename: R L
authors.initials: RL
authors.lastname: Cohen
authors.forename: X P
authors.initials: XP
authors.lastname: Xi
authors.forename: C W
authors.initials: CW
authors.lastname: Crowley
authors.forename: B K
authors.initials: BK
authors.lastname: Lucas
authors.forename: A D
authors.initials: AD
authors.lastname: Levinson
authors.forename: M A
authors.initials: MA
authors.lastname: Shuman
mesh_terms: D000230:Adenocarcinoma; D001483:Base Sequence; D001485:Basement Membrane / Q000378:metabolism; D001665:Binding Sites; D002460:Cell Line; D003110:Colonic Neoplasms; D005341:Fibrinolysin / Q000096:biosynthesis*; D005822:Genetic Vectors; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D015345:Oligonucleotide Probes; D010960:Plasminogen Activators / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D012334:RNA, Neoplasm / Q000235:genetics / Q000302:isolation & purification; D011956:Receptors, Cell Surface / Q000502:physiology*; D055293:Receptors, Urokinase Plasminogen Activator; D014162:Transfection; D014568:Urokinase-Type Plasminogen Activator / Q000235:genetics / Q000378:metabolism*
chemical_list: D015345:Oligonucleotide Probes; C524898:PLAUR protein, human; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D010960:Plasminogen Activators; D005341:Fibrinolysin; D014568:Urokinase-Type Plasminogen Activator