IntroductionThe urokinase-urokinase receptor (u-PA-u-PAR) system seems to play a crucial role in a number of biological processes, including local fibrinolysis, tumor invasion, angiogenesis, neointima and atherosclerotic plaque formation, inflammation, and matrix remodeling during wound healing and development.1-6 Binding of urokinase to its specific receptor provides cells with a localized proteolytic potential. It stimulates conversion of cell surface-bound plasminogen into active plasmin, which, in turn, is required for proteolytic degradation of basement membrane components, including fibronectin, collagen, laminin, and proteoglycan core proteins.7 Moreover, plasmin activates other matrix-degrading enzymes, such as matrix metalloproteinases.8 Overexpression of u-PA/u-PAR correlates with tumor invasion and metastasis formation,9-13 while reduction of cell-surface bound u-PA and inhibition of u-PAR expression leads to a significant decrease of invasive and metastatic activity.14 Specific antagonists that suppress binding of u-PA to u-PAR have been shown to inhibit cell-surface plasminogen activation, tumor growth, and angiogenesis both in vitro and in vivo models.15,16
Independently of its proteolytic activity, u-PA is implicated in many biological processes that seem to require u-PAR-mediated intracellular signal transduction, such as proliferation, chemotactic movement and adhesion, migration, and differentiation.17 Data obtained in the late 1980s indicated that u-PA not only provides cells with local proteolytic activity, but might also be capable of transducing signals to the cell.18-22 At that time, however, the u-PAR has just been isolated, cloned, and identified as a glycosylphosphatidylinositol (GPI)-linked protein and not a transmembrane protein. Signaling via the u-PAR was, therefore, regarded as being unlikely, and the effects of u-PA on cell proliferation18-22 were thought to be mediated by proteolytic activation of latent growth factors. The assumption of direct signaling via u-PAR was, in fact, considered controversial, until about 10 years later when a physical association between u-PAR and signaling proteins was found.23 From this report on, several proteins associated with u-PAR have been identified. Now, u-PAR seems to be part of a large “signalosome” associated and interacting with several proteins on both the outside and inside of the cell.
Modification of proteolytic activity matrix analysis (PrAMA) to measure ADAM10 and ADAM17 sheddase activities in cell and tissue lysates
