THE UROKINASE-TYPE PLASMINOGEN-ACTIVATOR RECEPTOR, A GPI-LINKED PROTEIN, IS LOCALIZED IN CAVEOLAE

The urokinase plasminogen activator receptor (uPAR), a glycosylphospha tidylinositol-linked glycoprotein, plays a central role in the regulat ion of pericellular proteolysis and participates in events leading to cell activation. Here, we demonstrate that uPAR, on a human melanoma c ell line, is localized in caveolae, flask-shaped microinvaginations of the plasma membrane found in a variety of cell types. Indirect immuno fluorescence with anti-uPAR antibodies on the melanoma cells showed a punctated staining pattern that accumulated to stretches along sides o f cell-cell contact and membrane ruffles. uPAR colocalized with caveol in, a characteristic protein in the coat of caveolae, as demonstrated by double staining with specific antibodies. Further, uPAR could be di rectly localized in caveolae by in vivo immunoelectron microscopy. Bot h uPAR and its ligand, uPA, were present in caveolae enriched low dens ity Triton X-100 insoluble complexes, as shown by immunoblotting. From such complexes, caveolin could be coprecipitated with uPAR-specific a ntibodies suggesting a close spatial association between uPAR and cave olin that might have implications for the signal transduction mediated by uPAR. Further, functional studies indicated that the localization of uPAR and its ligand in caveolae enhances pericellular plasminogen a ctivation, since treatment of the cells with drugs that interfere with the structural integrity of caveolae, such as nystatin, markedly redu ced cell surface plasmin generation. Thus, caveolae promote efficient cell surface plasminogen activation by clustering uPAR, uPA, and possi bly other protease receptors in one membrane compartment.