EFFECTS OF MEMBRANE-ASSOCIATED CATHEPSIN-B ON THE ACTIVATION OF RECEPTOR-BOUND PROUROKINASE AND SUBSEQUENT INVASION OF RECONSTITUTED BASEMENT-MEMBRANES

The present study was undertaken to assess the role of membrane-associ ated cathepsin B as an activator of receptor-bound single-chain urokin ase-type plasminogen activator (pro-uPA) and to determine the importan ce of receptor-bound uPA activity in the destruction of extracellular matrix by tumor cells with subsequent invasion through basement membra nes. Ovarian cancer HOC-I cells express pro-uPA/HMW-uPA and cathepsin B on their surface. uPAs are bound to a specific surface receptor, abo ut 30% of which is saturated. 60% of the receptor-bound uPA is pro-uPA . No reduction in the specific binding of biotinylated DFP-HMW-uPA was observed when cells were cultivated in the presence of E-64, a cystei ne proteinase inhibitor, for 24 h. Inhibition of cell-surface cathepsi n B activity was associated with a decrease in cell-bound uPA activity to undetectable levels, and > 95% of the membrane-associated uPA was pro-uPA in cells cultivated with E-64. This suggested that receptor-bo und pro-uPA cannot be converted to HMW-uPA in the absence of enzymatic ally-active cathepsin B. The significance of the expression of cell-su rface uPA activity regarding invasive potential was examined in an in- vitro Matrigel invasion assay. Decreased cell-surface uPA activity was associated with a decrease in invasive potential. These data support our hypothesis that membrane-associated cathepsin B may be important f or the conversion of pro-uPA to HMW-uPA and that receptor-bound uPA ac tivity constitutes an efficient mechanism which contributes to tumor c ell invasion. As HOC-1 cells produce both uPA and cathepsin B, the imp lications of tumor-cell-derived pro-uPA activation by cellular protein ase cathepsin B should be considered.