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-associated cathepsin B as an activator of receptor-bound single-chain urokinase-type plasminogen activator (pro-uPA) and to determine the importance of receptor-bound uPA activity in the destruction of extracellular matrix by tumor cells with subsequent invasion through basement membranes. Ovarian cancer HOC-I cells express pro-uPA/HMW-uPA and cathepsin B on their surface. uPAs are bound to a specific surface receptor, about 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 cysteine proteinase inhibitor, for 24 h. Inhibition of cell-surface cathepsin 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-bound pro-uPA cannot be converted to HMW-uPA in the absence of enzymatically-active cathepsin B. The significance of the expression of cell-surface 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 for the conversion of pro-uPA to HMW-uPA and that receptor-bound uPA activity constitutes an efficient mechanism which contributes to tumor cell invasion. As HOC-I cells produce both uPA and cathepsin B, the implications of tumor-cell-derived pro-uPA activation by cellular proteinase cathepsin B should be considered.
Kobayashi
H
H
Department of Obstetrics and Gynecology, Hamamatsu University School of Medicine, Shizuoka, Japan.
Moniwa
N
N
Sugimura
M
M
Shinohara
H
H
Ohi
H
H
Terao
T
T
D001485:Basement Membrane / Q000378:metabolism; D001710:Biotin; D002401:Cathepsin B / Q000378:metabolism*; D002460:Cell Line / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism*; D003094:Collagen; D015853:Cysteine Proteinase Inhibitors; D004338:Drug Combinations; D004789:Enzyme Activation; D006801:Humans; D007797:Laminin; D007930:Leucine / Q000031:analogs & derivatives; D011509:Proteoglycans; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D014407:Tumor Cells, Cultured / Q000378:metabolism*; D014568:Urokinase-Type Plasminogen Activator / Q000037:antagonists & inhibitors / Q000378:metabolism*
D015853:Cysteine Proteinase Inhibitors; D004338:Drug Combinations; D007797:Laminin; D011509:Proteoglycans; D011994:Recombinant Proteins; C059485:matrigel; D001710:Biotin; D003094:Collagen; D014568:Urokinase-Type Plasminogen Activator; D002401:Cathepsin B; D007930:Leucine; C024974:E 64; C067551:saruplase