BOUND PLASMINOGEN IS RATE-LIMITING FOR CELL-SURFACE-MEDIATED ACTIVATION OF PLASMINOGEN BY UROKINASE

The ability of U937 monocyte-like cells and KATO III cells (a human ga stric carcinoma line) to potentiate activation of plasminogen by singl e-chain urokinase-type plasminogen activator (scu-PA), as mediated by the cell receptor for urokinase (u-PAR), was compared. It was observed that, although the concentration of u-PAR on these cell lines differe d considerably (U937 cells: 5000 receptors/cell, K-d 0.35 nM; KATO III cells: 400 receptors/cell, K-d 0.85 nM), the rate of activation of pl asminogen by scu-PA in the presence of the same density of each cell l ine was equivalent. From data generated in the presence of increasing concentrations of scu-PA, the k(cat). for plasminogen activation in th e presence of each cell line was calculated and found to differ by 26- fold (0.36 s(-1) on U937 cells; 9.25 s(-1) on KATO III cells). However , the K-m for plasminogen with respect to the rate of formation of pla smin was lower than the K-d for binding (0.2 mu M compared with 0.5 mu M on U937 cells; 0.34 mu M compared with 1.6 mu M on KATO III cells). A rapid transformation from Glu-plasminogen (native plasminogen with N-terminal Glu) to Lys-plasminogen (plasmin-degraded plasminogen with primarily N-terminal Lys-77) occurred on the surface of U937 cells (un like KATO III cells), but this transition did not coincide with faster rates of plasminogen activation. From this evidence it is concluded t hat the accessibility of bound plasminogen acts to limit the rate of a ctivation by cell-bound urokinase. The significance of this proposal i s that the proteolytic potential of the cell-mediated activation of pl asminogen would be controlled by the accessibility of plasminogen for activation rather than by the concentration of u-PAR (the latter may a ct to localize proteolysis to appropriate domains on the surface of th e cell).