PLASMINOGEN ACTIVATION BY T-PA ON THE SURFACE OF HUMAN-MELANOMA CELLS IN THE PRESENCE OF ALPHA-2-MACROGLOBULIN SECRETION

Several human melanoma cell lines produced tissue-type plasminogen activator (t-PA), as detected by zymography and immunocapture assay of culture media and cell lysates. Urokinase (u-PA) was found at only ≤1% the level of t-PA. Acid eluates of the cell surface indicated that the melanoma cells had t-PA bound on their surface, but no u-PA, and also had a very low capacity to bind exogenous u-PA. After incubation of the melanoma cells with 10% plasminogen-depleted fetal calf serum and human plasminogen, bound plasmin activity could be eluted from the cell surface with tranexamic acid, an analogue of lysine. This indicated that plasminogen was activated on the cell surface. The cell-surface plasmin formation was inhibited by an anti-catalytic monoclonal antibody to human t-PA, and not by an anti-catalytic antibody to u-PA. The melanoma cells also synthesized and secreted α2-macroglobulin (α2M), as shown by α2M-specific mRNA in Northern blotting and detection of α2M protein in conditioned cell culture media. The media were found to inhibit u-PA but not t-PA. This inhibition was related to their α2M content, and immunoabsorption of α2M removed the inhibitory activity. These studies suggest that t-PA can bind to the surface of melanoma cells and generate surface-bound plasmin. Because t-PA and cell-bound plasmin are unaffected by α2M, t-PA may, in the case of melanoma cells, serve an analogous function to u-PA in supporting tumor cell invasion. © 1990 by The American Society for Cell Biology.