CELL SURFACE-MEDIATED ACTIVATION OF PROGELATINASE-A - DEMONSTRATION OF THE INVOLVEMENT OF THE C-TERMINAL DOMAIN OF PROGELATINASE-A IN CELL-SURFACE BINDING AND ACTIVATION OF PROGELATINASE-A BY PRIMARY FIBROBLASTS

We report that the isolated C-terminal domain of progelatinase A is in hibitory to the activation of this proenzyme by primary skin fibroblas t plasma membranes but is unable to inhibit organomercurial-induced se lf-cleavage and activation. Ligand binding studies demonstrate that fi broblasts stimulated with concanavalin A to activate progelatinase A h ave a significantly enhanced level of cell surface-associated progelat inase A. Tissue inhibitor of metalloproteinases-2 (TIMP-2), an effecti ve inhibitor of membrane-mediated progelatinase A activation, is able to abolish the enhanced level of cell surface-associated progelatinase A that occurs following stimulation. TIMP-1, a poor inhibitor of memb rane activation, is unable to inhibit the cell surface binding of prog elatinase A. The enhancement in the binding of I-125-progelatinase A t o fibroblasts following concanavalin A stimulation can be blocked by t he inclusion of excess C-terminal gelatinase A but not by a truncated form of gelatinase A lacking the C-terminal domain. Scatchard analysis of the binding of I-125-progelatinase A to concanavalin A-stimulated fibroblasts has identified 950 000 gelatinase binding sites per cell w ith a K-d of 1.3 x 10(-8) M. Analysis of non-stimulated fibroblasts ha s identified 500 000 sites per cell with a K-d of 2.6 x 10(-8) M. We p ropose that membrane-mediated activation of progelatinase A involves b inding of the proenzyme through its C-terminal domain to the cell surf ace and that TIMP-2 can inhibit activation by interaction with progela tinase A through the C-terminal domain, thus preventing binding of the proenzyme.