Intermolecular Autolytic Cleavage Can Contribute to the Activation of Progelatinase A by Cell Membranes

Abstract

Membrane-type matrix metalloproteinase (MT-MMP) messenger RNA and protein expression were shown to be elevated in human fibroblasts following treatment with concanavalin A, coincident with the induction of the ability to process progelatinase A.CHO cells transfected with the cDNA for MT-MMP were able to process both wild type progelatinase A and a catalytically inactive mutant, E375A progelatinase A. Both proenzymes were converted to a 68-kDa intermediate (reducing gels) form, but only the wild type enzyme was processed further to a 66-kDa end product. In contrast, both forms of progelatinase were processed via the 68-kDa intermediate to 66 kDa by concanavalin Astimulated fibroblasts.Further study of the processing of E375A progelatinase A by plasma membrane preparations from concanavalin A-stimulated fibroblasts showed that addition of active gelatinase A enhanced processing to the mature form. It was concluded that cell membrane-mediated activation of progelatinase A could be via a cascade involving both MT-MMP and intermolecular autolytic cleavage.

Cell surface binding and activation of gelatinase A induced by expression of membrane‐type‐1‐matrix metalloproteinase (MT1‐MMP)

“…4, lanes denoted 'E/A' and 'E/A+MT', respectively). This is consistent with the previous observation that a catalytically inactive mutant progelatinase A was converted to an intermediate form by incubation with CHO cells expressing MT1-MMP [25].…”

Type IV Collagen Induces Matrix Metalloproteinase 2 Activation in HT1080 Fibrosarcoma Cells

“…IB, lane 1). This latter band represents an intermediate activated form of MMP-2 which has been previously shown to result from a MT1-MMP-depen-dent proteolytic processing of pro-MMP-2 [20,31,32]. After 48 h, the amount of 62-kDa MMP-2 increased in both conditioned medium and cell extract [accounting for 10 and 35% of total MMP-2 activity, respectively (Figs.…”

“…It can be hypothesized that type IV collagen mediates its effect by bringing closer cell membrane-associated MT1-MMP/TIMP-2/MMP-2 complexes, therefore promoting the intermolecular autolytic cleavage of intermediate MMP-2 and thus leading to the generation of the fully mature species. Additionally, type IV collagen decreases the level of secreted TIMP-2, thus potentially furthering the processing of pro-MMP-2 which is known to be inhibited by high TIMP-2 concentrations [20,31,53]. It is worth noting that the TIMP-2 degradation process illustrated here potentially represents an additional regulatory mechanism of the proteolytic activity of the cells.…”

“…On the basis of the dual role of TIMP-2 during MMP-2 activation, promoting MMP-2 activation when present at low concentrations [20,29,31,54,63,64], but acting as an efficient inhibitor of this process when present at higher doses [19,20,26,31,53,[65][66][67], we hypothesized that modulations of TIMP-2 concentration could play important roles in the regulation of MMP-2 processing. Consequently, we investigated the modifications of both TIMP-2 concentration and distribution between conditioned media and cell extracts induced by MMP-2-activating treatments.…”

Low molecular weight inhibitors of matrix metalloproteinases can enhance the expression of matrix metalloproteinase‐2 (gelatinase A) without inhibiting its activation

“…This finding is interesting, because it has been suggested that, after the MT1-MMP-mediated conversion of the 72-kD form of MMP-2 into the 68-kD intermediate form, an autoproteolytic activation step is required for the final activation of MMP-2, resulting in the 66-kD form. [25][26][27] Our data indicate that a considerable fraction still is activated, despite the absence of active MMP-2. The higher level of the intermediate form of MMP-2 (68 kD) under these conditions suggests that this repre- sents the fraction activated by MMP-2 itself, indicating that, in addition to an autoproteolytic activation step, compensatory mechanisms may exist by which the intermediate form of MMP-2 may be converted into its fully activated, 66-kD form.…”