Matrix metalloprotease (MMP)-2 plays a key role in many biological and pathological processes related to cell migration, invasion, and mitogenesis. MMP-2 is synthesized as a zymogen that is activated through either a conformational change or proteolysis of the propeptide. Several activating enzymes for pro-MMP-2 have been proposed, including metalloproteases and serine proteases. The mechanism of pro-MMP-2 activation by metalloproteases is well established, and the most studied activation mechanism involves cleavage of the propeptide by membrane type 1-MMP (MT1-MMP). In contrast, serine protease activation has not been thoroughly studied, although studies suggest that MT1-MMP may be involved in activation by thrombin and plasmin. Here, we demonstrate that factor Xa mediates MT1-MMP-independent processing of pro-MMP-2 in vascular smooth muscle cells and endothelial cells. Factor Xa and thrombin directly cleaved the propeptide on the carboxyl terminal sides of the Arg 98 and Arg 101 residues, whereas plasmin only cleaved the propeptide downstream of Arg 101 . Moreover, processed MMP-2 showed enzymatic activity that was enhanced by intermolecular autoproteolytic processing at the Asn 109 -Tyr peptide bond. In addition to its role in activation, factor Xa rapidly degraded MMP-2, thereby restricting excessive MMP-2 activity. Thrombin also degraded MMP-2, but the degradation was reduced greatly under cell-associated conditions, resulting in an increase in processed MMP-2. Overall, factor Xa and thrombin regulate MMP-2 enzymatic activity through its activation and degradation. Thus, the net enzymatic activity results from a balance between MMP-2 activation and degradation. Matrix metalloprotease (MMP)3 -2 is a member of the zincdependent endopeptidase family, which comprises 24 enzymes (1). MMP-2 plays a key role in many biological and pathological processes, including organ growth, endometrial cycling, wound healing, bone remodeling, tumor invasion, and metastasis (2). This enzyme functions through proteolysis of non-structural extracellular molecules and components of the basement membrane, including type IV collagen, fibronectin, elastin, laminin, aggrecan, and fibrillin (3).Like most MMPs, MMP-2 is synthesized as a zymogen that is activated by conformational change (4) or proteolysis within the propeptide, which may involve membrane type MMPs (MT-MMPs) (5-9). The most studied activation mechanism for pro-MMP-2 is cleavage of the propeptide by MT1-MMP, which requires cooperative activity between MT1-MMP and tissue inhibitor of metalloprotease (TIMP)-2 (5, 10 -12). Serine proteases, such as thrombin, factor Xa, activated protein C, and plasmin as well as the cysteine protease legumain are all known activators of pro-MMP-2 (13-17). (25,26). Both proteases can also elicit endothelial cell and SMC migration through pro-MMP-2 activation and subsequent extracellular matrix degradation (13,27,28). However, despite studies suggesting that MT1-MMP is involved in thrombin-mediated activation of pro-MMP-2, a detailed mechanism...

Matrix metalloprotease‐2 is implicated in many biological processes and degrades extracellular and non‐extracellular matrix molecules. Matrix metalloprotease‐2 maintains a latent state through a cysteine–zinc ion pairing which, when disrupted, results in full enzyme activation. This pairing can be disrupted by a conformational change or cleavage within the propeptide. The best known activation mechanism for pro‐matrix metalloprotease‐2 occurs via cleavage of the propeptide by membrane type‐1 matrix metalloprotease. However, significant residual activation of pro‐matrix metalloprotease‐2 is seen in membrane type‐1 matrix metalloprotease knockout mice and in fibroblasts treated with metalloprotease inhibitors. These findings indicate the presence of a membrane type‐1 matrix metalloprotease‐independent activation mechanism for pro‐matrix metalloprotease‐2 in vivo, which prompted us to explore an alternative activation mechanism for pro‐matrix metalloprotese‐2. In this study, we demonstrate membrane type‐1 matrix metalloprotease‐independent propeptide processing of matrix metalloprotease‐2 in HEK293F and various tumor cell lines, and show that proprotein convertases can mediate the processing intracellularly as well as extracellularly. Furthermore, processed matrix metalloprotease‐2 exhibits enzymatic activity that is enhanced by intermolecular autolytic cleavage. Thus, our experimental data, taken together with the broad expression of proprotein convertases, suggest that the proprotein convertase‐mediated processing may be a general activation mechanism for pro‐matrix metalloprotease‐2 in vivo.