In order to investigate the role of matrix metalloproteinases (MMP) in tumor invasion and metastases, we have focused on the multilevel regulation of these enzymes. Studies have shown that in contrast with other members of the MMP enzyme family, 72 kDa gelatinase A levels are increased in response to TGFbeta1, are unaffected by the tumor promoting phorbol esters, and show elevated levels in colorectal, breast, thyroid, ovarian and bladder tumor tissues when compared with adjacent normal mucosa tissues. We have identified a cellular activation mechanism which is cell surface associated and specific for the 72 kDa gelatinase A enzyme, and which can be induced by pretreatment with phorbol esters or concanavalin A. This cellular activation mechanism does not affect other members of the collagenase gene family. This activation mechanism appears to require cell surface binding of the gelatinase A enzyme. We have now demonstrated that formation of the progelatinase A/TIMP-2 complex and subsequent binding to a putative TIMP-2 receptor are important steps in this cellular activation mechanism. In addition, we have characterized the kinetics of MMP-2 and TIMP-2 biosynthesis in order to determine when and where formation of the proenzyme-inhibitor complex occurs. These results suggest that the progelatinase A/TIMP-2 complex is unique in that binding of the inhibitor mediates the cellular activation of this protease. We are also examining the degradation of cell surface substrates by MMP-2. In addition, the protein biochemistry of pro-MMP-2 and TIMP-2 interactions are being studied using mutant recombinant enzymes and plasmon surface resonance techniques (BIAcore).
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