Role of Cell Surface Metalloprotease Mt1-Mmp in Epithelial Cell Migration over Laminin-5

Abstract

Laminin-5 (Ln-5) is an extracellular matrix substrate for cell adhesion and migration, which is found in many epithelial basement membranes. Mechanisms eliciting migration on Ln-5 need to be elucidated because of their relevance to tissue remodeling and cancer metastasis. We showed that exogenous addition of activated matrix metalloprotease (MMP) 2 stimulates migration onto Ln-5 in breast epithelial cells via cleavage of the γ2 subunit. To investigate the biological scope of this proteolytic mechanism, we tested a panel of cells, including colon and breast carcinomas, hepatomas, and immortalized hepatocytes, selected because they migrated or scattered constitutively in the presence of Ln-5. We found that constitutive migration was inhibited by BB94 or TIMPs, known inhibitors of MMPs. Limited profiling by gelatin zymography and Western blotting indicated that the ability to constitutively migrate on Ln-5 correlated with expression of plasma membrane bound MT1-MMP metalloprotease, rather than secretion of MMP2, since MMP2 was not produced by three cell lines (one breast and two colon carcinomas) that constitutively migrated on Ln-5. Moreover, migration on Ln-5 was reduced by MT1-MMP antisense oligonucleotides both in MMP2+ and MMP2− cell lines. MT1-MMP directly cleaved Ln-5, with a pattern similar to that of MMP2. The hemopexin-like domain of MMP2, which interferes with MMP2 activation, reduced Ln-5 migration in MT1-MMP+, MMP2+ cells, but not in MT1-MMP+, MMP2− cells. These results suggest a model whereby expression of MT1-MMP is the primary trigger for migration over Ln-5, whereas MMP2, which is activated by MT1-MMP, may play an ancillary role, perhaps by amplifying the MT1-MMP effects. Codistribution of MT1-MMP with Ln-5 in colon and breast cancer tissue specimens suggested a role for this mechanism in invasion. Thus, Ln-5 cleavage by MMPs may be a widespread mechanism that triggers migration in cells contacting epithelial basement membranes.

Membrane type-1 matrix metalloproteinase and TIMP-2 in tumor angiogenesis

“…MT1-MMP (MMP14) is the prototypic member of the MT-MMPs and its expression has been associated with a variety of cellular and developmental processes, as well as multiple pathophysiological conditions (Pepper, 2001;Yana and Seiki, 2002). MT1-MMP displays a broad spectrum of activity against ECM components such as type I and II collagen, fibronectin, vitronectin, laminin, fibrin and proteoglycan (d 'Ortho et al, 1997;Koshikawa et al, 2000). However, it has substrates that extend beyond ECM components such as cell surface molecules including CD44 (Kajita et al, 2001), pro α V integrin (Deryugina et al, 2002b) and transglutaminase (Belkin et al, 2001).…”

“…Indeed, MT1-MMP-deficient mice exhibit damages in skeletal development manifested by craniofacial dysmorphism, dwarfism, osteopenia and fibrosis (Holmbeck et al, 1999;Zhou et al, 2000). The expression of MT1-MMP has been described in processes involving cell migration (Koshikawa et al, 2000). Overexpression of MT1-MMP increases the number of experimental metastases (Tsunezuka et al, 1996).…”

“…MT1-MMP may function at least as a fibrinolytic and gelatinolytic enzyme (Hotary et al, 2000;d'Ortho et al, 1997). Cleavage of laminin-5 by MMP2 and MT1-MMP reveals a cryptic site that trigger epithelial cell motility (Gilles et al, 2001;Koshikawa et al, 2000). However, recent data indicate that MMPs and particularly MT1-MMP do far more to promote cancer progression than merely remove the physical barriers to migration and invasion (Egeblad and Werb, 2002;McCawley and Matrisian, 2000).…”

Laminin-5 offsets the efficacy of gefitinib (‘Iressa’) in hepatocellular carcinoma cells

“…Stock solutions were prepared at 20 mM in dimethyl sulphoxide (DMSO) and stored in aliquots at À201C. Purified Ln-5 was prepared as previously described (Koshikawa et al, 2000). Working dilutions were made in culture medium supplemented with 10% fetal bovine serum, 2 mM glutamine, 50 000 UL À1 penicillin and 80 mM streptomycin.…”

Aberrant expression of laminin gamma 2 chain and its prognostic significance in intrahepatic cholangiocarcinoma according to growth morphology

“…8 Furthermore, matrix metalloproteinase-2 (MMP2) and membrane-type 1 matrix metalloproteinase (MT1-MMP) are considered to be candidates for laminin gamma 2 processing protease. 26,27 Processing of the 155-kd gamma 2 chain involves a cleavage within domain III leading to a 105-kd polypeptide. It appears that the resultant degraded laminin gamma 2 chain is an active form that enhances epithelial cell migration.…”