Matrix metalloproteinase-7 (MMP-7) (also known as matrilysin-1) is secreted as a proenzyme (proMMP-7) and plays a key role in the degradation of various extracellular matrix (ECM) and non-ECM molecules after activation. To identify the binding proteins related to proMMP-7 activation, a human lung cDNA library was screened by yeast two-hybrid system using proMMP-7 as bait. We identified a candidate molecule CD151, which is a member of the transmembrane 4 superfamily. Complex formation of proMMP-7 with CD151 was demonstrated by immunoprecipitation of the molecules from CaR-1 cells, a human rectal carcinoma cell line, expressing both proMMP-7 and CD151, and CD151 stable transfectants incubated with proMMP-7. Yeast twohybrid assays using deletion mutants of proMMP-7 and CD151 suggested an interaction between the propeptide of proMMP-7 and the COOH-terminal extracellular loop of CD151. The binding activity of 125 I-labeled proMMP-7 to CD151 on the cell membranes was shown with CD151 stable transfectants. Laser-scanning confocal microscopy demonstrated that proMMP-7 colocalizes with CD151 on the cell membranes of CD151 stable transfectants and CaR-1 cells. In situ zymography using crosslinked carboxymethylated transferrin, a substrate of MMP-7, demonstrated proteinase activity on and around CD151 stable transfectants and CaR-1 cells, while the activity was abolished by their treatment with MMP inhibitors, anti-MMP-7 antibody or anti-CD151 antibody. In human lung adenocarcinoma tissues, colocalization of MMP-7 and CD151 was demonstrated on the carcinoma cells. Metalloproteinase activity was present in these tissues and could be inhibited by antibodies to MMP-7 or CD151. These data demonstrate for the first time that proMMP-7 is captured and activated on the cell membranes through interaction with CD151, and suggest the possibility that similar to the MT1-MMP/MMP-2 system, MMP-7 is involved in the pericellular activation mechanism mediated by CD151, a crucial step in proteolysis on the cell membranes under various pathophysiological conditions including cancer invasion and metastasis.

SUMMARY:In vitro, membrane-type matrix metalloproteinases (MT-MMP) are known to activate the zymogen of MMP-2 (proMMP-2, progelatinase A), which is one of the key MMP in joint destruction in rheumatoid arthritis. In the present study, we examined the production and activation of proMMP-2, and the expression of MT1-MMP, MT2-MMP, and MT3-MMP, their correlation with proMMP-2 activation, and their localization in rheumatoid synovial tissue. Using sandwich enzyme immunoassay and gelatin zymography techniques, proMMP-2 production levels and activation ratios were found to be significantly higher in rheumatoid synovium compared with normal synovium (p Ͻ 0.01). Quantitative RT-PCR analyses demonstrated that MT1-MMP and MT3-MMP were expressed in all rheumatoid synovial tissue (30 of 30 cases), but that the mean expression level of MT1-MMP was approximately 11-fold higher than MT3-MMP. Significant correlation was found between the mRNA expression level of MT1-MMP and the activation ratio of proMMP-2 (p Ͻ 0.01). In situ hybridization indicated that the hyperplastic lining cells of rheumatoid synovium expressed MT1-MMP. Immunohistochemistry demonstrated that MT1-MMP was co-localized with MMP-2 and with a tissue inhibitor of metalloproteinase-2, and was mainly located in the rheumatoid synovial lining cells. In situ zymography of rheumatoid synovium showed gelatinolytic activity, predominantly in the lining cell layer. This activity was blocked when incubated with BB94, a specific MMP inhibitor. These results demonstrate that MT1-MMP plays an important role in the activation of proMMP-2 in the rheumatoid synovial lining cell layer, and suggest that its activity may be involved in the cartilage destruction of rheumatoid arthritis. (Lab Invest 2000, 80:677-687).

No abstract

BACKGROUND The 5‐year survival rate of patients with submucosal (sm) esophageal carcinoma continues to be much worse than that with intramucosal (m) lesions. The reasons are unclear, but matrix metalloproteinases (MMPs) and membrane type matrix metalloproteinases (MT‐MMPs) are known to be involved in degradation of extracellular matrix macromolecules associated with tumor cell invasion. Expression of these enzymes may have some relation to the difference in survival between patients with sm and m esophageal carcinomas. METHODS Surgical specimens from 148 primary esophageal squamous cell carcinomas were examined for expression of MMP‐1, 2, 3, 7, and 9; tissue inhibitor of MMP (TIMP)‐1 and 2; MT1‐MMP; and MT2‐MMP by immunohistochemical staining. The results were compared with tumor progression and other clinicopathologic parameters. Localization of gelatinase activity was also confirmed in carcinoma cells by in situ zymography. RESULTS Expression of MMP‐7 and 9, expression of MT1‐MMP, and gelatinase activity were elevated in the carcinomas, correlating with depth of tumor invasion. Percentages of positive sm cases were 44.0%, 51.9%, and 52.0%, respectively, significantly higher than for the m cases. Percentages of venous invasion positive cases were 61.9%, 65.1%, and 65.1%, respectively. CONCLUSIONS Expression of MMP‐7 and 9 and MT1‐MMP is closely associated with invasion depth and venous invasion in esophageal squamous cell carcinomas. Cancer 2000;88:2201–9. © 2000 American Cancer Society.