title: Regulation of membrane-type-1 matrix metalloproteinase activity by its cytoplasmic domain.
abstract: Membrane-type-1 matrix metalloproteinase (MT1-MMP) has transmembrane and cytoplasmic domains, which target it to invasive fronts. We analyzed the role of the cytoplasmic tail by expressing wild type MT1-MMP and three mutants with progressively truncated C termini in human Bowes melanoma cells. We examined gelatinase A activation and the localization and processing of recombinant proteins in stable cell clones using gelatin zymography, immunoblotting, and immunofluorescence. Cell invasion was analyzed in vitro by Matrigel invasion assays. Gelatinase A was activated in all cell clones. However, the localization of MT1-MMP to the leading edge of migrating cells and cell invasion through Matrigel were strongly enhanced only in cells expressing either wild type or truncated MT1-MMP lacking 6 C-terminal amino acid residues (Delta577). Truncations of 10 or 16 amino acid residues in the cytoplasmic domain (Delta567 and Delta573, respectively) disturbed MT1-MMP localization. The expression of wild type and Delta577 MT1-MMPs induced also their cleavage to 43-kDa cell surface forms and the release of soluble, approximately 20-kDa N-terminal fragments containing the catalytic center. A synthetic MMP inhibitor but not a gelatinase inhibitor prevented the processing, suggesting that autocatalytic cleavage occurs. Purified soluble MT1-MMP was also autoproteolytically processed to 43- and 20-kDa forms in vitro. Our results indicate that the cytoplasmic domain has an important role in cell invasion by controlling both the targeting and degradation/turnover of MT1-MMP.
authors.affiliation: Departments of Virology and Pathology, Haartman Institute, University of Helsinki, FIN-00014 Helsinki, Finland.
authors.forename: K
authors.initials: K
authors.lastname: Lehti
authors.forename: H
authors.initials: H
authors.lastname: Valtanen
authors.forename: S A
authors.initials: SA
authors.lastname: Wickström
authors.forename: J
authors.initials: J
authors.lastname: Lohi
authors.forename: J
authors.initials: J
authors.lastname: Keski-Oja
mesh_terms: D003001:Cloning, Molecular; D003094:Collagen; D003593:Cytoplasm / Q000201:enzymology; D004338:Drug Combinations; D004789:Enzyme Activation; D006801:Humans; D007797:Laminin; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D053504:Matrix Metalloproteinases, Membrane-Associated; D008545:Melanoma; D008666:Metalloendopeptidases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D009361:Neoplasm Invasiveness; D011480:Protease Inhibitors / Q000494:pharmacology; D011509:Proteoglycans; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017384:Sequence Deletion; D014162:Transfection; D014407:Tumor Cells, Cultured
chemical_list: D004338:Drug Combinations; D007797:Laminin; D011480:Protease Inhibitors; D011509:Proteoglycans; D011994:Recombinant Proteins; C059485:matrigel; D003094:Collagen; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases; D020778:Matrix Metalloproteinase 2