The basement membrane protein laminin-5, a heterotrimer of laminin ␣3, ␤3, and ␥2 chains, potently promotes cellular adhesion and motility. It has been supposed that the carboxyl-terminal globular region of the ␣3 chain consisting of five distinct domains (G1 to G5) is important for its interaction with integrins. To clarify the function of each G domain, we transfected cDNAs for the full-length (wild type (WT)) and five deletion derivatives (⌬Gs) of the ␣3 chain into human fibrosarcoma cell line HT1080, which expressed and secreted the laminin ␤3 and ␥2 chains but not the ␣3 chain. The transfectants with the ␣3 chain cDNAs lacking G5 (⌬G 5 ), G4 -5 (⌬G 4 -5 ), G3-5 (⌬G 3-5 ), and G2-5 (⌬G 2-5 ) secreted laminin-5 variants at levels comparable to that with WT cDNA. However, the transfectant with the cDNA without any G domains (⌬G 1-5 ) secreted little laminin-5, suggesting that the G domains are essential for the efficient assembly and secretion of the heterotrimer ␣3␤3␥2. The transfectants with WT, ⌬G 5 , and ⌬G 4 -5 cDNAs survived in serum-free medium longer than those with ⌬G 3-5 , ⌬G 2-5 , and ⌬G 1-5 cDNAs. The transfectants with WT, ⌬G 5 , and ⌬G 4 -5 cDNAs secreted apparently the same size of laminin-5, which lacked G4 and G5 due to proteolytic cleavage between G3 and G4, and these laminin-5 forms potently promoted integrin ␣ 3 ␤ 1 -dependent cell adhesion and migration. However, the laminin-5 forms of ⌬G 3-5 and ⌬G 2-5 hardly promoted the cell adhesion and motility. These findings demonstrate that the G3 domain, but not the G4 and G5 domains, of the ␣3 chain is essential for the potent promotion of cell adhesion and motility by laminin-5.Laminins are a family of extracellular matrix proteins that are localized mainly in basement membranes and regulate various cellular functions such as adhesion, motility, growth, differentiation, and apoptosis through interaction with specific integrins on the cell surface (1, 2). The three subunits of laminins, designated ␣, ␤, and ␥ chains, form the well known cross-shaped structure linked together by disulfide bonds. Five ␣, three ␤, and three ␥ chains and at least 12 structural isoforms of laminin (laminin-1 to -12) having distinct chain combinations have been identified in human thus far (3, 4). Among these laminin isoforms, laminin-5, which consists of the laminin ␣3, ␤3, and ␥2 chains, is unique in the structure and biological activity. Laminin-5 was originally found as a keratinocyte-derived matrix protein named epiligrin, kalinin, or nicein (5-7) and a laminin-like cell scattering factor, ladsin, secreted by human gastric carcinoma cells (8). Laminin-5 has a feature lacking some domains found in the amino-terminal regions (or the short arms) of the three subunits of other laminin isoforms (2). The laminin ␣3 chain is found in laminin-6 (␣3␤1␥1) and laminin-7 (␣3␤2␥1) besides laminin-5, but the laminin ␤3 and ␥2 chains are found only in laminin-5. More interestingly, laminin-5 has much higher activity to promote adhesion, migration, and scattering of various types...

These proteinases include MMPs, ADAMs, type II transmembrane serine proteinases, and secretory serine proteinases such as plasminogen activators and trypsin. It is well known that some soluble and membrane-bound MMPs significantly contribute to tumor invasion and angiogenesis by degrading or processing extracellular matrix proteins and membrane proteins.(1-3)The type II transmembrane serine proteinase family is a newly identified superfamily of membrane-associated serine proteinases and consists of four subfamilies, including the hepsin subfamily and the matriptase subfamily.(4) Recently, considerable attention has been focused on matriptase, a member of the type II transmembrane serine proteinases superfamily. The matriptase family includes matripase/MT-SP1, matriptase-2, and matriptase-3. Matriptase contains a transmembrane domain, two CUB domains, four low-density lipoprotein-receptor domains and a serine protease domain.(5,6) Among the four family members, matriptase has been the most extensively characterized. Matriptase is expressed in a wide range of epithelial tissues, such as the epidermis, gastrointestinal tract and respiratory tract, and also in endothelial cells, neural cells and white blood cells. (7,8) Matriptase-deficient mice die within 48 h of birth due to a loss of the epithelial water barrier function, and they also exhibit profound abnormalities in follicular and thymic development. (9) Recent studies have suggested an association between defective matriptase regulation and/or activation and tumor development. Matriptase and its inhibitor HAI-1 are often co-expressed in cultured cancer cells and carcinomas as well as normal epithelial cells.(10-13) Co-expression of matriptase, HAI-1 and the HGFreceptor c-Met is correlated with poor prognosis of node-negative breast cancer patients.(12) In ovarian cancer, matriptase and HAI-1 are commonly co-expressed at an early stage, whereas the expression of HAI-1 is lost at stages III and IV, presumably leading to unopposed, active matriptase.(11) Matriptase activates PAR-2, latent HGF (pro-HGF) and pro-uPA. (14,15) Some animal studies have shown that matriptase promotes tumor growth. (16)(17)(18) For example, down-regulation of matriptase in human ovarian cancer cells by an antisense cDNA inhibits the activation of uPA and suppresses tumor growth in nude mice.(17) Furthermore, a recent study has shown that transgenic expression of matriptase in the epidermis of mice causes spontaneous squamous cell carcinoma.(19) However, it is still unclear how matriptase contributes to the development and malignant progression of human cancers.To clarify the roles of matriptase in tumor growth, here we overexpressed a matriptase cDNA in human stomach cancer cells. Our results demonstrate that the overexpression of matriptase strongly promotes tumor growth and angiogenesis in vivo.

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