Expression of a Human Hepatocyte Growth Factor/Scatter Factor eDNA in MDCK Epithelial Cells Influences Cell Morphology, Motility, and Anchorage-independent Growth

The addition of exogenous hepatocyte growth factor (HGF)/scatter factor (SF) to MDCK epithelial cells results in fibroblastic morphology and cell motility. We generated HGF/SF producing MDCK cells by transfection with an expression plasmid containing human HGF/SF eDNA. Production of HGF/SF by these cells induced a change from an epithelial to a fibroblastic morphology and increased cell motility. In addition, the HGF/SF producing cells acquired efficient anchorage-independent growth in soft agar but did not form tumors in nude mice. The morphological change and the stimulation of the anchorage-independent growth were prevented by anti-HGF/SF antibody, suggesting that the factor is secreted and then exerts its effects through cell surface receptors. C ELL dispersion and motility are required for normal embryogenesis and tissue remodeling, and are also important steps in the invasion of tumor cells (Rosen and Goldberg, 1989). The molecular mechanisms responsible for the induction of cell dispersion and cell motility remain unclear. Several factors affecting cell motility have been discovered (Rosen and Goldberg, 1989). Among them, scatter factor (SF), ~ a mesenchymal cell-derived protein, dissociates epithelial cell colonies to individual cells and stimulates the migration of epithelial cells (Stoker and Perryman, 1985; Stoker et al., 1987; Gherardi et al., 1989). Since implantation of SF-producing human fibmblasts into chick embryos caused abnormal development, it was suggested that SF is involvedin embryogenesis (Stem et al., 1990). Purified SF promotes the invasiveness into collagen matrices of a number of human carcinoma cell lines, suggesting that SF is involved in the invasion of tumor cells (Weidner et al., 1990). Recently, scatter factor has been found to be identical to hepatoeyte growth factor (HGF) (Weidner et al., 1991). HGF was originally identified as a potent mitogen for hepatocytes in primary culture (Gohda et al., 1988; Nakamura et al., 1987; Zarnegar and Michalopoulos, 1989). HGF is also mitogenic in melanocytes and endothelial and epithelial cells (Rubin et al., 1991). Moreover, the same factor was purified from the culture medium of human embryonic lung fibroblasts as a cytotoxie factor in some tumor cell lines (Higashio et al., 1990). Thus, HGF has a broad spectrum L Abbreviations used in this paper: HGF, hepatocyte growth factor; SF, scatter factor. of activities among its various target cells. To simplify the multiple names of this factor, we will refer to it as HGF/SF in this paper. We and others have cloned and sequenced the eDNA for human and rat HGF/SF (Miyazawa et al., 1989; Nakamura et al., 1989; Tashiro et al., 1990; Okajima et al., 1990). Human HGF/SF is synthesized as a single 90-kD precursor, whose NH2-terminal 31 amino acid residues probably function as a signal peptide (Yoshiyama et al., 1991). The precursor is glycosylated and cleaved at a specific proteolytic site to yield a 65-kD heavy chain and a 35-kD light chain, which are linked together by a disulfide bond. The dog kidney epithelial cell line, MDCK, is a highly sensitive target and has been used in a bioassay for the factor. In tissue culture, MDCK cells form tight epithelial monolayers with junctional complexes (McRoberts et al., 1981). When added to cultured MDCK cells, purified SF causes the breakdown of intercellular junctions and a transition from an epithelial to a fibroblastic morphology (Gherardi et al., 1989; Weidner et al., 1990). It also stimulates the motility of MDCK cells (Stoker, 1989). To determine the potential autocrine role of HGF/SF in the acquisition of scattering and motile phenotypes at the single cell level, we introduced a plasmid that directs the constitutive synthesis of human HGF/SF into MDCK epithelial cells. Cells producing human HGF/SF were converted morphologically to fibroblast-like scattered cells and became motile. These cells also acquired efficient growth property in soft agar. Since the morphological change and the stimulation of the growth in soft agar were prevented in the presence of antiHGF/SF antibody, the factor is secreted and then exerts its effects through receptors on the cell surface. 9 The Rockefeller University Press, 0021-9525/92/05/889/6 $2.00 The Journal of Ceil Biology, Volume 117, Number 4, May 1992 889-894 889 on M ay 2, 2016 jcb.rress.org D ow nladed fom Published May 15, 1992