Abstract
Little is known about the the signalling pathways driving the adenoma-to-carcinoma sequence in human colonic epithelial cells. Accumulation and activation of the src tyrosine kinase in colon cancer suggest a potential role of this oncogene in this early progression. Therefore, we introduced either activated src (m-src), polyoma-MT alone or combined with normal c-src in the adenoma PC/AA/C1 cell line (PC) to define the function and phenotypic transformations induced by these oncogenes in familial adenomatous polyposis (FAP) colonic epithelial cells. Functional expression of these oncoproteins induced the adenoma-to-carcinoma conversion, overexpression of the hepatocyte growth factor (HGF) receptor Met, but failed to confer invasiveness in vivo and in vitro, or to produce alterations in cell proliferation and differentiation. In contrast, PC-msrc cells became susceptible to the HGF-induced invasion of collagen gels and exhibited sustained activation of the pp60src tyrosine kinase and Tyr phosphorylation of the 120-kDa E-cadherin, which was further increased by HGF Transcripts of HGF were clearly identified by reverse transcription-polymerase chain reaction (RT-PCR) and Southern blot in the parental and transformed PC cells, suggesting an autocrine mechanism. Taken together, the data indicate that: (1) experimental activation of src and PyMT pathways directly induces tumorigenicity and Met upregulation in a colon adenoma cell line; (2) HGF-activated Met and src cooperate in inducing invasion; (3) in view of the molecular associations between catenins and cadherin or the tumour-suppressor gene product APC, the cell adhesion molecule E-cadherin may constitute a downstream effector of src and Met.
Highlights
Introduction of mutated src confer thehepatocyte growth factor (HGF)-dependent invasiveness in collagen gelsNo metastases were histologically evidenced in peritoneal cavity, liver or lungs in nude mice injected subcutaneously with the more aggressive PCPy/c-src cells, as well as the other oncogene-transfected cells
We characterized the tumorigenic potential and the pattern of differentiation and proliferation of these transfected PC/AA/C1 cell line (PC) AA/C I cell derivatives. Since both APC and pp6Osr have been shown to be localized or associated with cell-cell adhesion effectors or cytoskeletal components, we investigated the invasive phenotype of the parental and transfected PC cell lines in relation to the expression of the receptor, tyrosine kinase Met, and its ligand hepatocyte growth factor / scatter factor (HGF/SF) as known effectors of invasion (Guan and Shalloway, 1992; Rubinfield et al, 1993; Rosen et al, 1994; Smith et al, 1994)
Since the paracrine factor HGF secreted by stromal cells acts on the proliferation and the scattering of epithelial cells (Rosen et al, 1994), we investigated the invasive properties of the parental PC cells and their derivatives in the presence of HGF/SF
Summary
Introduction of mutated src confer theHGF-dependent invasiveness in collagen gelsNo metastases were histologically evidenced in peritoneal cavity, liver or lungs in nude mice injected subcutaneously with the more aggressive PCPy/c-src cells, as well as the other oncogene-transfected cells. The parental and transfected PC cell lines failed to invade into the collagen gel, like MCF-7/AZ cells, which were used as negative control (Table 1). PCmsrc cells became invasive into collagen gels after HGF treatment. These results indicate that HGF/SF alone is not sufficient to confer the invasive phenotype to PC cells, but that it cooperates with the activated pp6Osrc transduction pathway to induce invasion in PCmsrc. The absence of response of the other cell lines to HGF is not related to a dysfunction of the HGF receptor Met, since the treatment of the parental and oncogene-transfected PC cells is associated with tyrosine phosphorylation of the Met receptor, corresponding to its activation (data not shown). The parental PC cells and their derivatives were tested for their invasiveness using embryonic chick heart fragments (Table 1). Addition of HGF in this assay had no effect on the invasion properties of PC cell lines (data not shown)
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