PP60(C-SRC) IS A POSITIVE REGULATOR OF GROWTH FACTOR-INDUCED CELL SCATTERING IN A RAT BLADDER-CARCINOMA CELL-LINE

The NBT-II rat carcinoma cell line exhibits two mutually exclusive responses to FGF-I and EGF, entering mitosis at cell confluency while undergoing an epithelium-to-mesenchyme transition (EMT) when cultured at subconfluency. EMT is characterized by acquisition of cell motility, modifications of cell morphology, and cell dissociation correlating with the loss of desmosomes from cellular cortex. The pleiotropic effects of EGF and FGF-1 on NBT-II cells suggest that multiple signaling pathways may be activated. We demonstrate here that growth factor activation is linked to at least two intracellular signaling pathways. One pathway leading to EMT involves an early and sustained stimulation of pp60(c-src) kinase activity, which is not observed during the growth factor-induced entry into the cell cycle. Overexpression of normal c-src causes a subpopulation of cells to undergo spontaneous EMT and sensitizes the rest of the population to the scattering activity of EGF and FGF-1 without affecting their mitogenic responsiveness. Addition of cholera toxin, a cAMP-elevating agent, severely perturbs growth factor induction of EMT without altering pp60(c-src) activation, therefore demonstrating that cAMP blockade takes place downstream or independently of pp60(c-src). On the other hand, overexpression of a mutated, constitutively activated form of pp60(c-src) does not block cell dispersion while strongly inhibiting growth factor-induced entry into cell division. Moreover, stable transfection of a dominant negative mutant of c-src inhibits the scattering response without affecting mitogenesis induced by the growth factors. Altogether, these results suggest a role for pp60(c-src) in epithelial cell scattering and indicate that pp60(c-src) might contribute unequally to the two separate biological activities engendered by a single signal.