Growth factor-dependent activation of the Ras-Raf-MEK-MAPK pathway in the human pancreatic carcinoma cell line PANC-1 carrying activated K-ras: implications for cell proliferation and cell migration

Human ductal adenocarcinoma of the pancreas frequently carry activating poi nt mutations in the K-ras protooncogene. We have anal! sed the activity of the Ras-Raf-MEK-MAPK cascade in the human pancreatic carcinoma cell line PA NC-1 carrying an activating K-ras mutation. Serum-starved cells and cells g rown in medium with serum did not show constitutively activated c-Raf, MEK- 1, or p42 MAPK. Stimulation bf cells with epidermal growth factor (EGF) or fetal calf serum (FCS) resulted in activation of N-Ras, but not K-Ras, as w ell as activation of c-Raf, MEK-1, and p42 MAPK. Preincubation of serum-sta rved cells with MEK-1 inhibitor PD98059 abolished EGF- and FCS-induced MAPK activation, identifying MEK as the upstream activator of MAPK. PANC-1 cell s exhibited marked serum-dependence of anchorage-dependent and -independent cell growth as well as cell migration. EGF, alone or in combination with i nsulin and transferrin, did not induce cell proliferation of serum-starved PANC-1 cells, indicating that activation of MAPK alone was not sufficient t o induce cell proliferation, FCS-induced DNA synthesis was inhibited by 40% by the MEK-1 inhibitor. On the other hand, treatment with either FCS or EG F alone resulted in marked, MEK-dependent increase of directed cell migrati on. Collectively, our results show that the activating K-ras mutation in PA NC-1 cells does not result in constitutively increased Raf-MEK-MAPK signali ng. Signal transduction via the Ras-Raf-MEK-MAPK cascade is maintained in t hese cells and is required for growth factor-induced cell proliferation and directed cell migration.