NIC/CSL TRANSCRIPTIONAL ACTIVITY IS REGULATED BY THE MEK/ERK PATHWAY IN HUMAN PANCREATIC CANCER CELLS

Abstract

Background: Notch are transmembrane receptors which, upon ligand binding, are cleaved thus releasing a Notch intracellular domain (NIC). NIC then translocates into the nucleus where it binds the DNA-binding protein CSL to induce expression of target genes. The Notch pathway is normally inactive in the adult pancreas. However, recent findings revealed that this pathway is reactivated during pancreatic carcinogenesis. The mechanisms by which Notch activity is regulated are still unknown. Aim: To investigate the role of the Mek/Erk pathway in Notch activity regulation. Methods: The MIA PaCa-2 human pancreatic cancer cell line was used. The specific Mek inhibitor U0126 was used while the phorbol-ester PMA was used to stimulate the Mek/Erk pathway. NIC/CSL transcriptional activity was measured by luciferase assays using the CSL-luciferase reporter gene. Overexpression of Notch1 and NIC cDNAs were used to up-regulate NIC/CSL transcriptional activity. Results: 1-Inhibition of the Mek/Erk pathway down-regulated equally (by 50%) the basal and the Notch- and NIC-induced NIC/CSL transcriptional activity. 2-Conversely, strong and sustained activation of the Mek/Erk pathway by PMA increased by 3-fold the basal and Notch- and NIC-induced NIC/CSL transcriptional activity, 3-an effect that was completely blocked by addition of U0126. 4-No modulation in NIC expression was observed following U0126 or PMA treatment. Conclusion: Taking together, these results suggest that the Mek/Erk pathway can directly affects NIC/CSL transcriptional activity independently of an impact on NIC expression.