Figure 1

Sequestration of β-catenin by GFP-tagged E-cadherin lead to up-regulation of CLU levels in colon carcinoma cells. Changes in CLU protein levels were investigated in response to over-expression of GFP tagged E-cadherin and GFP as control. (A) Immunofluorescence was used to show the effects of transient over-expression of GFP tagged E-cadherin, which binds β-catenin, in LS174T and HCT116 colon carcinoma cell lines. In the left panel it is shown that the GFP-cyt-E-cadherin fusion protein efficiently sequesters β-catenin from the nucleus whereupon Wnt signaling is abrogated. In transfected cells (white arrow heads) β-catenin has a perinuclear localization contrasting untransfected cells (white arrows) where it is uniformly distributed. Shown in the right panel is the up-regulation of cytoplasmic CLU protein which follows the abrogation of Wnt signaling induced by the GFP-cyt-E-cadherin fusion protein. CLU is up-regulated in GFP-cyt-E-cadherin transfected cells (yellow arrow heads) but not in untransfected cells nor in cells transfected with GFP alone. (B) Western blot with cell lysates from LS174T cells 24 hr after transient transfection with GFP-cyt-E-cadherin and GFP as a control. β-actin was used as a loading control. CLU protein levels are up-regulated and c-MYC protein levels are down-regulated as a consequence of over-expressing GFP-cyt-E-cadherin but not GFP alone. WB: western blot, ab: Antibody.