title: Regulation of β-catenin by t-DARPP in upper gastrointestinal cancer cells.
abstract: BACKGROUND
  Truncated dopamine and cyclic-AMP-regulated phosphoprotein (t-DARPP) is frequently overexpressed in gastrointestinal malignancies. In this study, we examined the role of t-DARPP in regulating β-catenin.
  
  
  RESULTS
  The pTopFlash construct that contains multiple TCF/LEF-binding sites was used as a measure of β-catenin/TCF transcription activity. Gastric (AGS, MKN28) and esophageal (FLO-1) adenocarcinoma cancer cell lines that lack t-DARPP expression were utilized to establish stable and transient in vitro expression models of t-DARPP. The expression of t-DARPP led to a significant induction of the pTOP reporter activity, indicative of activation of β-catenin/TCF nuclear signaling. Immunofluorescence assays supported this finding and showed accumulation and nuclear translocation of β-catenin in cells expressing t-DARPP. These cells had a significant increase in their proliferative capacity and demonstrated up-regulation of two transcription targets of β-catenin/TCF: Cyclin D1 and c-MYC. Because phosphorylated GSK-3β is inactive and loses its ability to phosphorylate β-catenin and target it towards degradation by the proteasome, we next examined the levels of phospho-GSK-3β. These results demonstrated an increase in phospho-GSK-3β and phospho-AKT. The knockdown of endogenous t-DARPP in MKN45 cancer cells demonstrated a reversal of the signaling events. To examine whether t-DARPP mediated GSK-3β phosphorylation in an AKT-dependent manner, we used a pharmacologic inhibitor of PI3K/AKT, LY294002, in cancer cells expressing t-DARPP. This treatment abolished the phosphorylation of AKT and GSK-3β leading to a reduction in β-catenin, Cyclin D1, and c-MYC protein levels.
  
  
  CONCLUSIONS
  Our findings demonstrate, for the first time, that t-DARPP regulates β-catenin/TCF activity, thereby implicating a novel oncogenic signaling in upper gastrointestinal cancers.
authors: Department of Surgery, Vanderbilt University Medical Center, Nashville, Tennessee, USA.
authors: Bhavatarini
authors: B
authors: Vangamudi
authors: Shoumin
authors: S
authors: Zhu
authors: Mohammed
authors: M
authors: Soutto
authors: Abbes
authors: A
authors: Belkhiri
authors: Wael
authors: W
authors: El-Rifai
mesh_terms: D001665:Binding Sites / Q000235:genetics; D015153:Blotting, Western; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D049109:Cell Proliferation; D002867:Chromones / Q000494:pharmacology; D051937:Dopamine and cAMP-Regulated Phosphoprotein 32 / Q000235:genetics* / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005770:Gastrointestinal Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D015972:Gene Expression Regulation, Neoplastic; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D009025:Morpholines / Q000494:pharmacology; D009154:Mutation; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D034622:RNA Interference; D015398:Signal Transduction / Q000187:drug effects; D051820:TCF Transcription Factors / Q000235:genetics* / Q000378:metabolism; D041742:Upper Gastrointestinal Tract / Q000378:metabolism / Q000473:pathology; D051176:beta Catenin / Q000235:genetics* / Q000378:metabolism
chemical_list: D002867:Chromones; D051937:Dopamine and cAMP-Regulated Phosphoprotein 32; D004791:Enzyme Inhibitors; D038362:Glycogen Synthase Kinase 3; D000071679:Glycogen Synthase Kinase 3 beta; D008156:Luciferases; D009025:Morpholines; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D020033:Protein Isoforms; D051057:Proto-Oncogene Proteins c-akt; D051820:TCF Transcription Factors; D051176:beta Catenin; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C000605506:GSK3B protein, human