Inappropriate WNT/β-catenin signal transduction has been implicated in various haematological malignancies for its ability to sustain proliferation of B- and T-cell immature progenitors as well as the self-renewal of haematopoietic stem cells (HSCs). Inactivating mutations of β-catenin, Adenomatous Polyposis Coli (APC) or Axin genes, identified in epithelial colorectal cancer (CRC), are known to prevent the glycogen synthase kinase-3β (GSK-3β)-dependent phosphorylation of β-catenin in specific N-terminal serine/threonine (Ser/Thr) residues. This promotes stabilization of β-catenin protein and its subsequent nuclear accumulation. In Chronic Myelogenous Leukemia (CML), a disease caused by constitutive activation of the tyrosine kinase (TK) Bcr/Abl, a marked β-catenin nuclear staining has been documented although the molecular causes underlying WNT pathway deregulation remain undefined. The purposes of this study were to study the molecular mechanisms by which the oncogenic Bcr/Abl promotes β-catenin accumulation in CML and the requirement of β-catenin in Bcr/Abl-transforming ability. In a human CML-established KU812 cell line and fresh CML patient samples β-catenin was constitutively tyrosine (Tyr)-phosphorylated and physically associated to Bcr/Abl and to the TCF-4 (T-cell factor-4) nuclear transcription factor. Inhibition of Bcr/Abl but not of Src tyrosine kinase activity prevented β-catenin Tyr-modification increasing its protein degradation rate in leukemic cells as judged by analysis of 35S-incorporation in β-catenin (pulse-chase assay). Accordingly, phosphorylation of β-catenin Tyr-86 and Tyr-654 was observed in HEK-293T harboring activated Bcr/Abl correlating with increased β-catenin cellular levels. Imatinib, a Bcr-Abl antagonist, affected β-catenin signaling function disrupting β-catenin/TCF-4 nuclear complexes as well as promoting a caspase-dependent cleavage of β-catenin in Bcr/Abl+ cells undergoing apoptosis. Silencing of β-catenin by siRNA decreased the expression of cyclin D1, a TCF4-target gene, and synergistically with Imatinib reduced CML-Bcr/Abl+ KU812 cell-growth. Although β-catenin was also detectable in its Ser/Thr-phosphorylated form (APC/Axin/GSK3β-dependent) which is more sensitive to proteosomal degradation, the β-catenin recruited by Bcr/Abl never showed this modification. Pull-down experiments revealed that β-catenin Tyr-phosphorylation occurring constitutively in Bcr/Abl+ cells prevented β-catenin-Axin/GSK3β interaction thus exerting a dominant effect on β-catenin protein stabilization. In conclusion, these results establish a functional link between Bcr/Abl kinase activity and a post-translational control of nuclear function of β-catenin which contribute to extend the lifespan of leukemic cells during CML progression.
[Proc Amer Assoc Cancer Res, Volume 47, 2006]