β-catenin is involved in N-cadherin–dependent adhesion, but not in canonical Wnt signaling in E2A-PBX1–positive B acute lymphoblastic leukemia cells

ObjectiveThe t(1;19)(q23;13) translocation, resulting in the production of the E2A-PBX1 chimeric protein, is a common nonrandom translocation in pediatric B-lineage acute lymphoblastic leukemia (B-ALL). The E2A-PBX1 chimeric protein activates expression of several genes, including Wnt16. In the present study, we explored the role of Wnt16 and β-catenin in t(1;19) B-ALL cells.Materials and MethodsCanonical Wnt signaling was measured by TOPflash activity. Localization of β-catenin in the cell membrane and its involvement in leukemia-stroma interaction were studied by confocal microscopy. Adhesion to N-cadherin was analyzed by adding 3H-thymidin–labeled cells to N-cadherin–coated wells.ResultsIn contrast to previous reports, we detected no effects on cell viability or proliferation upon modulation of the Wnt16 levels. Moreover, despite high levels of Wnt16 and β-catenin, the cells had very low levels of canonical Wnt signaling. Instead, β-catenin was located in the cell membrane along with N-cadherin. E2A-PBX1–positive leukemia cells adhered strongly to bone marrow stroma cells, and we showed that adherence junctions stained strongly for both proteins. Moreover, knockdown of β-catenin reduced the adhesion of E2A-PBX1–positive leukemia cells to N-cadherin, suggesting that β-catenin and N-cadherin play a central role in homotypic cell-to-cell adhesion and in leukemia–stroma adhesion. Interestingly, knockdown of Wnt16 by small interfering RNA reduced the level of N-cadherin.ConclusionWnt16 does not activate canonical Wnt signaling in E2A-PBX1–positive cells. Instead, β-catenin is involved in N-cadherin–dependent adherence junctions, suggesting for the first time that leukemia–stroma interactions may be mediated via an N-cadherin–dependent mechanism.