Artificially accumulated β-catenin inhibits proliferation and induces neurite extension of neuroblastoma cell line NB-1 via up-regulation of trkA

Recent evidence suggests an association between up-regulation of β-catenin/Wnt signaling pathway and neuronal differentiation of neuroblastoma. We overexpressed β-catenin into a human neuroblastoma cell line NB-1 and observed its effect on cellular morphology, growth potential and alteration in a known differentiation related gene, trkA. Expression plasmids containing wild-type and mutated forms of β-catenin gene were transfected into NB-1 cells, using liposome-based transfection method. The mutated forms were a deletion of three nucleotides of codon 45 and a large deletion involving the whole exon 3. In the transient transfection model, cell viability assay demonstrated significant negative effect of mutated β-catenin transfection, but not wild-type, on the cell proliferation. To investigate impacts of β-catenin overexpression in detail, a stable transfection model was established. Clones with comparable expression of β-catenin at the mRNA level were selected. Only the selected clones with mutated form of β-catenin exhibited neurite extension pattern and stunned cell proliferation, in association with higher accumulation of total cellular β-catenin protein as evidenced by Western blot and immunocytochemistry. Cell cycle progression demonstrated significantly higher G0-G1 fraction in each stable cell clone with β-catenin expression plasmid. In addition, retarded G1/S transition was observed exclusively in the cell clones with mutated form. Concomitantly with overexpressed β-catenin, up-regulations of trkA and Ha-ras were also identified. Our study suggests a potential availability of β-catenin/Wnt signaling pathway as a target of molecular manipulation for treatment of high-risk neuroblastoma and a potential association between the pathway and the trkA/neurotrophin cascades.