ShRNA-mediated gene silencing of β-catenin inhibits growth of human colon cancer cells

AIM: To observe the gene silencing mediated by the specific shRNA targeted against beta-catenin and its effect on cell proliferation and cycle distribution in the human colon cancer cell line Colo205. METHODS: Two shRNA plasmid vectors against beta-catenin were constructed and transfected into Colo205 cells with Lipofectamine (TM) 2000. The down-regulations of beta-catenin, c-myc and cyclinD1 expressions were detected by RT-PCR and western blot analysis. The cell proliferation inhibitions were determined by MTT assay and soft agar colony formation assay. The effect of these two beta-catenin shRNAs on cell cycle distribution and apoptosis was examined by flow cytometry. RESULTS: These two shRNA vectors targeted against p-catenin efficiently suppressed the expression of p-catenin and its down stream genes, c-myc and cyclinD1. The expression inhibition rates were around 40%-50% either at the mRNA or at the protein level. The shRNA-mediated gene silencing of beta-catenin resulted in significant inhibition of cell growth both on the culture plates and in the soft agar. Moreover, the cancer cells showed significant G(0)/G(1) arrest and increased apoptosis at 72 h post transfection due to gene silencing. CONCLUSION: These specific shRNAs targeted against p-catenin could have a gene silencing effect and block the WNT signaling pathway. They could inhibit cell growth, increase apoptosis, and induce cell cycle arrest in Colo205 cells. ShRNA interference against beta-catenin is of potential value in gene therapy of colon cancer. (C) 2007 WJG. All rights reserved.