RT-PCR results of Fz4 mRNA expression in normal mucosa-derived NCM460 cells and the colon cancer cell lines HCT116 and RKO following exposure to Wnt-producing cells in co-culture or cells transfected with a non-Wnt-producing pCMV control plasmid. The Fz4 RT-PCR product was 832 bp. All experiments were performed simultaneously with actin primers, producing a 544 bp product for quality control. Fz4 mRNA was downregulated by Wnt2 in normal colonic mucosa-derived NCM460 cells but not in the colon cancer cell lines. No effect on Fz4 expression following exposure to Wnt5a was seen. A. NCM460, HCT116 and RKO cells co-cultured using inserts with sham-transfected cells. B. NCM460, HCT116 and RKO co-cultured with Wnt2-producing cells. C. NCM460 co-cultured with Wnt5a-producing cells. D. Wnt 2 and Wnt5a production by transfected RKO; Western blot. A non-specific band is present in all 3 lanes, including control. The Wnt2 and Wnt5a specific bands are annotated with an arrow.