FIGURE 8.

Induction and repression of Wnt3a target genes require the canonical β-catenin pathway. A, TOPFLASH assay to establish complete inhibition of Wnt/β-catenin signaling with IWR-1 and quercetin. NIH3T3 cells were treated for 24 h with DMSO (Control), IWR-1 at 100 μm, or quercetin at 500 μm either in control medium (DMEM/10% FBS) or in DMEM/10% FBS plus 500 ng/ml Wnt3a-RP. IWR-1 and quercetin at these concentrations completely blocked Wnt3a/β-catenin-induced TOPFLASH activity in NIH3T3 cells. B, NIH3T3 cells were treated with Wnt3a-RP (500 ng/ml) for 36 h in the presence or absence of IWR-1 and quercetin at 100 and 500 μm, respectively, followed by RNA preparation and qRT-PCR. The expression levels of three positively regulated (Tnfsf9, Mrgprf, and Prickle1) and three negatively regulated (Lcn2, Casp12, and Arrdc4) Wnt3a target genes were analyzed as described in the legend for Fig. 3. Complete inhibition of Wnt/β-catenin signaling eliminated both the induction and repression of target genes by Wnt3a-RP (*, p < 0.05).