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. 2023 Apr 1;299(5):104667. doi: 10.1016/j.jbc.2023.104667

Figure 5.

Figure 5

FOXD1 and FOXI1 have discrete functions in the Wnt pathway.A, TOPflash assay in 293T. Where indicated, cells were treated with 5 ng/ml recombinant human R-spondin 3 (Rspo3) or 10 nM porcupine inhibitor LGK974. B, TOPflash assay in 293T ΔLRP5/6. Where indicated, cells were transfected with constitutively active LRP6 (LRP6ΔE1-4) to uncouple Wnt/β-catenin pathway activation from LRP5/6 engagement by WNT proteins. FOXQ1 and FOXB2 were included as positive and negative controls, respectively. Data were normalized to the corresponding empty vector control. C, TOPflash assay in 293T following WNT7B or RECK depletion. D, WNT1 and WNT7B promoter reporter assay in 293T. E, TOPflash assay in 293T. Where indicated, cells were treated with 5 ng/ml recombinant human R-spondin 3 (Rspo3) or 10 nM LGK974. F, immunoblot of nonphosphorylated, active β-catenin (ABC) levels in Wnt3a-treated 293T following FOXI expression. The relative ratio of ABC versus HSP70 housekeeping control is indicated below. G, TOPflash assay in HCT116. Where indicated, cells were treated with 100 nM proteasome inhibitor bortezomib. Data in (B) and (G) were analyzed using an unpaired Welch’s t test with Bonferroni-Hochberg correction for multiple testing. Data in (C) and (D) were analyzed using Dunnett’s post hoc test against EV or siControl following one-way ANOVA (∗∗∗p < 0.001, ∗p < 0.05). RLA, relative luciferase activity. See also Fig. S7.