Fig. 5: PLD6 upregulates Wnt/β-catenin signaling pathways.

a The protein levels of β-catenin and its target genes were assessed in the indicated cells after treatment with recombinant Wnt3a (150 ng ml⁻¹) for 6 h. b The expression levels of β-catenin and its target genes were analyzed in PLD6-KO Caco-2 cells using qRT–PCR. c The cells were cotransfected with the TOP/Flash or FOP/Flash reporter constructs and Renilla luciferase. After Wnt3a treatment for 4 h (or no treatment), the cell lysates were collected to measure luciferase activity. d A ChIP assay was performed using an anti-β-catenin antibody to demonstrate β-catenin binding to its target gene promoters. e The β-catenin stability was evaluated in Caco-2 and HCT116 cells treated with cycloheximide (CHX) (10 μg ml⁻¹) for the indicated time points. f The HEK293T cells were treated with a single-guide RNA targeting PLD6 (sgPLD6) for 16 h to deplete PLD6, followed by treatment with or without MG132 (10 μM) for an additional 6 h. g The confocal microscopy images of PLD6-depleted Caco-2 cells and PLD6-overexpressing cells showing β-catenin localization (green) and nuclear colocalization (blue). The representative images were obtained from at least three fields. The results represent at least three independent experiments. Scale bar, 100 μm. Statistical significance: *P < 0.05, **P < 0.01, ***P < 0.001.