Figure 5.

VBP1 regulates TCF/LEFs protein stability through pVHL.A, TCF7L2 protein levels in control or VBP1-overexpressing 786-O cells. Similar results were obtained from three experiments. B, TCF7L2 protein levels in control or VBP1-knockdown 786-O cells. Similar results were obtained from three experiments. C, the association between endogenous TCF7L2 and pVHL in control or VBP1-overexpressing cells. HEK293T cells were transfected with FLAG-tagged VBP1. After 48 h, the cell extracts were immunoprecipitated with an anti-TCF7L2 antibody. The immunoprecipitates and the inputs were analyzed by Western blotting with indicated antibody. D and E, the association between endogenous TCF7L2 and pVHL in control or VBP1-knockdown HEK293T (D) and HCT116 (E) cells. Cell extracts were immunoprecipitated with an anti-TCF7L2 antibody. The immunoprecipitates and the inputs were analyzed by Western blotting with indicated antibody. Similar results were obtained from three experiments. F and G, the transcriptional levels of indicated Wnt target genes in VBP1-overexpressing (F) or -depleted (G) HCT116 cells were analyzed by qRT-PCR. FLAG-VBP1 or endogenous VBP1 protein levels in VBP1-overexpressing or -depleted HCT116 cells were analyzed by Western blotting. Data are from 3–5 independent experiments with individual data points shown. H, representative images of colony formation assay were shown. I, quantification of the colony number shown in (H). Data are from four independent experiments with individual data points shown. Values are means ± S.D. Unpaired t test, two-tailed. *p < 0.05; **p < 0.01; ns, nonsignificant.