1. Fig. 5 β-Catenin/Tcf7l2 complex recruits Zic5 to regulate gene expression.
β-Catenin/Tcf7l2 complex recruits Zic5 to regulate gene expression. (A) MudPIT analysis of the purification of 3×Flag-Zic5 and 3×Flag-Zic2 in HCT116 cells from nuclear extracts. (B and C) Validation of Tcf7l2 and β-catenin co-immunoprecipitation with Zic5 or Zic2 by Western blot. VT, vector control. (D) Tcf7l2 but not β-catenin directly binds to Zic5 and Zic2 in the in vitro binding assay. IB, immunoblotting. (E) C terminus of Zic5 is required to be directly associated with the β-catenin/Tcf7l2 complex. (F and G) CTNNB1 or TCF7L2 knockdown in HCT116 cells by distinct short hairpin RNAs (shRNAs). The protein levels of β-catenin, Tcf7l2, Zic2, and Zic5 were examined by Western Blot. (H) Average plot and (I) heat maps showing the global binding of Zic5 in shControl cells and shCTNNB1 or shTCF7L2 cells from −1 to +1 kb centered on Zic5 peaks in shControl cells. (J) Boxplot depicts the normalized reads coverage for Zic5 peaks in shControl cells and shCTNNB1 or shTCF7L2 cells. The P values were calculated with the Wilcoxon signed-rank test. (K and L) Track examples showing the Zic5 binding at the GLUT1 gene promoter region with Tcf7l2 or β-catenin knockdown. IgG, immunoglobulin G.
Zibo Zhao et al. Sci Adv 2019;5:eaax0698
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