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. 2012 Jul 6;31(16):3384–3397. doi: 10.1038/emboj.2012.181

Figure 1.

Figure 1

DP1 exerts both positive and negative roles in Wnt/β-catenin signalling. (A) TOP-FLASH activities and cytosolic β-catenin level were measured in HEK293T cells. Flag–DP1 was transfected and cells were treated with Wnt3a conditioned medium (Wnt3a-CM) for 20 h. Error bars indicate the standard deviations of three independent analyses. P<0.01. (B) Knockdown efficiencies of two siDP1 were measured by immunoblotting with anti-DP1 antibody. (C) TOP-FLASH activities were measured in cells transfected with siDP1 and treated with Wnt3a-CM. siGFP was transfected as a control. Data represent average values from one representative experiment performed in triplicate. Error bars indicate standard deviations of triplicate. (D) Quantitative real-time PCR analysis for the expression of Wnt-target genes (Axin2, c-Jun and LEF1) after knockdown of DP1 by siRNA in HEK293 cells treated with or without Wnt3a-CM. The quantities of indicated mRNA were normalized by β-actin. Data represent average values from one representative experiment performed in triplicate. Error bars indicate standard deviations of triplicate. (E) Knockdown of DP1 hampered Wnt3a-mediated stabilization of cytosolic β-catenin in HEK293 cells. (F) TOP-FLASH activities were measured in cells treated with Wnt3a-CM or transfected with plasmids as indicated. Data represent average values from one representative experiment performed in triplicate. Error bars indicate standard deviations of triplicate. (G) TOP-FLASH activities were measured in cells transfected with indicated plasmids. Data represent average values from one representative experiment performed in triplicate. Error bars indicate standard deviations of triplicate. Significant differences were evaluated either by P<0.001 or by P<0.005. (H) Axis duplication was assessed in Xenopus tadpole embryo. mRNAs for xWnt8 (4 pg), xDsh (500 pg), β-catenin (500 pg) and DP1 (1 ng with β-catenin or 2 ng each with xWnt8 and xDsh) were injected as indicated. Arrows indicate duplicated axis. Each number of analysed embryos (n) is indicated.