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. Author manuscript; available in PMC: 2013 Oct 1.
Published in final edited form as: Biochim Biophys Acta. 2012 Jul 16;1823(10):1807–1814. doi: 10.1016/j.bbamcr.2012.07.004

Fig. 5.

Fig. 5

β-catenin/Tcf dependent transcription regulates MAP4K1 expression, JNK activation, and AP-1 dependent transcription. (A) Over-expression of dnTcf4 inhibits the expression of MAP4K1. GEO-shPdcd4 cells transfected with pcDNA4/Max (control) or pcDNA4-dnTcf4 (dnTcf4) plasmid along with pMACS K+.II plasmid were collected for extracting RNA or making cell lysates. The dnTcf expressing cells were enriched by H-2Kk antibody conjugated beads as described in Materials and methods. Total RNA was isolated and used in qPCR to examine the mRNA level of c-myc and map4k1. The ratio of c-myc/GAPDH and map4k1/GAPDH in cells transfected with pcDNA4/Max is designated as 100%. Two independent experiments were performed with 3 replicates for each sample. The data are shown and expressed as mean ± SD. The asterisk indicates a significant difference compared with control cells as determined by one-way ANOVA (P<0.05). (B) Over-expression of dnTcf4 inhibits map4k1 promoter activity. Indicated amount of pcDNA4-dnTcf4 plasmid was transfected into GEO-shPdcd4 cells while the total DNA for each transfection was maintained at 0.6 µg by adding pcDNA4/Max vector DNA. The activity of GEO-shPdcd4 cells transfected with 0 µg of dnTcf4 expression plasmid is designated as 100%. Three independent experiments were performed with 5 replicates for each sample. The represented data are shown and expressed as mean ± SD (n = 5). The asterisk denotes a significant difference compared with cells transfected with 0 µg of dnTcf4 expression plasmid as determined by one-way ANOVA (P<0.001). (C) Over-expression of dnTcf4 inhibits the activation of the JNK pathway. Western blot analysis was performed using the cell lysates from (A) with the indicated antibodies. The ratio of target protein/GAPDH in control cells is designated as 1.0. (D) Over-expression of dnTcf4 suppresses AP-1 dependent transcription. Indicated amount of pcDNA4-dnTcf4 plasmid was transfected into GEO-shPdcd4 cells while the total DNA for each transfection was maintained at 0.4 µg by adding pcDNA4/Max vector DNA The activity of GEO-shPdcd4 cells transfected with 0 µg of dnTcf4 expression plasmid is designated as 100%. Three independent experiments were performed with 5 replicates for each sample. The represented data are shown and expressed as mean ± SD (n = 5). The asterisk denotes a significant difference compared with cells transfected with 0 µg of dnTcf4 expression plasmid as determined by one-way ANOVA (P<0.01).