Silencing of Wnt components reduces TTF-1 levels and required for the basal expression of <i>TTF-1</i>.

<p><b>A, B and C:</b> TPC-1 cells were transfected with 50 nM of siRNAs targeting LRP6, β-catenin, LEF1 or TCF-4, and a non-targeting siRNA sequence (siNT) as control. mRNA expression of <i>LRP6</i>, <i>β-catenin</i>, <i>LEF1</i>, <i>TCF4</i> (<b>A</b>) and <i>TTF-1</i> (<b>B</b>) were analyzed by real-time PCR and normalized to <i>RPL13A</i> mRNA, then recorded as the fold change compared to the non targeting siRNA (siNT). <b>A</b>. Significant inhibition of <i>LRP6</i>, <i>β-catenin</i>, <i>LEF1 and TCF4</i> was found for cells transfected with siRNAs targeting the corresponding genes compared with those transfected with the scrambled sequence. (<b>B</b>) Significant inhibition of <i>TTF-1</i> in cells transfected with siRNAs targeting LRP6, β-catenin, LEF1 or TCF-4. (<b>C</b>) Western blot data showing the effect of siRNAs targeting LRP6, β-catenin, LEF1 and TCF-4 on TTF-1 protein expression, β-actin was used as internal control (images from one experiment representative of three independent experiments) (<b>D</b>) TPC-1 cells were co-transfected with TOP-Flash-Luc vectors or with the TTF-1-luc-2.11 and dominant-negative plasmids (LRP6Dn, DEPXDn, β-cateninDn, TCFDn) and luciferase activity was assessed 24 h later. Results were expressed as percent of transcriptional activity as described in Material and Methods section. Columns represent three independent experiments and bar the standard deviation of the mean.