Figure 3. Grb2 synergizes with downstream components of the wnt signaling pathway.

(A) Wnt3a expressed by one population of cells synergizes with Grb2 in a second population. HEK293 cells were transfected with either Wnt3a or GFP expression plasmids and mixed with cells transfected with GFP or Grb2 expression plasmids along with sTF. Luciferase activity was assessed at 24 hours. * – p<0.02 compared to GFP alone. (B) Grb2 synergizes with CA-LRP6. HEK293 were transfected with sTF, (CA) LRP6 and Grb2 and incubated for 24 hrs before harvesting for luciferase assay. * – effect of CA-LRP6 + Grb2 differs significantly from effect of either alone: p<0.01. (C) As for (B) except CA-β-catenin (S33Y) was used. * – effect of CA-β-catenin + Grb2 differs significantly from effect of either alone: p<0.01. (D) As for (B) except LEF1 was used. * – effect of LEF1 + Grb2 differs significantly from effect of either alone: p<0.01. (E) Grb2 and CA-β-catenin cooperate to induce the MMP9 promoter. HEK293 cells were transfected with MMP9-Luciferase reporter along with CA β-catenin, and/or Grb2. Phorbol ester (PMA, 25 ng/mL) was used as a positive control and was added 3 hrs post transfection. Cells were harvested 24 hrs later and analyzed for luciferase activity. * – p<0.01, relative to GFP; ** – p<0.02, relative to Grb2 or CA-β-catenin alone. (F) Grb2 and LEF1 cooperate to induce the MMP9 promoter. Transfections as described in (E). * – p<0.05, relative to GFP; ** – p<0.01 relative to Grb2 or LEF1 alone. All luciferase data are normalized to total protein content. Mean and standard error of the mean are shown. Significance assessed by Student's t-test.