FIGURE 4.

Enhanced phosphorylation and Wnt/β-catenin signaling activation of LRP6 with defective tyrosine-based motifs. A, lysates of cells expressing WT or LRP6 mutants were subjected to Western blot analysis and probed with anti-LRP6 or anti-phospho-LRP6 (p-LRP6) (Ser-1490) antibodies. Ymt, tyrosine mutant; Smt, serine/threonine mutant. B, the ratios of phospho-LRP6 versus total LRP6 (p-LRP6/LRP6)were compared between WT and tyrosine mutant. C–E, cells expressing WT or mutant LRP6 were transfected with TOPFlash/FOPFlash plasmids and were subsequently incubated with control CM or Wnt3a CM for 16 h. Wnt/β-catenin signaling activity was evaluated with TOPFlash TCF-luciferase reporter (C and E) or GST-E-cadherin pulldown assays (D). To clearly illustrate the signal activation in the presence or absence of Wnt3a stimulation, the activities were plotted together (C) or separately (E). Data are representative of three independent experiments. Data represent mean ± S.D. N.S., not significant; **, p < 0.01.