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. Author manuscript; available in PMC: 2017 Feb 27.
Published in final edited form as: Development. 2007 Dec 12;135(2):367–375. doi: 10.1242/dev.013540

Fig. 3. Dvl is required for Wnt-induced LRP6 phosphorylation and acts via the DIX and PDZ domains.

Fig. 3

(A) Reduction of Dvl proteins diminished Wnt3a-induced Lrp6 (endogenous) phosphorylation. MEFs lacking Dvl1 and Dvl2 (Dvl1−/−;Dvl2−/−) (see Fig. S1 in the supplementary material) were infected with each of the four different lentiviral shRNAs against mouse Dvl3. After 3 days, the cells were treated with Wnt3a CM or control CM for 1 hour. Dvl3 protein level were drastically reduced by shRNA2, 3 or 4. Dvl3 protein exhibited typical Wnt-induced mobility shift (due to phosphorylation). shRNA2 and 4 were used in further experiments and yielded identical results. (B) The wild-type Dvl2 rescued LRP6 (endogenous) phosphorylation in Dvl3 knockdown Dvl1−/−;Dvl2−/− MEFs. Dvl1−/−;Dvl2−/− MEFs stably expressing Dvl2 (lanes 5 to 8) or the control vector (lanes 1 to 4) were pooled and infected with the lentiviral Dvl3 shRNA as indicated, then treated with Wnt3a or control CM for 1 hour as indicated. The exogenously expressed Dvl2 was detected by the Flag tag. (C) Dvl2ΔDEP, but neither Dvl2ΔDIX nor Dvl2ΔPDZ, rescued the Wnt3a-induced Lrp6 (endogenous) phosphorylation in Dvl3 knockdown Dvl1−/−;Dvl2−/− MEFs. Dvl1−/−;Dvl2−/− MEFs stably expressing the control vector (lanes 1–4), Dvl2ΔDIX (lanes 9–12), Dvl2ΔPDZ (lanes 5–8) or Dvl2ΔDEP (lanes 13–16) were infected with the lentiviral Dvl3 shRNA as indicated, and treated with Wnt3a CM or control CM for 1 hour as indicated. The expression of Dvl2 mutants was detected by the Flag tag.