Figure 7.

Drg1 modulation of Daam1 activity is critical for multiciliation. (A) Drg1 knockdown reduces the Dvl2–Daam1 interaction. The cocktail of the indicated mRNAs and MOs was injected into embryos, and co-IPs were conducted with embryos harvested at stage 20. MO-resistant Drg1-V5 expression rescues decreased the Dvl2–Daam1 interaction upon Drg1 knockdown. (B) Drg1 knockdown decreases the Daam1–RhoA interaction. The indicated mRNAs and morpholinos were coinjected into embryos, which were collected at stage 20 for co-IP analysis. (C) C-Daam1 expression rescues reduced acetylated tubulin staining upon Drg1 knockdown. The indicated mRNAs and MOs were injected to two ventral blastomeres at the four-cell stage. For C-Daam1 expression, 2.5 pg of HA-tagged C-Daam1 encoding plasmid DNAs were injected. The embryos fixed at stage 27 were used for immunostaining. Expression levels of Daam1-WT and C-Daam1 were tested by Western blot. Relative acetylated tubulin intensity is quantified (image n = 40 from 15 embryos for each condition). ****, P < 0.0001, one-way ANOVA. Scale bars, 50 µm. (D) C-Daam1 expression restores the formation of the apical actin meshwork that was decreased upon Drg1 knockdown. The indicated mRNAs and MOs were coinjected into two ventral blastomeres of four-cell-stage embryos and fixed at stage 27 for phalloidin and acetylated tubulin staining. ****, P < 0.0001, one-way ANOVA; n = 40. Scale bars, 5 µm. Error bars indicate ± SD. n.s., not significant.