XB-ART-38157Nat Genet. July 1, 2008; 40 (7): 871-9.
Dishevelled controls apical docking and planar polarization of basal bodies in ciliated epithelial cells.
The planar cell polarity (PCP) signaling system governs many aspects of polarized cell behavior. Here, we use an in vivo model of vertebrate mucociliary epithelial development to show that Dishevelled (Dvl) is essential for the apical positioning of basal bodies. We find that Dvl and Inturned mediate the activation of the Rho GTPase specifically at basal bodies, and that these three proteins together mediate the docking of basal bodies to the apical plasma membrane. Moreover, we find that this docking involves a Dvl-dependent association of basal bodies with membrane-bound vesicles and the vesicle-trafficking protein, Sec8. Once docked, basal bodies again require Dvl and Rho for the planar polarization that underlies directional beating of cilia. These results demonstrate previously undescribed functions for PCP signaling components and suggest that a common signaling apparatus governs both apical docking and planar polarization of basal bodies.
PubMed ID: 18552847
PMC ID: PMC2771675
Article link: Nat Genet.
Grant support: R01 GM074104-04 NIGMS NIH HHS , R01 GM074104-04S1 NIGMS NIH HHS , R01 GM076507-04 NIGMS NIH HHS , R01 GM074104 NIGMS NIH HHS , R01 GM076507 NIGMS NIH HHS , R01 GM074104-04S1 NIGMS NIH HHS , R01 GM074104 NIGMS NIH HHS , R01 GM076507 NIGMS NIH HHS , R01 GM074104-04 NIGMS NIH HHS , R01 GM076507-04 NIGMS NIH HHS
Genes referenced: cetn1 dvl1 dvl2 exoc4 intu rho rho.2 tjp1 tuba4b tubg1 tuba8
Antibodies referenced: Cetn1 Ab1 Dvl2 Ab2 Dvl3 Ab1 Tjp1 Ab1 Tuba4b Ab2 Tubg1 Ab3
Morpholinos referenced: dvl1 MO1 dvl2 MO1 dvl3 MO1 p2rx1 MO1
Article Images: [+] show captions
|Figure 1. Dvl is essential for ciliogenesis in multi-ciliated cells. (a) The epidermis of the Xenopus embryo is composed of multi-ciliated cells and mucus-secreting cells. Ciliated cells, labeled by alpha-tubulin immunostaining (red), are evenly spaced between mucus-secreting cells. Polarized beating of cilia generates directional flow across the epithelium16, 26. (b) Normal ciliated cell from a control-injected embryo (injected with a mismatched morpholino). (c) Defective cilia outgrowth in Dvl1–Dvl3 double morphants. (d) Wider field view of control (mismatch morpholino–injected) embryo. (e) Wider field view of Dvl1–Dvl3 double morphant embryo showing many affected cells. Image redisplayed with permission from Macmillan Publishers Ltd.|
|Figure 4 - Dvl localizes near the base of cilia in multi-ciliated cells. (a–c) Dvl2 immunostaining shows enrichment at cell membranes throughout the epidermis and enrichment at the apical surface of ciliated cells, as indicated by immunostaining for centrin (b,c). Scale bar, 15 mum. (d) Signal for Dvl2 immunostaining (red) localizes immediately adjacent to the basal body, as indicated by Centrin2-GFP (green). Scale bar, 1 mum. (e) Signal for Dvl2 immunostaining (red) localizes to the center of the region marked by CLAMP-GFP (green); data in Supplementary Figure 4 indicate that CLAMP-GFP labels the ciliary rootlet. Scale bar, 1 mum. (f) The C terminus of Dvl2 is sufficient to drive localization to basal body in multi-ciliated cells. Stage 27 embryos are shown throughout. Scale bar, 5 um. Image redisplayed with permission from Macmillan Publishers Ltd.|
|Figure 8 - Dvl and Rho govern directional fluid flow across the ciliated epidermis. (a) Control embryos were grown until stage 26, and latex beads were added to the media to visualize fluid flow over the epidermis. The movement of beads was recorded and representative beads were tracked. Representative bead movements were tracked from the horizontal myoseptum, as indicated by the colored lines. Beads move from dorso-anterior to ventral-posterior uniformly in control embryos. (b) DN-RhoA expression severely disrupts directional flow; beads were turning and swirling frequently. (c) Xdd1 expression severely disrupts directional flow; beads were turning and swirling frequently. (d) The directionality of beads was quantified as the ratio of the linear distance between the first and last point of a track and the total distance along the track. A straight line would be a value of 1.00. Error bars, s.d. (e) DN-RhoA or Xdd1 reduces the rate of bead movement across the epidermis. Defects in polarized flow were evident in that even beads moving in comparatively straight paths moved at a greatly reduced rate across experimental embryos. Error bars, s.d. (f) Still images from a high-speed time-lapse movie of beating cilia labeled with tau-GFP. (g) Still images from a high-speed time-lapse movie of beating cilia expressing Xdd1; frequency is comparable to controls. Image redisplayed with permission from Macmillan Publishers Ltd.|