September 1, 2009;
Bicaudal C, a novel regulator of Dvl signaling abutting RNA-processing bodies, controls cilia orientation and leftward flow.
Polycystic diseases and left
(LR) axis malformations are frequently linked to cilia
defects. Renal cysts also arise in mice and frogs lacking Bicaudal C (BicC
), a conserved RNA-binding protein containing K-homology (KH) domains and a sterile alpha motif (SAM). However, a role for BicC
function has not been demonstrated. Here, we report that targeted inactivation of BicC
(LR) asymmetry by disrupting the planar alignment of motile cilia
required for cilia
-driven fluid flow. Furthermore, depending on its SAM domain, BicC
can uncouple Dvl2
signaling from the canonical Wnt pathway, which has been implicated in antagonizing planar cell polarity (PCP). The SAM domain concentrates BicC
in cytoplasmic structures harboring RNA-processing bodies (P-bodies) and Dvl2
. These results suggest a model whereby BicC
links the orientation of cilia
with PCP, possibly by regulating RNA silencing in P-bodies.
[+] show captions
Fig. 3. Knockdown of xBicC alters laterality, cilia polarization and leftward flow in Xenopus laevis. (A-B′) Expression of xBicC in the gastrocoel roof plate (GRP; outlined by dotted lines in A and B), floor plate (arrow in A′,B′,B′) and epithelial lining of the circumblastoporal collar (cbc) at flow stages 16 (A) and 18/19 (B), as shown by whole-mount in situ hybridization analysis of dorsal explants (ventral views). Levels of sections in A′, A′, B′, B′ are indicated by dashed lines. ar, archenteron roof; n, notochord; so, somite; black arrowhead, xBicC-positive somitic cells. (C,C′) Neural tube closure defects in embryos injected with xBicC MO1/2 (C′) compared with Co-MO-injected specimen (C). Asterisks indicate closed (C) and open (C′) neural tubes. (D,D′) Delayed closure of the neural tube in a unilaterally injected embryo with xBicC MO1/2 and lineage tracer DsRed. Dorsal (D) and posterior (D′) view of superimposed fluorescence and bright-field images (midline indicated by dashed line, neural fold by yellow dotted line, floor plate by white dotted line). bp, blastopore. (E) Summary of marker gene expression patterns (Xnr1, top; Pitx2, bottom). Numbers indicate number of treated embryos. (F,J) Scanning electron microscopy analysis of GRP cells at stage 18 in Co-MO (F) and xBicC MO1/2 (J) injected embryos revealed predominantly posteriorly polarized cilia in control (F) and intermingled cilia orientation in xBicC morphant specimens. Scale bar: 10 μm. (G,K) Markedly reduced proportion of polarized cilia in xBicC MO1/2 (K) compared with Co-MO (G) injected embryos. The border of the GRP area relevant for flow is indicated by dashed lines. (H,I,L,M) Flow analysis in Co-MO (H,I) and xBicC MO1/2 (L,M) injected embryos. (H,L) Trajectories of beads displayed as GTTs of 25 second length (cf. color gradient bar in H). Red lines mark approximate limits of targeted areas, as visualized by co-injected DsRed; white dashed lines indicate border of the GRP. Note the presence of left (1), right (2) and circling (3) GTTs in L. (I,M) Frequency distributions of trajectory directionalities at the GRP of Co-MO (I) and xBicC MO1/2 (M) injected specimens. Solid circles represent the 50% boundary, dashed circles mark maximum frequency in histogram specified in percent. n, number of particles above threshold; ρ, quality of flow. Note the leftward direction of 60% of trajectories in (I) compared with the about equal distribution of trajectories in M, and the overall much reduced number of particles above the threshold of 2.5 μm/second in M. (N) Summary of flow analysis from 54 explants. Classification of embryos with flow into categories I-IV based onρ . Numbers indicate the number of analyzed explants.