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Ciliogenesis defects in embryos lacking inturned or fuzzy function are associated with failure of planar cell polarity and Hedgehog signaling

Abstract

The vertebrate planar cell polarity (PCP) pathway has previously been found to control polarized cell behaviors rather than cell fate. We report here that disruption of Xenopus laevis orthologs of the Drosophila melanogaster PCP effectors inturned (in) or fuzzy (fy) affected not only PCP-dependent convergent extension but also elicited embryonic phenotypes consistent with defective Hedgehog signaling. These defects in Hedgehog signaling resulted from a broad requirement for Inturned and Fuzzy in ciliogenesis. We show that these proteins govern apical actin assembly and thus control the orientation, but not assembly, of ciliary microtubules. Finally, accumulation of Dishevelled and Inturned near the basal apparatus of cilia suggests that these proteins function in a common pathway with core PCP components to regulate ciliogenesis. Together, these data highlight the interrelationships between cell polarity, cellular morphogenesis, signal transduction and cell fate specification.

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Figure 1: Phenotypes of Xint morphant embryos reflect a loss of Hedgehog signaling.
Figure 2: Loss of Hedgehog target gene expression in Xint morphants.
Figure 3: Morpholino knockdown of Xfy phenocopies that of Xint.
Figure 4: Convergent extension defects in Xint morphants.
Figure 5: Xint and Xfy regulate convergent extension and interact with the PCP cascade.
Figure 6: Xint is required for ciliogenesis.
Figure 7: Xint controls cytoskeletal organization during ciliogenesis.
Figure 8: Inturned and Dishevelled accumulate at the apical surface of ciliated epidermal cells.

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References

  1. Wallingford, J.B. Neural tube closure and neural tube defects: Studies in animal models reveal known knowns and known unknowns. Am. J. Med. Genet. C Semin. Med. Genet. 135, 59–68 (2005).

    Article  Google Scholar 

  2. Mlodzik, M. Planar cell polarization: do the same mechanisms regulate Drosophila tissue polarity and vertebrate gastrulation? Trends Genet. 18, 564–571 (2002).

    Article  CAS  Google Scholar 

  3. Adler, P.N. & Lee, H. Frizzled signaling and cell-cell interactions in planar polarity. Curr. Opin. Cell Biol. 13, 635–640 (2001).

    Article  CAS  Google Scholar 

  4. Feiguin, F., Hannus, M., Mlodzik, M. & Eaton, S. The ankyrin repeat protein Diego mediates Frizzled-dependent planar polarization. Dev. Cell 1, 93–101 (2001).

    Article  CAS  Google Scholar 

  5. Jenny, A., Reynolds-Kenneally, J., Das, G., Burnett, M. & Mlodzik, M. Diego and Prickle regulate Frizzled planar cell polarity signalling by competing for Dishevelled binding. Nat. Cell Biol. 7, 691–697 (2005).

    Article  CAS  Google Scholar 

  6. Simons, M. et al. Inversin, the gene product mutated in nephronophthisis type II, functions as a molecular switch between Wnt signaling pathways. Nat. Genet. 37, 537–543 (2005).

    Article  CAS  Google Scholar 

  7. Watanabe, D. et al. The left-right determinant Inversin is a component of node monocilia and other 9+0 cilia. Development 130, 1725–1734 (2003).

    Article  CAS  Google Scholar 

  8. Ross, A.J. et al. Disruption of Bardet-Biedl syndrome ciliary proteins perturbs planar cell polarity in vertebrates. Nat. Genet. 37, 1135–1140 (2005).

    Article  CAS  Google Scholar 

  9. Corbit, K.C. et al. Vertebrate Smoothened functions at the primary cilium. Nature 437, 1018–1021 (2005).

    Article  CAS  Google Scholar 

  10. Huangfu, D. et al. Hedgehog signalling in the mouse requires intraflagellar transport proteins. Nature 426, 83–87 (2003).

    Article  CAS  Google Scholar 

  11. Liu, A., Wang, B. & Niswander, L.A. Mouse intraflagellar transport proteins regulate both the activator and repressor functions of Gli transcription factors. Development 132, 3103–3111 (2005).

    Article  CAS  Google Scholar 

  12. Huangfu, D. & Anderson, K.V. Cilia and Hedgehog responsiveness in the mouse. Proc. Natl. Acad. Sci. USA 102, 11325–11330 (2005).

    Article  CAS  Google Scholar 

  13. Curtin, J.A. et al. Mutation of Celsr1 disrupts planar polarity of inner ear hair cells and causes severe neural tube defects in the mouse. Curr. Biol. 13, 1129–1133 (2003).

    Article  CAS  Google Scholar 

  14. Han, Y.G., Kwok, B.H. & Kernan, M.J. Intraflagellar transport is required in Drosophila to differentiate sensory cilia but not sperm. Curr. Biol. 13, 1679–1686 (2003).

    Article  CAS  Google Scholar 

  15. Collier, S. & Gubb, D. Drosophila tissue polarity requires the cell-autonomous activity of the fuzzy gene, which encodes a novel transmembrane protein. Development 124, 4029–4037 (1997).

    CAS  PubMed  Google Scholar 

  16. Park, W.J., Liu, J., Sharp, E.J. & Adler, P.N. The Drosophila tissue polarity gene inturned acts cell autonomously and encodes a novel protein. Development 122, 961–969 (1996).

    CAS  PubMed  Google Scholar 

  17. Lee, H. & Adler, P.N. The function of the frizzled pathway in the Drosophila wing is dependent on inturned and fuzzy. Genetics 160, 1535–1547 (2002).

    CAS  PubMed  PubMed Central  Google Scholar 

  18. Draper, B.W., Morcos, P.A. & Kimmel, C.B. Inhibition of zebrafish fgf8 pre-mRNA splicing with morpholino oligos: a quantifiable method for gene knockdown. Genesis 30, 154–156 (2001).

    Article  CAS  Google Scholar 

  19. Hu, D. & Helms, J.A. The role of sonic hedgehog in normal and abnormal craniofacial morphogenesis. Development 126, 4873–4884 (1999).

    CAS  PubMed  Google Scholar 

  20. Chiang, C. et al. Cyclopia and defective axial patterning in mice lacking Sonic hedgehog gene function. Nature 383, 407–413 (1996).

    Article  CAS  Google Scholar 

  21. Ekker, S.C. et al. Distinct expression and shared activities of members of the hedgehog gene family of Xenopus laevis. Development 121, 2337–2347 (1995).

    CAS  PubMed  Google Scholar 

  22. Hallonet, M., Hollemann, T., Pieler, T. & Gruss, P. Vax1, a novel homeobox-containing gene, directs development of the basal forebrain and visual system. Genes Dev. 13, 3106–3114 (1999).

    Article  CAS  Google Scholar 

  23. Wada, N. et al. Hedgehog signaling is required for cranial neural crest morphogenesis and chondrogenesis at the midline in the zebrafish skull. Development 132, 3977–3988 (2005).

    Article  CAS  Google Scholar 

  24. Jeong, J., Mao, J., Tenzen, T., Kottmann, A.H. & McMahon, A.P. Hedgehog signaling in the neural crest cells regulates the patterning and growth of facial primordia. Genes Dev. 18, 937–951 (2004).

    Article  CAS  Google Scholar 

  25. Trueb, L. & Hanken, J. Skeletal development in Xenopus laevis (Anura: Pipidae). J. Morphol. 214, 1–41 (1992).

    Article  CAS  Google Scholar 

  26. Lopez, S.L. et al. Notch activates sonic hedgehog and both are involved in the specification of dorsal midline cell-fates in Xenopus. Development 130, 2225–2238 (2003).

    Article  CAS  Google Scholar 

  27. Pichon, B., Taelman, V., Kricha, S., Christophe, D. & Bellefroid, E.J. XHRT-1, a hairy and Enhancer of split related gene with expression in floor plate and hypochord during early Xenopus embryogenesis. Dev. Genes Evol. 212, 491–495 (2002).

    Article  CAS  Google Scholar 

  28. Take-uchi, M., Clarke, J.D. & Wilson, S.W. Hedgehog signalling maintains the optic stalk-retinal interface through the regulation of Vax gene activity. Development 130, 955–968 (2003).

    Article  CAS  Google Scholar 

  29. Wallingford, J.B. & Harland, R.M. Xenopus dishevelled signaling regulates both neural and mesodermal convergent extension: parallel forces elongating the body axis. Development 128, 2581–2592 (2001).

    CAS  PubMed  Google Scholar 

  30. Wallingford, J.B. & Harland, R.M. Neural tube closure requires Dishevelled-dependent convergent extension of the midline. Development 129, 5815–5825 (2002).

    Article  CAS  Google Scholar 

  31. Keller, R., Shih, J. & Sater, A. The cellular basis of the convergence and extension of the Xenopus neural plate. Dev. Dyn. 193, 199–217 (1992).

    Article  CAS  Google Scholar 

  32. Keller, R. et al. Mechanisms of convergence and extension by cell intercalation. Phil. Trans. R. Soc. Lond. B 355, 897–922 (2000).

    Article  CAS  Google Scholar 

  33. Kimmel, C.B. et al. The shaping of pharyngeal cartilages during early development of the zebrafish. Dev. Biol. 203, 245–263 (1998).

    Article  CAS  Google Scholar 

  34. Topczewski, J. et al. The zebrafish glypican knypek controls cell polarity during gastrulation movements of convergent extension. Dev. Cell 1, 251–264 (2001).

    Article  CAS  Google Scholar 

  35. Nishikawa, S., Hirata, J. & Sasaki, F. Fate of ciliated epidermal cells during early development of Xenopus laevis using whole-mount immunostaining with an antibody against chondroitin 6-sulfate proteoglycan and anti-tubulin: transdifferentiation or metaplasia of amphibian epidermis. Histochemistry 98, 355–358 (1992).

    Article  CAS  Google Scholar 

  36. Assheton, R. Notes on the ciliation of the ectoderm of the amphibian embryo. Q. J. Microsc. Sci. 38, 465–484 (1896).

    Google Scholar 

  37. Kim, J.C. et al. The Bardet-Biedl protein BBS4 targets cargo to the pericentriolar region and is required for microtubule anchoring and cell cycle progression. Nat. Genet. 36, 462–470 (2004).

    Article  CAS  Google Scholar 

  38. Kulaga, H.M. et al. Loss of BBS proteins causes anosmia in humans and defects in olfactory cilia structure and function in the mouse. Nat. Genet. 36, 994–998 (2004).

    Article  CAS  Google Scholar 

  39. Chailley, B., Nicolas, G. & Laine, M.C. Organization of actin microfilaments in the apical border of oviduct ciliated cells. Biol. Cell. 67, 81–90 (1989).

    Article  CAS  Google Scholar 

  40. Boisvieux-Ulrich, E., Laine, M.C. & Sandoz, D. Cytochalasin D inhibits basal body migration and ciliary elongation in quail oviduct epithelium. Cell Tissue Res. 259, 443–454 (1990).

    Article  CAS  Google Scholar 

  41. Boisvieux-Ulrich, E. & Sandoz, D. Determination of ciliary polarity precedes differentiation in the epithelial cells of quail oviduct. Biol. Cell. 72, 3–14 (1991).

    Article  CAS  Google Scholar 

  42. Tamm, S. & Tamm, S.L. Development of macrociliary cells in Beroë. J. Cell Sci. 89, 67–80 (1988).

    PubMed  Google Scholar 

  43. Adler, P.N., Zhu, C. & Stone, D. Inturned localizes to the proximal side of wing cells under the instruction of upstream planar polarity proteins. Curr. Biol. 14, 2046–2051 (2004).

    Article  CAS  Google Scholar 

  44. Hagiwara, H., Ohwada, N. & Takata, K. Cell biology of normal and abnormal ciliogenesis in the ciliated epithelium. Int. Rev. Cytol. 234, 101–141 (2004).

    Article  Google Scholar 

  45. Eaton, S., Wepf, R. & Simons, K. Roles for Rac1 and Cdc42 in planar polarization and hair outgrowth in the wing of Drosophila. J. Cell Biol. 135, 1277–1289 (1996).

    Article  CAS  Google Scholar 

  46. Murdoch, J.N., Doudney, K., Paternotte, C., Copp, A.J. & Stanier, P. Severe neural tube defects in the loop-tail mouse result from mutation of Lpp1, a novel gene involved in floor plate specification. Hum. Mol. Genet. 10, 2593–2601 (2001).

    Article  CAS  Google Scholar 

  47. Konig, G. & Hausen, P. Planar polarity in the ciliated epidermis of Xenopus embryos. Dev. Biol. 160, 355–368 (1993).

    Article  CAS  Google Scholar 

  48. Kramer-Zucker, A.G. et al. Cilia-driven fluid flow in the zebrafish pronephros, brain and Kupffer's vesicle is required for normal organogenesis. Development 132, 1907–1921 (2005).

    Article  CAS  Google Scholar 

  49. Otto, E.A. et al. Mutations in INVS encoding inversin cause nephronophthisis type 2, linking renal cystic disease to the function of primary cilia and left-right axis determination. Nat. Genet. 34, 413–420 (2003).

    Article  CAS  Google Scholar 

  50. Pan, J., Wang, Q. & Snell, W.J. Cilium-generated signaling and cilia-related disorders. Lab. Invest. 85, 452–463 (2005).

    Article  CAS  Google Scholar 

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Acknowledgements

The authors thank J. Reiter and R. Harland for critical discussions during this work. We also thank J. Hayes, S. Peyrot, A. Ellis for technical assistance; J. Sisson and J. Gross for critical reading; T. Holleman for the xVax1 plasmid; H. El-Hodiri for the xZic3 plasmid. J.B.W. dedicates this paper to J. Cruz. This work is supported by awards from the National Institute of General Medical Sciences/US National Institutes of Health and the Burroughs Wellcome Fund.

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Correspondence to John B Wallingford.

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Supplementary information

Supplementary Fig. 1

Sequence and expression pattern of Xint. (PDF 1450 kb)

Supplementary Fig. 2

Additional Xint morphant phenotypes. (PDF 2199 kb)

Supplementary Fig. 3

Sequence and expression pattern of Xenopus fuzzy. (PDF 1991 kb)

Supplementary Fig. 4

Quantification of convergent extennsion in Xint and Xfy morphants. (PDF 71 kb)

Supplementary Fig. 5

Xfy controls cytoskeletal organization during ciliogenesis. (PDF 890 kb)

Supplementary Table 1

Sequences of primer and morpholino-oligonucleotides used in this study. (PDF 23 kb)

Supplementary Video 1

Ciliated epidermal cell in control embryo. (MOV 409 kb)

Supplementary Video 2

Ciliated epidermal cell in Xint morphant. (MOV 321 kb)

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Park, T., Haigo, S. & Wallingford, J. Ciliogenesis defects in embryos lacking inturned or fuzzy function are associated with failure of planar cell polarity and Hedgehog signaling. Nat Genet 38, 303–311 (2006). https://doi.org/10.1038/ng1753

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