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CEP162 is an axoneme-recognition protein promoting ciliary transition zone assembly at the cilia base

Nature Cell Biology volume 15, pages 591–601 (2013)Cite this article

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Abstract

The transition zone is a specialized compartment found at the base of cilia, adjacent to the centriole distal end, where axonemal microtubules are heavily crosslinked to the surrounding membrane to form a barrier that gates the ciliary compartment. A number of ciliopathy molecules have been found to associate with the transition zone, but factors that directly recognize axonemal microtubules to specify transition zone assembly at the cilia base remain unclear. Here, through quantitative centrosome proteomics, we identify an axoneme-associated protein, CEP162 (KIAA1009), tethered specifically at centriole distal ends to promote transition zone assembly. CEP162 interacts with core transition zone components, and mediates their association with microtubules. Loss of CEP162 arrests ciliogenesis at the stage of transition zone assembly. Abolishing its centriolar tethering, however, allows CEP162 to stay on the growing end of the axoneme and ectopically assemble transition zone components at cilia tips. This generates extra-long cilia with strikingly swollen tips that actively release ciliary contents into the extracellular environment. CEP162 is thus an axoneme-recognition protein pre-tethered at centriole distal ends before ciliogenesis to promote and restrict transition zone formation specifically at the cilia base.

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Figure 1: Identification of CEP162 as a microtubule-binding protein localized at centriole distal ends.
Figure 2: CEP162 is required for ciliogenesis.
Figure 3: Loss of CEP162 blocks ciliogenesis at the stage of transition zone assembly.
Figure 4: CEP162 is tethered at centriole distal ends to recognize axonemes.
Figure 5: CEP162 interacts with CEP290 and mediates its association with microtubules.
Figure 6: Untethered CEP162 promotes ectopic recruitment of transition zone components to cilia tips.
Figure 7: Cilia tips modified by CEP162 swell exceedingly and discharge ciliary contents.

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References

  1. Sorokin, S. Centrioles and the formation of rudimentary cilia by fibroblasts and smooth muscle cells. J. Cell Biol. 15, 363–377 (1962).

    Article  CAS  Google Scholar 

  2. Sorokin, S. P. Reconstructions of centriole formation and ciliogenesis in mammalian lungs. J. Cell Sci. 3, 207–230 (1968).

    CAS  PubMed  Google Scholar 

  3. Eggenschwiler, J. T. & Anderson, K. V. Cilia and developmental signaling. Annu. Rev. Cell Dev. Biol. 23, 345–373 (2007).

    Article  CAS  Google Scholar 

  4. Rohatgi, R. & Snell, W. J. The ciliary membrane. Curr. Opin. Cell Biol. 22, 541–546 (2010).

    Article  CAS  Google Scholar 

  5. Schmidt, K. N. et al. Cep164 mediates vesicular docking to the mother centriole during early steps of ciliogenesis. J. Cell Biol. 199, 1083–1101 (2012).

    Article  CAS  Google Scholar 

  6. Tanos, B. E. et al. Centriole distal appendages promote membrane docking, leading to cilia initiation. Gen. Dev. 27, 163–168 (2013).

    Article  CAS  Google Scholar 

  7. Gilula, N. B. & Satir, P. The ciliary necklace. A ciliary membrane specialization. J. Cell Biol. 53, 494–509 (1972).

    Article  CAS  Google Scholar 

  8. Ishikawa, H. & Marshall, W. F. Ciliogenesis: building the cell’s antenna. Nat. Rev. Mol. Cell Biol. 12, 222–234 (2011).

    Article  CAS  Google Scholar 

  9. Pedersen, L. B. & Rosenbaum, J. L. Intraflagellar transport (IFT) role in ciliary assembly, resorption and signalling. Curr. Top. Dev. Biol. 85, 23–61 (2008).

    Article  CAS  Google Scholar 

  10. Hu, Q. et al. A septin diffusion barrier at the base of the primary cilium maintains ciliary membrane protein distribution. Science 329, 436–439 (2010).

    Article  CAS  Google Scholar 

  11. Kee, H. L. et al. A size-exclusion permeability barrier and nucleoporins characterize a ciliary pore complex that regulates transport into cilia. Nat. Cell Biol. 14, 431–437 (2012).

    Article  CAS  Google Scholar 

  12. Anderson, R. G. The three-dimensional structure of the basal body from the rhesus monkey oviduct. J. Cell Biol. 54, 246–265 (1972).

    Article  CAS  Google Scholar 

  13. Chih, B. et al. A ciliopathy complex at the transition zone protects the cilia as a privileged membrane domain. Nat. Cell Biol. 14, 61–72 (2012).

    Article  CAS  Google Scholar 

  14. Reiter, J. F., Blacque, O. E. & Leroux, M. R. The base of the cilium: roles for transition fibres and the transition zone in ciliary formation, maintenance and compartmentalization. EMBO Rep. 13, 608–618 (2012).

    Article  CAS  Google Scholar 

  15. Sang, L. et al. Mapping the NPHP-JBTS-MKS protein network reveals ciliopathy disease genes and pathways. Cell 145, 513–528 (2011).

    Article  CAS  Google Scholar 

  16. Williams, C. L. et al. MKS and NPHP modules cooperate to establish basal body/transition zone membrane associations and ciliary gate function during ciliogenesis. J. Cell Biol. 192, 1023–1041 (2011).

    Article  CAS  Google Scholar 

  17. Garcia-Gonzalo, F. R. et al. A transition zone complex regulates mammalian ciliogenesis and ciliary membrane composition. Nat. Gene. 43, 776–784 (2011).

    Article  CAS  Google Scholar 

  18. Winkelbauer, M. E., Schafer, J. C., Haycraft, C. J., Swoboda, P. & Yoder, B. K. The C. elegans homologs of nephrocystin-1 and nephrocystin-4 are cilia transition zone proteins involved in chemosensory perception. J. Cell Sci. 118, 5575–5587 (2005).

    Article  CAS  Google Scholar 

  19. Czarnecki, P. G. & Shah, J. V. The ciliary transition zone: from morphology and molecules to medicine. Trends Cell Biol. 22, 201–210 (2012).

    Article  CAS  Google Scholar 

  20. Craige, B. et al. CEP290 tethers flagellar transition zone microtubules to the membrane and regulates flagellar protein content. J. Cell Biol. 190, 927–940 (2010).

    Article  CAS  Google Scholar 

  21. Wang, W. J., Soni, R. K., Uryu, K. & Tsou, M. F. The conversion of centrioles to centrosomes: essential coupling of duplication with segregation. J. Cell Biol. 193, 727–739 (2011).

    Article  CAS  Google Scholar 

  22. Ong, S. E. et al. Stable isotope labeling by amino acids in cell culture, SILAC, as a simple and accurate approach to expression proteomics. Mol. Cell. Proteom. 1, 376–386 (2002).

    Article  CAS  Google Scholar 

  23. Spektor, A., Tsang, W. Y., Khoo, D. & Dynlacht, B. D. Cep97 and CP110 suppress a cilia assembly program. Cell 130, 678–690 (2007).

    Article  CAS  Google Scholar 

  24. Singla, V., Romaguera-Ros, M., Garcia-Verdugo, J. M. & Reiter, J.F. Ofd1, a human disease gene, regulates the length and distal structure of centrioles. Dev. Cell 18, 410–424 (2010).

    Article  CAS  Google Scholar 

  25. Tsang, W. Y. et al. CP110 suppresses primary cilia formation through its interaction with CEP290, a protein deficient in human ciliary disease. Dev. Cell 15, 187–197 (2008).

    Article  CAS  Google Scholar 

  26. Otto, E. A. et al. Candidate exome capture identifies mutation of SDCCAG8 as the cause of a retinal-renal ciliopathy. Nat. Genet. 42, 840–850 (2010).

    Article  CAS  Google Scholar 

  27. Tsang, W. Y. et al. Cep76, a centrosomal protein that specifically restrains centriole reduplication. Dev. Cell 16, 649–660 (2009).

    Article  CAS  Google Scholar 

  28. Jakobsen, L. et al. Novel asymmetrically localizing components of human centrosomes identified by complementary proteomics methods. EMBO J. 30, 1520–1535 (2011).

    Article  CAS  Google Scholar 

  29. Leon, A., Omri, B., Gely, A., Klein, C. & Crisanti, P. QN1/KIAA1009: a new essential protein for chromosome segregation and mitotic spindle assembly. Oncogene 25, 1887–1895 (2006).

    Article  CAS  Google Scholar 

  30. Fry, A. M. et al. C-Nap1, a novel centrosomal coiled-coil protein and candidate substrate of the cell cycle-regulated protein kinase Nek2. J. Cell Biol. 141, 1563–1574 (1998).

    Article  CAS  Google Scholar 

  31. Graser, S. et al. Cep164, a novel centriole appendage protein required for primary cilium formation. J. Cell Biol. 179, 321–330 (2007).

    Article  CAS  Google Scholar 

  32. Snell, W. J. Mating in Chlamydomonas: a system for the study of specific cell adhesion. I. Ultrastructural and electrophoretic analyses of flagellar surface components involved in adhesion. J. Cell Biol. 68, 48–69 (1976).

    Article  CAS  Google Scholar 

  33. Baldari, C. T. & Rosenbaum, J. Intraflagellar transport: it’s not just for cilia anymore. Curr. Opin. Cell Biol. 22, 75–80 (2010).

    Article  CAS  Google Scholar 

  34. Habedanck, R., Stierhof, Y. D., Wilkinson, C. J. & Nigg, E. A. The Polo kinase Plk4 functions in centriole duplication. Nat. Cell Biol. 7, 1140–1146 (2005).

    Article  CAS  Google Scholar 

  35. Tsou, M. F. & Stearns, T. Mechanism limiting centrosome duplication to once per cell cycle. Nature 442, 947–951 (2006).

    Article  CAS  Google Scholar 

  36. Valente, E. M. et al. Mutations in CEP290, which encodes a centrosomalprotein, cause pleiotropic forms of Joubert syndrome. Nat. Genet. 38, 623–625 (2006).

    Article  CAS  Google Scholar 

  37. Sakaguchi, T., Kikuchi, Y., Kuroiwa, A., Takeda, H. & Stainier, D. Y. The yolk syncytial layer regulates myocardial migration by influencing extracellular matrix assembly in zebrafish. Development 133, 4063–4072 (2006).

    Article  CAS  Google Scholar 

  38. Huang, C. J., Tu, C. T., Hsiao, C. D., Hsieh, F. J. & Tsai, H. J. Germ-line transmission of a myocardium-specific GFP transgene reveals critical regulatory elementsin the cardiac myosin light chain 2 promoter of zebrafish. Dev. Dyn. 228, 30–40 (2003).

    Article  CAS  Google Scholar 

  39. Kimmel, C. B., Ballard, W. W., Kimmel, S. R., Ullmann, B. & Schilling, T. F. Stages of embryonic development of the zebrafish. Dev. Dyn. 203, 253–310 (1995).

    Article  CAS  Google Scholar 

  40. Long, S., Ahmad, N. & Rebagliati, M. The zebrafish nodal-related gene southpaw is required for visceral and diencephalic left–right asymmetry. Development 130, 2303–2316 (2003).

    Article  CAS  Google Scholar 

  41. Gao, C., Wang, G., Amack, J. D. & Mitchell, D. R. Oda16/Wdr69 is essential for axonemal dynein assembly and ciliary motility during zebrafish embryogenesis. Dev. Dyn. 239, 2190–2197 (2010).

    Article  CAS  Google Scholar 

  42. Robu, M. E. et al. p53 activation by knockdown technologies. PLoS Gene. 3, e78 (2007).

    Article  Google Scholar 

Download references

Acknowledgements

We are grateful to K. Anderson and her laboratory at Memorial Sloan-Kettering Cancer Center, USA, for reagents and antibodies. We thank L. Gunther, G. Perumal and F. Macaluso at the Analytical Imaging Center of Albert Einstein College of Medicine for assistance with transmission and scanning electron microscopy; K. Uryu at Rockefeller university and N. Lampen at MSKCC for assisting with the usage of electron microscopes; F. Foley at SUNY Upstate and D. Gutierrez at MSKCC for technical assistance and zebrafish management; and A. Hall, Z. Bao and C. Haynes at MSKCC for comments on the manuscript. This work was supported by the National Institutes of Health grants HL095690 to J.D.A. and GM088253 to M-F.B.T.

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Authors and Affiliations

  1. Cell Biology Program, Memorial Sloan-Kettering Cancer Center, New York, New York 10065, USA

    Won-Jing Wang, Rajesh Soni & Meng-Fu Bryan Tsou

  2. Department of Cell and Developmental Biology, State University of New York, Upstate Medical University, Syracuse, New York 13210, USA

    Hwee Goon Tay & Jeffrey D. Amack

  3. Analytical Imaging Facility, Albert Einstein College of Medicine, Bronx, New York 10461, USA

    Geoffrey S. Perumal & Frank P. Macaluso

  4. Developmental Biology Program, Memorial Sloan-Kettering Cancer Center, New York, New York 10065, USA

    Mary G. Goll

  5. Department of Medicine, Harvard Medical School, Boston, Massachusetts 02115, USA

    John M. Asara

  6. Mass Spectrometry Core, Beth Israel Deaconess Medical Center, Boston, Massachusetts 02115, USA

    John M. Asara

  7. Weill Cornell Graduate School of Medical Sciences, Cornell University, New York, New York 10065, USA

    Meng-Fu Bryan Tsou

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Contributions

W-J.W. and M-F.B.T. designed experiments and analysed data. W-J.W. performed most of the experiments. W-J.W., R.S. and J.M.A. did the quantitative centrosome proteomics. W-J.W. and R.S. prepared purified centrosomes. J.M.A. carried out SILAC mass spectrometry and analysed the data. The zebrafish works were carried out by H.G.T. and J.D.M. M.G.G. helped with initial zebrafish experiments. G.S.P., F.P.M. and W-J.W. performed the correlative light and scanning electron microscopy. M-F.B.T. and W-J.W. wrote the manuscript.

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Correspondence to Meng-Fu Bryan Tsou.

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Ciliary contents are actively released from cilia tips modified with CEP162.

RPE1 cells expressing GFP-tagged CEP162tN1C1C2 were serum-starved for 24 h, and imaged by time-lapse fluorescence microscopy (2 min time interval). Images shown in movies were generated from maximum intensity projections of a z-stack. (MOV 697 kb)

Bursts of cilia tips modified by CEP162.

Images were collected and processed from RPE1 cells expressing GFP-tagged CEP162tN1C1C2 as described in Supplementary Video S1. (MOV 578 kb)

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Wang, WJ., Tay, H., Soni, R. et al. CEP162 is an axoneme-recognition protein promoting ciliary transition zone assembly at the cilia base. Nat Cell Biol 15, 591–601 (2013). https://doi.org/10.1038/ncb2739

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