The dynein regulatory complex is required for ciliary motility and otolith biogenesis in the inner ear

Article metrics


In teleosts, proper balance and hearing depend on mechanical sensors in the inner ear. These sensors include actin-based microvilli and microtubule-based cilia that extend from the surface of sensory hair cells and attach to biomineralized ‘ear stones’ (or otoliths)1. Otolith number, size and placement are under strict developmental control, but the mechanisms that ensure otolith assembly atop specific cells of the sensory epithelium are unclear. Here we demonstrate that cilia motility is required for normal otolith assembly and localization. Using in vivo video microscopy, we show that motile tether cilia at opposite poles of the otic vesicle create fluid vortices that attract otolith precursor particles, thereby biasing an otherwise random distribution to direct localized otolith seeding on tether cilia. Independent knockdown of subunits for the dynein regulatory complex and outer-arm dynein disrupt cilia motility, leading to defective otolith biogenesis. These results demonstrate a requirement for the dynein regulatory complex in vertebrates and show that cilia-driven flow is a key epigenetic factor in controlling otolith biomineralization.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Figure 1: gas8 is expressed in ciliated tissues.
Figure 2: gas8 morphants exhibit developmental defects.
Figure 3: Gas8 is required for tether cilia motility.
Figure 4: Tether cilia motility drives otolith biogenesis.


  1. 1

    Sollner, C. & Nicolson, T. in Biomineralization: From Biology to Biotechnology and Medical Application 2nd edn (ed. Bauerlein, E.) 229–242 (Wiley-VCH, 2004)

  2. 2

    Satir, P. & Christensen, S. T. Overview of structure and function of mammalian cilia. Annu. Rev. Physiol. 69, 377–400 (2007)

  3. 3

    Bisgrove, B. W. & Yost, H. J. The roles of cilia in developmental disorders and disease. Development 133, 4131–4143 (2006)

  4. 4

    Badano, J. L., Mitsuma, N., Beales, P. L. & Katsanis, N. The ciliopathies: an emerging class of human genetic disorders. Annu. Rev. Genomics Hum. Genet. 7, 125–148 (2006)

  5. 5

    Fliegauf, M., Benzing, T. & Omran, H. When cilia go bad: cilia defects and ciliopathies. Nature Rev. Mol. Cell Biol. 8, 880–893 (2007)

  6. 6

    Riley, B. B., Zhu, C., Janetopoulos, C. & Aufderheide, K. J. A critical period of ear development controlled by distinct populations of ciliated cells in the zebrafish. Dev. Biol. 191, 191–201 (1997)

  7. 7

    Haddon, C. & Lewis, J. Early ear development in the embryo of the zebrafish, Danio rerio . J. Comp. Neurol. 365, 113–128 (1996)

  8. 8

    Hughes, I., Thalmann, I., Thalmann, R. & Ornitz, D. M. Mixing model systems: using zebrafish and mouse inner ear mutants and other organ systems to unravel the mystery of otoconial development. Brain Res. 1091, 58–74 (2006)

  9. 9

    Nicolson, T. The genetics of hearing and balance in zebrafish. Annu. Rev. Genet. 39, 9–22 (2005)

  10. 10

    Huang, B., Ramanis, Z. & Luck, D. J. Suppressor mutations in Chlamydomonas reveal a regulatory mechanism for Flagellar function. Cell 28, 115–124 (1982)

  11. 11

    Piperno, G., Mead, K. & Shestak, W. The inner dynein arms I2 interact with a “dynein regulatory complex” in Chlamydomonas flagella. J. Cell Biol. 118, 1455–1463 (1992)

  12. 12

    Gardner, L. C., O’Toole, E., Perrone, C. A., Giddings, T. & Porter, M. E. Components of a “dynein regulatory complex” are located at the junction between the radial spokes and the dynein arms in Chlamydomonas flagella. J. Cell Biol. 127, 1311–1325 (1994)

  13. 13

    Ralston, K. S., Lerner, A. G., Diener, D. R. & Hill, K. L. Flagellar motility contributes to cytokinesis in Trypanosoma brucei and is modulated by an evolutionarily conserved dynein regulatory system. Eukaryot. Cell 5, 696–711 (2006)

  14. 14

    Hutchings, N. R., Donelson, J. E. & Hill, K. L. Trypanin is a cytoskeletal linker protein and is required for cell motility in African trypanosomes. J. Cell Biol. 156, 867–877 (2002)

  15. 15

    Rupp, G. & Porter, M. E. A subunit of the dynein regulatory complex in Chlamydomonas is a homologue of a growth arrest-specific gene product. J. Cell Biol. 162, 47–57 (2003)

  16. 16

    Baron, D. M., Ralston, K. S., Kabututu, Z. P. & Hill, K. L. Functional genomics in Trypanosoma brucei identifies evolutionarily conserved components of motile flagella. J. Cell Sci. 120, 478–491 (2007)

  17. 17

    Hill, K. L., Hutchings, N. R., Grandgenett, P. M. & Donelson, J. E. T lymphocyte triggering factor of African trypanosomes is associated with the flagellar fraction of the cytoskeleton and represents a new family of proteins that are present in several divergent eukaryotes. J. Biol. Chem. 275, 39369–39378 (2000)

  18. 18

    Ralston, K. S. & Hill, K. L. Trypanin, a component of the flagellar dynein regulatory complex, is essential in bloodstream form African trypanosomes. PLoS Pathogens 2 10.1371/journal.ppat.0020101 (2006)

  19. 19

    Bekker, J. M. et al. Direct interaction of Gas11 with microtubules: Implications for the dynein regulatory complex. Cell Motil. Cytoskeleton 64, 461–473 (2007)

  20. 20

    Colantonio, J. R. et al. Expanding the role of the dynein regulatory complex to non-axonemal functions: association of GAS11 with the golgi apparatus. Traffic 7, 538–548 (2006)

  21. 21

    Yeh, S. D. et al. Isolation and properties of Gas8, a growth arrest-specific gene regulated during male gametogenesis to produce a protein associated with the sperm motility apparatus. J. Biol. Chem. 277, 6311–6317 (2002)

  22. 22

    Whitmore, S. A. et al. Characterization and screening for mutations of the growth arrest-specific 11 (GAS11) and C16orf3 genes at 16q24.3 in breast cancer. Genomics 52, 325–331 (1998)

  23. 23

    Bok, J., Brunet, L. J., Howard, O., Burton, Q. & Wu, D. K. Role of hindbrain in inner ear morphogenesis: analysis of Noggin knockout mice. Dev. Biol. 311, 69–78 (2007)

  24. 24

    Leger, S. & Brand, M. Fgf8 and Fgf3 are required for zebrafish ear placode induction, maintenance and inner ear patterning. Mech. Dev. 119, 91–108 (2002)

  25. 25

    Mowbray, C., Hammerschmidt, M. & Whitfield, T. T. Expression of BMP signalling pathway members in the developing zebrafish inner ear and lateral line. Mech. Dev. 108, 179–184 (2001)

  26. 26

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

  27. 27

    Sullivan-Brown, J. et al. Zebrafish mutations affecting cilia motility share similar cystic phenotypes and suggest a mechanism of cyst formation that differs from pkd2 morphants. Dev. Biol. 314, 261–275 (2008)

  28. 28

    van Rooijen, E. et al. LRRC50, a conserved ciliary protein implicated in polycystic kidney disease. J. Am. Soc. Nephrol. 19, 1128–1138 (2008)

  29. 29

    Kawakami, Y., Raya, A., Raya, R. M., Rodriguez-Esteban, C. & Belmonte, J. C. Retinoic acid signalling links left–right asymmetric patterning and bilaterally symmetric somitogenesis in the zebrafish embryo. Nature 435, 165–171 (2005)

  30. 30

    Shastry, B. S. Mammalian cochlear genes and hereditary deafness. Microb. Comp. Genomics 5, 61–69 (2000)

  31. 31

    Westerfield, M. The Zebrafish Book: A Guide for the Laboratory Use of Zebrafish (Danio rerio) (Univ. Oregon Press, 1993)

  32. 32

    Curwen, V. et al. The Ensembl automatic gene annotation system. Genome Res. 14, 942–950 (2004)

  33. 33

    Chen, J. N. & Fishman, M. C. Zebrafish tinman homolog demarcates the heart field and initiates myocardial differentiation. Development 122, 3809–3816 (1996)

  34. 34

    Drees, B. L. et al. A protein interaction map for cell polarity development. J. Cell Biol. 154, 549–576 (2001)

Download references


We are grateful to R. Crosbie for discussions and encouragement throughout the course of the project. We thank I. Drummond and C. Nguyen for discussions and comments on the work. We are grateful to L. Trinh, O. Bricaud and A. Collazo for sharing reagents and for providing probes, as well as to all the members of the Fraser laboratory for discussions, in particular M. Liebling and W. Supatto for sharing Matlab scripts and comments. We are grateful to Z. P. Kabututu for performing the lrrc50 reverse transcriptase PCR experiments. J.V. was supported by a fellowship from the Human Frontier Science Program, D.W. was supported by the NIH Medical Scientist Training Program at UCLA/Caltech. J.R.C. was supported by NIH RSDA training grant no. M07185 and a Warsaw Fellowship from the MIMG Department at UCLA. A.D.L is supported by an NSF fellowship. This work was supported by NIH grant R01 HL081799 (J.C.), NIH grant R01AI52348 and Beckman Young Investigator Award (K.L.H.).

Author Contributions J.R.C., J.V., D.W. and K.L.H. designed the experiments and interpreted the results. J.R.C, J.V. and D.W. conducted the experiments. J.V. and D.W. developed the equipment and systems for and performed in vivo video imaging for the quantitative flow study and analyzed the data with J.R.C., S.F. and K.L.H. A.D.L. assisted with in situ hybridization. A.D.L. and J.C. provided guidance on gas8 morpholino injections. The manuscript was written by J.R.C., J.V., D.W. and K.L.H. All authors discussed the results and commented on the manuscript.

Author information

Correspondence to Kent L. Hill.

Supplementary information

Supplementary Figures and legends

This file contains Supplementary Figures S1-S6 with legends, and legends for Supplementary Movies 1-11b. (PDF 2347 kb)

Supplementary movie 1

Supplemental movie 1. Three dimensional display of a control inner ear at stage 27 hpf after immunofluorescence labelling of cilia with acetylated tubulin antibody. (MOV 1270 kb)

Supplementary movie 2

Supplemental movie 2. Three dimensional display of a gas11 morphant inner ear at stage 27 hpf after acetylated tubulin immunofluorescence labelling. (MOV 1304 kb)

Supplementary movie 3

Supplemental movie 3. Video shows tether cilia motility in control embryo. (MOV 1303 kb)

Supplementary movie 4

Supplemental movie 4. Video shows tether cilia motility in control embryo. (MOV 1496 kb)

Supplementary movie 5

Supplemental movie 5. Video shows tether cilia motility in control embryo. (MOV 2952 kb)

Supplementary movie 6

Supplemental movie 6. Video shows short cilia in control embryo are not motile. (MOV 5902 kb)

Supplementary movie 7

Supplemental movie 7. Video shows tether cilia in a gas11 morphant embryo are not motile. (MOV 831 kb)

Supplementary movie 8

Supplemental movie 8. Video shows tether cilia in a gas11 morphant embryo are not motile. (MOV 8050 kb)

Supplementary movie 9a

Supplemental movie 9a. High speed video microscopy of tether cilia in a control embryo at 24 hpf showing vortices in the vicinity of the tether cilia and particle propelling along the growing otolith. (MOV 1690 kb)

Supplementary movie 9b

Supplemental movie 9b. Same as 9a with particle tracking. (MOV 10688 kb)

Supplementary movie 10a

Supplemental movie 10a. High speed video microscopy of the inner ear in a control embryo at 23 hpf showing high displacement of otolith precursors in the vicinity of tether cilia and low displacement away from tether cilia. (MOV 3109 kb)

Supplementary movie 10b

Supplemental movie 10b. Same as supplemental movie 10a with particle tracking. (MOV 1591 kb)

Supplementary movie 11a

Supplemental movie 11a. High speed video microscopy of tether cilia in a gas11 morphant at 25 hpf showing very low particle displacement in the vicinity of tether cilia. (MOV 1357 kb)

Supplementary movie 11b

Supplemental movie 11b. Same as 11a with particle tracking. (MOV 14789 kb)

PowerPoint slides

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Colantonio, J., Vermot, J., Wu, D. et al. The dynein regulatory complex is required for ciliary motility and otolith biogenesis in the inner ear. Nature 457, 205–209 (2009) doi:10.1038/nature07520

Download citation

Further reading


By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.