Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Clathrin is required for the function of the mitotic spindle

Abstract

Clathrin has an established function in the generation of vesicles that transfer membrane and proteins around the cell1,2,3,4. The formation of clathrin-coated vesicles occurs continuously in non-dividing cells5, but is shut down during mitosis6, when clathrin concentrates at the spindle apparatus7,8. Here, we show that clathrin stabilizes fibres of the mitotic spindle to aid congression of chromosomes. Clathrin bound to the spindle directly by the amino-terminal domain of clathrin heavy chain. Depletion of clathrin heavy chain using RNA interference prolonged mitosis; kinetochore fibres were destabilized, leading to defective congression of chromosomes to the metaphase plate and persistent activation of the spindle checkpoint. Normal mitosis was rescued by clathrin triskelia but not the N-terminal domain of clathrin heavy chain, indicating that stabilization of kinetochore fibres was dependent on the unique structure of clathrin. The importance of clathrin for normal mitosis may be relevant to understanding human cancers that involve gene fusions of clathrin heavy chain.

This is a preview of subscription content, access via your institution

Relevant articles

Open Access articles citing this article.

Access options

Buy article

Get time limited or full article access on ReadCube.

$32.00

All prices are NET prices.

Figure 1: Clathrin was targeted to the mitotic spindle of NRK cells.
Figure 2: Clathrin was targeted to the mitotic spindle via the N-terminal domain of the heavy chain.
Figure 3: Inhibition of clathrin-mediated endocytosis did not disrupt mitosis.
Figure 4: Depletion of clathrin results in destabilized kinetochore fibres, defective congression of chromosomes and prolonged activation of the spindle checkpoint.
Figure 5: Full-length CHC, but not CHC N-terminal domain, is sufficient to rescue the mitotic defects found in cells depleted of endogenous CHC.

References

  1. Kirchhausen, T. Clathrin. Annu. Rev. Biochem. 69, 699–727 (2000)

    CAS  Article  Google Scholar 

  2. Brodsky, F. M., Chen, C. Y., Knuehl, C., Towler, M. C. & Wakeham, D. E. Biological basket weaving: formation and function of clathrin-coated vesicles. Annu. Rev. Cell Dev. Biol. 17, 517–568 (2001)

    CAS  Article  Google Scholar 

  3. Robinson, M. S. Adaptable adaptors for coated vesicles. Trends Cell Biol. 14, 167–174 (2004)

    CAS  Article  Google Scholar 

  4. Fotin, A. et al. Molecular model for a complete clathrin lattice from electron cryomicroscopy. Nature 432, 573–579 (2004)

    ADS  CAS  Article  Google Scholar 

  5. Gaidarov, I., Santini, F., Warren, R. A. & Keen, J. H. Spatial control of coated-pit dynamics in living cells. Nature Cell Biol. 1, 1–7 (1999)

    CAS  Article  Google Scholar 

  6. Warren, G. Membrane partitioning during cell division. Annu. Rev. Biochem. 62, 323–348 (1993)

    CAS  Article  Google Scholar 

  7. Maro, B., Johnson, M. H., Pickering, S. J. & Louvard, D. Changes in the distribution of membranous organelles during mouse early development. J. Embryol. Exp. Morphol. 90, 287–309 (1985)

    CAS  PubMed  Google Scholar 

  8. Okamoto, C. T., McKinney, J. & Jeng, Y. Y. Clathrin in mitotic spindles. Am. J. Physiol. Cell Physiol. 279, C369–C374 (2000)

    CAS  Article  Google Scholar 

  9. Sutherland, H. G. et al. Large-scale identification of mammalian proteins localized to nuclear sub-compartments. Hum. Mol. Genet. 10, 1995–2011 (2001)

    CAS  Article  Google Scholar 

  10. Compton, D. A. Spindle assembly in animal cells. Annu. Rev. Biochem. 69, 95–114 (2000)

    CAS  Article  Google Scholar 

  11. Yao, X., Abrieu, A., Zheng, Y., Sullivan, K. F. & Cleveland, D. W. CENP-E forms a link between attachment of spindle microtubules to kinetochores and the mitotic checkpoint. Nature Cell Biol. 2, 484–491 (2000)

    CAS  Article  Google Scholar 

  12. Mack, G. J. & Compton, D. A. Analysis of mitotic microtubule-associated proteins using mass spectrometry identifies astrin, a spindle-associated protein. Proc. Natl Acad. Sci. USA 98, 14434–14439 (2001)

    ADS  CAS  Article  Google Scholar 

  13. Motley, A., Bright, N. A., Seaman, M. N. & Robinson, M. S. Clathrin-mediated endocytosis in AP-2-depleted cells. J. Cell Biol. 162, 909–918 (2003)

    CAS  Article  Google Scholar 

  14. Hinrichsen, L., Harborth, J., Andrees, L., Weber, K. & Ungewickell, E. J. Effect of clathrin heavy chain- and α-adaptin-specific small inhibitory RNAs on endocytic accessory proteins and receptor trafficking in HeLa cells. J. Biol. Chem. 278, 45160–45170 (2003)

    CAS  Article  Google Scholar 

  15. Skibbens, R. V., Skeen, V. P. & Salmon, E. D. Directional instability of kinetochore motility during chromosome congression and segregation in mitotic newt lung cells: a push-pull mechanism. J. Cell Biol. 122, 859–875 (1993)

    CAS  Article  Google Scholar 

  16. Waters, J. C., Skibbens, R. V. & Salmon, E. D. Oscillating mitotic newt lung cell kinetochores are, on average, under tension and rarely push. J. Cell Sci. 109, 2823–2831 (1996)

    CAS  PubMed  Google Scholar 

  17. Cleveland, D. W., Mao, Y. & Sullivan, K. F. Centromeres and kinetochores: from epigenetics to mitotic checkpoint signaling. Cell 112, 407–421 (2003)

    CAS  Article  Google Scholar 

  18. Howell, B. J., Hoffman, D. B., Fang, G., Murray, A. W. & Salmon, E. D. Visualization of Mad2 dynamics at kinetochores, along spindle fibers, and at spindle poles in living cells. J. Cell Biol. 150, 1233–1250 (2000)

    CAS  Article  Google Scholar 

  19. Ungewickell, E. & Branton, D. Assembly units of clathrin coats. Nature 289, 420–422 (1981)

    ADS  CAS  Article  Google Scholar 

  20. ter Haar, E., Harrison, S. C. & Kirchhausen, T. Peptide-in-groove interactions link target proteins to the β-propeller of clathrin. Proc. Natl Acad. Sci. USA 97, 1096–1100 (2000)

    ADS  CAS  Article  Google Scholar 

  21. Miele, A. E., Watson, P. J., Evans, P. R., Traub, L. M. & Owen, D. J. Two distinct interaction motifs in amphiphysin bind two independent sites on the clathrin terminal domain β-propeller. Nature Struct. Mol. Biol. 11, 242–248 (2004)

    CAS  Article  Google Scholar 

  22. Hepler, P. K., McIntosh, J. R. & Cleland, S. Intermicrotubule bridges in mitotic spindle apparatus. J. Cell Biol. 45, 438–444 (1970)

    CAS  Article  Google Scholar 

  23. Kirchhausen, T., Harrison, S. C. & Heuser, J. Configuration of clathrin trimers: evidence from electron microscopy. J. Ultrastruct. Mol. Struct. Res. 94, 199–208 (1986)

    CAS  Article  Google Scholar 

  24. Jallepalli, P. V. & Lengauer, C. Chromosome segregation and cancer: cutting through the mystery. Nature Rev. Cancer 1, 109–117 (2001)

    CAS  Article  Google Scholar 

  25. Pulford, K., Morris, S. W. & Turturro, F. Anaplastic lymphoma kinase proteins in growth control and cancer. J. Cell. Physiol. 199, 330–358 (2004)

    CAS  Article  Google Scholar 

  26. Argani, P. et al. A novel CLTC-TFE3 gene fusion in pediatric renal adenocarcinoma with t(X;17)(p11.2;q23). Oncogene 22, 5374–5378 (2003)

    CAS  Article  Google Scholar 

  27. Bobanovic, L. K., Royle, S. J. & Murrell-Lagnado, R. D. P2X receptor trafficking in neurons is subunit specific. J. Neurosci. 22, 4814–4824 (2002)

    CAS  Article  Google Scholar 

  28. Simpson, F. et al. A novel adaptor-related protein complex. J. Cell Biol. 133, 749–760 (1996)

    CAS  Article  Google Scholar 

  29. Royle, S. J., Bobanovic, L. K. & Murrell-Lagnado, R. D. Identification of a non-canonical tyrosine-based endocytic motif in an ionotropic receptor. J. Biol. Chem. 277, 35378–35385 (2002)

    CAS  Article  Google Scholar 

  30. Bright, N. A., Reaves, B. J., Mullock, B. M. & Luzio, J. P. Dense core lysosomes can fuse with late endosomes and are re-formed from the resultant hybrid organelles. J. Cell Sci. 110, 2027–2040 (1997)

    CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank W. C. Earnshaw, G. Fang, G. Ihrke, A. P. Jackson and M. S. Robinson for their gifts of antibodies, plasmids and cells. We also thank J. W. Raff and M. S. Robinson for useful discussion. This work was supported by the MRC and the Human Frontiers Science Program (grant to L.L.). N.A.B. was funded by the MRC.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Stephen J. Royle.

Ethics declarations

Competing interests

The authors declare that they have no competing financial interests.

Supplementary information

Supplementary Notes

Contains Supplementary Results, Supplementary Figure Legends, Supplementary Tables and Supplementary Methods. (DOC 83 kb)

Supplementary Figure S1

Clathrin was targeted to the mitotic spindle of NRK cells. (JPG 523 kb)

Supplementary Figure S2

Clathrin light chains were targeted to the mitotic spindle. (JPG 149 kb)

Supplementary Figure S3

Clathrin remains associated with the spindle apparatus after extraction of soluble proteins. (JPG 264 kb)

Supplementary Figure S4

The association of clathrin with microtubules was not via coated membranes. (JPG 427 kb)

Supplementary Figure S5

Knockdown of clathrin heavy chain using RNAi. (JPG 435 kb)

Supplementary Figure S6

Clathrin depletion did not significantly alter spindle morphology. (JPG 170 kb)

Supplementary Figure S7

Mitotic defects in clathrin-depleted cells. (JPG 302 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Royle, S., Bright, N. & Lagnado, L. Clathrin is required for the function of the mitotic spindle. Nature 434, 1152–1157 (2005). https://doi.org/10.1038/nature03502

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature03502

This article is cited by

Comments

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.

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing