Letter | Published:

Kinetochore components are required for central spindle assembly

Nature Cell Biology volume 17, pages 697705 (2015) | Download Citation

  • A Corrigendum to this article was published on 30 June 2015

This article has been updated

Abstract

A critical structure poised to coordinate chromosome segregation with division plane specification is the central spindle that forms between separating chromosomes after anaphase onset1,2. The central spindle acts as a signalling centre that concentrates proteins essential for division plane specification and contractile ring constriction3. However, the molecular mechanisms that control the initial stages of central spindle assembly remain elusive. Using Caenorhabditis elegans zygotes, we found that the microtubule-bundling protein SPD-1PRC1 and the motor ZEN-4MKLP-1 are required for proper central spindle structure during its elongation4,5,6,7,8,9. In contrast, we found that the kinetochore controls the initiation of central spindle assembly. Specifically, central spindle microtubule assembly is dependent on kinetochore recruitment of the scaffold protein KNL-1, as well as downstream partners BUB-1, HCP-1/2CENP-F and CLS-2CLASP; and is negatively regulated by kinetochore-associated protein phosphatase 1 activity. This in turn promotes central spindle localization of CLS-2CLASP and initial central spindle microtubule assembly through its microtubule polymerase activity. Together, our results reveal an unexpected role for a conserved kinetochore protein network in coupling two critical events of cell division: chromosome segregation and cytokinesis.

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Change history

  • 08 June 2015

    In the version of this Letter originally published, the following sentence was omitted from the Acknowledgements: 'B.L. is supported by a post-doctoral fellowship from FRM (ARF20140129055).' This has been corrected in the online versions of the Letter.

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Acknowledgements

We thank all members of the Tran/Paoletti, Pintard, Doye and Dumont laboratories for support and advice. We are grateful to P. Moussounda and P. Feynerol for providing technical support. We thank C. Janke and N. Tavernier for their help with protein purification. We thank A. Desai and the CGC for worm strains. We thank Y. Kohara for the zen-4MKLP1 cDNA (yk35d10). We are grateful to M. Shirasu-Hiza for critical reading of the manuscript. T.K. is supported by R01-GM074215 (awarded to A. Desai). This work was supported by grants from the ANR (ANR-09-RPDOC-005-01), the FRM (AJE201112) and the Mairie de Paris (Emergence) to J.D., and NIH DP2 OD008773 to J.C.C. B.L. is supported by a post-doctoral fellowship from FRM (ARF20140129055).

Author information

Author notes

    • Gilliane Maton
    •  & Frances Edwards

    These authors contributed equally to this work.

Affiliations

  1. Institut Jacques Monod, CNRS, UMR 7592, University Paris Diderot, Sorbonne Paris Cité F-75205 Paris, France

    • Gilliane Maton
    • , Frances Edwards
    • , Benjamin Lacroix
    • , Marine Stefanutti
    • , Kimberley Laband
    • , Tiffany Lieury
    •  & Julien Dumont
  2. Ludwig Institute for Cancer Research/Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, California 92093, USA

    • Taekyung Kim
  3. Université Montpellier, CRBM, CNRS UMR 5237, 34293 Montpellier, France

    • Julien Espeut
  4. Columbia University, Department of Pathology and Cell Biology, New York, New York 10033, USA

    • Julie C. Canman

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Contributions

All experiments were conceived by J.D. with input from G.M. and F.E. Experiments were primarily performed and analysed by G.M. and F.E. Biochemistry experiments were performed by B.L. Most of the transgenic strains used here were constructed by M.S. and K.L. T.L. developed the automated central spindle tracking and quantification software. T.K. constructed and provided the OD971 strain. J.C.C. and J.E. constructed and shared several strains used here. G.M., F.E., J.C.C. and J.D. prepared the figures and wrote the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Julien Dumont.

Integrated supplementary information

Supplementary information

PDF files

  1. 1.

    Supplementary Information

    Supplementary Information

Videos

  1. 1.

    Supplementary Video 1

    Movie montage of the first embryonic division in (Left) GFP::TBB-2β−Tubulinf (yellow) and mCherry::H2B (magenta) or (Right) GFP::AIR-2AuroraB, GFP::γ-Tubulin (green) and mCherry::H2B (magenta) expressing embryos in control and after SPD-1PRC1 or ZEN-4MKLP1 depletion. Images, which are the maximum projection of 4 z-sections, were collected every 10 s and played back at 60x real time (6 images per second) with time 0 corresponding to anaphase onset.

  2. 2.

    Supplementary Video 2

    Movie montage of the first embryonic division in (Left) GFP::TBB-2β−Tubulin (yellow) and mCherry::H2B (magenta) or (Right) GFP::AIR-2AuroraB, GFP::γ-Tubulin (green) and mCherry::H2B (magenta) expressing embryos in GPR-1/2 depletion or GPR-1/2 and SPD-1PRC1, ZEN-4MKLP1 or CLS-2CLASP co-depletion. Images, which are the maximum projection of 4 z-sections, were collected every 10 s and played back at 60x real time (6 images per second) with time 0 corresponding to anaphase onset.

  3. 3.

    Supplementary Video 3

    Movie montage of the first embryonic division in (Left) GFP::TBB-2β−Tubulin (yellow) and mCherry::H2B (magenta) or (Right) GFP::AIR-2AuroraB, GFP::γ-Tubulin (green) and mCherry::H2B (magenta) expressing embryos in control and after NDC-80, ZWL-1ZWILCH or BUB-1 depletion. Images, which are the maximum projection of 4 z-sections, were collected every 10 s and played back at 60x real time (6 images per second) with time 0 corresponding to anaphase onset.

  4. 4.

    Supplementary Video 4

    Movie montage of the first embryonic division in β-Tubulin::GFP (yellow) and KNL-1::RFP (magenta) expressing embryos in control and after different CLS-2CLASP depletion conditions (28h or 32h post RNAi injection). Images, which are single z-sections were collected every second and played back at 6x real time (6 images per second) with time 0 corresponding to anaphase onset.

  5. 5.

    Supplementary Video 5

    Movie montage of the first embryonic division in (Left) GFP::TBB-2β−Tubulin (yellow) and mCherry::H2B (magenta) or (Right) GFP::AIR-2AuroraB, GFP::γ-Tubulin (green) and mCherry::H2B (magenta) expressing embryos in control and after partial HCP-1/2CENP-F or CLS-2CLASP depletion. Images, which are the maximum projection of 4 z-sections, were collected every 10 s and played back at 60x real time (6 images per second) with time 0 corresponding to anaphase onset.

  6. 6.

    Supplementary Video 6

    Movie montage of the first embryonic division in (Left) GFP::TBB-2β−Tubulin (yellow) and mCherry::H2B (magenta) or (Right) GFP::AIR-2AuroraB, GFP::γ-Tubulin (green) and mCherry::H2B (magenta) expressing embryos and either KNL-1 WT::RFP, KNL-1 Δ85-505::RFP or KNL-1 RRASA::RFP in absence of endogenous KNL-1. Images, which are the maximum projection of 4 z-sections were collected every 10 s and played back at 60x real time (6 images per second) with time 0 corresponding to anaphase onset.

  7. 7.

    Supplementary Video 7

    Movie montage of the first embryonic division in (Left) GFP::TBB-2β−Tubulin (yellow) and mCherry::H2B (magenta) or (Right) GFP::AIR-2AuroraB, GFP::γ-Tubulin (green) and mCherry::H2B (magenta) expressing embryos and either KNL-1 WT::RFP or KNL-1 RRASA::RFP in absence of endogenous KNL-1 or/and KLP-7MCAK. Images, which are the maximum projection of 4 z-sections were collected every 10 s and played back at 60x real time (6 images per second) with time 0 corresponding to anaphase onset.

  8. 8.

    Supplementary Video 8

    Movie montage of the first embryonic division in embryos expressing mCherry::TBB-2β−Tubulin (yellow) and GFP::CLS-2CLASP (magenta) WT (left) or 3A (right). Images, which are single z-sections, were collected every 10 s and played back at 60x real time (6 images per second) with time 0 corresponding to anaphase onset.

  9. 9.

    Supplementary Video 9

    Movie montage of the first embryonic division in embryos expressing GFP::EBP-1EB1 (green) and either KNL-1 WT::RFP, KNL-1 Δ85-505::RFP or KNL-1 RRASA::RFP (magenta) in absence of endogenous KNL-1 or KNL-1 WT::RFP (magenta) in absence of endogenous CLS-2CLASP. Images, which are single z-sections, were collected every 4 s and played back at 24x real time (6 images per second) with time 0 corresponding to anaphase onset.

  10. 10.

    Supplementary Video 10

    Movie montage of the first embryonic division in embryos expressing mCherry::H2B (magenta) and either GFP::TBB-2β−Tubulin or CLS-2CLASP ::GFP or SPD-1PRC1 ::GFP (green). Images, which are the maximum projection of 4 z-sections were collected every 10 s and played back at 60x real time (6 images per second) with time 0 corresponding to anaphase onset.

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DOI

https://doi.org/10.1038/ncb3150

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