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Probing the mechanical architecture of the vertebrate meiotic spindle

Nature Methods volume 6, pages 167–172 (2009)Cite this article

Abstract

Accurate chromosome segregation during meiosis depends on the assembly of a microtubule-based spindle of proper shape and size. Current models for spindle-size control focus on reaction diffusion–based chemical regulation and balance in activities of motor proteins. Although several molecular perturbations have been used to test these models, controlled mechanical perturbations have not been possible. Here we report a piezoresistive dual cantilever–based system to test models for spindle-size control and examine the mechanical features, such as deformability and stiffness, of the vertebrate meiotic spindle. We found that meiotic spindles prepared in Xenopus laevis egg extracts were viscoelastic and recovered their original shape in response to small compression. Larger compression resulted in plastic deformation, but the spindle adapted to this change, establishing a stable mechanical architecture at different sizes. The technique we describe here may also be useful for examining the micromechanics of other cellular organelles.

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Figure 1: Force measurements using a force-sensing cantilever.
Figure 2: Elastic response of meiotic spindle to small deformations.
Figure 3: Elastic response of meiotic spindle to the pole-to-pole compression.
Figure 4: Irreversible changes of the meiotic spindle shape owing to large deformations.
Figure 5: Transitions of meiotic spindle between different sizes driven by mechanical perturbations.

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Acknowledgements

This work was supported in part by Grant-in-Aid for Young Scientists (B) and Grant-in-Aid for Scientific Research on Priority Areas (T.I.), and Grant-in-Aid for Scientific Research (A), The 21st Century Center Of Excellence program and “Establishment of Consolidated Research Institute for Advanced Science and Medical Care” from the Ministry of Education, Culture, Sports, Science and Technology of Japan (S.I.), and a Research Grant from the Human Frontier Science Program (S.I. and T.M.K.). T.M.K. also acknowledges support from National Institutes of Health–National Institute of General Medical Sciences (GM65933).

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Author notes

  1. Jedidiah Gaetz

    Present address: Present address: Department of Human Genetics, University of Chicago, 929 East 57th Street, Chicago, Illinois 60637, USA.,

Authors and Affiliations

  1. Department of Physics, Faculty of Science and Engineering, Waseda University, 3-4-1 Okubo, Shinjuku, Tokyo, 169-8555, Japan

    Takeshi Itabashi, Jun Takagi, Yuta Shimamoto & Shin'ichi Ishiwata

  2. Advanced Research Institute for Science and Engineering, Waseda University, 3-4-1 Okubo, Shinjuku, Tokyo, 169-8555, Japan

    Takeshi Itabashi & Shin'ichi Ishiwata

  3. Laboratory of Chemistry and Cell Biology, Rockefeller University, 1230 York Avenue, Mail Box 202, New York, 10065, New York, USA

    Yuta Shimamoto, Jedidiah Gaetz & Tarun M Kapoor

  4. Graduate School of Information Science and Technology, The University of Tokyo, 7-3-1 Hongo, Bunkyo, Tokyo, 113-8656, Japan

    Hiroaki Onoe, Kenta Kuwana & Isao Shimoyama

Authors
  1. Takeshi Itabashi
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Contributions

T.I. performed the experiments and data analysis. J.T. provided considerable experimental assistance. T.I., T.M.K. and S.I. wrote the manuscript. H.O., K.K. and I.S. contributed to design and provide the piezo-resistive cantilevers. J.G. and Y.S. contributed to the initial project planning and experiment design. All authors discussed the results and commented on the manuscript.

Corresponding authors

Correspondence to Tarun M Kapoor or Shin'ichi Ishiwata.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–4 (PDF 613 kb)

Supplementary Video 1

Time-lapse observation of fluorescent meiotic spindle during small deformation. The meiotic spindle, which was sandwiched between the manipulating (left) and the force-sensing (right) cantilevers, was compressed perpendicular to the pole-to-pole axis by moving the manipulating cantilever horizontally with an amplitude of 2 μm on this image plane. Images were acquired every 1 s. Force-dependent deformability and the force response of the spindle are shown in Fig. 2 and Supplementary Fig. 1. Scale bar is 10 μm. (MOV 706 kb)

Supplementary Video 2

Time-lapse observation of fluorescent meiotic spindle during pole-to-pole compression. The meiotic spindle, which was sandwiched between the manipulating (left) and the force-sensing (right) cantilevers, was compressed to the pole-to-pole axis by moving the manipulating cantilever horizontally with an amplitude of 2 to 10-μm range in periodic cycles. Images were acquired every 1 s. Force-dependent deformability and the force response of the spindle at an amplitude of 2 μm are shown in Fig. 3. Scale bar is 10 μm. (MOV 4517 kb)

Supplementary Video 3

Time-lapse observation of fluorescent meiotic spindle during large deformation. The meiotic spindle, which was sandwiched between the manipulating (left) and the force-sensing (right) cantilevers, was compressed perpendicular to the pole-to-pole axis by moving the manipulating cantilever horizontally with an amplitude of 8 to 12-μm range in successive cycles. Images were acquired every 1 s. Force-dependent deformability and the force response of the spindle are shown in Fig. 4 and Supplementary Fig. 3. Scale bar is 10 μm. (MOV 4850 kb)

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Itabashi, T., Takagi, J., Shimamoto, Y. et al. Probing the mechanical architecture of the vertebrate meiotic spindle. Nat Methods 6, 167–172 (2009). https://doi.org/10.1038/nmeth.1297

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