Letter | Published:

Asymmetric division of contractile domains couples cell positioning and fate specification

Nature volume 536, pages 344348 (18 August 2016) | Download Citation

Abstract

During pre-implantation development, the mammalian embryo self-organizes into the blastocyst, which consists of an epithelial layer encapsulating the inner-cell mass (ICM) giving rise to all embryonic tissues1. In mice, oriented cell division, apicobasal polarity and actomyosin contractility are thought to contribute to the formation of the ICM2,3,4,5. However, how these processes work together remains unclear. Here we show that asymmetric segregation of the apical domain generates blastomeres with different contractilities, which triggers their sorting into inner and outer positions. Three-dimensional physical modelling of embryo morphogenesis reveals that cells internalize only when differences in surface contractility exceed a predictable threshold. We validate this prediction using biophysical measurements, and successfully redirect cell sorting within the developing blastocyst using maternal myosin (Myh9)-knockout chimaeric embryos. Finally, we find that loss of contractility causes blastomeres to show ICM-like markers, regardless of their position. In particular, contractility controls Yap subcellular localization6, raising the possibility that mechanosensing occurs during blastocyst lineage specification. We conclude that contractility couples the positioning and fate specification of blastomeres. We propose that this ensures the robust self-organization of blastomeres into the blastocyst, which confers remarkable regulative capacities to mammalian embryos.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

References

  1. 1.

    , , & A self-organization framework for symmetry breaking in the mammalian embryo. Nature Rev. Mol. Cell Biol. 14, 452–459 (2013)

  2. 2.

    , , , & Initiation of Hippo signaling is linked to polarity rather than to cell position in the pre-implantation mouse embryo. Development 141, 2813–2824 (2014)

  3. 3.

    et al. Cortical tension allocates the first inner cells of the mammalian embryo. Dev. Cell 34, 435–447 (2015)

  4. 4.

    et al. Polarity-dependent distribution of angiomotin localizes Hippo signaling in preimplantation embryos. Curr. Biol. 23, 1181–1194 (2013)

  5. 5.

    , & Orientation of mitotic spindles during the 8- to 16-cell stage transition in mouse embryos. PLoS ONE 4, e8171 (2009)

  6. 6.

    et al. Role of YAP/TAZ in mechanotransduction. Nature 474, 179–183 (2011)

  7. 7.

    From mouse egg to mouse embryo: polarities, axes, and tissues. Annu. Rev. Cell Dev. Biol. 25, 483–512 (2009)

  8. 8.

    , , & Limited predictive value of blastomere angle of division in trophectoderm and inner cell mass specification. Development 141, 2279–2288 (2014)

  9. 9.

    & The foundation of two distinct cell lineages within the mouse morula. Cell 24, 71–80 (1981)

  10. 10.

    et al. PKCλ in liver mediates insulin-induced SREBP-1c expression and determines both hepatic lipid content and overall insulin sensitivity. J. Clin. Invest. 112, 935–944 (2003)

  11. 11.

    et al. Par-aPKC-dependent and -independent mechanisms cooperatively control cell polarity, Hippo signaling, and cell positioning in 16-cell stage mouse embryos. Dev. Growth Differ. 57, 544–556 (2015)

  12. 12.

    , , , & Pulsatile cell-autonomous contractility drives compaction in the mouse embryo. Nature Cell Biol. 17, 849–855 (2015)

  13. 13.

    Changes in cell dimensions and intercellular contacts during cleavage-stage cell cycles in mouse embryonic cells. J. Embryol. Exp. Morphol. 58, 231–249 (1980)

  14. 14.

    & Forces in tissue morphogenesis and patterning. Cell 153, 948–962 (2013)

  15. 15.

    et al. A nonapoptotic cell death process, entosis, that occurs by cell-in-cell invasion. Cell 131, 966–979 (2007)

  16. 16.

    & Stochastic patterning in the mouse pre-implantation embryo. Development 134, 4219–4231 (2007)

  17. 17.

    & Cell interactions influence the fate of mouse blastomeres undergoing the transition from the 16- to the 32-cell stage. Dev. Biol. 95, 211–218 (1983)

  18. 18.

    & Simulation of biological cell sorting using a two-dimensional extended Potts model. Phys. Rev. Lett. 69, 2013–2016 (1992)

  19. 19.

    The Differential Interfacial Tension Hypothesis (DITH): a comprehensive theory for the self-rearrangement of embryonic cells and tissues. J. Biomech. Eng. 124, 188–197 (2002)

  20. 20.

    , , & The structure and stability of multiple micro-droplets. Soft Matter 8, 7269–7278 (2012)

  21. 21.

    , & Multimaterial mesh-based surface tracking. ACM Trans. Graph. 33, 112 (2014)

  22. 22.

    et al. Tensile forces govern germ-layer organization in zebrafish. Nature Cell Biol. 10, 429–436 (2008)

  23. 23.

    et al. Adhesion functions in cell sorting by mechanically coupling the cortices of adhering cells. Science 338, 253–256 (2012)

  24. 24.

    et al. Confinement-optimized three-dimensional T cell amoeboid motility is modulated via myosin IIA-regulated adhesions. Nature Immunol. 11, 953–961 (2010)

  25. 25.

    et al. Nonmuscle myosin II isoform and domain specificity during early mouse development. Proc. Natl Acad. Sci. USA 107, 14645–14650 (2010)

  26. 26.

    et al. A mechanical checkpoint controls multicellular growth through YAP/TAZ regulation by actin-processing factors. Cell 154, 1047–1059 (2013)

  27. 27.

    , & Mechanical strain induces E-cadherin-dependent Yap1 and β-catenin activation to drive cell cycle entry. Science 348, 1024–1027 (2015)

  28. 28.

    et al. Contractile forces sustain and polarize hematopoiesis from stem and progenitor cells. Cell Stem Cell 14, 81–93 (2014)

  29. 29.

    et al. Lifeact mice for studying F-actin dynamics. Nature Methods 7, 168–169 (2010)

  30. 30.

    , , , & A global double-fluorescent Cre reporter mouse. Genesis 45, 593–605 (2007)

  31. 31.

    et al. Nuclear reprogramming: kinetics of cell cycle and metabolic progression as determinants of success. PLoS ONE 7, e35322 (2012)

  32. 32.

    et al. Expression of Cre recombinase in mouse oocytes: a means to study maternal effect genes. Genesis 26, 110–112 (2000)

  33. 33.

    et al. Conditional ablation of nonmuscle myosin II-B delineates heart defects in adult mice. Circ. Res. 105, 1102–1109 (2009)

Download references

Acknowledgements

We are grateful to the Hiiragi laboratory members and the European Molecular Biology Laboratory (EMBL) animal facility for their support. We thank F. Da and C. Batty for discussions on the simulations. We thank Y. Bellaïche for comments on an earlier version of the manuscript. Marie Curie individual fellowships under FP7 and H2020 programs support J.-L.M., H.T. and R.N. under Research Executive Agency grant agreements 329044, 656306 and 326701, respectively. H.T. acknowledges support from the Bettencourt-Schueller and Joachim Herz foundations. The Hiiragi laboratory is supported by EMBL, the European Research Council and VolkswagenStifftung.

Author information

Author notes

    • Jean-Léon Maître
    •  & Björn Eismann

    Present addresses: Mechanics of Mammalian Development Group, Institut Curie, CNRS UMR 3215, INSERM U934, 26, rue d’Ulm, 75248 Paris Cedex 05, France (J.-L.M.); Bioquant, Im Neuenheimer Feld 267, 69120 Heidelberg, Germany (B.E.).

    • Hervé Turlier
    •  & Rukshala Illukkumbura

    These authors contributed equally to this work.

Affiliations

  1. European Molecular Biology Laboratory, Meyerhofstrasse 1, 69117 Heidelberg, Germany

    • Jean-Léon Maître
    • , Hervé Turlier
    • , Rukshala Illukkumbura
    • , Björn Eismann
    • , Ritsuya Niwayama
    • , François Nédélec
    •  & Takashi Hiiragi

Authors

  1. Search for Jean-Léon Maître in:

  2. Search for Hervé Turlier in:

  3. Search for Rukshala Illukkumbura in:

  4. Search for Björn Eismann in:

  5. Search for Ritsuya Niwayama in:

  6. Search for François Nédélec in:

  7. Search for Takashi Hiiragi in:

Contributions

J.-L.M. designed the project and experiments, and wrote the manuscript with input from all authors. J.-L.M. and B.E. performed and analysed the tension and lineage mapping experiments. J.-L.M. and R.I. performed and analysed the remaining experiments. R.N. helped with image analysis of the periodic contractions. H.T. designed the physical model and performed the simulations with help from F.N. T.H. supervised the study and helped design the project.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Jean-Léon Maître or Takashi Hiiragi.

Reviewer Information Nature thanks D. Discher, P.-F. Lenne, B. Plusa and the other anonymous reviewer(s) for their contribution to the peer review of this work.

Extended data

Supplementary information

PDF files

  1. 1.

    Supplementary Information

    This file contains Supplementary Text and Data, Supplementary Figures 1-2 and additional references.

Videos

  1. 1.

    Periodic contractions in a polarized 8-cell stage blastomere

    Montage of mTmG (left) and local curvature measurement (right) of a polarized 8-cell stage blastomere showing periodic contractions. Time-lapse imaging consists of an image taken every 5 s and is displayed at 10 fps. Scale bar 10 μm.

  2. 2.

    Periodic contractions of in a 16-cell stage doublet

    Montage of mTmG (left) and local curvature measurement (right) of a doublet of a polarized and unpolarized 16-cell stage blastomeres showing periodic contractions. Time-lapse imaging consists of an image taken every 5 s and is displayed at 5 fps. Scale bar 10 μm.

  3. 3.

    Numerical simulation of the compaction and internalization of a doublet

    Numerical simulation of a cell doublet undergoing compaction (compaction parameter α decreasing from 0.9 to 0.25) followed by internalization (tension asymmetry parameter δ increasing from 1.0 to 1.6 with α kept at 0.25).

  4. 4.

    Numerical simulation of the compaction of a 16-cell embryo followed by the internalization of one cell

    Numerical simulation of a 16-cell embryo undergoing compaction (compaction parameter α decreasing from 0.9 to 0.25) followed by the internalization of one cell (tension asymmetry parameter δ increasing from 1.0 to 1.6 with α kept at 0.25).

  5. 5.

    Internalization of an asymmetric doublet

    Montage of mTmG (left), LifeAct-GFP (middle), merged image (right) of a polarized 8-cell stage blastomere undergoing asymmetric division to form an doublet of a polarized and unpolarized blastomere. The top panels show a single confocal slice whereas maximum projections of all slices are shown on the bottom. The polarized blastomere envelops the unpolarized one before the next division, which gives rise to a mini-blastocyst equivalent to a 32-cell stage embryo composed of four blasotmeres. Time-lapse imaging consists of an image taken every 30 min and is displayed at 5 fps. Scale bar 10 μm.

  6. 6.

    Surface tension and lineage mapping

    Fluorescent imaging of a H2B-GFP (magenta) and mTmG (green) expressing embryo from the 8- to 32-cell stage. Bright-field imaging shows the micropipette used for surface tension measurement. Time-lapse imaging consists of an image taken every 30 min and is displayed at 5 fps except when surface tension measurements and cell positions in the blastocyst are shown. Scale bar 10 μm.

  7. 7.

    Pre-implantation development of mzMyh10 embryos

    Bright-field imaging of maternal zygotic Myh10 knockout (mzMyh10) embryos developing from the 4- to 32-cell stage. Time-lapse imaging consists of an image taken every 30 min and is displayed at 10 fps. Scale bar 10 μm.

  8. 8.

    Development of a mMy9h embryo

    Fluorescent (green) and bright-field (grey) imaging of a mMyh9 mTmG embryo developing from the 4- to 32-cell stage. Time-lapse imaging consists of an image taken every 30 min and is displayed at 10 fps. Scale bar 10 μm.

  9. 9.

    Internalization of a WT blastomere within a mMyh9 host

    Fluorescent imaging of a WT mG (magenta) blastomere internalizing a mMyh9 mTmG (green) host. Time-lapse imaging consists of an image taken every 30 min and is displayed at 5 fps. Scale bar 10 μm.

  10. 10.

    Numerical simulation of the internalization of a cell of higher contractility grafted onto a 16-cell compacted embryo

    Numerical simulation the internalization of a cell of higher contractility (fixed tension asymmetry parameter δ = 1.6 and compaction parameter decreasing from α = 0.8 to 0.25, both relative to the embryo) grafted onto a homogeneous and compacted 16-cell stage embryo (fixed internal compaction parameter α = 0.25).

  11. 11.

    Spreading of a mMyh9 blastomere onto a WT host

    Fluorescent imaging of a mMyh9 mTmG (green) blastomere grafted onto a WT mG (magenta) host. Time-lapse imaging consists of an image taken every 30 min and is displayed at 5 fps. Scale bar 10 μm.

  12. 12.

    Numerical simulation of the spreading of a cell of lower contractility grafted onto a compacted 16-cell stage embryo

    Numerical simulation of the spreading of a cell of lower contractility (fixed tension asymmetry parameter δ = 0.5 and compaction parameter decreasing from α = 0.8 to 0.25, both relative to the embryo) grafted onto a homogeneous and compacted 16-cell stage embryo (fixed internal compaction parameter α = 0.25).

  13. 13.

    Internalization of a WT blastomere within a WT host after cell division

    Fluorescent imaging of a WT mTmG (magenta) blastomere internalizing a WT mG (cyan) host. Time-lapse imaging consists of an image taken every 30 min and is displayed at 5 fps. Scale bar 10 μm.

About this article

Publication history

Received

Accepted

Published

DOI

https://doi.org/10.1038/nature18958

Further reading

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.