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Actin dynamics modulate mechanosensitive immobilization of E-cadherin at adherens junctions

Abstract

Mechanical stress is increasingly being shown to be a potent modulator of cell–cell junctional morphologies in developmental and homeostatic processes. Intercellular force sensing is thus expected to be an important regulator of cell signalling and tissue integrity. In particular, the interplay between myosin contractility, actin dynamics and E-cadherin recruitment largely remains to be uncovered. We devised a suspended cell doublet assay to quantitatively assess the correlation between myosin II activity and local E-cadherin recruitment. The single junction of the doublet exhibited a stereotypical morphology, with E-cadherin accumulating into clusters of varied concentrations at the rim of the circular contact. This local recruitment into clusters derived from the sequestration of E-cadherin through a myosin-II-driven modulation of actin turnover. We exemplify how the regulation of actin dynamics provides a mechanism for the mechanosensitive response of cell contacts.

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Figure 1: E-cad herin (E-cad) organizes into stable clusters at the contact rim in an actin anchoring-dependent manner.
Figure 2: The contact is under tension.
Figure 3: E-cad dynamic recruitment following spontaneous deformation.
Figure 4: Mapping of E-cad key partners.
Figure 5: Comparative E-cad and actin dynamics on the same junction.
Figure 6: Myosin II activity and F-actin stability regulate the level of E-cad recruitment.
Figure 7: Comparative E-cad and actin dynamics following modulation of myosin II activity and F-actin stability.

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References

  1. Kan, N. G. et al. Gene replacement reveals a specific role for E-cadherin in the formation of a functional trophectoderm. Development 134, 31–41 (2007).

    Article  CAS  PubMed  Google Scholar 

  2. Gumbiner, B. A functional assay for proteins involved in establishing an epithelial occluding barrier: Identification of a uvomorulin-like polypeptide. J. Cell Biol. 102, 457–468 (1986).

    Article  CAS  PubMed  Google Scholar 

  3. Takeichi, M. The cadherins: Cell-cell adhesion molecules controlling animal morphogenesis. Development 102, 639–655 (1988).

    CAS  PubMed  Google Scholar 

  4. Schmalhofer, O., Brabletz, S. & Brabletz, T. E-cadherin, beta-catenin, and ZEB1 in malignant progression of cancer. Cancer Metastasis Rev. 28, 151–166 (2009).

    Article  CAS  PubMed  Google Scholar 

  5. Stepniak, E., Radice, G. L. & Vasioukhin, V. Adhesive and signaling functions of cadherins and catenins in vertebrate development. Cold Spring Harb. Perspect. Biol. 1, a002949 (2009).

    Article  PubMed  PubMed Central  Google Scholar 

  6. Huang, R. Y-J., Guilford, P. & Thiéry, J-P. Early events in cell adhesion and polarity during epithelial-mesenchymal transition. J. Cell Sci. 125, 4417–4422 (2012).

    Article  CAS  PubMed  Google Scholar 

  7. De Beco, S., Amblard, F. & Coscoy, S. New insights into the regulation of E-cadherin distribution by endocytosis. Inter. Rev. Cell Mol. Biol. 295, 63–108 (2012).

    Article  CAS  Google Scholar 

  8. Lecuit, T. & Le Goff, L. Orchestrating size and shape during morphogenesis. Nature 450, 189–192 (2007).

    Article  CAS  PubMed  Google Scholar 

  9. Lecuit, T., Lenne, P-F. & Munro, E. Force generation, transmission, and integration during cell and tissue morphogenesis. Annu. Rev. Cell Dev. Biol. 27, 157–184 (2011).

    Article  CAS  PubMed  Google Scholar 

  10. Käfer, J., Hayashi, T., Marée, A. F. M., Carthew, R. W. & Graner, F. Cell adhesion and cortex contractility determine cell patterning in the Drosophila retina. Proc. Natl Acad. Sci. USA 104, 18549–18554 (2007).

    Article  PubMed  PubMed Central  Google Scholar 

  11. Manning, M. L., Foty, R. A., Steinberg, M. S. & Schoetz, E-M. Coaction of intercellular adhesion and cortical tension specifies tissue surface tension. Proc. Natl Acad. Sci. USA 107, 12517–12522 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Shewan, A. M. Myosin 2 is a key rho kinase target necessary for the local concentration of E-cadherin at cell–cell contacts. Mol. Biol. Cell 16, 4531–4542 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Miyake, Y. et al. Actomyosin tension is required for correct recruitment of adherens junction components and zonula occludens formation. Exp. Cell Res. 312, 1637–1650 (2006).

    Article  CAS  PubMed  Google Scholar 

  14. Krendel, M. et al. Myosin-dependent contractile activity of the actin cytoskeleton modulates the spatial organization of cell–cell contacts in cultured epitheliocytes. Proc. Natl Acad. Sci. USA 96, 9666–9670 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Taguchi, K., Ishiuchi, T. & Takeichi, M. Mechanosensitive EPLIN-dependent remodeling of adherens junctions regulates epithelial reshaping. J. Cell Biol. 194, 643–656 (2011).

    Article  PubMed  PubMed Central  Google Scholar 

  16. Rauzi, M., Lenne, P-F. & Lecuit, T. Planar polarized actomyosin contractile flows control epithelial junction remodelling. Nature 468, 1110–1114 (2010).

    Article  CAS  PubMed  Google Scholar 

  17. Adams, C. L., Chen, Y. T., Smith, S. J. & James Nelson, W. Mechanisms of epithelial cell–cell adhesion and cell compaction revealed by high-resolution tracking of E-cadherin–green fluorescent protein. J. Cell Biol. 142, 1105 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Vasioukhin, V., Bauer, C., Yin, M. & Fuchs, E. Directed actin polymerization is the driving force for epithelial cell–cell adhesion. Cell 100, 209–219 (2000).

    Article  CAS  PubMed  Google Scholar 

  19. Ehrlich, J. S., Hansen, M. D. H. & Nelson, W. J. Spatio-temporal regulation of Rac1 localization and lamellipodia dynamics during epithelial cell–cell adhesion. Dev. Cell 3, 259–270 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Yamada, S. & Nelson, W. J. Localized zones of Rho and Rac activities drive initiation and expansion of epithelial cell–cell adhesion. J. Cell Biol. 178, 517–527 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Priya, R., Yap, A. S. & Gomez, G. A. E-cadherin supports steady-state Rho signaling at the epithelial zonula adherens. Differentiation 86, 133–140 (2013).

    Article  CAS  PubMed  Google Scholar 

  22. Wu, S. K. et al. Cortical F-actin stabilization generates apical-lateral patterns of junctional contractility that integrate cells into epithelia. Nat. Cell Biol. 16, 167–178 (2014).

    Article  CAS  PubMed  Google Scholar 

  23. Harrison, O. J. et al. The extracellular architecture of adherens junctions revealed by crystal structures of type i cadherins. Structure 19, 244–256 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Wu, Y. et al. Cooperativity between trans and cis interactions in cadherin-mediated junction formation. Proc. Natl Acad. Sci. USA 107, 17592–17597 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Hong, S., Troyanovsky, R. B. & Troyanovsky, S. M. Binding to F-actin guides cadherin cluster assembly, stability, and movement. J. Cell Biol. 201, 131–143 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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

    Article  PubMed  Google Scholar 

  27. Yonemura, S., Wada, Y., Watanabe, T., Nagafuchi, A. & Shibata, M. α-Catenin as a tension transducer that induces adherens junction development. Nat. Cell Biol. 12, 1–22 (2010).

    Article  Google Scholar 

  28. Cavey, M., Rauzi, M., Lenne, P-F. & Lecuit, T. A two-tiered mechanism for stabilization and immobilization of E-cadherin. Nature 453, 751–756 (2008).

    Article  CAS  PubMed  Google Scholar 

  29. Ladoux, B. et al. Strength dependence of cadherin-mediated adhesions. Biophys. J. 98, 534–542 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Tabdili, H. et al. Cadherin-dependent mechanotransduction depends on ligand identity but not affinity. J. Cell Sci. 125, 4362–4371 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Dufour, S., Beauvais-Jouneau, A., Delouvée, A. & Thiery, J. P. Differential function of N-cadherin and cadherin-7 in the control of embryonic cell motility. J. Cell Biol. 146, 501–516 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Chu, Y. S. Force measurements in E-cadherin-mediated cell doublets reveal rapid adhesion strengthened by actin cytoskeleton remodeling through Rac and Cdc42. J. Cell Biol. 167, 1183–1194 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Truong Quang, B-A., Mani, M., Markova, O., Lecuit, T. & Lenne, P-F. Principles of E-cadherin supramolecular organization in vivo. Curr. Biol. 23, 2197–2207 (2013).

    Article  CAS  PubMed  Google Scholar 

  34. De Beco, S., Gueudry, C., Amblard, F. & Coscoy, S. Endocytosis is required for E-cadherin redistribution at mature adherens junctions. Proc. Natl Acad. Sci. USA 106, 7010–7015 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Maruthamuthu, V., Sabass, B., Schwarz, U. S. & Gardel, M. L. Cell–ECM traction force modulates endogenous tension at cell–cell contacts. Proc. Natl Acad. Sci. USA 108, 4708–4713 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Liu, Z. et al. Mechanical tugging force regulates the size of cell–cell junctions. Proc. Natl Acad. Sci. USA 107, 9944–9949 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Tinevez, J-Y. et al. Role of cortical tension in bleb growth. Proc. Natl Acad. Sci. USA 106, 18581–18586 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Maddugoda, M. P., Crampton, M. S., Shewan, A. M. & Yap, A. S. Myosin VI and vinculin cooperate during the morphogenesis of cadherin cell cell contacts in mammalian epithelial cells. J. Cell Biol. 178, 529–540 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Chang, Y-C., Nalbant, P., Birkenfeld, J., Chang, Z-F. & Bokoch, G. M. GEF-H1 couples nocodazole-induced microtubule disassembly to cell contractility via RhoA. Mol. Biol. Cell 19, 2147–2153 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

This work was supported by MBI seed funding (NRF grant). V.V. acknowledges additional support from the joint Singapore/France research laboratory LIA CAFS. J.P.T acknowledges IMCB A-star core funding.

Author information

Authors and Affiliations

Authors

Contributions

W.E. performed the experiments and handled the data analysis. B.A. prepared the microwells. L.L.Y. prepared the plasmids. W.E., J.P.T. and V.V. conceived the experiments. W.E. and V.V. wrote the manuscript. V.V. supervised the work.

Corresponding author

Correspondence to V. Viasnoff.

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Competing interests

The authors declare no competing financial interests.

Integrated supplementary information

Supplementary Figure 1 Digital procedure for rotation compensation and comparison of E-cadherin junctional distribution for different cell types.

(a) Top view (TV1-4) and side view (SV1-4) of a 3D stack used for determining the coordinates of the contact plane. TV1-SV1: deconvoluted stack as imaged in confocal microscopy. TV2-SV2: deconvoluted stack after rotation along the z-axis. TV3-SV3: deconvoluted stack in the referential of the contact. TV4-SV4: raw data in the referential of the contact (scalebar 5 μm). (b) Kymograph of a portion of the E-cad ring before and after image registration (scale bar 5 μm). (c) Typical ring of a MDCK-E-cadGFP doublet (n = 24 cell doublets, 4 days) (left) and an embryonic stem cell E-cadGFP doublet (n = 26 cells, 3 days) (right) (scalebar 5 μm). (d) Western blot of alpha catenin for control S180 cells and cells after alpha catenin knockdown. Cells used to look at the doublets had all the known phenotype of alpha catenin knockdown junctions displaying transient and dynamic cadherin clusters.

Supplementary Figure 2 Measurements of doublet deformation.

(a) Five rings and their anisotropy vector (top line). Same rings after rescaling and alignment of the anisotropy (bottom line) (scalebar 5 μm). (b) Superimposition of n = 5, 20 and 110 rings after image processing taken. (c) Side view schematic of the doublet deformation. (d) Images of the 2 slices taken 1.5 μm above and below the contact used to determine the contour of the cell (top and bottom). Image of the junction with anisotropy and deformation vectors (middle) (scalebar 5 μm). (e) Extreme cases where d1 and d2 have opposite directions (isotrope ring) and same direction (anisotrope ring) (scalebar 5 μm). (f) Anisotropy orientation as a function of the deformation orientation taken during contact formation and at steady state (n = 28 doublets with a total number of 572 time-points).

Supplementary Figure 3 Typical distribution of different junctional proteins.

Typical distributions at the junction of E-cad, phospho-MLC, -catenin, p120, actin, EPLIN, myosin VI and vinculin. The S180-Ecad-GFP cells were transfected with different junctional proteins tagged with m-cherry. The distribution of these proteins are compared to that of E-cad. A total of 116 doublets were observed (scalebar 5 μm).

Supplementary Figure 4 FRAP measurements on E-cadherins and actin.

(a) Typical E-cad FRAP signal for a doublet treated with Y27632 at 5 μM, in control conditions and treated with nocodazole at 10 μM. (b) E-cad recovery time for doublets treated with latrunculin at 1 μM (n = 15 doublets), with Y27632 at 5 μM (n = 21 doublets), in control conditions (n = 24 doublets), with nocodazole at 10 μM (n = 15 doublets) and with jasplakinolide at 100 nM (n = 12 doublets). The p-values are calculated from unpaired Students t-tests. (c) Typical actin FRAP signal for a doublet treated with Y27632 at 5 μM, in control conditions and treated with nocodazole at 10 μM. (d) E-cad recovery time in the dim region and the bright region of the same doublet (n = 22 doublets). The p-value is calculated from a paired Students t-test.

Supplementary Figure 5 Features of the E-cadherin distribution at the junction and cortical tension of single cells.

Data in graphs ac correspond to the same drugs and concentrations: Y27632 at 2 μM (n = 49 doublets), 5 μM (n = 24 doublets), 10 μM (n = 44 doublets) and 20 μM (n = 23 doublets); blebbistatin at 1 μM (n = 22 doublets), 5 μM (n = 17 doublets) and 50 μM (n = 12 doublets); nocodazole at 10 μM (n = 40 doublets); latrunculin at 1 μM (n = 11 doublets), and 5 μM (n = 22 doublets) and jasplakinolide at 100 nM (n = 18 doublets) (*, P < 0.05, **, P < 0.01, ***, P < 0.001, ****, P < 10 − 4). P-values are calculated from paired Students t-test. The whole distributions of the data values are also shown. a Average full-width at half maximum (FWHM) of the E-cad puncta on the dim side and on the bright side in control conditions (grey) and upon drug treatment. The error bars represent the standard deviation. The whole distribution is also shown. b Inter-cluster distance, contact radius and mean E-cadherin signal along the ring in control conditions (grey) and upon drug treatment. The same statistical tests as a. were applied. c E-cadherin density on the ring, at the center of the contact and on the membrane (as depicted in the schematic) in control conditions (grey) and upon drug treatment. d Image of a single cell deformed by negative pressure (left). Cortical tension of single cells for different drug treatment: latrunculin 1 μM (n = 8 doublets), Y27632 5 μM (n = 8 doublets), control (n = 12 doublets), nocodazole 10 μM (n = 9 doublets) and jasplakinolide at 100 nM (n = 10 doublets) (right).

Supplementary Figure 6 Model of E-cadherin immobilization by actin turnover.

We propose that the immobilization of E-cadherin is regulated by the turnover dynamics of the underlying actin cortex. In turn the turnover dynamics is modulated by the tension of the cortex generated by the actomyosin contractility. This mechanism provides a regulatory pathway for enhanced adhesion in cortex under tension.

Supplementary information

Supplementary Information

Supplementary Information (PDF 2565 kb)

Digital correction of residual rotations.

Illustration of the different stages of correction for the residual rotations of the doublets. The doublets are imaged by confocal imaging with z-stacks spaced by 0.5 μm during the expansion of the contact (E-cad labeled with GFP). The movie displays the maximum intensity projection. From left to right: raw image, correction for azimuthal rotation, additional correction for rotation in the plane of the junction. (AVI 2277 kb)

Protocol for digital correction during contact expansion.

llustration of the effect of correction (Movie 1) projected along the ZY plane. (AVI 426 kb)

Dynamics of contact formation.

Simultaneous dynamics of E-cad (green) and actin (magenta) localization at the rim of the contact during the contact expansion. Time t = 0 corresponds to the initial physical contact between both cells. (AVI 887 kb)

Cell contact shrinkage upon myosin II inhibition.

Dynamics of the shrinkage of the contact after inhibition of myosin II activity by Y27632 at 10 μM. (Ecad GFP). (AVI 195 kb)

Time correlation of cadherin anisotropy and doublet deformation.

Correlated time variations of the E-cad distribution anisotropy a (red) and the doublet deformation at the level of the contact d (green). (AVI 259 kb)

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Engl, W., Arasi, B., Yap, L. et al. Actin dynamics modulate mechanosensitive immobilization of E-cadherin at adherens junctions. Nat Cell Biol 16, 584–591 (2014). https://doi.org/10.1038/ncb2973

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