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Mapping the dynamics of force transduction at cell-cell junctions of epithelial clusters.

Ng MR, Besser A, Brugge JS, Danuser G - Elife (2014)

Bottom Line: We developed computational and experimental approaches to quantify, with both sub-cellular and multi-cellular resolution, the dynamics of force transmission in cell clusters.Applying this technology to spontaneously-forming adherent epithelial cell clusters, we found that basal force fluctuations were coupled to E-cadherin localization at the level of individual cell–cell junctions.Importantly, force transmission through a cell required coordinated modulation of cell-matrix adhesion and actomyosin contractility in the cell and its neighbors.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Harvard Medical School, Boston, United States.

ABSTRACT
Force transduction at cell–cell adhesions regulates tissue development, maintenance and adaptation. We developed computational and experimental approaches to quantify, with both sub-cellular and multi-cellular resolution, the dynamics of force transmission in cell clusters. Applying this technology to spontaneously-forming adherent epithelial cell clusters, we found that basal force fluctuations were coupled to E-cadherin localization at the level of individual cell–cell junctions. At the multi-cellular scale, cell–cell force exchange depended on the cell position within a cluster, and was adaptive to reconfigurations due to cell divisions or positional rearrangements. Importantly, force transmission through a cell required coordinated modulation of cell-matrix adhesion and actomyosin contractility in the cell and its neighbors. These data provide insights into mechanisms that could control mechanical stress homeostasis in dynamic epithelial tissues, and highlight our methods as a resource for the study of mechanotransduction in cell–cell adhesions [corrected].

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Mosaic downregulation of paxillin, talin-1, and myosin-IIA.(A) Western blot showing downregulation of paxillin in cells transfected with siPax. (B) E-cadherin-GFP-expressing cell pairs without (left) or with paxillin downregulation (marked by red nuclei). (C) Cell–cell force magnitudes at junctions between control or siPax cells. (D) Sum of traction force magnitudes exerted by individual control or paxillin-downregulated cells in cell pairs. (E) Western blot showing downregulation of talin-1 in cells transfected with siTln1. (F) E-cadherin-GFP-expressing cell pairs without (left) or with talin-1 downregulation (marked by red nuclei). (G) Cell–cell force magnitudes at junctions between control or siTln1 cells. (H) Sum of traction force magnitudes exerted by individual control or talin-1-downregulated cells in cell pairs. (I) Western blot showing knock-down of myosin-IIA in cells transfected with shRNA targeting the protein. (J) E-cadherin-GFP-expressing cell pairs without (left) or with myosin-IIA downregulation (marked by red nuclei). (K) Cell-cell force magnitudes at junctions between control or myosin-IIA-downregulated cells. (L) Sum of traction force magnitudes exerted by individual control or myosin-IIA-downregulated cells in cell pairs. (M) Sum of cell–cell force magnitudes at individual control or talin-1-downregulated cells with various degrees of connectivity. (N) Sum of cell–cell force magnitudes at individual control or myosin-IIA-downregulated cells with various degrees of connectivity. N = number of distinct junctions or cells measured; n = total number of measurements from N junctions or cells. ***p << 0.05.DOI:http://dx.doi.org/10.7554/eLife.03282.020
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fig9s1: Mosaic downregulation of paxillin, talin-1, and myosin-IIA.(A) Western blot showing downregulation of paxillin in cells transfected with siPax. (B) E-cadherin-GFP-expressing cell pairs without (left) or with paxillin downregulation (marked by red nuclei). (C) Cell–cell force magnitudes at junctions between control or siPax cells. (D) Sum of traction force magnitudes exerted by individual control or paxillin-downregulated cells in cell pairs. (E) Western blot showing downregulation of talin-1 in cells transfected with siTln1. (F) E-cadherin-GFP-expressing cell pairs without (left) or with talin-1 downregulation (marked by red nuclei). (G) Cell–cell force magnitudes at junctions between control or siTln1 cells. (H) Sum of traction force magnitudes exerted by individual control or talin-1-downregulated cells in cell pairs. (I) Western blot showing knock-down of myosin-IIA in cells transfected with shRNA targeting the protein. (J) E-cadherin-GFP-expressing cell pairs without (left) or with myosin-IIA downregulation (marked by red nuclei). (K) Cell-cell force magnitudes at junctions between control or myosin-IIA-downregulated cells. (L) Sum of traction force magnitudes exerted by individual control or myosin-IIA-downregulated cells in cell pairs. (M) Sum of cell–cell force magnitudes at individual control or talin-1-downregulated cells with various degrees of connectivity. (N) Sum of cell–cell force magnitudes at individual control or myosin-IIA-downregulated cells with various degrees of connectivity. N = number of distinct junctions or cells measured; n = total number of measurements from N junctions or cells. ***p << 0.05.DOI:http://dx.doi.org/10.7554/eLife.03282.020

Mentions: (A) Schematic of temporal cross-correlation analysis of force fluctuations at opposing cell–cell junctions. To determine the extent of force transmission from one cell–cell junction across a cell (depicted here in red) to the next junction or to the cell substrate, force fluctuations at one cell–cell interface i of the cell-of-interest are correlated with the fluctuations of the vector sum of cell–cell forces at all remaining cell–cell junctions of this cell or with the fluctuations of the negative residual traction force of the cell, respectively. See ‘Materials and methods’ for details. (B) Cross-correlation analysis results for control cells on 8 kPa or 35 kPa substrates and for cells with downregulation of paxillin (siPax), talin-1 (siTln1), or myosin-IIA (shMyoIIA) in mosaic cell clusters on 8 kPa substrates. See Figure 9—figure supplement 1. (C) Mosaic cell cluster with two siTln1-treated cells (red nuclei). (D) Graphical network representation of the cluster at the same time point. See Video 5 for full time lapse sequence. (E) Time courses of x-component of junctional forces (junction 2, magenta; junction 3, cyan) and residual traction force (black) in target cell 1 (cf. graphical network in D).


Mapping the dynamics of force transduction at cell-cell junctions of epithelial clusters.

Ng MR, Besser A, Brugge JS, Danuser G - Elife (2014)

Mosaic downregulation of paxillin, talin-1, and myosin-IIA.(A) Western blot showing downregulation of paxillin in cells transfected with siPax. (B) E-cadherin-GFP-expressing cell pairs without (left) or with paxillin downregulation (marked by red nuclei). (C) Cell–cell force magnitudes at junctions between control or siPax cells. (D) Sum of traction force magnitudes exerted by individual control or paxillin-downregulated cells in cell pairs. (E) Western blot showing downregulation of talin-1 in cells transfected with siTln1. (F) E-cadherin-GFP-expressing cell pairs without (left) or with talin-1 downregulation (marked by red nuclei). (G) Cell–cell force magnitudes at junctions between control or siTln1 cells. (H) Sum of traction force magnitudes exerted by individual control or talin-1-downregulated cells in cell pairs. (I) Western blot showing knock-down of myosin-IIA in cells transfected with shRNA targeting the protein. (J) E-cadherin-GFP-expressing cell pairs without (left) or with myosin-IIA downregulation (marked by red nuclei). (K) Cell-cell force magnitudes at junctions between control or myosin-IIA-downregulated cells. (L) Sum of traction force magnitudes exerted by individual control or myosin-IIA-downregulated cells in cell pairs. (M) Sum of cell–cell force magnitudes at individual control or talin-1-downregulated cells with various degrees of connectivity. (N) Sum of cell–cell force magnitudes at individual control or myosin-IIA-downregulated cells with various degrees of connectivity. N = number of distinct junctions or cells measured; n = total number of measurements from N junctions or cells. ***p << 0.05.DOI:http://dx.doi.org/10.7554/eLife.03282.020
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4300730&req=5

fig9s1: Mosaic downregulation of paxillin, talin-1, and myosin-IIA.(A) Western blot showing downregulation of paxillin in cells transfected with siPax. (B) E-cadherin-GFP-expressing cell pairs without (left) or with paxillin downregulation (marked by red nuclei). (C) Cell–cell force magnitudes at junctions between control or siPax cells. (D) Sum of traction force magnitudes exerted by individual control or paxillin-downregulated cells in cell pairs. (E) Western blot showing downregulation of talin-1 in cells transfected with siTln1. (F) E-cadherin-GFP-expressing cell pairs without (left) or with talin-1 downregulation (marked by red nuclei). (G) Cell–cell force magnitudes at junctions between control or siTln1 cells. (H) Sum of traction force magnitudes exerted by individual control or talin-1-downregulated cells in cell pairs. (I) Western blot showing knock-down of myosin-IIA in cells transfected with shRNA targeting the protein. (J) E-cadherin-GFP-expressing cell pairs without (left) or with myosin-IIA downregulation (marked by red nuclei). (K) Cell-cell force magnitudes at junctions between control or myosin-IIA-downregulated cells. (L) Sum of traction force magnitudes exerted by individual control or myosin-IIA-downregulated cells in cell pairs. (M) Sum of cell–cell force magnitudes at individual control or talin-1-downregulated cells with various degrees of connectivity. (N) Sum of cell–cell force magnitudes at individual control or myosin-IIA-downregulated cells with various degrees of connectivity. N = number of distinct junctions or cells measured; n = total number of measurements from N junctions or cells. ***p << 0.05.DOI:http://dx.doi.org/10.7554/eLife.03282.020
Mentions: (A) Schematic of temporal cross-correlation analysis of force fluctuations at opposing cell–cell junctions. To determine the extent of force transmission from one cell–cell junction across a cell (depicted here in red) to the next junction or to the cell substrate, force fluctuations at one cell–cell interface i of the cell-of-interest are correlated with the fluctuations of the vector sum of cell–cell forces at all remaining cell–cell junctions of this cell or with the fluctuations of the negative residual traction force of the cell, respectively. See ‘Materials and methods’ for details. (B) Cross-correlation analysis results for control cells on 8 kPa or 35 kPa substrates and for cells with downregulation of paxillin (siPax), talin-1 (siTln1), or myosin-IIA (shMyoIIA) in mosaic cell clusters on 8 kPa substrates. See Figure 9—figure supplement 1. (C) Mosaic cell cluster with two siTln1-treated cells (red nuclei). (D) Graphical network representation of the cluster at the same time point. See Video 5 for full time lapse sequence. (E) Time courses of x-component of junctional forces (junction 2, magenta; junction 3, cyan) and residual traction force (black) in target cell 1 (cf. graphical network in D).

Bottom Line: We developed computational and experimental approaches to quantify, with both sub-cellular and multi-cellular resolution, the dynamics of force transmission in cell clusters.Applying this technology to spontaneously-forming adherent epithelial cell clusters, we found that basal force fluctuations were coupled to E-cadherin localization at the level of individual cell–cell junctions.Importantly, force transmission through a cell required coordinated modulation of cell-matrix adhesion and actomyosin contractility in the cell and its neighbors.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Harvard Medical School, Boston, United States.

ABSTRACT
Force transduction at cell–cell adhesions regulates tissue development, maintenance and adaptation. We developed computational and experimental approaches to quantify, with both sub-cellular and multi-cellular resolution, the dynamics of force transmission in cell clusters. Applying this technology to spontaneously-forming adherent epithelial cell clusters, we found that basal force fluctuations were coupled to E-cadherin localization at the level of individual cell–cell junctions. At the multi-cellular scale, cell–cell force exchange depended on the cell position within a cluster, and was adaptive to reconfigurations due to cell divisions or positional rearrangements. Importantly, force transmission through a cell required coordinated modulation of cell-matrix adhesion and actomyosin contractility in the cell and its neighbors. These data provide insights into mechanisms that could control mechanical stress homeostasis in dynamic epithelial tissues, and highlight our methods as a resource for the study of mechanotransduction in cell–cell adhesions [corrected].

Show MeSH
Related in: MedlinePlus