Multicellularity in animals requires dynamic maintenance of cell-cell contacts. Intercellularly ligated cadherins recruit numerous proteins to form supramolecular complexes that connect with the actin cytoskeleton and support force transmission. However, the molecular organization within such structures remains unknown. Here we mapped protein organization in cadherin-based adhesions by super-resolution microscopy, revealing a multi -compartment nanoscale architecture, with the plasma -membrane-proximal cadherin-catenin compartment segregated from the actin cytoskeletal compartment, bridged by an interface zone containing vinculin. Vinculin position is determined by alpha-catenin, and following activation, vinculin can extend similar to 30 nm to bridge the cadherin-catenin and actin compartments, while modulating the nanoscale positions of the actin regulators zyxin and VASP. Vinculin conformational activation requires tension and tyrosine phosphorylation, regulated by Abl kinase and PTP1B phosphatase. Such modular architecture provides a structural framework for mechanical and biochemical signal integration by vinculin, which may differentially engage cadherin-catenin complexes with the actomyosin machinery to regulate cell adhesions.
Coordination between Intra- and Extracellular Forces Regulates Focal Adhesion Dynamics
Bibhu Ranjan Sarangi,
Mukund Gupta,
Bryant L. Doss,
Nicolas Tissot,
France Lam,
Rene-Marc Mege,
Nicolas Borghi,
Benoit Ladoux
Focal adhesions (FAs) are important mediators of cell substrate interactions. One of their key functions is the transmission of forces between the intracellular acto-myosin network and the substrate. However, the relationships between cell traction forces, FA architecture, and molecular forces within FAs are poorly understood. Here, by combining Forster resonance energy transfer (FRET)-based molecular force biosensors with micropillar-based traction force sensors and high-resolution fluorescence microscopy, we simultaneously map molecular tension across vinculin, a key protein in FAs, and traction forces at FAs. Our results reveal strong spatiotemporal correlations between vinculin tension and cell traction forces at FAs throughout a wide range of substrate stiffnesses. Furthermore, we find that molecular tension within individual FAs follows a biphasic distribution from the proximal (toward the cell nucleus) to distal end (toward the cell-edge). Using super-resolution imaging, we show that such a distribution relates to that of FA proteins. On the basis of our experimental data, we propose a model in which FA dynamics results from tension changes along the FAs.
Epithelial Cell Packing Induces Distinct Modes of Cell Extrusions
Leyla Kocgozlu,
Thuan Beng Saw,
Phuong Anh Le,
Ivan Yow,
Murat Shagirov,
Eunice Wong,
Rene-Marc Mege,
Chwee Teck Lim,
Yusuke Toyama, et al.
Current Biology
26
(21)
2942-2950
(2016)
| Journal
The control of tissue growth, which is a key to maintain the protective barrier function of the epithelium, depends on the balance between cell division and cell extrusion rates [1, 2]. Cells within confluent epithelial layers undergo cell extrusion, which relies on cell-cell interactions [3] and actomyosin contractility [4, 5]. Although it has been reported that cell extrusion is also dependent on cell density [6, 7], the contribution of tissue mechanics, which is tightly regulated by cell density [8-12], to cell extrusion is still poorly understood. By measuring the multicellular dynamics and traction forces, we show that changes in epithelial packing density lead to the emergence of distinct modes of cell extrusion. In confluent epithelia with low cell density, cell extrusion is mainly driven by the lamellipodia-based crawling mechanism in the neighbor non-dying cells in connection with large-scale collective movements. As cell density increases, cell motion is shown to slow down, and the role of a supracellular actomyosin cable formation and its contraction in the neighboring cells becomes the preponderant mechanism to locally promote cell extrusion. We propose that these two distinct mechanisms complement each other to ensure proper cell extrusion depending on the cellular environment. Our study provides a quantitative and robust framework to explain how cell density can influence tissue mechanics and in turn regulate cell extrusion mechanisms.
Mechanics of epithelial tissues during gap closure
Simon Begnaud,
Tianchi Chen,
Delphine Delacour,
Rene-Marc Mege,
Benoit Ladoux
Current Opinion in Cell Biology
42
52-62
(2016)
| Journal
The closure of gaps is crucial to maintaining epithelium integrity during developmental and repair processes such as dorsal closure and wound healing. Depending on biochemical as well as physical properties of the microenvironment, gap closure occurs through assembly of multicellular actin-based contractile cables and/or protrusive activity of cells lining the gap. This review discusses the relative contributions of 'pursestring' and cell crawling mechanisms regulated by cell-substrate and cell-cell interactions, cellular mechanics and physical constraints from the environment.
Remodeling the zonula adherens in response to tension and the role of afadin in this response
Wangsun Choi,
Bipul R. Acharya,
Gregoire Peyret,
Marc-Antoine Fardin,
Rene-Marc Mege,
Benoit Ladoux,
Alpha S. Yap,
Alan S. Fanning,
Mark Peifer
Journal of Cell Biology
213
(2)
243-260
(2016)
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Morphogenesis requires dynamic coordination between cell cell adhesion and the cytoskeleton to allow cells to change shape and move without losing tissue integrity. We used genetic tools and superresolution microscopy in a simple model epithelial cell line to define how the molecular architecture of cell-cell zonula adherens (ZA) is modified in response to elevated contractility, and how these cells maintain tissue integrity. We previously found that depleting zonula occludens 1 (ZO-1) family proteins in MDCK cells induces a highly organized contractile actomyosin array at the ZA. We find that ZO knockdown elevates contractility via a Shroom3/Rho-associated, coiled-coil containing protein kinase (ROCK) pathway. Our data suggest that each bicellular border is an independent contractile unit, with actin cables anchored end-on to cadherin complexes at tricellular junctions. Cells respond to elevated contractility by increasing junctional afadin. Although ZO/afadin knockdown did not prevent contractile array assembly, it dramatically altered cell shape and barrier function in response to elevated contractility. We propose that afadin acts as a robust protein scaffold that maintains ZA architecture at tricellular junctions.
Single cell rigidity sensing: A complex relationship between focal adhesion dynamics and large-scale actin cytoskeleton remodeling
Many physiological and pathological processes involve tissue cells sensing the rigidity of their environment. In general, tissue cells have been shown to react to the stiffness of their environment by regulating their level of contractility, and in turn applying traction forces on their environment to probe it. This mechanosensitive process can direct early cell adhesion, cell migration and even cell differentiation. These processes require the integration of signals over time and multiple length scales. Multiple strategies have been developed to understand force- and rigidity-sensing mechanisms and much effort has been concentrated on the study of cell adhesion complexes, such as focal adhesions, and cell cytoskeletons. Here, we review the major biophysical methods used for measuring cell-traction forces as well as the mechanosensitive processes that drive cellular responses to matrix rigidity on 2-dimensional substrates.
Inference of Internal Stress in a Cell Monolayer
Vincent Nier,
Shreyansh Jain,
Chwee Teck Lim,
Shuji Ishihara,
Benoit Ladoux,
Philippe Marcq
Biophysical Journal
110
(7)
1625-1635
(2016)
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We combine traction force data with Bayesian inversion to obtain an absolute estimate of the internal stress field of a cell monolayer. The method, Bayesian inversion stress microscopy, is validated using numerical simulations performed in a wide range of conditions. It is robust to changes in each ingredient of the underlying statistical model. Importantly, its accuracy does not depend on the rheology of the tissue. We apply Bayesian inversion stress microscopy to experimental traction force data measured in a narrow ring of cohesive epithelial cells, and check that the inferred stress field coincides with that obtained by direct spatial integration of the traction force data in this quasi one-dimensional geometry.
Mechanical confinement triggers glioma linear migration dependent on formin FHOD3
Pascale Monzo,
Yuk Kien Chong,
Charlotte Guetta-Terrier,
Anitha Krishnasamy,
Sharvari R. Sathe,
Evelyn K. F. Yim,
Wai Hoe Ng,
Beng Ti Ang,
Carol Tang, et al.
Molecular Biology of the Cell
27
(8)
1246-1261
(2016)
| Journal
Glioblastomas are extremely aggressive brain tumors with highly invasive properties. Brain linear tracks such as blood vessel walls constitute their main invasive routes. Here we analyze rat C6 and patient-derived glioma cell motility in vitro using micropatterned linear tracks to mimic blood vessels. On laminin-coated tracks (3-10 mu m), these cells used an efficient saltatory mode of migration similar to their in vivo migration. This saltatory migration was also observed on larger tracks (50-400 mu m in width) at high cell densities. In these cases, the mechanical constraints imposed by neighboring cells triggered this efficient mode of migration, resulting in the formation of remarkable antiparallel streams of cells along the tracks. This motility involved microtubule-dependent polarization, contractile actin bundles and dynamic paxillin-containing adhesions in the leading process and in the tail. Glioma linear migration was dramatically reduced by inhibiting formins but, surprisingly, accelerated by inhibiting Arp2/3. Protein expression and phenotypic analysis indicated that the formin FHOD3 played a role in this motility but not mDia1 or mDia2. We propose that glioma migration under confinement on laminin relies on formins, including FHOD3, but not Arp2/3 and that the low level of adhesion allows rapid antiparallel migration.
Front-Rear Polarization by Mechanical Cues: From Sincle Cells to Tissues
Benoit Ladoux,
Rene-Marc Mege,
Xavier Trepat
Trends in Cell Biology
26
(6)
420-433
(2016)
| Journal
Directed cell migration is a complex process that involves front-rear polarization, characterized by cell adhesion and cytoskeleton-based protrusion, retraction, and contraction of either a single cell or a cell collective. Single cell polarization depends on a variety of mechanochemical signals including external adhesive cues, substrate stiffness, and confinement. In cell ensembles, coordinated polarization of migrating tissues results not only from the application of traction forces on the extracellular matrix but also from the transmission of mechanical stress through intercellular junctions. We focus here on the impact of mechanical cues on the establishment and maintenance of front rear polarization from single cell to collective cell behaviors through local or large-scale mechanisms.
Kontakt
Abteilung Mechanobiologie von Geweben Prof. Benoît Ladoux Principal Investigator
Max-Planck-Zentrum für Physik und Medizin Kussmaulallee 2 91054 Erlangen
