Formation of adherens junctions leads to the emergence of a tissue-level tension in epithelial monolayers (original) (raw)
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Spontaneous spatial correlation of elastic modulus in jammed epithelial monolayers observed by AFM
Biophysical Journal
For isolated single cells on a substrate, the intracellular stiffness, which is often measured as the Young's modulus, E, by atomic force microscopy (AFM), depends on the substrate rigidity. However, little is known about how the E of cells is influenced by the surrounding cells in a cell population system in which cells physically and tightly contact adjacent cells. In this study, we investigated the spatial heterogeneities of E in a jammed epithelial monolayer in which cell migration was highly inhibited, allowing us to precisely measure the spatial distribution of E in large-scale regions by AFM. The AFM measurements showed that E can be characterized using two spatial correlation lengths: the shorter correlation length, l S , is within the single cell size, whereas the longer correlation length, l L , is longer than the distance between adjacent cells and corresponds to the intercellular correlation of E. We found that l L decreased significantly when the actin filaments were disrupted or calcium ions were chelated using chemical treatments, and the decreased l L recovered to the value in the control condition after the treatments were washed out. Moreover, we found that l L decreased significantly when E-cadherin was knocked down. These results indicate that the observed long-range correlation of E is not fixed within the jammed state but inherently arises from the formation of a large-scale actin filament structure via E-cadherin-dependent cell-cell junctions.
Nature of active forces in tissues: how contractile cells can form extensile monolayers
Actomyosin machinery endows cells with contractility at a single cell level. However, at a tissue scale, cells can show either contractile or extensile behaviour based on the direction of pushing or pulling forces due to neighbour interactions or substrate interactions. Previous studies have shown that a monolayer of fibroblasts behaves as a contractile system1 while a monolayer of epithelial cells2,3 or neural crest cells behaves as an extensile system.4 How these two contradictory sources of force generation can coexist has remained unexplained. Through a combination of experiments using MDCK (Madin Darby Canine Kidney) cells, and in-silico modeling, we uncover the mechanism behind this switch in behaviour of epithelial cell monolayers from extensile to contractile as the weakening of intercellular contacts. We find that this switch in active behaviour also promotes the buildup of tension at the cell-substrate interface through an increase in actin stress fibers and higher tractio...
Characterizing the mechanics of cultured cell monolayers
One-cell-thick monolayers are the simplest tissues in multicellular organisms, yet they fulfill critical roles in development and normal physiology. In early development, embryonic morphogenesis results largely from monolayer rearrangement and deformation due to internally generated forces. Later, monolayers act as physical barriers separating the internal environment from the exterior and must withstand externally applied forces. Though resisting and generating mechanical forces is an essential part of monolayer function, simple experimental methods to characterize monolayer mechanical properties are lacking. Here, we describe a system for tensile testing of freely suspended cultured monolayers that enables the examination of their mechanical behavior at multi-, uni-, and subcellular scales. Using this system, we provide measurements of monolayer elasticity and show that this is two orders of magnitude larger than the elasticity of their isolated cellular components. Monolayers could withstand more than a doubling in length before failing through rupture of intercellular junctions. Measurement of stress at fracture enabled a first estimation of the average force needed to separate cells within truly mature monolayers, approximately ninefold larger than measured in pairs of isolated cells. As in single cells, monolayer mechanical properties were strongly dependent on the integrity of the actin cytoskeleton, myosin, and intercellular adhesions interfacing adjacent cells. High magnification imaging revealed that keratin filaments became progressively stretched during extension, suggesting they participate in monolayer mechanics. This multiscale study of monolayer response to deformation enabled by our device provides the first quantitative investigation of the link between monolayer biology and mechanics.
Nature Materials
Actomyosin machinery endows cells with contractility at a single cell level. However, within a monolayer, cells can be contractile or extensile based on the direction of pushing or pulling forces exerted by their neighbours or on the substrate. It has been shown that a monolayer of fibroblasts behaves as a contractile system while epithelial or neural progentior monolayers behave as an extensile system. Through a combination of cell culture experiments and in silico modeling, we reveal the mechanism behind this switch in extensile to contractile as the weakening of intercellular contacts. This switch promotes the buildup of tension at the cell-substrate interface through an increase in actin stress fibers and traction forces. This is accompanied by mechanotransductive changes in vinculin and YAP activation. We further show that contractile and extensile differences in cell activity sort cells in mixtures, uncovering a generic mechanism for pattern formation during cell competition, and morphogenesis. Main text The ability of cell monolayers to self-organize, migrate and evolve depends crucially on the interplay between cell-matrix and cell-cell interactions [1-4] which controls various phenomena including tissue morphogenesis [5, 6], epithelial-mesenchymal transition [1], wound healing and tumor progression [7]. Cells are active systems, engines that operate away from thermal equilibrium, transducing chemical energy into motion. Single isolated cells generate contractile force dipoles: the resultant of the forces due to actomyosin contraction, pulling on focal adhesion sites on the substrate, is typically a pair of approximately equal and opposite forces acting inwards along the cellular long axis [8] (Figure 1a). It is reasonable to expect that contractile particles also generate contractile behaviour in the monolayer [9]. However, at the collective cell level, epithelial monolayers [10, 11] and a monolayer of neural progentior cells display
The role of actomyosin contractility at epithelial adherens junctions has been extensively studied. However, little is known about how external forces are integrated to establish epithelial cell and organ shape in vivo. We use the Drosophila follicle epithelium to investigate how tension at adherens junctions is regulated to integrate external forces arising from changes in germline size and shape. We find that overall tension in the epithelium decreases despite pronounced growth of enclosed germline cells, suggesting that the epithelium relaxes to accommodate growth. However, we find local differences in adherens junction tension correlate with apposition to germline nurse cells or the oocyte. We demonstrate that medial Myosin II coupled to corrugating adherens junctions resists nurse cell-derived forces and thus maintains apical surface areas and cuboidal cell shapes. Furthermore, medial reinforcement of the apical surface ensures cuboidal-to-columnar cell shape transitions and im...
A new strategy to measure intercellular adhesion forces in mature cell-cell contacts
Scientific Reports, 2017
Intercellular adhesion plays a major role in tissue development and homeostasis. Yet, technologies to measure mature cell-cell contacts are not available. We introduce a methodology based on fluidic probe force microscopy to assess cell-cell adhesion forces after formation of mature intercellular contacts in cell monolayers. With this method we quantify that L929 fibroblasts exhibit negligible cell-cell adhesion in monolayers whereas human endothelial cells from the umbilical artery (HUAECs) exert strong intercellular adhesion forces per cell. We use a new in vitro model based on the overexpression of Muscle Segment Homeobox 1 (MSX1) to induce Endothelial-to-Mesenchymal Transition (EndMT), a process involved in cardiovascular development and disease. We reveal how intercellular adhesion forces in monolayer decrease significantly at an early stage of EndMT and we show that cells undergo stiffening and flattening at this stage. This new biomechanical insight complements and expands th...
A new approach to measure forces at junction vertices in an epithelium
2021
The mechanical properties of cell-cell junctions are critical for the stability of an epithelium. Cell-cell junction ablation experiments are classically used as a readout for junctional mechanics. However, without the knowledge of the viscoelastic properties of the microenvironment of the ablated junction, tensile junctional forces cannot be measured. Here we combine laser ablation with intracellular microrheology and develop a model to measure tensile forces exerted on cell-cell junctions. We show that the overexpression of the proto-oncogene atypical Protein Kinase C iota (aPKCi) in a single cell within a normal epithelium induces a gradient of junctional tension with neighbouring cells. Our method allows us to demonstrate that junctions contacting the aPKCi-overexpressing cell display a mechanical asymmetry which correlates with the levels of E-cadherin and P-MLC2. Measuring intracellular viscoelasticity is crucial for accurate measurements of cell-cell junction mechanics in the...
Mechanical adaptions of collective cells nearby free tissue boundaries
Journal of Biomechanics, 2020
Mechanical adaptions of cells, including stiffness variation, cytoskeleton remodeling, motion coordination, and shape changing, are essential for tissue morphogenesis, wound healing, and malignant progression. In this paper, we take confluent monolayers of Madin-Darby canine kidney (MDCK) and mouse myoblast (C2C12) cells as model systems to probe how cells collectively adapt their mechanical features in response to a free tissue boundary. We show that the free boundary not only can trigger unjamming transition but also induces cell fluidization nearby the boundary. The Young's moduli of cells near the boundary are found to be much lower than those of interior cells. We demonstrate that the stiffness of cells in monolayers with a free tissue boundary exhibits negative dependence on the projected cell area, in contrast to previous studies where cells were found to stiffen as cellular area increases in a confluent MDCK monolayer without boundary. In addition, the free tissue boundary may activate cell remodeling, rendering volume enlargement and cell-specified cytoskeleton organization. Our study emphasizes the important role of geometrical boundary in regulating biomechanical properties of cell aggregates.