François Graner

Mechanical Control of Morphogenesis by Fat/Dachsous/Four-Jointed Planar Cell Polarity Pathway.

The Right Move During development, epithelial tissues deform to give rise to functional tissues and organs. How gene expression controls local cell mechanical properties to drive tissue deformation remains poorly understood. Bosveld et al. (p. 724, published online 12 April) have uncovered how the conserved Fat/Dachsous/Four-jointed signaling pathway controls local mechanical cell properties to generate global tissue contraction in Drosophila epithelial tissue. Polarized proto-cadherin and myosin induce an anisotropic tension at cell junctions and thereby shape epithelial tissue. During animal development, several planar cell polarity (PCP) pathways control tissue shape by coordinating collective cell behavior. Here, we characterize by means of multiscale imaging epithelium morphogenesis in the Drosophila dorsal thorax and show how the Fat/Dachsous/Four-jointed PCP pathway controls morphogenesis. We found that the proto-cadherin Dachsous is polarized within a domain of its tissue-wide expression gradient. Furthermore, Dachsous polarizes the myosin Dachs, which in turn promotes anisotropy of junction tension. By combining physical modeling with quantitative image analyses, we determined that this tension anisotropy defines the pattern of local tissue contraction that contributes to shaping the epithelium mainly via oriented cell rearrangements. Our results establish how tissue planar polarization coordinates the local changes of cell mechanical properties to control tissue morphogenesis.

PTEN controls junction lengthening and stability during cell rearrangement in epithelial tissue.

Planar cell rearrangements control epithelial tissue morphogenesis and cellular pattern formation. They lead to the formation of new junctions whose length and stability determine the cellular pattern of tissues. Here, we show that during Drosophila wing development the loss of the tumor suppressor PTEN disrupts cell rearrangements by preventing the lengthening of newly formed junctions that become unstable and keep on rearranging. We demonstrate that the failure to lengthen and to stabilize is caused by the lack of a decrease of Myosin II and Rho-kinase concentration at the newly formed junctions. This defect results in a heterogeneous cortical contractility at cell junctions that disrupts regular hexagonal pattern formation. By identifying PTEN as a specific regulator of junction lengthening and stability, our results uncover how a homogenous distribution of cortical contractility along the cell cortex is restored during cell rearrangement to control the formation of epithelial cellular pattern.

The role of fluctuations and stress on the effective viscosity of cell aggregates

Cell aggregates are a tool for in vitro studies of morphogenesis, cancer invasion, and tissue engineering. They respond to mechanical forces as a complex rather than simple liquid. To change an aggregates shape, cells have to overcome energy barriers. If cell shape fluctuations are active enough, the aggregate spontaneously relaxes stresses (“fluctuation-induced flow”). If not, changing the aggregates shape requires a sufficiently large applied stress (“stress-induced flow”). To capture this distinction, we develop a mechanical model of aggregates based on their cellular structure. At stress lower than a characteristic stress τ*, the aggregate as a whole flows with an apparent viscosity η*, and at higher stress it is a shear-thinning fluid. An increasing cell–cell tension results in a higher η* (and thus a slower stress relaxation time tc). Our constitutive equation fits experiments of aggregate shape relaxation after compression or decompression in which irreversibility can be measured; we find tc of the order of 5 h for F9 cell lines. Predictions also match numerical simulations of cell geometry and fluctuations. We discuss the deviations from liquid behavior, the possible overestimation of surface tension in parallel-plate compression measurements, and the role of measurement duration.

Two-dimensional flow of foam around a circular obstacle: local measurements of elasticity, plasticity and flow

We investigate the two-dimensional flow of a liquid foam around a circular obstacle by measuring all the local fields necessary to describe this flow: velocity, pressure, and bubble deformations and rearrangements. We show how our experimental set-up, a quasi-two-dimensional ‘liquid pool’ system, is adapted to the determination of these fields: the velocity and bubble deformations are easy to measure from two-dimensional movies, and the pressure can be measured by exploiting a specific feature of this system, a two-dimensional effective compressibility. To describe accurately neighbour swapping (so-called ‘T1’ processes), we propose a new, tensorial descriptor. All these quantities are evaluated via an averaging procedure that we justify by showing that the fluctuations of the fields are essentially Gaussian. The flow is extensively studied in a reference experimental case; the velocity presents an overshoot in the wake of the obstacle, and the pressure is maximum at the leading side and minimal at the trailing side. The study of the elastic deformations and of the velocity gradients shows that the transition between plug flow and yielded regions is smooth. Our tensorial description of T1s highlights their correlation both with the bubble deformations and the velocity gradients. A salient feature of the flow, notably for the velocity and T1 distribution, is a marked fore–aft asymmetry, the signature of the elastic behaviour of the foam. We show that the results do not change qualitatively when various control parameters (flow rate, bubble area, fluid fraction, bulk viscosity, obstacle size and boundary conditions) vary, identifying a robust quasi-static regime. These results are discussed in the framework of the foam rheology literature. A movie is available with the online version of the paper.

Experimental growth law for bubbles in a moderately Wet 3D liquid foam

We used x-ray tomography to characterize the geometry of all bubbles in a liquid foam of average liquid fraction phi(l) approximately 17% and to follow their evolution, measuring the normalized growth rate G=V(-1/3) dV/dt for 7000 bubbles. While G does not depend only on the number of faces of a bubble, its average over f-faced bubbles scales as G(f) approximately f - f(0) for large fs at all times. We discuss the dispersion of G and the influence of V and phi(l) on G.

Mechanical state, material properties and continuous description of an epithelial tissue.

During development, epithelial tissues undergo extensive morphogenesis based on coordinated changes of cell shape and position over time. Continuum mechanics describes tissue mechanical state and shape changes in terms of strain and stress. It accounts for individual cell properties using only a few spatially averaged material parameters. To determine the mechanical state and parameters in the Drosophila pupa dorsal thorax epithelium, we severed in vivo the adherens junctions around a disc-shaped domain comprising typically a hundred cells. This enabled a direct measurement of the strain along different orientations at once. The amplitude and the anisotropy of the strain increased during development. We also measured the stress-to-viscosity ratio and similarly found an increase in amplitude and anisotropy. The relaxation time was of the order of 10 s. We propose a space–time, continuous model of the relaxation. Good agreement with experimental data validates the description of the epithelial domain as a continuous, linear, visco-elastic material. We discuss the relevant time and length scales. Another material parameter, the ratio of external friction to internal viscosity, is estimated by fitting the initial velocity profile. Together, our results contribute to quantify forces and displacements, and their time evolution, during morphogenesis.

Comparative study of non-invasive force and stress inference methods in tissue

In the course of animal development, the shape of tissue emerges in part from mechanical and biochemical interactions between cells. Measuring stress in tissue is essential for studying morphogenesis and its physical constraints. For that purpose, a possible new approach is force inference (up to a single prefactor) from cell shapes and connectivity. It is non-invasive and can provide space-time maps of stress in a whole tissue, unlike existing methods. To validate this approach, three force-inference methods, which differ in their approach of treating indefiniteness in an inverse problem between cell shapes and forces, were compared. Tests using two artificial and two experimental data sets consistently indicate that our Bayesian force inference, by which cell-junction tensions and cell pressures are simultaneously estimated, performs best in terms of accuracy and robustness. Moreover, by measuring the stress anisotropy and relaxation, we cross-validated the force inference and the global annular ablation of tissue, each of which relies on different prefactors. A practical choice of force-inference methods in different systems of interest is discussed.Graphical abstract

Two-dimensional flow of foam around an obstacle: force measurements.

A Stokes experiment for foams is proposed. It consists of a two-dimensional flow of a foam, confined between a water subphase and a top plate, around a fixed circular obstacle. We present systematic measurements of the drag exerted by the flowing foam on the obstacle versus various separately controlled parameters: flow rate, bubble volume, bulk viscosity, obstacle size, shape, and boundary conditions. We separate the drag into two contributions: an elastic one (yield drag) at vanishing flow rate and a fluid one (viscous coefficient) increasing with flow rate. We quantify the influence of each control parameter on the drag. The results exhibit in particular a power-law dependence of the drag as a function of the bulk viscosity and the flow rate with two different exponents. Moreover, we show that the drag decreases with bubble size and increases proportionally to the obstacle size. We quantify the effect of shape through a dimensional drag coefficient, and we show that the effect of boundary conditions is small.

Anti-Inertial Lift in Foams: A Signature of the Elasticity of Complex Fluids

To understand the mechanics of a complex fluid such as a foam we propose a model experiment (a bidimensional flow around an obstacle) for which an external sollicitation is applied, and a local response is measured, simultaneously. We observe that an asymmetric obstacle (cambered airfoil profile) experiences a downwards lift, opposite to the lift usually known (in a different context) in aerodynamics. Correlations of velocity, deformations, and pressure fields yield a clear explanation of this inverse lift, involving the elasticity of the foam. We argue that such an inverse lift is likely common to complex fluids with elasticity.

Modulation of junction tension by tumor suppressors and proto-oncogenes regulates cell-cell contacts.

Tumor suppressors and proto-oncogenes play crucial roles in tissue proliferation. Furthermore, de-regulation of their functions is deleterious to tissue architecture and can result in the sorting of somatic rounded clones minimizing their contact with surrounding wild-type (wt) cells. Defects in the shape of somatic clones correlate with defects in proliferation, cell affinity, cell-cell adhesion, oriented cell division and cortical contractility. Combining genetics, live-imaging, laser ablation and computer simulations, we aim to analyze whether distinct or similar mechanisms can account for the common role of tumor suppressors and proto-oncogenes in cell-cell contact regulation. In Drosophila epithelia, the tumor suppressors Fat (Ft) and Dachsous (Ds) regulate cell proliferation, tissue morphogenesis, planar cell polarity and junction tension. By analyzing the evolution over time of ft mutant cells and clones, we show that ft clones reduce their cell-cell contacts with the surrounding wt tissue in the absence of concomitant cell divisions and over-proliferation. This contact reduction depends on opposed changes of junction tensions in the clone bulk and its boundary with neighboring wt tissue. More generally, either clone bulk or boundary junction tension is modulated by the activation of Yorkie, Myc and Ras, yielding similar contact reductions with wt cells. Together, our data highlight mechanical roles for proto-oncogene and tumor suppressor pathways in cell-cell interactions. Highlighted article: Time-lapse imaging studies to follow cell behavior and dynamics in Drosophila epithelia reveal mechanical roles for tumor suppressors and proto-oncogenes.