Roles of cell adhesion and mechanics in cell and tissue polarization – POLCAM

We have explored the influence of cell adhesion-mediated mechanical constraints on the emergence of cytoskeleton anisotropy (F-actin and microtubules) and front-rear cell polarization (lamellipodial activity, acto-myosin flow, polarization of organelles, polarization of traction forces) in simplified 1D cell models obtained by plating mammalian epithelial cell (MDCK) doublets and cell ensembles on micropatterned substrates (lines, circles or disks).
We determined the combined influences of substrate stiffness and of cadherin-catenin complexes linked to the acto-myosin cytoskeleton. Substrate stiffness has been controlled by the composition of polyacrylamide gels used as substrate that in addition allow to measure actual forces exerted by the cells. Intercellular junctions have been modulated by silencing or overexpressing E-cadherin and mutant forms of E-cadherin deficient for anchoring to acto-myosin. The role of the actomyosin has been investigated thanks to pharmacological perturbation or silencing of Myosin II isoform expression. Thereby, we have testes our working hypothesis that forces transmission at cell substrate and cell-cell contacts may support cytoskeleton symmetry breaking and polarization not only at the single cell level but in cell doublets and larger cell collectives. Quantitative data from these experiments have allow to feed theoretical models to describe the emergence of small cell assembly polarization.

Our two major results obtained during this project are: 1) the discovery of a new mode of polarization and mechanical coupling between collectively migrating cells in a closed system with strong developmental physiological implications (Nat. Phys 2020), 2) the demonstration of the role of active forces in the sorting of cell populations behaving like nematic particles (Nat. Mater. 2021). We have also deepened our understanding of the mechanisms of: 1) symmetry breaking in single cells, 2) formation of intercellular junctions, 3) and repair of epithelial injuries.

Overall, even if we were not able to identify novel biomechanical signaling pathways linking cell-cell adhesion and single cell polarity in complex cell population, our research allowed to reach more than all of the objectives that we identified initially. Our publications are already well cited indicating that they had already a significant impact in the field. The ongoing development of optogenetic tools are expected to allow us now to further progress in our dissection of associated mechanosignalling pathways.
The investigation of these core mechanisms of cell mechanics provided both fundamental biological and biophysical insights in collective cell behaviour processes. The better understanding how cells individually or collectively probe and respond to mechanical properties of their environment extend our current understanding of mechanotransduction at the cell and tissue scales. The novel model of mechanical coupling between collectively migrating epithelial cells allow a better understanding of the mode of migration undertaken by cell during development and tissue homeostatic renewal as discussed in our recent review for Current Opinion in Cell Biology. Our results also specify the critical roles of mechanics in the establishment of the body plan, whose deregulation associates with an array of human pathologies.

Funding for this project has resulted in the publication of at least 5 major original publications (Elife 2019, Nat. Phys. 2020, 2x Nat. Comm. 2021, Nat. Mater, 2021) reporting discoveries at the heart of POLCAM objectives. It also produced results published in 17 other original publications. We also synthesized our work in 3 journal publications. A key piece of our findings is still being re-evaluated for publication and available on BioRxiv: www.biorxiv.org/content/10.1101/2022.03.10.483785v1.

Front-rear polarization results from a symmetry breaking in cell organization required for efficient cell migration. Motile cells have however evolved and optimized different motile modes, for example single and collective cell migration. The same motility mechanisms are utilized by malignant cells during cancer progression. Cell motility both of isolated cells and when moving collectively, is therefore of fundamental importance in many fields of biology, and the emergence of front-rear cell polarity during these processes is the open question we aim to address in this proposal. Cell migration and polarization have recently attracted a large amount of interest in the field of biological physics, as it is a clear example where a key biological phenomenon is closely linked to physics concepts such as forces, friction and flows. From the physics point of view, cell motility is a prime example of self-propulsion, where self-organization of internally produced forces at the microscopic scale gives rise to a breaking of symmetry at the cell scale, and polarized motion. The problem of cell motility is therefore a subject where biology and physics meet, and where experimental data can be used to construct quantitative theoretical models that are based on physical concepts.
If symmetry breaking and maintenance of front-rear polarity of single migrating cells and of cells at the leading edge of a migrating tissue begin to be well understood, the situation is far more complex for cells migrating in the bulk of a compact tissue such as an epithelium. The complexity of migration patterns and the maintenance of physical interactions between neighbouring cells add a level of complexity that hindered so far a comprehensive biological and physical understanding of the processes of symmetry breaking and polarization at the scale of the tissues and/or of individual cells within the collective. The objective of our interdisciplinary project is to decipher, from the viewpoints of both physics and biology, how force transmission and biomechanical signalling at cell-cell contacts support multi-scale polarization in collectively migrating cell ensembles. To achieve this, our consortium combines a multidisciplinary range of skills comprising microfabrication, experimental cell biology, functional live cell imaging, optogenetic and biophysics with theoretical modelling. We aim at identifying how the force generating cytoskeleton and intercellular force transmission participates to cell polarization and coordination during epithelial collective cell migration in simplified model systems in vitro. We will thus explore the emergence of front-rear cell polarization and coordinated polarization from single cell up to multicellular assemblies. We will: 1) analyse how mechanical coupling at cell contacts induces symmetry breaking beyond the single cell level, 2) characterize coordinated polarization emerging during collective cell migration, and 3) determine how intercellular mechano-chemical cues regulate polarization at the tissue scale. We expect our results will provide new concepts on emerging properties of multicellular assembly as well as a quantitative framework to understand some of the general principles that govern the establishment of the body plan and tissue repair, and how their deregulation may lead to disease.