Assembly, dynamics and remodeling of epithelial cell junctions throughout development.

Cell junctions play key roles in epithelial integrity. During development, when epithelia undergo extensive morphogenesis, these junctions must be remodeled in order to maintain mechanochemical barriers and ensure the cohesion of the tissue. In this Review, we present a comprehensive and integrated description of junctional remodeling mechanisms in epithelial cells during development, from embryonic to adult epithelia. We largely focus on Drosophila, as quantitative analyses in this organism have provided a detailed characterization of the molecular mechanisms governing cell topologies, and discuss the conservation of these mechanisms across metazoans. We consider how changes at the molecular level translate to tissue-scale irreversible deformations, exploring the composition and assembly of cellular interfaces to unveil how junctions are remodeled to preserve tissue homeostasis during cell division, intercalation, invagination, ingression and extrusion.

Toward a predictive understanding of epithelial cell death.

Epithelial cell death is highly prevalent during development and tissue homeostasis. While we have a rather good understanding of the molecular regulators of programmed cell death, especially for apoptosis, we still fail to predict when, where, how many and which specific cells will die in a tissue. This likely relies on the much more complex picture of apoptosis regulation in a tissular and epithelial context, which entails cell autonomous but also non-cell autonomous factors, diverse feedback and multiple layers of regulation of the commitment to apoptosis. In this review, we illustrate this complexity of epithelial apoptosis regulation by describing these different layers of control, all demonstrating that local cell death probability is a complex emerging feature. We first focus on non-cell autonomous factors that can locally modulate the rate of cell death, including cell competition, mechanical input and geometry as well as systemic effects. We then describe the multiple feedback mechanisms generated by cell death itself. We also outline the multiple layers of regulation of epithelial cell death, including the coordination of extrusion and regulation occurring downstream of effector caspases. Eventually, we propose a roadmap to reach a more predictive understanding of cell death regulation in an epithelial context.

Crk mediates Csk-Hippo signaling independently of Yap tyrosine phosphorylation to induce cell extrusion.

Src family kinases (SFKs), including Src, Fyn and Yes, play important roles in development and cancer. Despite being first discovered as the Yes-associated protein, the regulation of Yap by SFKs remains poorly understood. Here, through single-cell analysis and genetic lineage tracing, we show that the pan-epithelial ablation of C-terminal Src kinase (Csk) in the lacrimal gland unleashes broad Src signaling but specifically causes extrusion and apoptosis of acinar progenitors at a time when they are shielded by myoepithelial cells from the basement membrane. Csk mutants can be phenocopied by constitutively active Yap and rescued by deleting Yap or Taz, indicating a significant functional overlap between Src and Yap signaling. Although Src-induced tyrosine phosphorylation has long been believed to regulate Yap activity, we find that mutating these tyrosine residues in both Yap and Taz fails to perturb mouse development or alleviate the Csk lacrimal gland phenotype. In contrast, Yap loses Hippo signaling-dependent serine phosphorylation and translocates into the nucleus in Csk mutants. Further chemical genetics studies demonstrate that acute inhibition of Csk enhances Crk/CrkL phosphorylation and Rac1 activity, whereas removing Crk/CrkL or Rac1/Rap1 ameliorates the Csk mutant phenotype. These results show that Src controls Hippo-Yap signaling through the Crk/CrkL-Rac/Rap axis to promote cell extrusion.

Cell extrusion - a novel mechanism driving neural crest cell delamination.

Neural crest cells (NCC) comprise a heterogeneous population of cells with variable potency, that contribute to nearly every tissue and organ system throughout the body. Considered unique to vertebrates, NCC are transiently generated within the dorsolateral region of the neural plate or neural tube, during neurulation. Their delamination and migration are crucial events in embryo development as the differentiation of NCC is heavily influenced by their final resting locations. Previous work in avian and aquatic species has shown that NCC delaminate via an epithelial-mesenchymal transition (EMT), which transforms these stem and progenitor cells from static polarized epithelial cells into migratory mesenchymal cells with fluid front and back polarity. However, the cellular and molecular drivers facilitating NCC delamination in mammals are poorly understood. We performed live timelapse imaging of NCC delamination in mouse embryos and discovered a group of cells that exit the neuroepithelium as isolated round cells, which then halt for a short period prior to acquiring the mesenchymal migratory morphology classically associated with most delaminating NCC. High magnification imaging and protein localization analyses of the cytoskeleton, together with measurements of pressure and tension of delaminating NCC and neighboring neuroepithelial cells, revealed these round NCC are extruded from the neuroepithelium prior to completion of EMT. Furthermore, we demonstrate that cranial NCC are extruded through activation of the mechanosensitive ion channel, PIEZO1, a key regulator of the live cell extrusion pathway, revealing a new role for PIEZO1 in neural crest cell development. Our results elucidating the cellular and molecular dynamics orchestrating NCC delamination support a model in which high pressure and tension in the neuroepithelium results in activation of the live cell extrusion pathway and delamination of a subpopulation of NCC in parallel with EMT. This model has broad implications for our understanding of cell delamination in development and disease.