VE-Cadherin and Endothelial Adherens Junctions: Active Guardians of Vascular Integrity

Epithelia are robust tissues that support the structure of embryos and organs and serve as effective barriers against pathogens. Epithelia also chemically separate different physiological environments. These vital functions require tight association between cells through the assembly of junctions that mechanically stabilize the tissue. Remarkably, epithelia are also dynamic and can display a fluid behavior. Cells continuously die or divide, thereby allowing functional tissue homeostasis. Epithelial cells can change shape or intercalate as tissues deform during morphogenesis. We review the mechanical basis of tissue robustness and fluidity, with an emphasis on the pivotal role of junction dynamics. Tissue fluidity emerges from local active stresses acting at cell interfaces and allows the maintenance of epithelial organization during morphogenesis and tissue renewal.

https://science.sciencemag.org/content/sci/340/6137/1185.full.pdf

Mechanics of epithelial tissue homeostasis and morphogenesis.

Classical cadherins are transmembrane proteins at the core of intercellular adhesion complexes in cohesive metazoan tissues. The extracellular domain of classical cadherins forms intercellular bonds with cadherins on neighboring cells, whereas the cytoplasmic domain recruits catenins, which in turn associate with additional cytoskeleton binding and regulatory proteins. Cadherin/catenin complexes are hypothesized to play a role in the transduction of mechanical forces that shape cells and tissues during development, regeneration, and disease. Whether mechanical forces are transduced directly through cadherins is unknown. To address this question, we used a Forster resonance energy transfer (FRET)-based molecular tension sensor to test the origin and magnitude of tensile forces transmitted through the cytoplasmic domain of E-cadherin in epithelial cells. We show that the actomyosin cytoskeleton exerts pN-tensile force on E-cadherin, and that this tension requires the catenin-binding domain of E-cadherin and αE-catenin. Surprisingly, the actomyosin cytoskeleton constitutively exerts tension on E-cadherin at the plasma membrane regardless of whether or not E-cadherin is recruited to cell–cell contacts, although tension is further increased at cell–cell contacts when adhering cells are stretched. Our findings thus point to a constitutive role of E-cadherin in transducing mechanical forces between the actomyosin cytoskeleton and the plasma membrane, not only at cell–cell junctions but throughout the cell surface.

https://www.pnas.org/content/pnas/109/31/12568.full.pdf

E-cadherin is under constitutive actomyosin-generated tension that is increased at cell–cell contacts upon externally applied stretch

Cancer-associated fibroblasts (CAFs) promote tumour invasion and metastasis. We show that CAFs exert a physical force on cancer cells that enables their collective invasion. Force transmission is mediated by a heterophilic adhesion involving N-cadherin at the CAF membrane and E-cadherin at the cancer cell membrane. This adhesion is mechanically active; when subjected to force it triggers β-catenin recruitment and adhesion reinforcement dependent on α-catenin/vinculin interaction. Impairment of E-cadherin/N-cadherin adhesion abrogates the ability of CAFs to guide collective cell migration and blocks cancer cell invasion. N-cadherin also mediates repolarization of the CAFs away from the cancer cells. In parallel, nectins and afadin are recruited to the cancer cell/CAF interface and CAF repolarization is afadin dependent. Heterotypic junctions between CAFs and cancer cells are observed in patient-derived material. Together, our findings show that a mechanically active heterophilic adhesion between CAFs and cancer cells enables cooperative tumour invasion.

/pdf/a-mechanically-active-heterotypic-e-cadherin-n-cadherin-56geajz9xt.pdf

A mechanically active heterotypic E-cadherin/N-cadherin adhesion enables fibroblasts to drive cancer cell invasion

Linkage between the adherens junction (AJ) and the actin cytoskeleton is required for tissue development and homeostasis. In vivo findings indicated that the AJ proteins E-cadherin, β-catenin, and the filamentous (F)-actin binding protein αE-catenin form a minimal cadherin-catenin complex that binds directly to F-actin. Biochemical studies challenged this model because the purified cadherin-catenin complex does not bind F-actin in solution. Here, we reconciled this difference. Using an optical trap-based assay, we showed that the minimal cadherin-catenin complex formed stable bonds with an actin filament under force. Bond dissociation kinetics can be explained by a catch-bond model in which force shifts the bond from a weakly to a strongly bound state. These results may explain how the cadherin-catenin complex transduces mechanical forces at cell-cell junctions.

/pdf/the-minimal-cadherin-catenin-complex-binds-to-actin-1veqg66ip8.pdf

The minimal cadherin-catenin complex binds to actin filaments under force

To remodel endothelial cell–cell adhesion, inflammatory cytokine- and angiogenic growth factor–induced signals impinge on the vascular endothelial cadherin (VE-cadherin) complex, the central component of endothelial adherens junctions. This study demonstrates that junction remodeling takes place at a molecularly and phenotypically distinct subset of VE-cadherin adhesions, defined here as focal adherens junctions (FAJs). FAJs are attached to radial F-actin bundles and marked by the mechanosensory protein Vinculin. We show that endothelial hormones vascular endothelial growth factor, tumor necrosis factor α, and most prominently thrombin induced the transformation of stable junctions into FAJs. The actin cytoskeleton generated pulling forces specifically on FAJs, and inhibition of Rho-Rock-actomyosin contractility prevented the formation of FAJs and junction remodeling. FAJs formed normally in cells expressing a Vinculin binding-deficient mutant of α-catenin, showing that Vinculin recruitment is not required for adherens junction formation. Comparing Vinculin-devoid FAJs to wild-type FAJs revealed that Vinculin protects VE-cadherin junctions from opening during their force-dependent remodeling. These findings implicate Vinculin-dependent cadherin mechanosensing in endothelial processes such as leukocyte extravasation and angiogenesis.

/pdf/vinculin-associates-with-endothelial-ve-cadherin-junctions-2zzj6e670n.pdf

Vinculin associates with endothelial VE-cadherin junctions to control force-dependent remodeling

The integrity of the endothelial barrier is controlled by the combined action of chemical and mechanical signaling systems. Permeability-regulating factors signal through small GTPases to regulate the architecture of the cytoskeleton and this has a strong impact on the morphology and stability of VE-cadherin-based cell-cell junctions. The details of how structural and mechanical properties of the actin cytoskeleton influence cell-cell adhesion and how this impacts the dynamic regulation of the endothelial barrier, are beginning to be elucidated. In this review, we discuss the physical and regulatory interactions between the VE-cadherin complex and the actomysoin cytoskeleton, as they are the main determinants of cell-cell adhesion and the mechanical architecture of the cytoskeleton. We discuss, based on recent in vitro data, how a balance between Linear Adherens Junctions, paralleled by cortical actin bundles and Focal Adherens Junctions, connected to radial action bundles, determines endothelial barrier function. We discuss how small GTPases control this balance by regulating the spatial organization and mechanics of actomyosin. We propose a hypothetical model of how biochemical and mechanical signals cooperate locally, at the actomyosin-adhesion interface to open and re-seal the barrier in a rapid and controlled manner.

Mechanical control of the endothelial barrier.

Mechanical forces are integrated at cadherin-based adhesion complexes to regulate morphology and strength of cell-cell junctions and organization of associated F-actin. A central mechanosensor at the cadherin complex is α-catenin, whose stretching recruits vinculin to regulate adhesion strength. The identity of the F-actin regulating signals that are also activated by mechanical forces at cadherin-based junctions has remained elusive. Here we identify the actin-regulators VASP, zyxin and TES as members of punctate, tensile cadherin-based junctions called Focal Adherens Junctions (FAJ) and show that they display mechanosensitive recruitment similar to that of vinculin. However, this recruitment is not altered by destroying or over-activating the α-catenin/vinculin module. Structured Illumination Microscopy (SIM) indicates that these tension sensitive proteins concentrate at locations within FAJs that are distinct from the core cadherin complex proteins. Furthermore, localization studies using mutated versions of VASP and zyxin indicate that these two proteins require binding to each other in order to localize to the FAJs. We conclude that there are multiple force sensitive modules present at the FAJ that are activated at distinct locations along the cadherin-F-actin axis and regulate specific aspects of junction dynamics.

/pdf/vasp-zyxin-and-tes-are-tension-dependent-members-of-focal-46v1x1x7an.pdf

Joppe Oldenburg

Papers

Vinculin associates with endothelial VE-cadherin junctions to control force-dependent remodeling

Mechanical control of the endothelial barrier.

VASP, zyxin and TES are tension-dependent members of Focal Adherens Junctions independent of the α-catenin-vinculin module