Force-dependent binding of vinculin to α-catenin regulates cell-cell contact stability and collective cell behavior.

The shaping of a multicellular body and repair of adult tissues require fine--tuning of cell adhesion, cell mechanics, and intercellular transmission of mechanical load. Adherens junctions (AJs) are the major intercellular junctions by which cells sense and exert mechanical force on each other. However, how AJs adapt to mechanical stress and how this adaptation contributes to cell-cell cohesion and eventually to tissue-scale dynamics and mechanics remains largely unknown. Here, by analyzing the tension-dependent recruitment of vinculin, α-catenin, and F-actin as a function of stiffness, as well as the dynamics of GFP-tagged wild-type and mutated α-catenins, altered for their binding capability to vinculin, we demonstrate that the force-dependent binding of vinculin stabilizes α-catenin and is responsible for AJ adaptation to force. Challenging cadherin complexes mechanical coupling with magnetic tweezers, and cell-cell cohesion during collective cell movements, further highlight that tension-dependent adaptation of AJs regulates cell-cell contact dynamics and coordinated collective cell migration. Altogether, these data demonstrate that the force-dependent α-catenin/vinculin interaction, manipulated here by mutagenesis and mechanical control, is a core regulator of AJ mechanics and long-range cell-cell interactions.

Force-dependent conformational switch of α-catenin controls vinculin binding.

Force sensing at cadherin-mediated adhesions is critical for their proper function. α-Catenin, which links cadherins to actomyosin, has a crucial role in this mechanosensing process. It has been hypothesized that force promotes vinculin binding, although this has never been demonstrated. X-ray structure further suggests that α-catenin adopts a stable auto-inhibitory conformation that makes the vinculin-binding site inaccessible. Here, by stretching single α-catenin molecules using magnetic tweezers, we show that the subdomains MI vinculin-binding domain (VBD) to MIII unfold in three characteristic steps: a reversible step at ~5 pN and two non-equilibrium steps at 10-15 pN. 5 pN unfolding forces trigger vinculin binding to the MI domain in a 1:1 ratio with nanomolar affinity, preventing MI domain refolding after force is released. Our findings demonstrate that physiologically relevant forces reversibly unfurl α-catenin, activating vinculin binding, which then stabilizes α-catenin in its open conformation, transforming force into a sustainable biochemical signal.

Symmetry breaking in spore germination relies on an interplay between polar cap stability and spore wall mechanics.

The morphogenesis of single cells depends on their ability to coordinate surface mechanics and polarity. During germination, spores of many species develop a polar tube that hatches out of a rigid outer spore wall (OSW) in a process termed outgrowth. However, how these awakening cells reorganize to stabilize this first growth axis remains unknown. Here, using quantitative experiments and modeling, we reveal the mechanisms underlying outgrowth in fission yeast. We find that, following an isotropic growth phase during which a single polarity cap wanders around the surface, outgrowth occurs when spores have doubled their volume, concomitantly with the stabilization of the cap and a singular rupture in the OSW. This rupture happens when OSW mechanical stress exceeds a threshold, releases the constraints of the OSW on growth, and stabilizes polarity. Thus, outgrowth exemplifies a self-organizing morphogenetic process in which reinforcements between growth and polarity coordinate mechanics and internal organization.

Adhesive interactions of N-cadherin limit the recruitment of microtubules to cell-cell contacts through organization of actomyosin.

Adhesive interactions of cadherins induce crosstalk between adhesion complexes and the actin cytoskeleton, allowing strengthening of adhesions and cytoskeletal organization. The underlying mechanisms are not completely understood, and microtubules (MTs) might be involved, as for integrin-mediated cell-extracellular-matrix adhesions. Therefore, we investigated the relationship between N-cadherin and MTs by analyzing the influence of N-cadherin engagement on MT distribution and dynamics. MTs progressed less, with a lower elongation rate, towards cadherin adhesions than towards focal adhesions. Increased actin treadmilling and the presence of an actomyosin contractile belt, suggested that actin relays inhibitory signals from cadherin adhesions to MTs. The reduced rate of MT elongation, associated with reduced recruitment of end-binding (EB) proteins to plus ends, was alleviated by expression of truncated N-cadherin, but was only moderately affected when actomyosin was disrupted. By contrast, destabilizing actomyosin fibers allowed MTs to enter the adhesion area, suggesting that tangential actin bundles impede MT growth independently of MT dynamics. Blocking MT penetration into the adhesion area strengthened cadherin adhesions. Taken together, these results establish a crosstalk between N-cadherin, F-actin and MTs. The opposing effects of cadherin and integrin engagement on actin organization and MT distribution might induce bias of the MT network during cell polarization.

α-Catenin and vinculin cooperate to promote high E-cadherin-based adhesion strength.

Maintaining cell cohesiveness within tissues requires that intercellular adhesions develop sufficient strength to support traction forces applied by myosin motors and by neighboring cells. Cadherins are transmembrane receptors that mediate intercellular adhesion. The cadherin cytoplasmic domain recruits several partners, including catenins and vinculin, at sites of cell-cell adhesion. Our study used force measurements to address the role of αE-catenin and vinculin in the regulation of the strength of E-cadherin-based adhesion. αE-catenin-deficient cells display only weak aggregation and fail to strengthen intercellular adhesion over time, a process rescued by the expression of αE-catenin or chimeric E-cadherin·αE-catenins, including a chimera lacking the αE-catenin dimerization domain. Interestingly, an αE-catenin mutant lacking the modulation and actin-binding domains restores cadherin-dependent cell-cell contacts but cannot strengthen intercellular adhesion. The expression of αE-catenin mutated in its vinculin-binding site is defective in its ability to rescue cadherin-based adhesion strength in cells lacking αE-catenin. Vinculin depletion or the overexpression of the αE-catenin modulation domain strongly decreases E-cadherin-mediated adhesion strength. This supports the notion that both molecules are required for intercellular contact maturation. Furthermore, stretching of cell doublets increases vinculin recruitment and α18 anti-αE-catenin conformational epitope immunostaining at cell-cell contacts. Taken together, our results indicate that αE-catenin and vinculin cooperatively support intercellular adhesion strengthening, probably via a mechanoresponsive link between the E-cadherin·ß-catenin complexes and the underlying actin cytoskeleton.