Desmosome-anchored intermediate filaments facilitate tension-sensitive RhoA signaling for epithelial homeostasis

Abstract

Epithelia are subject to diverse forms of mechanical stress during development and post-embryonic life. They possess multiple mechanisms to preserve tissue integrity against tensile forces, which characteristically involve specialized cell-cell adhesion junctions coupled to the cytoskeleton. Desmosomes connect to intermediate filaments (IF) via desmoplakin (DP) 1,2, while the E-cadherin complex links to the actomyosin cytoskeleton in adherens junctions (AJ) 3. These distinct adhesion-cytoskeleton systems support different strategies to preserve epithelial integrity, especially against tensile stress. IFs coupled to desmosomes can passively respond to tension by strain-stiffening 4-10, whereas for AJs a variety of mechanotransduction mechanisms associated with the E-cadherin apparatus itself 11,12, or proximate to the junctions 13, can modulate the activity of its associated actomyosin cytoskeleton by cell signaling. We now report a pathway where these systems collaborate for active tension-sensing and epithelial homeostasis. We found that DP was necessary for epithelia to activate RhoA at AJ on tensile stimulation, an effect that required its capacity to couple IF to desmosomes. DP exerted this effect by facilitating the association of Myosin VI with E-cadherin, the mechanosensor for the tension-sensitive RhoA pathway at AJ 12. This connection between the DP-IF system and AJ-based tension-sensing promoted epithelial resilience when contractile tension was increased. It further facilitated epithelial homeostasis by allowing apoptotic cells to be eliminated by apical extrusion. Thus, active responses to tensile stress in epithelial monolayers reflect an integrated response of the IF- and actomyosin-based cell-cell adhesion systems.

Caveola mechanotransduction reinforces the cortical cytoskeleton to promote epithelial resilience

“…Overall, our data indicate that the junctional cortex is enhanced by activation of PtdIns (4, 5)P 2 - FMNL2 signalling when caveolae disassemble in response to mechanical stress. In considering the potential function of this response, we hypothesized that such cortical reinforcement might increase the ability of cell-cell junctions to resist disruptive mechanical stress (Acharya et al ., 2018; Nanavati et al , 2023). To test this, we stimulated cellular contractility with calyculin A, which increases mechanical tension on cell-cell adhesions, but eventually causes contacts to break.…”

“…This role for cortical reinforcement in epithelial resilience is consistent with the wellcharacterized contribution of F-actin to cadherin adhesion. Association of classical cadherins with Factin is essential for effective adhesion (Noordstra et al, 2023) and junctional cohesion is enhanced by actin assembly and stabilization (Engl et al, 2014;Kovacs et al, 2011;Lenne et al, 2021). Therefore, signalling to F-actin would provide a pathway for caveola to reinforce cell-cell junctions against mechanical stresses.…”

“…Our results thus identify caveola mechanotransduction as one of a number of mechanisms that epithelial cells possess to detect and respond to mechanical stresses. At AJ, other mechanoprotective mechanisms include catch bonds in the adhesive ectodomain of classical cadherins as well as at their interface with cortical actin filaments (Noordstra et al, 2023); and tension-activated RhoA signalling based on the assembly of an E-cadherin-Myosin VI mechanosensor (Acharya et al, 2018;Duszyc et al, 2021). This raises questions about how these diverse mechanisms may be integrated.…”

“…To corroborate the results of recoil experiments, we then assayed changes in the conformation of the E-cadherin-associated protein, a-catenin, as an index of molecular-level tension at AJ. Application of mechanical tension causes conformational unfolding of a-catenin, exposing cryptic epitopes including one in the central M-domain that is recognised by the a18 monoclonal antibody (mAb) (Noordstra et al, 2023;Yonemura et al, 2010). We therefore measured a18 mAb fluorescence intensity normalised for total junctional a-catenin levels (a18/pan-a-catenin ratio) as a proxy for molecular-level tension applied to the cadherin-catenin complex.…”

“…For example, the cadherin molecular complex in AJ contains mechanosensitive elements, such as the adhesive binding domain of E-cadherin, the cytosolic adapter protein, a-catenin, and the cadherin-associated protein, Myosin VI. Mechanical forces transmitted via E-cadherin can induce conformational changes in a-catenin to promote vinculin and F-actin binding (Noordstra et al, 2023), while recruitment of Myosin VI engages a signal transduction apparatus that activates RhoA at AJ (Acharya et al, 2018).…”