Research Article

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

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Science  31 Oct 2014:
Vol. 346, Issue 6209, 1254211
DOI: 10.1126/science.1254211

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Structured Abstract

INTRODUCTION

Cadherins are an ancient class of transmembrane proteins that are essential for the formation of multicellular tissues in metazoans. Cadherins link intercellular adhesions to the cellular cytoskeleton, but how they are connected specifically to actin filaments is a hotly debated issue. Genetic and cell culture experiments indicate that E-cadherin, β-catenin, and the actin filament binding protein αE-catenin form a minimal cadherin-catenin complex that binds to the actin cytoskeleton directly in epithelial tissues. However, experiments with purified proteins showed that a stable cadherin-catenin complex can be reconstituted, but it does not bind strongly to actin filaments in solution. Nevertheless, cell culture experiments indicated that the cadherin-catenin complex is under constitutive actomyosin-generated tension and that this connection is required for mechanotransduction at cadherin-based adhesions. Here, we tested the hypothesis that tension is required to stabilize a linkage between the cadherin-catenin complex and actin filaments, and clarify how the cadherin-catenin complex could interact directly with the actin cytoskeleton in cells.

Embedded Image

Two-state catch bond model of cadherin-catenin/F-actin interactions. Force stabilizes the cadherin-catenin/F-actin bond by shifting it from a weakly to a strongly bound state. The force dependence of the connection between the cadherin-catenin complex and actin f laments may explain the mechanosensitivity of cadherin-mediated intercellular adhesions.

RATIONALE

We developed an optical trap–based assay to measure the lifetime of cadherin-catenin complex/actin filament bonds under tension. An actin filament was attached to two optically trapped beads and suspended above purified cadherin-catenin complexes immobilized on a glass coverslip surface that was precoated with glass microspheres. The coverslip was mounted on a motorized stage. This spatial arrangement was informed by electron tomography of cell-cell junctions, which showed actin filaments parallel to the plasma membrane. Tension was applied to cadherin-catenin complex/actin bonds by moving the sample stage back and forth parallel to the actin filament; if the immobilized cadherin-catenin complexes bound the actin filament, the attached beads were displaced from the optical trap. The lifetime of the bond was measured from the time series of the force exerted on the trapped beads. Kinetic models were fit to bond lifetime distributions with respect to applied force.

RESULTS

We observed robust, reproducible cadherin-catenin complex/actin filament binding under force in optical trap–based experiments. Bond lifetime distributions had a biphasic dependence on force. The mean lifetimes increased from ~60 ms at low force to ~1.2 s at ~10 pN, after which they decreased. A two-state catch bond model is consistent with the biphasic mean lifetime distribution and the presence of two distinct lifetime subpopulations. In this model, bonds between a cadherin-catenin complex and an actin filament form in a weakly bound state and quickly dissociate, but rapidly transition to a strongly bound state as applied force increases. Long lifetimes are achieved in this state until higher forces accelerate dissociation from the strongly bound state (see the figure).

CONCLUSION

Our data and kinetic model reconcile previous in vitro and in vivo work by demonstrating that the cadherin-catenin complex binds robustly to actin filaments under force. Thus, it seems that direct cadherin-catenin complex/actin filament binding was not detected in previous solution-based assays because bonds were not strengthened by tension. The two bound states in our model may correspond to different conformational states of αE-catenin, consistent with previous observations that αE-catenin may undergo changes in conformation in response to actomyosin-generated cytoskeletal tension. Our model of cadherin-catenin complex/ actin filament bond dissociation, combined with previous evidence of cooperative binding of αE-catenin to actin filaments, indicates that the linkage is self-reinforcing and that its stability is dynamically regulated by mechanical force during tissue development and maintenance.

Pulling me apart only makes me stronger

Tension transmitted between neighboring cells can exert profound effects on cell proliferation, differentiation, and tissue organization. Exactly how intercellular mechanical tension is sensed at the molecular level is unknown. One attractive hypothesis is that a linkage between the cell-cell adhesion molecule E-cadherin, its binding partners α- and β-catenin, and actin filaments may act as a tension sensor. However, how this linkage is established at the molecular level is not known. Buckley et al. used optical tweezers to determine how mechanical load influences interactions of the cadherin/catenin complex with single actin filaments. The data support a model in which force shifts the interaction from a force-independent, weakly bound state to a highly force-sensitive, strongly bound state. The findings may explain how cells maintain tissue integrity while still being able to move and change shape.

Science, this issue p. 10.1126/science.1254211

Abstract

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.

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