Distinct inter-domain interactions of dimeric versus monomeric α-catenin link cell junctions to filaments

Abstract

Attachment between cells is crucial for almost all aspects of the life of cells. These inter-cell adhesions are mediated by the binding of transmembrane cadherin receptors of one cell to cadherins of a neighboring cell. Inside the cell, cadherin binds β-catenin, which interacts with α-catenin. The transitioning of cells between migration and adhesion is modulated by α-catenin, which links cell junctions and the plasma membrane to the actin cytoskeleton. At cell junctions, a single β-catenin/α-catenin heterodimer slips along filamentous actin in the direction of cytoskeletal tension which unfolds clustered heterodimers to form catch bonds with F-actin. Outside cell junctions, α-catenin dimerizes and links the plasma membrane to F-actin. Under cytoskeletal tension, α-catenin unfolds and forms an asymmetric catch bond with F-actin. To understand the mechanism of this important α-catenin function, we determined the 2.7 Å cryogenic electron microscopy (cryoEM) structures of filamentous actin alone and bound to human dimeric α-catenin. Our structures provide mechanistic insights into the role of the α-catenin interdomain interactions in directing α-catenin function and suggest a bivalent mechanism. Further, our cryoEM structure of human monomeric α-catenin provides mechanistic insights into α-catenin autoinhibition. Collectively, our structures capture the initial α-catenin interaction with F-actin before the sensing of force, which is a crucial event in cell adhesion and human disease.

Plakophilin-3 Binds the Membrane and Filamentous Actin without Bundling F-Actin

“…Size exclusion chromatography and multi-angle light scattering (SEC-MALS) analyses were carried out with purified plakophilin-3, as described previously [51,52]. Briefly, protein samples were passed through a Superdex 200 10/300 GL column, pre-equilibrated with 20 mM Tris (pH 7.5), 150 mM NaCl, and 1 mM dithiothreitol, using an Agilent 1260 Infinity series HPLC system connected with a Dawn-Heleos II multi-angle light-scattering detector (Wyatt Technology) and OptiLab T-rex differential refractive index detector (Wyatt Technology).…”

“…All cryoEM data were recorded with a JEOL cryoARM300 operated at 300 kV with a K3 detector (Gatan Inc., Warrendale, PA, USA). The condenser aperture was set to 50 µm with an Omega in-column energy filter, a slit width of 20 eV, and a zero-loss peak that aligned every six hours [51]. Automated image acquisition was performed using SerialEM [56] at a nominal magnification of ×60,000 and in the super-resolution mode with a pixel size of 0.36 Å.…”