Nanomechanics of the Cadherin Ectodomain

Abstract

Cadherins form a large family of calcium-dependent cell-cell adhesion receptors involved in development, morphogenesis, synaptogenesis, differentiation, and carcinogenesis through signal mechanotransduction using an adaptor complex that connects them to the cytoskeleton. However, the molecular mechanisms underlying mechanotransduction through cadherins remain unknown, although their extracellular region (ectodomain) is thought to be critical in this process. By single molecule force spectroscopy, molecular dynamics simulations, and protein engineering, here we have directly examined the nanomechanics of the C-cadherin ectodomain and found it to be strongly dependent on the calcium concentration. In the presence of calcium, the ectodomain extends through a defined ("canalized") pathway that involves two mechanical resistance elements: a mechanical clamp from the cadherin domains and a novel mechanostable component from the interdomain calcium-binding regions ("calcium rivet") that is abolished by magnesium replacement and in a mutant intended to impede calcium coordination. By contrast, in the absence of calcium, the mechanical response of the ectodomain becomes largely "decanalized" and destabilized. The cadherin ectodomain may therefore behave as a calcium-switched "mechanical antenna" with very different mechanical responses depending on calcium concentration (which would affect its mechanical integrity and force transmission capability). The versatile mechanical design of the cadherin ectodomain and its dependence on extracellular calcium facilitate a variety of mechanical responses that, we hypothesize, could influence the various adhesive properties mediated by cadherins in tissue morphogenesis, synaptic plasticity, and disease. Our work represents the first step toward the mechanical characterization of the cadherin system, opening the door to understanding the mechanical bases of its mechanotransduction.The acquisition of specific machinery for cell adhesion represented a crucial step in the evolution of metazoan organisms, enabling multicellularity and the development of tissues. There are four major classes of cell adhesion receptors: the immunoglobulin (Ig)-like superfamily, the cadherin superfamily, selectins, and integrins (1). Because of their location at the cell-cell interface, these receptors are often subject to mechanical stress (2, 3). However, to date only the mechanical properties of Ig and fibronectin type III domains have been investigated using atomic force microscopy (AFM) 5 -based single molecule force spectroscopy (SMFS) and protein engineering (4).Cadherins can be defined (with some exceptions) as a superfamily of calcium (Ca 2ϩ )-dependent homophilic cell-cell adhesion proteins that mediate most cell adhesion in vertebrates, playing a crucial role in morphogenesis, synaptogenesis, tissue architecture, tissue repair, and carcinogenesis (5-7). Still, certain cadherins are heterophilic or/and connect two membranes from the same cell. At least in the case of the socalled "classical" cadh...

A Partial Calcium-Free Linker Confers Flexibility to Inner-Ear Protocadherin-15

“…Because Ca 2+ is important for the mechanical strength of cadherins (Oroz et al, 2011; Sotomayor and Schulten, 2008; Sotomayor et al, 2005), we investigated how the partial Ca 2+ -free EC3-4 linker impacts the strength of PCDH15 using steered molecular dynamics (SMD) simulations. In these simulations, we stretched EC3-5 by applying forces through springs attached to the N- and C-termini, which moved in opposite directions at constant speeds of 10, 1, and 0.1 nm/ns (Grubmuller, 2005; Isralewitz et al, 2001; Sotomayor and Schulten, 2007).…”

“…In the absence of Ca 2+ , cadherin linkers become flexible, allowing adjacent EC repeats to move relative to one another (Cailliez and Lavery, 2005; Haussinger et al, 2002; Pokutta et al, 1994; Sotomayor and Schulten, 2008). In addition, cadherin mechanical strength is significantly reduced in the absence of Ca 2+ (Oroz et al, 2011; Sotomayor and Schulten, 2008; Sotomayor et al, 2005). Tip links are also Ca 2+ -sensitive, as removal of extracellular Ca 2+ eliminates them and abolishes transduction currents in hair cells (Assad et al, 1991; Vollrath et al, 2007; Zhao et al, 1996).…”

An elastic element in the protocadherin-15 tip link of the inner ear

“…Calcium is known to help prevent mechanical unfolding in cadherins3654, and mutations that alter cadherin calcium-coordinating residues cause deafness (see Supplementary Discussion and Supplementary Fig. 16).…”

Sensing sound: molecules that orchestrate mechanotransduction by hair cells

“…However, high-resolution electron microscopic (EM) images suggest that tip links are stiff and buckle under strain [79]. Modeling of the classical C-cadherin extracellular domain suggests high stiffness in the presence of Ca 2+ and limited elasticity without Ca 2+ [80, 81]. Modeling studies based on the structure of the CDH23 EC1-EC2 fragment attempted to provide estimates of the tip-link stiffness [72].…”

“…Mechanical properties of tip-link cadherins have not been studied, but the nanomechanics of C-cadherin has been experimentally analyzed [81]. Single molecule force spectroscopy confirms Ca 2+ induces ectodomain rigidification.…”