Homophilic adhesion of E-cadherin occurs by a co-operative two-step interaction of N-terminal domains.

Abstract

Cluster formation of E‐cadherin on the cell surface is believed to be of major importance for cell‐cell adhesion. To mimic this process the extracellular part of mouse E‐cadherin (ECAD) was recombinantly fused to the assembly domain of rat cartilage oligomeric matrix protein (COMP), resulting in the chimeric protein ECAD‐COMP. The COMP domain formed a five‐stranded alpha‐helical coiled‐coil. This enabled the formation of a pentameric ECAD with bundled C‐termini and free N‐termini. The pentameric protein construct ECAD‐COMP and the monomeric ECAD were expressed in human embryonal kidney 293 cells. Electron microscopy, analytical ultracentrifugation, solid phase binding and cell attachment assays revealed that pentamers showed strong self‐association and cell attachment, whereas monomers exhibited no activity. At the high internal concentration in the pentamer the N‐terminal EC1 domains of two E‐cadherin arms interact to form a ring‐like structure. Then the paired domains interact with a corresponding pair from another pentamer. None of the four other extracellular domains of E‐cadherin is involved in this interaction. Based on these results, an in vivo mechanism is proposed whereby two N‐terminal domains of neighbouring E‐cadherins at the cell surface first form a pair, which binds with high affinity to a similar complex on another cell. The strong dependence of homophilic interactions on C‐terminal clustering points towards a regulation of E‐cadherin mediated cell‐cell adhesion via lateral association.

Lateral self‐assembly of E‐cadherin directed by cooperative calcium binding

“…2, right). This observation on Ca 2+ -loaded Ecadl2 is consistent with previous crystallographic and electron microscopy studies showing that calcium extends and rigidities the extracellular segment of E-cadherin [7] and that lateral association of adjacent extracellular segments occurs at the N-terminal tip of the molecule [8].…”

Cadherin Adhesion: Mechanisms and Molecular Interactions

“…Interestingly, this is the approximate length of a cadherin extracellular domain if all the repeats would be in a straight line, one after the other. However, as noted earlier, electron microscopy of the extracellular domain of Ecadherin and the crystal structure of the full-length extracellular domain of C-cadherin depict the cadherin extracellular domain as a bent rod (Pokutta et al 1994;Tomschy et al 1996;Ahrens et al 2002;Boggon et al 2002). Due to the curve of the extracellular domain, the trans-adhesion dimer proposed by Boggon et al (2002) would occur between surfaces approximately 25 nm apart.…”

“…First, further work on ECADCOMP showed that adhesive pentamers oligomerized through the N-terminal regions of E-cadherin (Tomschy et al 1996;Koch et al 1999;Pertz et al 1999). It is unclear, however, exactly which parts of the N-terminus are involved in adhesion.…”

“…In their studies on Ca ++ -dependent adhesion, Tomschy et al (1996) and Pertz et al (1999) describe association of E-cadherin extracellular domains within a single pentamer before extracellular domains in different pentamers adhere. They base this on the observation that ring-like structures are found in some isolated pentamers at high Ca ++ concentrations, but they are always seen as the adherent cadherins when two or more pentamers associate.…”

Direct molecular force measurements of multiple adhesive interactions between cadherin ectodomains

“…Because the linear dimension of the full-length extracellular domain is 22 Ϯ 1 nm (9,21,22), and the length of the DSIDA linker is ca. 1 nm, the proteins should interdigitate completely at D Ͻ 25 Ϯ 1 nm (9,21,22). We therefore attribute the repulsive force at D Ͻ 25 nm to steric interactions between the proteins and the opposing bilayer surface.…”