Cadherin cis and trans interactions are mutually cooperative

Abstract

Significance Cadherin-mediated intercellular adhesion involves both cis and trans interactions between cadherin extracellular domains. It has been hypothesized that the combination of these cis and trans interactions is critical for the formation of macroscopic adhesions. Here, we have used single-molecule microscopy to show that cis and trans interactions are mutually cooperative, in that cis interactions directly increase the stability of trans bonds and trans interactions similarly increase the stability of cis bonds. Furthermore, we show that this cooperativity directly results in cadherin accumulation to form macroscopic intermembrane junctions. This cooperativity occurs at the molecular level and suggests that trans dimers are allosterically stabilized by individual cis interactions. These results provide mechanistic insight into cadherin junction assembly, independent of cytoplasmic interactions. Cadherin transmembrane proteins are responsible for intercellular adhesion in all biological tissues and modulate tissue morphogenesis, cell motility, force transduction, and macromolecular transport. The protein-mediated adhesions consist of adhesive trans interactions and lateral cis interactions. Although theory suggests cooperativity between cis and trans bonds, direct experimental evidence of such cooperativity has not been demonstrated. Here, the use of superresolution microscopy, in conjunction with intermolecular single-molecule Förster resonance energy transfer, demonstrated the mutual cooperativity of cis and trans interactions. Results further demonstrate the consequent assembly of large intermembrane junctions, using a biomimetic lipid bilayer cell adhesion model. Notably, the presence of cis interactions resulted in a nearly 30-fold increase in trans-binding lifetimes between epithelial-cadherin extracellular domains. In turn, the presence of trans interactions increased the lifetime of cis bonds. Importantly, comparison of trans-binding lifetimes of small and large cadherin clusters suggests that this cooperativity is primarily due to allostery. The direct quantitative demonstration of strong mutual cooperativity between cis and trans interactions at intermembrane adhesions provides insights into the long-standing controversy of how weak cis and trans interactions act in concert to create strong macroscopic cell adhesions.