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Epithelial cadherin (Ecadherin) is responsible for the intercellular cohesion of epithelial tissues. It forms lateral clusters within adherens cell-cell junctions, but its association state outside these clusters is unknown. Here, we use a quantitative Forster resonance energy transfer (FRET) approach to show that Ecadherin forms constitutive dimers and that these dimers exist independently of the actin cytoskeleton or cytoplasmic proteins. The dimers are stabilized by intermolecular contacts that occur along the entire length of Ecadherin, with the intracellular domains having a surprisingly strong favorable contribution. We further show that Ecadherin mutations and calcium depletion induce structural alterations that propagate from the N terminus all the way to the C terminus, without destabilizing the dimeric state. These findings provide context for the interpretation of Ecadherin adhesion experiments. They also suggest that early events of adherens junction assembly involve interactions between from preformed Ecadherin dimers.

Receptor Tyrosine Kinases (RTKs) conduct biochemical signals upon dimerization in the membrane plane. While RTKs are generally known to be activated in response to ligand binding, many of these receptors are capable of forming unliganded dimers that are likely important intermediates in the signaling process. All 58 RTKs consist of an extracellular domain, a transmembrane (TM) domain, and an intracellular domain which includes a juxtamembrane (JM) sequence and a kinase domain. Here we investigate directly the effect of the JM domain on unliganded dimer stability of FGFR3, a receptor that is critically important for skeletal development. The data suggest that FGFR3 unliganded dimers are stabilized by receptor-receptor contacts that involve the JM domains. The contribution is significant, as it is similar in magnitude to the stabilizing contribution of a pathogenic mutation and the repulsive contribution of the extracellular domain. Furthermore, we show that the effects of the JM domain and a TM pathogenic mutation on unliganded FGFR3 dimer stability are additive. We observe that the JM-mediated dimer stabilization occurs when the JM domain is linked to FGFR3 TM domain and not simply anchored to the plasma membrane. These results point to a coordinated stabilization of the unliganded dimeric state of FGFR3 by its JM and TM domains via a mechanism that is distinctly different from the case of another well studied receptor, EGFR.

Missense mutations that introduce or remove cysteine residues in receptor tyrosine kinases are believed to cause pathologies by stabilizing the active receptor tyrosine kinase dimers. However, the magnitude of this stabilizing effect has not been measured for full-length receptors. Here, we characterize the dimer stabilities of three full-length fibroblast growth factor receptor (FGFR) mutants harboring pathogenic cysteine substitutions: the C178S FGFR1 mutant, the C342R FGFR2 mutant, and the C228R FGFR3 mutant. We find that the three mutations stabilize the FGFR dimers. We further see that the mutations alter the configuration of the FGFR transmembrane dimers. Thus, both aberrant dimerization and perturbed dimer structure likely contribute to the pathological phenotypes arising due to these mutations.