Cadherins are calcium-dependent adhesion molecules responsible for the establishment of tight cell–cell contacts. p120 catenin (p120ctn) binds to the cytoplasmic domain of cadherins in the juxtamembrane region, which has been implicated in regulating cell motility. It has previously been shown that overexpression of p120ctn induces a dendritic morphology in fibroblasts (Reynolds, A.B., J. Daniel, Y. Mo, J. Wu, and Z. Zhang. 1996. Exp. Cell Res. 225:328–337.). We show here that this phenotype is suppressed by coexpression of cadherin constructs that contain the juxtamembrane region, but not by constructs lacking this domain. Overexpression of p120ctn disrupts stress fibers and focal adhesions and results in a decrease in RhoA activity. The p120ctn-induced phenotype is blocked by dominant negative Cdc42 and Rac1 and by constitutively active Rho-kinase, but is enhanced by dominant negative RhoA. p120ctn overexpression increased the activity of endogenous Cdc42 and Rac1. Exploring how p120ctn may regulate Rho family GTPases, we find that p120ctn binds the Rho family exchange factor Vav2. The behavior of p120ctn suggests that it is a vehicle for cross-talk between cell–cell junctions and the motile machinery of cells. We propose a model in which p120ctn can shuttle between a cadherin-bound state and a cytoplasmic pool in which it can interact with regulators of Rho family GTPases. Factors that perturb cell–cell junctions, such that the cytoplasmic pool of p120ctn is increased, are predicted to decrease RhoA activity but to elevate active Rac1 and Cdc42, thereby promoting cell migration.

The formation of cell-cell adherens junctions is a cadherin-mediated process associated with reorganization of the actin cytoskeleton. Because Rho family GTPases regulate actin dynamics, we investigated whether cadherin-mediated adhesion regulates the activity of RhoA, Rac1, and Cdc42. Confluent epithelial cells were found to have elevated Rac1 and Cdc42 activity but decreased RhoA activity when compared with low density cultures. Using a calcium switch method to manipulate junction assembly, we found that induction of cell-cell junctions increased Rac1 activity, and this was inhibited by E-cadherin function-blocking antibodies. Using the same calcium switch procedure, we found little effect on RhoA activity during the first hour of junction assembly. However, over several hours, RhoA activity significantly decreased. To determine whether these effects are mediated directly through cadherins or indirectly through engagement of other surface proteins downstream from junction assembly, we used a model system in which cadherin engagement is induced without cell-cell contact. For these experiments, Chinese hamster ovary cells expressing C-cadherin were plated on the extracellular domain of C-cadherin immobilized on tissue culture plates. Whereas direct cadherin engagement did not stimulate Cdc42 activity, it strongly inhibited RhoA activity but increased Rac1 activity. Deletion of the C-cadherin cytoplasmic domain abolished these effects.

Recent evidence supports a role for EphB receptor tyrosine kinases as tumour suppressors in colorectal and prostate cancer. However, it is unclear how these receptors inhibit cancer cell tumorigenicity - an activity that is highly unusual for a family of receptor tyrosine kinases. Here, we report that the EphB4 receptor can behave as a tumour suppressor in a mouse xenograft model of breast cancer when stimulated by its ligand, ephrin-B2. In breast cancer cells, EphB4 activates an antioncogenic pathway involving Abl family tyrosine kinases and the Crk adaptor protein. This Abl-Crk pathway inhibits breast cancer cell viability and proliferation in addition to motility and invasion, and also downregulates the pro-invasive matrix metalloprotease, MMP-2. Consistent with these effects, EphB4 and the Abl-Crk pathway are constitutively active in non-transformed mammary epithelial cells. These findings identify a novel Eph receptor signalling pathway with tumour-suppressor activity and predict that therapeutic intervention to activate EphB4 signalling will inhibit tumour progression.

Two prominent characteristics of epithelial cells, apical-basal polarity and a highly ordered cytoskeleton, depend on the existence of precisely localized protein complexes associated with the apical plasma membrane, and on a separate machinery that regulates the spatial order of actin assembly. ERM (ezrin, radixin, moesin) proteins have been proposed to link transmembrane proteins to the actin cytoskeleton in the apical domain, suggesting a structural role in epithelial cells, and they have been implicated in signalling pathways. Here, we show that the sole Drosophila ERM protein Moesin functions to promote cortical actin assembly and apical-basal polarity. As a result, cells lacking Moesin lose epithelial characteristics and adopt invasive migratory behaviour. Our data demonstrate that Moesin facilitates epithelial morphology not by providing an essential structural function, but rather by antagonizing activity of the small GTPase Rho. Thus, Moesin functions in maintaining epithelial integrity by regulating cell-signalling events that affect actin organization and polarity. Furthermore, our results show that there is negative feedback between ERM activation and activity of the Rho pathway.

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Cadherins are transmembrane receptors that mediate cell-cell adhesion in epithelial cells. A number of changes occur during cadherin-mediated junction formation, one of which is a rearrangement of the actin cytoskeleton. Key regulators of actin cytoskeletal dynamics in cells are the Rho family of GTPases. We have demonstrated in previous studies that cadherin signaling suppresses RhoA activity and activates Rac1. The signaling events downstream of cadherins that modulate the activity of Rho family proteins remain unknown. Here we have identified a pathway by which RhoA becomes inactivated by cadherins. To determine whether cadherins regulate RhoA through activation of a GTPase-activating protein (GAP) for RhoA, we used constitutively active RhoA to isolate activated GAPs. Using this assay, we have identified the RhoA-specific GAP, p190RhoGAP, downstream from engaged cadherins. We found that cadherin engagement induced tyrosine phosphorylation of p190RhoGAP and increased its binding to p120RasGAP. The increased precipitation of p190RhoGAP with 63LRhoA was blocked by addition of PP2 suggesting that Src family kinases are required downstream from cadherin signaling. The inhibition of RhoA activity by cadherins was antagonized by expression of a dominant negative p190RhoGAP. Taken together, these data demonstrate that p190RhoGAP activity is critical for RhoA inactivation by cadherins.

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