Regulation of E-cadherin/catenin association by tyrosine phosphorylation

Alteration of cadherin-mediated cell-cell adhesion is frequently associated to tyrosine phosphorylation of p120- and beta-catenins. We have examined t he role of this modification in these proteins in the control of beta-caten in/E-cadherin binding using in vitro assays with recombinant proteins. Reco mbinant pp60(c-src) efficiently phosphorylated both catenins in vitro, with stoichiometries of 1.5 and 2.0 mol of phosphate/mol of protein for beta-ca tenin and p120-catenin, respectively. pp60(c-src) phosphorylation had oppos ing effects on the affinities of beta-catenin and p120 for the cytosolic do main of E-cadherin; it decreased tin the case of beta-catenin) or increased (for p120) catenin/E-cadherin binding. How ever, a role for p120-catenin i n the modulation of beta-catenin/E-cadherin binding was not observed, since addition of phosphorylated p120-catenin did not modify the affinity of pho sphorylated (or unphosphorylated) beta-catenin far E-cadherin, The phosphor ylated Tyr residues were identified as Tyr-86 and Tyr-654. Experiments usin g point mutants in these two residues indicated that, although Tyr-86 was a better substrate for pp60(c-src), only modification of Tyr-654 was relevan t for the interaction with E-cadherin. Transient transfections of different mutants demonstrated that Tyr-654 is phosphorylated in conditions in which adherens junctions are disrupted and evidenced that binding of beta-cateni n to E-cadherin in vivo is controlled by phosphorylation of beta-catenin Ty r-654.