Phosphorylation of N-Cadherin-associated Cortactin by Fer Kinase Regulates N-Cadherin Mobility and Intercellular Adhesion Strength

Abstract

Cortactin regulates the strength of nascent N-cadherin-mediated intercellular adhesions through a tyrosine phosphorylation-dependent mechanism. Currently, the functional significance of cortactin phosphorylation and the kinases responsible for the regulation of adhesion strength are not defined. We show that the nonreceptor tyrosine kinase Fer phosphorylates cadherin-associated cortactin and that this process is involved in mediating intercellular adhesion strength. In wild-type fibroblasts N-cadherin ligation-induced transient phosphorylation of Fer, indicating that junction formation activates Fer kinase. Tyrosine phosphorylation of cortactin after N-cadherin ligation was strongly reduced in fibroblasts expressing only catalytically inactive Fer (D743R), compared with wild-type cells. In wild-type cells, Ncadherin-coated bead pull-off assays induced fourfold greater endogenous N-cadherin association than in D743R cells. Fluorescence recovery after photobleaching showed that GFP-N-cadherin mobility at nascent contacts was 50% faster in wild-type than D743R cells. In shear wash-off assays, nascent intercellular adhesion strength was twofold higher in wild-type than D743R cells. Cortactin recruitment to adhesions was independent of Fer kinase activity, but was impacted by N-cadherin ligation-provoked Rac activation. We conclude that N-cadherin ligation induces Rac-dependent cortactin recruitment and Fer-dependent cortactin phosphorylation, which in turn promotes enhanced mobilization and interaction of surface expressed N-cadherin in contacting cells.

Small Rho GTPase-mediated actin dynamics at endothelial adherens junctions

“…22 In line with this finding, a rapid increase in Rac1 activity upon VE-cadherin homotypic adhesion was detected biochemically, which was induced using beads coated with the VE-cadherin ectodomain. Thus, as was shown for E-, M-and N-cadherin, [64][65][66][67] we showed that VEcadherin can signal in an outside-in fashion to activate Rac1, providing a bidirectional feedback mechanism. 22 Of note, ongoing local remodeling of cell-cell junctions is observed even in apparently stable endothelial monolayers.…”

Oncogenic Src requires a wild-type counterpart to regulate invadopodia maturation

“…The variations in binding intensity that arise in SYF and SYF +/+ cells expressing Src527F indicate a fundamental mechanistic difference in the concentration of phosphotyrosine binding sites for several SH2-domain-containing proteins that are likely to have key roles in invadopodia maturation. Interestingly, tyrosine-phosphorylated cortactin is known to interact with several high-intensity 'hits', including Arg, Abl (Boyle et al, 2007), Fer (El Sayegh et al, 2005), Crk (Bougneres et al, 2004) and Nck (Tehrani et al, 2007), consistent with its role in invadopodia maturation. Experiments to elucidate additional proteins involved in these signaling complexes are currently underway.…”

Making and breaking contacts: the cellular biology of cadherin regulation

“…It should be noted, however, that many non-specific manuevers that inhibit endocytosis can influence cadherin function by alternative mechanisms. For example, hypertonic stress induces tyrosine phosphorylation of cortactin [22], which contributes to cadherin junction integrity [23,24] (Helwani et al, unpublished). These disparate models thus remain to be resolved and warrant further investigation.…”