Junctional Adhesion Molecule A Interacts with Afadin and PDZ-GEF2 to Activate Rap1A, Regulate β1 Integrin Levels, and Enhance Cell Migration

Abstract

Junctional adhesion molecule-A (JAM-A) is a transmembrane tight junction protein that has been shown to regulate barrier function and cell migration through incompletely understood mechanisms. We have previously demonstrated that JAM-A regulates cell migration by dimerization of the membrane-distal immunoglobulin-like loop and a C-terminal postsynaptic density 95/disc-large/zona occludens (PDZ) binding motif. Disruption of dimerization resulted in decreased epithelial cell migration secondary to diminished levels of ␤1 integrin and active Rap1. Here, we report that JAM-A is physically and functionally associated with the PDZ domain-containing molecules Afadin and PDZ-guanine nucleotide exchange factor (GEF) 2, but not zonula occludens (ZO)-1, in epithelial cells, and these interactions mediate outside-in signaling events. Both Afadin and PDZ-GEF2 colocalized and coimmunoprecipitated with JAM-A. Furthermore, association of PDZ-GEF2 with Afadin was dependent on the expression of JAM-A. Loss of JAM-A, Afadin, or PDZ-GEF2, but not ZO-1 or PDZ-GEF1, similarly decreased cellular levels of activated Rap1, ␤1 integrin protein, and epithelial cell migration. The functional effects observed were secondary to decreased levels of Rap1A because knockdown of Rap1A, but not Rap1B, resulted in decreased ␤1 integrin levels and reduced cell migration. These findings suggest that JAM-A dimerization facilitates formation of a complex with Afadin and PDZ-GEF2 that activates Rap1A, which regulates ␤1 integrin levels and cell migration.

Loss of AF6/afadin, a marker of poor outcome in breast cancer, induces cell migration, invasiveness and tumor growth

“…This result tends to generalize the impact of loss of afadin expression in tumors. However, for an unclear reason, this result contrasts with a recently published work (Severson et al, 2009). This could be attributed to the origin of the cell line used.…”

“…More recently, afadin has been described as a regulator of cell migration. However, afadin contribution as positive or negative regulator of migration is still largely controversial (Lorger and Moelling, 2006;Miyata et al, 2009;Severson et al, 2009). The role of afadin in oncogenesis has been documented in some acute lymphoid and myeloid leukemias.…”

miR-145-dependent targeting of Junctional Adhesion Molecule A and modulation of fascin expression are associated with reduced breast cancer cell motility and invasiveness

“…Thus, in our study, the role of JAM-A in growth-factor-mediated signalling (Naik et al, 2003) may have contributed to the modulation of cell motility. Although our data are in line with the majority of published studies, suggesting an inhibition of cell motility and invasiveness upon loss of JAM-A function (Naik et al, 2003;Cera et al, 2009;McSherry et al, 2009;Severson et al, 2009), they also show that JAM-A function is context-dependent, and may be modulated by the presence or absence of cellcell-contact and growth factors, integrin expression patterns and matrix substrates. In addition to JAM-A, miR-145-dependent regulation of fascin may have contributed to the migratory phenotype.…”

Junctional adhesion molecule-A is co-expressed with HER2 in breast tumors and acts as a novel regulator of HER2 protein degradation and signaling

“…This adds to increasing evidence that JAM-A regulates protein turnover through proteasomal degradation, as JAM-A knockdown has previously been shown to similarly reduce the protein expression of β1-integrin (McSherry et al, 2011;McSherry et al, 2009;Severson et al, 2009) and αV-and α5-integrin (McSherry et al, 2011). Further studies are required to determine the specific proteasomal components implicated, but it is intriguing to speculate that JAM-A may also have the capability to likewise regulate the expression of other growth factor receptors/adhesion molecules relevant to breast cancer progression.…”