Junctional Adhesion Molecule A Interacts with Afadin and PDZ-GEF2 to Activate Rap1A, Regulate β1 Integrin Levels, and Enhance Cell Migration
Junctional Adhesion Molecule A Interacts with Afadin and PDZ-GEF2 to Activate Rap1A, Regulate β1 Integrin Levels, and Enhance Cell Migration
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DOI：
10.1091/mbc.e08-10-1014
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发表时间：
2009-04-01
影响因子：
3.3
通讯作者：
Parkos, Charles A.
中科院分区：
文献类型：
--
作者：
Severson, Eric A.;Lee, Winston Y.;Parkos, Charles A.
连接粘附分子-A (JAM-A) 是一种跨膜紧密连接蛋白，已被证明可以通过不完全了解的机制调节屏障功能和细胞迁移。我们之前已经证明，JAM-A 通过膜远端免疫球蛋白样环的二聚化和 C 端突触后密度 95/椎间盘大/闭塞带 (PDZ) 结合基序来调节细胞迁移。二聚化的破坏导致上皮细胞迁移减少，继发于β1整合素和活性Rap1水平的降低。在此，我们报道 JAM-A 在物理和功能上与上皮细胞中包含 PDZ 结构域的分子 Afadin 和 PDZ-鸟嘌呤核苷酸交换因子 (GEF) 2 相关，但与闭锁小带 (ZO)-1 无关，并且这些相互作用介导由外向内的信号转导事件。 Afadin 和 PDZ-GEF2 均与 JAM-A 共定位并共免疫沉淀。此外，PDZ-GEF2 与 Afadin 的关联依赖于 JAM-A 的表达。 JAM-A、Afadin 或 PDZ-GEF2 的缺失，但 ZO-1 或 PDZ-GEF1 的缺失，同样会降低活化 Rap1、β1 整联蛋白的细胞水平和上皮细胞迁移。观察到的功能效应是 Rap1A 水平降低的继发因素，因为 Rap1A（而非 Rap1B）的敲低会导致 β1 整合素水平降低并减少细胞迁移。这些发现表明，JAM-A 二聚化促进与 Afadin 和 PDZ-GEF2 形成复合物，从而激活 Rap1A，从而调节 β1 整合素水平和细胞迁移。
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 beta 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, beta 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 beta 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 beta 1 integrin levels and cell migration.