Monteiro Ana C, Luissint Anny-Claude, Sumagin Ronen, Lai Caroline, Vielmuth Franziska, Wolf Mattie F, Laur Oskar, Reiss Kerstin, Spindler Volker, Stehle Thilo, Dermody Terence S, Nusrat Asma, Parkos Charles A
Epithelial Pathobiology and Mucosal Inflammation Research Unit, Department of Pathology and Laboratory Medicine, Emory University School of Medicine, Atlanta, GA 30322.
Department of Pathology, Microbiology, and Immunology, Vanderbilt University School of Medicine, Nashville, TN 37232Elizabeth B. Lamb Center for Pediatric Research, Vanderbilt University School of Medicine, Nashville, TN 37232.
Mol Biol Cell. 2014 May;25(10):1574-85. doi: 10.1091/mbc.E14-01-0018. Epub 2014 Mar 26.
Junctional adhesion molecule-A (JAM-A) is a tight junction-associated signaling protein that regulates epithelial cell proliferation, migration, and barrier function. JAM-A dimerization on a common cell surface (in cis) has been shown to regulate cell migration, and evidence suggests that JAM-A may form homodimers between cells (in trans). Indeed, transfection experiments revealed accumulation of JAM-A at sites between transfected cells, which was lost in cells expressing cis- or predicted trans-dimerization null mutants. Of importance, microspheres coated with JAM-A containing alanine substitutions to residues 43NNP45 (NNP-JAM-A) within the predicted trans-dimerization site did not aggregate. In contrast, beads coated with cis-null JAM-A demonstrated enhanced clustering similar to that observed with wild-type (WT) JAM-A. In addition, atomic force microscopy revealed decreased association forces in NNP-JAM-A compared with WT and cis-null JAM-A. Assessment of effects of JAM-A dimerization on cell signaling revealed that expression of trans- but not cis-null JAM-A mutants decreased Rap2 activity. Furthermore, confluent cells, which enable trans-dimerization, had enhanced Rap2 activity. Taken together, these results suggest that trans-dimerization of JAM-A occurs at a unique site and with different affinity compared with dimerization in cis. Trans-dimerization of JAM-A may thus act as a barrier-inducing molecular switch that is activated when cells become confluent.
连接粘附分子A（JAM-A）是一种与紧密连接相关的信号蛋白，可调节上皮细胞的增殖、迁移和屏障功能。已证明，在共同细胞表面（顺式）上的JAM-A二聚化可调节细胞迁移，并且有证据表明JAM-A可能在细胞之间形成同型二聚体（反式）。事实上，转染实验显示JAM-A在转染细胞之间的位点积累，而在表达顺式或预测的反式二聚化无效突变体的细胞中这种积累消失。重要的是，用含有预测反式二聚化位点内43NNP45残基丙氨酸取代的JAM-A包被的微球不会聚集。相反，用顺式无效JAM-A包被的珠子表现出增强的聚集，类似于野生型（WT）JAM-A观察到的情况。此外，原子力显微镜显示，与WT和顺式无效JAM-A相比，NNP-JAM-A中的缔合作用力降低。对JAM-A二聚化对细胞信号传导影响的评估表明，反式而非顺式无效JAM-A突变体的表达降低了Rap2活性。此外，能够进行反式二聚化的汇合细胞具有增强的Rap2活性。综上所述，这些结果表明，与顺式二聚化相比，JAM-A的反式二聚化发生在一个独特的位点，且具有不同的亲和力。因此，JAM-A的反式二聚化可能作为一种屏障诱导分子开关，在细胞汇合时被激活。
