Jab1, a novel protease-activated receptor-2 (PAR-2)-interacting protein, is involved in PAR-2-induced activation of activator protein-1
Jab1, a novel protease-activated receptor-2 (PAR-2)-interacting protein, is involved in PAR-2-induced activation of activator protein-1
复制标题
DOI：
10.1074/jbc.m510784200
复制
发表时间：
2006-03-24
影响因子：
4.8
通讯作者：
Reiser, G
中科院分区：
文献类型：
--
作者：
Luo, WB;Wang, YF;Reiser, G
蛋白酶激活受体-2（PAR-2）是胰蛋白酶和类胰蛋白酶的G蛋白偶联受体，在多种系统中发挥重要的生理和病理功能。然而，与PAR-1不同，PAR-2介导的细胞内信号转导几乎不为人所知。在这里，使用酵母双杂交筛选与人脑cDNA文库，我们确定了一个相互作用的合作伙伴的人PAR-2，Jun激活域结合蛋白1（Jab 1）。谷胱甘肽S-转移酶下拉试验在体外，并在体内通过免疫共沉淀试验证实的相互作用。Jab 1也被证明是共定位与PAR-2在两个转染HEK 293细胞和正常的原代人星形胶质细胞的双重免疫荧光染色。进一步的实验表明，PAR-2的多个胞内结构域是与Jab 1相互作用所必需的。然后，我们表明，PAR-2的激动剂刺激破坏的相互作用，这可以防止由抑制剂的受体内吞作用氧化苯胂，但不是由溶酶体蛋白酶抑制剂ZPAD。重要的是，我们发现PAR-2的激活诱导Jab 1从质膜到胞质溶胶的重新分布，但不影响Jab 1的表达。此外，Jab 1介导PAR-2诱导的c-Jun活化，随后增加激活蛋白-1的活化。使用Jab 1小干扰RNA的功能丧失研究表明，Jab 1敲低阻断PAR-2诱导的激活蛋白-1激活。总之，我们的数据表明，Jab 1是一个重要的效应，介导一种新的信号转导途径PAR-2依赖的基因表达。
Protease-activated receptor-2 (PAR-2), a G protein-coupled receptor for trypsin and tryptase, exerts important physiological and pathological functions in multiple systems. However, unlike PAR-1, the PAR-2-mediated intracellular signal transductions are hardly known. Here, using yeast two-hybrid screening with a human brain cDNA library, we identified an interacting partner of human PAR-2, the Jun activation domain-binding protein 1 (Jab1). The interaction was confirmed by glutathione S-transferase pull-down assays in vitro, and by co-immunoprecipitation assays in vivo. Jab1 was also shown to be colocalized with PAR-2 in both transfected HEK293 cells and in normal primary human astrocytes by double immunofluorescence staining. Further experiments demonstrated that multiple intracellular domains of PAR-2 are required for the interaction with Jab1. We then showed that agonist stimulation of PAR-2 disrupted the interaction, which could be prevented by the inhibitor of receptor endocytosis phenylarsine oxide, but not by the lysosomal protease inhibitor ZPAD. Importantly, we found that activation of PAR-2 induced the redistribution of Jab1 from the plasma membrane to the cytosol, but did not influence expression of Jab1. Furthermore, Jab1 mediated PAR-2-induced c-Jun activation, which was followed by increased activation of activator protein-1. Loss-of-function studies, using Jab1 small interfering RNA, demonstrated that Jab1 knockdown blocked PAR-2-induced activator protein-1 activation. Taken together, our data demonstrate that Jab1 is an important effector that mediates a novel signal transduction pathway for PAR-2-dependent gene expression.