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Study of secretory granule proteins associated with syntaxin 2 in the rat parotid gland

大鼠腮腺中突触蛋白2相关分泌颗粒蛋白的研究

基本信息

批准号：
14571780
负责人：
IMAI Akane
金额：
$ 2.05万
依托单位：
The Nippon Dental University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2004
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14571780/
关键词：
parotid gland secretory granule Munc18 syntaxin Rab27 Slac2-c Slp4-a amylase release Munc18-3 SNARE Rabタンパク質 VAMP 細胞内局在相互作用

项目摘要

Small GTPase Rab is a large family of putative membrane trafficking proteins, and each member is thought to regulate a specific type(s) of membrane trafficking. However, little is known about the involvement of Rab protein(s) in secretory granule exocytosis in exocrine cells or the molecular mechanism underlying this process. Here we show that Rab27B, a closely related isoform of Rab27A that regulates lysosome-related granule exocytosis in cytotoxic T lymphocytes, is abundantly expressed on amylase-containing secretory granules in rat parotid gland acinar cells. We also identify the putative Rab27B effector protein, Slac2-c (Slp homologue lacking C2 domains-c)/MyRIP, which was originally described as a myosin Va/VIIa and actin binding protein, in rat parotid glands. The results of subcellular fractionation, immunoprecipitation, and immunohistochemical studies indicate that the Rab27B-Slac2-c complex is formed on secretory granules in vivo. The introduction of either a specific Rab27 binding domain (i.e., a recombinant Slp homology domain of Slac2-b that specifically binds Rab27A/B but not other Rabs) or functionally blocking antibodies that specifically disrupt Rab27B-Slac2-c complex in vitro strongly inhibited isoproterenol-stimulated amylase release from streptolysin O-permeabilized parotid acinar cells. Our results indicate that the Rab27B-Slac2-c complex is an important constituent of secretory granule exocytosis in parotid acinar cells.

小GTTRab是一个推定的膜运输蛋白的大家族，并且每个成员被认为调节特定类型的膜运输。然而，很少有人知道Rab蛋白（S）参与外分泌细胞中的分泌颗粒胞吐作用或该过程的分子机制。在这里，我们表明，Rab 27 B，Rab 27 A的一个密切相关的亚型，调节溶酶体相关的颗粒胞吐在细胞毒性T淋巴细胞，大量表达在大鼠腮腺腺泡细胞的含淀粉酶的分泌颗粒。我们还在大鼠腮腺中鉴定了推定的Rab 27 B效应蛋白Slac 2-c（缺乏C2结构域-c的Slp同源物）/MyRIP，它最初被描述为肌球蛋白Va/VIIa和肌动蛋白结合蛋白。亚细胞分级分离、免疫沉淀和免疫组化研究的结果表明，Rab 27 B-Slac 2-c复合物在体内的分泌颗粒上形成。引入特异性Rab 27结合结构域（即，特异性结合Rab 27 A/B但不结合其它Rabs的Slac 2-B的重组Slp同源结构域）或在体外特异性破坏Rab 27 B-Slac 2-c复合物的功能性阻断抗体强烈抑制异丙肾上腺素刺激的淀粉酶从链球菌溶血素O-透化的腮腺腺泡细胞释放。我们的研究结果表明，Rab 27 B-Slac 2-c复合物是腮腺腺泡细胞分泌颗粒胞吐的重要组成部分。