Munc18-1 is critical for plasma membrane localization of syntaxin1 but not of SNAP-25 in PC12 cells
Munc18-1 is critical for plasma membrane localization of syntaxin1 but not of SNAP-25 in PC12 cells
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DOI：
10.1091/mbc.e07-07-0662
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发表时间：
2008-02-01
影响因子：
3.3
通讯作者：
Sugita, Shuzo
中科院分区：
文献类型：
--
作者：
Arunachalam, Lakshmanan;Han, Liping;Sugita, Shuzo
虽然Munc 18 -1最初被鉴定为syntaxin 1相互作用蛋白，但这种相互作用的生理意义仍不清楚。事实上，最近对Munc 18 -1突变体的研究表明，Munc 18 -1在分泌囊泡的对接中起着关键作用，不依赖于syntaxin 1的调节。在这里，我们研究了Munc 18 -1在突触融合蛋白1定位的作用，通过产生稳定的神经内分泌细胞系，其中Munc 18 -1强烈下调。在这些细胞中，分泌能力以及致密核心囊泡的对接显著降低。更重要的是，不仅syntaxin 1的表达水平降低，但syntaxin 1在质膜上的定位也受到严重干扰。错误定位的syntaxin 1主要位于细胞的核周区域，在那里它是高度共定位与分泌颗粒蛋白II，致密核心囊泡的标志蛋白。相反，SNAP-25的表达水平和质膜定位不受影响。此外，突触融合蛋白1的定位和分泌能力恢复后，转染介导的重新引入Munc 18 -1。我们的研究结果表明，内源性Munc 18 -1在神经内分泌细胞中突触融合蛋白1的质膜定位中起着至关重要的作用，因此需要解释Munc 18 -1突变表型是在错误定位的突触融合蛋白1。
Although Munc18-1 was originally identified as a syntaxin1-interacting protein, the physiological significance of this interaction remains unclear. In fact, recent studies of Munc18-1 mutants have suggested that Munc18-1 plays a critical role for docking of secretory vesicles, independent of syntaxin1 regulation. Here we investigated the role of Munc18-1 in syntaxin1 localization by generating stable neuroendocrine cell lines in which Munc18-1 was strongly down-regulated. In these cells, the secretion capability, as well as the docking of dense-core vesicles, was significantly reduced. More importantly, not only was the expression level of syntaxin1 reduced, but the localization of syntaxin1 at the plasma membrane was also severely perturbed. The mislocalized syntaxin1 resided primarily in the perinuclear region of the cells, in which it was highly colocalized with Secretogranin II, a marker protein for dense-core vesicles. In contrast, the expression level and the plasma membrane localization of SNAP-25 were not affected. Furthermore, the syntaxin1 localization and the secretion capability were restored upon transfection-mediated reintroduction of Munc18-1. Our results indicate that endogenous Munc18-1 plays a critical role for the plasma membrane localization of syntaxin1 in neuroendocrine cells and therefore necessitates the interpretation of Munc18-1 mutant phenotypes to be in terms of mislocalized syntaxin1.