Meltrin β (ADAM19) mediates ectodomain shedding of Neuregulin β1 in the Golgi apparatus: fluorescence correlation spectroscopic observation of the dynamics of ectodomain shedding in living cells
Meltrin β (ADAM19) mediates ectodomain shedding of Neuregulin β1 in the Golgi apparatus: fluorescence correlation spectroscopic observation of the dynamics of ectodomain shedding in living cells
复制标题
DOI：
10.1111/j.1365-2443.2007.01060.x
复制
发表时间：
2007-03
期刊：
影响因子：
2.1
通讯作者：
Tomoichi Yokozeki;S. Wakatsuki;K. Hatsuzawa;R. Black;I. Wada;A. Sehara-Fujisawa
中科院分区：
文献类型：
--
作者：
Tomoichi Yokozeki;S. Wakatsuki;K. Hatsuzawa;R. Black;I. Wada;A. Sehara-Fujisawa
膜锚定的神经调节蛋白β-1以可溶性因子的形式脱落其胞外结构域。已知有两种属于去整合素和金属蛋白酶(ADAM)家族的酶能裂解神经调节蛋白β1。一种是肿瘤坏死因子-α转换酶(TACE/ADAM17)。另一家是美林β(ADAM19)。与我们的预期不同的是，亚当蛋白酶的脱落发生在细胞表面，在这里，我们发现Meltrinβ介导了高尔基体中神经调节蛋白β1的胞外脱落。在发育中的感觉神经元中，Meltrinβ定位于高尔基体及其周围。亚细胞分级显示，Meltrinβ在高尔基体富集组分中产生了可溶性的神经调节蛋白β1，而在较轻的组分中回收了经TACE裂解的神经调节蛋白β1。为了检验在活细胞的高尔基体中是否发生了Meltrinβ介导的胞外结构域脱落，我们利用了切割产物与膜锚定前体蛋白不同的扩散特性。荧光相关光谱是目前最灵敏的检测毫微∼在体内亚毫秒扩散的方法。蛋白酶活性的Meltrinβ导致高尔基体中绿色荧光蛋白标记的神经调节蛋白β1的自相关功能发生变化，表明该细胞器中的神经调节蛋白β1分子从膜锚定形式转变为可溶性形式。高尔基体是梅尔特林β加工神经调节蛋白β1的场所。
Membrane‐anchored Neuregulin β1 sheds its ectodomain as soluble factors. Two proteases that belong to a disintegrin and metalloprotease (ADAM) family are known to cleave Neuregulin β1. One is tumor necrosis factor‐α converting enzyme (TACE/ADAM17). The other is Meltrin β (ADAM19). Against our expectation that shedding by ADAM proteases occurs at the cell surface, here we found that Meltrin β mediates the ectodomain shedding of Neuregulin β1 in the Golgi apparatus. Meltrin β was localized in and around the Golgi apparatus in developing sensory neurons. Subcellular fractionation revealed that Meltrin β generated soluble Neuregulin β1 in Golgi‐enriched fractions while TACE‐cleaved Neuregulin β1 was recovered in lighter fractions. To examine whether Meltrin β‐mediated ectodomain shedding occurs in the Golgi apparatus in living cells, we took advantage of different diffusion properties of cleavage products from those of membrane‐anchored precursor proteins. Fluorescence correlation spectroscopy (FCS) is the most sensitive method to determine milli∼submillisecond diffusion in vivo. Protease‐active Meltrin β caused a shift in autocorrelation function in FCS of green fluorescent protein (GFP)‐tagged Neuregulin β1 in the Golgi apparatus, suggesting a conversion of Neuregulin β1 molecules from membrane‐anchored to soluble forms in that organelle. The Golgi apparatus is a site of processing Neuregulin β1 by Meltrin β.