A novel calcium-regulated membrane guanylate cyclase transduction system in the olfactory neuroepithelium
A novel calcium-regulated membrane guanylate cyclase transduction system in the olfactory neuroepithelium
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DOI：
10.1021/bi0108406
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发表时间：
2001-10-09
期刊：
影响因子：
2.9
通讯作者：
Sharma, RK
中科院分区：
文献类型：
--
作者：
Duda, T;Jankowska, A;Sharma, RK
这份报告定义了一个钙调节的膜鸟苷酸环化酶转导系统的嗅觉感觉神经元，这是气味转导的网站纤毛的身份。大鼠神经上皮层的膜部分表现出钙离子依赖性鸟苷酸环化酶活性，这是消除通过添加EGTA。这表明，环化酶并不代表一个杆外段鸟苷酸环化酶（ROS-GC），这是由游离Ca 2+抑制。这种解释得到了Ca 2+结合蛋白GCAP（鸟苷酸环化酶激活蛋白）研究的支持，GCAP在缺乏Ca 2+的情况下刺激光感受器ROS-GC。他们没有刺激嗅觉神经上皮细胞膜鸟苷酸环化酶。嗅神经上皮细胞含有一种钙结合蛋白，神经钙蛋白，它刺激环化酶的钙依赖性的方式。该环化酶是从神经上皮细胞克隆的，发现其结构与先前克隆的称为GC-D的环化酶相同。环化酶在异源细胞系统中表达，并在重组神经钙蛋白存在下用其Ca 2+依赖性活性重建。重组的环化酶模仿天然酶。免疫细胞化学研究表明，鸟苷酸环化酶与神经钙蛋白共存于纤毛的顶端区域。缺失分析表明，神经钙蛋白调节结构域位于环化酶的C-末端区域。研究结果建立了一种新的Ca 2+依赖性膜鸟苷酸环化酶转导系统的嗅觉上皮纤毛的生化，分子和功能的身份，提示嗅觉神经上皮鸟苷酸环化酶调节的机制从根本上不同于光转导相关的ROS-GC。
This report defines the identity of a calcium-regulated membrane guanylate cyclase transduction system in the cilia of olfactory sensory neurons, which is the site of odorant transduction. The membrane fraction of the neuroepithelial layer of the rat exhibited Ca2+-dependent guanylate cyclase activity, which was eliminated by the addition of EGTA. This indicated that the cyclase did not represent a rod outer segment guanylate cyclase (ROS-GC), which is inhibited by free Ca2+. This interpretation was supported by studies with the Ca2+ binding proteins, GCAPs (guanylate cyclase activating proteins), which stimulate photoreceptor ROS-GC in the absence of Ca2+. They did not stimulate the olfactory neuroepithelial membrane guanylate cyclase. The olfactory neuroepithelium contained a Ca2+ binding protein, neurocalcin, which stimulated the cyclase in a Ca2+-dependent fashion. The cyclase was cloned from the neuroepithelium and was found to be identical in structure to that of the previously cloned cyclase termed GC-D. The cyclase was expressed in a heterologous cell system, and was reconstituted with its Ca2+-dependent activity in the presence of recombinant neurocalcin. The reconstituted cyclase mimicked the native enzyme. Immunocytochemical studies showed that the guanylate cyclase coexists with neurocalcin in the apical region of the cilia. Deletion analysis showed that the neurocalcin-regulated domain resides at the C-terminal region of the cyclase. The findings establish the biochemical, molecular, and functional identity of a novel Ca2+-dependent membrane guanylate cyclase transduction system in the cilia of the olfactory epithelium, suggesting a mechanism of the olfactory neuroepithelial guanylate cyclase regulation fundamentally distinct from the phototransduction-linked ROS-GC.