Interaction of neuronal nitric-oxide synthase with α1-syntrophin in rat brain
Interaction of neuronal nitric-oxide synthase with α1-syntrophin in rat brain
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DOI：
10.1074/jbc.274.17.11736
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发表时间：
1999-04-23
影响因子：
4.8
通讯作者：
Nagai, K
中科院分区：
文献类型：
--
作者：
Hashida-Okumura, A;Okumura, N;Nagai, K
神经元一氧化氮合酶（nNOS）具有PSD-95/ Dlg/ZO-1 （PDZ）结构域，可与多种蛋白相互作用。已知nNOS与哺乳动物大脑中的PSD-95和相关蛋白PSD-93相互作用，并与骨骼肌中的α 1-syntrophin相互作用。本研究利用谷胱甘肽s -转移酶大鼠nNOS融合蛋白与Sepharose偶联的亲和柱从牛脑中纯化了一种nNOS相互作用蛋白，并通过微测序鉴定其为α 1-syntrophin。用抗这些蛋白的抗体对大鼠胚胎脑神经细胞原代培养物进行免疫染色，发现nNOS和α 1-syntrophin在神经元细胞体和神经突中共定位。免疫组化分析显示，大鼠下丘脑视交叉上核（SCN）和室旁核高度表达nNOS-和α 1-syntrophin样免疫反应物质。在SCN中，共聚焦显微镜检测到nNOS-和α 1-syntrophin样免疫反应物质共定位于相同的神经元。这些结果表明，脑内nNOS与SCN和室旁核特定神经元中的α 1-syntrophin相互作用，这种相互作用可能在这些神经元的功能中起生理作用。
Neuronal nitric-oxide synthase (nNOS) has a PSD-95/ Dlg/ZO-1 (PDZ) domain that can interact with multiple proteins. nNOS has been known to interact with PSD-95 and a related protein, PSD-93, in brain and with alpha 1-syntrophin in skeletal muscle in mammals. In this study, we have purified an nNOS-interacting protein from bovine brain using an affinity column made of Sepharose conjugated with glutathione S-transferase-rat nNOS fusion protein and identified it as alpha 1-syntrophin by microsequencing. Immunostaining of primary cultures of rat embryonic brain neuronal cells with antibodies against these proteins showed that nNOS and alpha 1-syntrophin were colocalized in neuronal cell bodies and neurites. Immunohistochemical analysis indicated that the nNOS- and alpha 1-syntrophin-like immunoreactive substances were highly expressed in the rat hypothalamic suprachiasmatic nucleus (SCN) and paraventricular nucleus. In the SCN, nNOS- and alpha 1-syntrophin-like immunoreactive substances were colocalized in the same neurons as detected by confocal microscopy. These results indicate that nNOS in brain interacts with alpha 1-syntrophin in specific neurons of the SCN and paraventricular nucleus and that this interaction might play a physiological role in functions of these neurons.