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首页> 外文期刊>Protoplasma: An International Journal of Cell Biology >Detection and localization of a putative cyclic-GMP-activated channel protein in the protozoan ciliate Stentor coeruleus
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Detection and localization of a putative cyclic-GMP-activated channel protein in the protozoan ciliate Stentor coeruleus

机译：推定的环状GMP激活的通道蛋白在原生动物纤毛Stentor蓝藻中的检测和定位

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摘要

Immunoblotting and immunocytochemical assays were employed to identify and localize a channel protein activated by cyclic GMP (cGMP) in the protozoan ciliate Stentor coeruleus. Analysis of whole-cell homogenate with antibodies raised against the alpha-subunit of the cGMP-activated channel protein from bovine rod outer segments and against cGMP revealed four major protein bands with molecular masses of 40 kDa, 63 kDa, and over 120 kDa, which bound cGMP. However. only a cGMP-binding protein of 63 kDa, corresponding to the alpha-subunit of the cGMP-activated ion channel protein from bovine rod outer segments, was found in the ciliate cortex fraction. The functional cGMP-activated channel protein was also shown to be present in the cortex fraction of S. coeruleus by patch-clamp measurements of artificial liposomes. Incorporation of the cortex fraction into liposomes resulted in the appearance of ion channel activity related to cGMP. The reconstituted protein channels were strongly inhibited by 1-cis-diltiazem, a known potent blocker of many types of cyclic-nucleotide-activated channels. The results presented here are the first demonstration of the existence and localization of a putative cGMP-activated channel protein in the ciliate S. coendetts. Cyclic-nucleotide-activated channel proteins are nonspecific cation channels which mediate the receptor potentials in photoreceptor cells and in cells of the olfactory epithelium. Oil the basis of these data. we suggest that the 63 kDa protein identified in Stentor coeruleus is also a cGMP-activated ion channel and that it may be involved as an effector in the photosensory transduction pathway leading to the motile photophobic response in this ciliate protist.
机译：免疫印迹法和免疫细胞化学法被用于鉴定和定位由环状GMP（cGMP）激活的通道蛋白在原生动物纤毛Stentor蓝藻中。用针对来自牛杆外段的cGMP激活通道蛋白的α亚基和cGMP产生的抗体对全细胞匀浆进行分析，发现了四个主要蛋白带，分子量分别为40 kDa，63 kDa和超过120 kDa，绑定的cGMP。然而。在纤毛状皮质部分中仅发现了一个63 kDa的cGMP结合蛋白，该蛋白与来自牛杆外部的cGMP激活的离子通道蛋白的α亚基相对应。通过人工脂质体的膜片钳测量，还显示了功能性cGMP激活的通道蛋白存在于蓝藻的皮质部分中。皮质部分掺入脂质体导致出现与cGMP有关的离子通道活性。重构的蛋白质通道受到1-cis-地尔硫卓的强烈抑制，1-cis-地尔硫卓是许多类型的环状核苷酸激活通道的已知有效阻断剂。这里介绍的结果是纤毛S. coendetts中假定的cGMP激活通道蛋白的存在和定位的第一个证明。环核苷酸激活的通道蛋白是非特异性阳离子通道，可在感光细胞和嗅觉上皮细胞中介导受体电位。石油是这些数据的基础。我们建议在蓝藻支架中鉴定出的63 kDa蛋白质也是cGMP激活的离子通道，它可能作为光感官转导途径的效应子参与该纤毛虫原色中的运动性畏光反应。

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