In a ciliate Paramecium, the presence of water channels on the membrane of contractile vacuole has long been predicted by both morphological and physiological data, however, to date either the biochemical or the molecular biological data have not been provided. In the present study, to examine the presence of aquaporin in Paramecium, we carried out RT‐PCR with degenerated primers designed based on the ParameciumDB, and an aquaporin cDNA (aquaporin 1, aqp1) with a full‐length ORF encoding 251 amino acids was obtained from Paramecium multimicronucleatum by using RACE. The deduced amino acid sequence of AQP1 had NPA‐NPG motifs, and the prediction of protein secondary structure by CNR5000 and hydropathy plot showed the presence of six putative transmembrane domains and five connecting loops. Phylogenetic analysis results showed that the amino acid sequence of AQP1 was close to that of the Super‐aquaporin group. The AQP1‐GFP fusion protein clearly demonstrated the subcellular localization of AQP1 on the contractile vacuole complex, except for the decorated spongiome membrane. The functional analyses of aqp1 were done by RNA interference‐based gene silencing, using an established feeding method. The aqp1 was found to be crucial for the total fluid output of the cell, the function of contractile vacuole membranes.

中文翻译：

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功能性 Aqp1 基因产物定位于草履虫多微核菌的收缩液泡复合体

在纤毛虫草履虫中，形态学和生理学数据早就预测了收缩液泡膜上存在水通道，但迄今为止，尚未提供生化或分子生物学数据。在本研究中，为了检测草履虫中水通道蛋白的存在，我们使用基于草履虫DB设计的简并引物和具有编码 251 个氨基酸的全长 ORF的水通道蛋白 cDNA（水通道蛋白 1，aqp1）进行 RT-PCR从草履虫 多微核中获得通过使用 RACE。推断的 AQP1 氨基酸序列具有 NPA-NPG 基序，通过 CNR5000 和亲水图预测蛋白质二级结构显示存在六个假定的跨膜结构域和五个连接环。系统发育分析结果表明，AQP1的氨基酸序列与Super-aquaporin组的氨基酸序列接近。除了装饰的海绵体膜外，AQP1-GFP 融合蛋白清楚地证明了 AQP1 在收缩液泡复合体上的亚细胞定位。aqp1的功能分析是通过基于 RNA 干扰的基因沉默，使用既定的喂养方法完成的。所述AQP1被发现是用于小区的总流体输出，收缩液泡膜的功能是至关重要的。