National Engineering Research Center of Marine Facilities Aquaculture, Marine Science College, Zhejiang Ocean University, Zhoushan, Zhejiang 316022, People's Republic of China.
Institute of Biochemistry, College of LifeSciences, Zijingang Campus, Zhejiang University, Hangzhou, Zhejiang 310058, People's Republic of China.
Sci Rep. 2017 Jan 6;7:40247. doi: 10.1038/srep40247.
Serotonin (5-HT) is an important neurotransmitter and neuromodulator that controls a variety of sensory and motor functions through 5-HT receptors (5-HTRs). The 5-HT4R subfamily is linked to Gs proteins, which activate adenylyl cyclases (ACs), and is involved in many responses in peripheral organs. In this study, the 5-HT4R from Apostichopus japonicus (Aj5-HT4R) was identified and characterised. The cloned full-length Aj5-HT4R cDNA is 1,544 bp long and contains an open reading frame 1,011 bp in length encoding 336 amino acid proteins. Bioinformatics analysis of the Aj5-HT4R protein indicated this receptor was a member of class A G protein coupled receptor (GPCR) family. Further experiments using Aj5-HT4R-transfected HEK293 cells demonstrated that treatment with 5-HT triggered a significant increase in intracellular cAMP level in a dose-dependent manner and induced a rapid internalisation of Aj5-HT4R fused with enhanced green fluorescent protein (Aj5-HT4R-EGFP) from the cell surface into the cytoplasm. In addition, the transcriptional profiles of Aj5-HT4R in aestivating A. japonicas and phosphofructokinase (AjPFK) in 5-HT administrated A. japonicus have been analysed by real-time PCR assays. Results have led to a basic understanding of Aj5-HT4R in A. japonicus, and provide a foundation for further exploration of the cell signaling and regulatory functions of this receptor.
血清素（5-HT）是一种重要的神经递质和神经调质，通过 5-HT 受体（5-HTRs）控制各种感觉和运动功能。5-HT4R 亚家族与 Gs 蛋白相连，激活腺苷酸环化酶（ACs），并参与外周器官的许多反应。在这项研究中，鉴定并表征了日本刺参（Aj5-HT4R）的 5-HT4R。克隆的全长 Aj5-HT4R cDNA 长 1544bp，包含一个 1011bp 的开放阅读框，编码 336 个氨基酸的蛋白质。Aj5-HT4R 蛋白的生物信息学分析表明，该受体是 A 类 G 蛋白偶联受体（GPCR）家族的成员。进一步使用 Aj5-HT4R 转染的 HEK293 细胞进行的实验表明，5-HT 处理以剂量依赖性方式触发细胞内环腺苷酸（cAMP）水平的显著增加，并诱导与增强型绿色荧光蛋白（Aj5-HT4R-EGFP）融合的 Aj5-HT4R 从细胞表面快速内化到细胞质中。此外，通过实时 PCR 分析了休眠的日本刺参中的 Aj5-HT4R 和 5-HT 处理的日本刺参中的磷酸果糖激酶（AjPFK）的 Aj5-HT4R 转录谱。结果基本了解了日本刺参中的 Aj5-HT4R，并为进一步探索该受体的细胞信号转导和调节功能提供了基础。
