Molecular cloning and mRNA expression analysis of two GH secretagogue receptor transcripts in orange-spotted grouper (Epinephelus coioides)
Molecular cloning and mRNA expression analysis of two GH secretagogue receptor transcripts in orange-spotted grouper (Epinephelus coioides)
复制标题
斜带石斑鱼两种 GH 促分泌素受体转录本的分子克隆及 mRNA 表达分析
DOI：
10.1677/joe-08-0325
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发表时间：
2008-11-01
影响因子：
4
通讯作者：
Lin, Haoran
中科院分区：
文献类型：
--
作者：
Chen, Ting;Tang, Zhiguo;Lin, Haoran
生长激素促分泌素受体（GH secretagogue receptor，GHSR）是生长激素释放肽（ghrelin）的受体，生长激素释放肽是一种循环的生长激素释放和食欲诱导激素。本文从硬骨鱼类斜带石斑鱼（Epineoderus coioides）中克隆了两个GhsrcDNA，即gpGhsr 1a和gpGhsr 1b。gpGHSK 1a全长1512 bp，开放阅读框（ORF）编码383个氨基酸，含7个跨膜结构域; gpGHSK 1b全长1703 bp，开放阅读框编码303个氨基酸，含5个跨膜结构域。cDNA和基因序列的比较表明，这两个转录本是一个单一的gpGhsr基因的两种选择性剪接形式。RT-PCR检测两种gpGhsr mRNA的组织分布和个体发育。gpGHSK 1a主要在脑和垂体中表达，而gpGHSR 1b表达更广泛。在胚胎和幼虫发育过程中，gpGhsr 1b mRNA先于gpGhsr 1a mRNA出现。此外，对脑进行的定量实时PCR显示，这两种转录本在脑垂体中的表达水平最高。gpGHSR 1a的表达水平普遍高于gpGHSR 1b。通过原位杂交，在石斑鱼脑中也广泛检测到GHSR表达细胞，其分布范围比以前在哺乳动物中的报道更广。最后，体外研究表明，在垂体和下丘脑的gpGHSR转录本的表达被GH和ghrelin下调，但不被去酰基ghrelin，这表明GHSR反馈调节也存在于硬骨鱼。
GH secretagogue receptor (GHSR) is the receptor of ghrelin, a circulating GH-releasing and appetite-inducing hormone. In this paper, two Ghsr cDNAs, gpGhsr1a and gpGhsr1b, were identified and characterized in a teleost, the orange-spotted grouper (Epinephelus coioides). The gpGHSK1a is 1512bp in length with an open reading frame (ORF) that encodes a protein of 383 amino acids with seven transmembrane (TM) domains, while the 1703 bp gpGHSK1b contains an ORF encoding for 303 amino acids with five TM domains. Comparison between cDNA and gene sequences showed that the two transcripts are two alternative splicing forms of a single gpGhsr gene. Tissue distribution and ontogeny of two gpGhsr mRNAs were examined by RT-PCR. The gpGHSK1a is mainly expressed in brain and pituitary gland, when compared with a more widespread expression of gpGHSR1b. During embryonic and larval development, the gpGhsr1b mRNA appears before the gpGhsr1a mRNA. Furthermore, quantitative real-time PCR performed on brain showed that both transcripts have the highest expression level in the pituitary gland. The expression level of gpGHSR1a was generally higher than that of gpGHSR1b. GHSR expressing cells were also detected widely in grouper brain by in situ hybridization, with a broader distribution than previous reports in mammals. Finally, an in vitro study showed that expression of both gpGHSR transcripts in pituitary and hypothalamus is downregulated by GH and ghrelin but not by des-acyl ghrelin, and this suggests that feedback regulation of GHSR also exists in teleostean fishes.