The pituitary hormone prolactin (PRL) regulates salt and water homeostasis by altering ion retention and water uptake through peripheral osmoregulatory organs. To understand the role of PRL and its receptor (PRLR) in hypoosmoregulation of turbot (Scophthalmus maximus), we characterized the PRL and PRLR gene and analyzed the tissue distribution of the two genes and their gene transcriptional patterns in the main expressed tissues under long-term and short-term low salt stress. The PRL cDNA is 1486 bp in length, incorporating an ORF of 636 bp with a putative primary structure of 211 residues. And the PRLR cDNA is 2849 bp in length, incorporating an ORF of 1944 bp with a putative primary structure of 647 residues. The deduced amino acid sequences of these two genes shared highly conserved structures with those from other teleosts. Quantitative real-time PCR results showed that PRL transcripts were strongly expressed in the pituitary and very weakly in brain, but were hardly expressed in other tissues. PRLR transcripts were most abundant in the kidney, to a lesser extent in the gill, intestine, brain, and spleen, and at low levels in the pituitary and other tissues examined. The expression of PRL in the pituitary increased after short-term or long-term low salt stress, and the highest expression level appeared 12 h after stress (P < 0.05). And there is no significant difference between both low salt group (5 ppt and 10 ppt) at each sampling point. The variation of PRLR expression in gill under short-term low salt stress is similar to that of PRL gene in pituitary, with highest value in 12 h (P < 0.05). However, the expression under long-term low salt stress was significantly higher than control group even than 12 h group under 5 ppt (P < 0.05). The expression of PRLR in the kidney increased first and then decreased after low salt stress, and the highest value also appeared in 12 h after stress and there was no significant difference between the salinity groups. After long-term low salt stress, the expression level also increased significantly (P < 0.05), but it was flat with 24 h, which was lower than 12 h. The variation of PRLR expression in the intestine was basically consistent with that in the kidney. The difference was that the expression level of 24 h after stress in the 5 ppt group was significantly higher than that of the 10 ppt group (P < 0.05). After a comprehensive analysis of the expression levels of the two genes, it can be found that the expression level increased and peaked at 12 h after short-term low salt stress, indicating that this time point is the key point for the regulation of turbot in response to low salt stress. This also provides very important information for studying the osmotic regulation of turbot. In addition, our results also showed that the expression of PRLR was stable in the kidney and intestine after long-term low salt stress, while the expression in the gill was much higher than short-term stress. It suggested that PRL and its receptors mainly exert osmotic regulation function in the gill under long-term low salt stress. At the same time, such a result also brings a hint for the low salt selection of turbot, focusing on the regulation of ion transport in the gill.

中文翻译：

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大菱t（Scophthalmus maximus）的PRL和PRLR的克隆，分子表征及其对短期和长期低盐胁迫的响应表达。

垂体激素催乳激素（PRL）通过改变离子保留和通过周围渗透调节器官的水分吸收来调节盐和水的体内稳态。为了了解PRL及其受体（PRLR）在比目鱼低渗调节中的作用，我们对PRL和PRLR基因进行了表征，并分析了长时分下主要表达组织中这两个基因的组织分布及其基因转录方式。长期和短期低盐胁迫。PRL cDNA的长度为1486 bp，并整合了636 bp的ORF，并具有推测的211个残基的一级结构。PRLR cDNA的长度为2849 bp，整合了1944 bp的ORF，并具有647个残基的推定一级结构。推导的这两个基因的氨基酸序列与其他硬骨鱼具有高度保守的结构。实时定量PCR结果显示，PRL转录本在脑垂体中表达强烈，在脑中非常弱，而在其他组织中几乎不表达。PRLR转录本在肾脏中含量最高，在ill，肠，脑和脾脏中含量较低，而在垂体和其他组织中含量较低。短期或长期低盐胁迫后垂体中PRL的表达增加，应激后12 h表达最高（P <0.05）。低盐组（5 ppt和10 ppt）在每个采样点之间没有显着差异。短期低盐胁迫下g中PRLR表达的变化与垂体中PRL基因的变化相似，在12 h时最高（P <0.05）。然而，长期低盐胁迫下5 ppt下的表达甚至明显高于对照组，甚至高于12 h组（P <0.05）。低盐胁迫后，肾脏中PRLR的表达先升高后降低，最高值也在胁迫后12 h出现，盐度组之间无显着差异。长期低盐胁迫后，表达水平也显着增加（P <0.05），但在24 h时则保持稳定，低于12 h。肠道PRLR表达的变化与肾脏基本一致。差异在于5 ppt组应激后24 h的表达水平显着高于10 ppt组（P <0.05）。对两个基因的表达水平进行全面分析后，可以发现，表达水平在短期低盐胁迫后12 h达到峰值，表明该时间点是响应低盐胁迫调节大菱t的关键。这也为研究大菱t的渗透调节提供了非常重要的信息。此外，我们的结果还表明，长期低盐胁迫后，PRLR在肾脏和肠中的表达稳定，而g中的表达远高于短期胁迫。提示长期低盐胁迫下PRL及其受体主要在ill中发挥渗透调节作用。同时，这样的结果也暗示了菱salt的低盐选择，着重于ill中离子迁移的调控。