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低氧条件对黄斑鲈(Perca flavescens)幼鱼缺氧诱导因子-1α基因的表达无影响。

The expression of hypoxia-inducible factor-1α gene is not affected by low-oxygen conditions in yellow perch (Perca flavescens) juveniles.

作者信息

Kwasek Karolina, Rimoldi Simona, Cattaneo Anna Giulia, Parker Timothy, Dabrowski Konrad, Terova Genciana

机构信息

School of Environment and Natural Resources, Ohio State University, Columbus, OH, 43210, USA.

Department of Biotechnology and Life Sciences, University of Insubria, Via Dunant, 3, 21100, Varese, Italy.

出版信息

Fish Physiol Biochem. 2017 Jun;43(3):849-862. doi: 10.1007/s10695-017-0340-9. Epub 2017 Jan 18.

Abstract

Hypoxia can affect various fish populations, including yellow perch Perca flavescens, which is an economically and ecologically important species in Lake Erie, a freshwater system that often experiences hypoxia in the hypolimnetic part of the lake. Fish, similarly to mammals, possess molecular oxygen sensor-hypoxia-inducible factor-1 (HIF-1), a transcription factor that can affect expression of many downstream genes related to animal growth and locomotion, protein synthesis, as well as ATP and amino acid metabolism. HIF-1 is a heterodimer, which consists of two subunits: oxygen-sensitive and oxygen-insensitive subunits, α and β, respectively. In this study, we report first on the molecular cloning and sequencing of P. flavescens HIF-1α. The full-length complementary DNA (cDNA) was isolated and submitted to the GenBank with accession number KT783483. It consists of 3529 base pairs (bp) carrying a single open-reading frame that encompasses 2250 bp of the coding region, 247 bp of the 5' untranslated region (UTR), and 1032 bp of the 3' UTR. The "de novo" prediction of the 3D structure of HIF-1α protein, which consists of 749 amino acids, is presented, too. We then utilized One-Step Taqman® real-time RT-PCR technology to monitor changes in HIF-1α messenger RNA (mRNA) copies in response to chronic hypoxic stress. An experiment was conducted using 14-day post-swim-up stage yellow perch larvae with uninflated swim bladders. This experiment included three treatment groups: hypoxia, mid-hypoxia, and normoxia, in four replicates (four tanks per treatment) with the following dissolved oxygen levels: 3, 4, and >7 mg O/L, respectively. At the end (2 weeks) and in the middle (1 week) of the experiment, fish from each tank were sampled for body measurements and molecular biology analysis. The results showed no differences in survival (∼90%) between treatment groups. Oxygen concentration was lowered to 3.02 ± 0.15 (mean ± SE) mg O/L with no adverse effect on fish survival. The highest growth rate was observed in the normoxic group. A similar trend was observed with fish body length. The growth rate of fish declined with decreasing water-dissolved oxygen. The number of HIF-1α mRNA copies was not significantly different between hypoxic, mid-hypoxic, and normoxic conditions, and this was true for fish obtained in the middle and at the end of the experiment. Graphical abstract.

摘要

缺氧会影响各种鱼类种群,包括黄鲈(Perca flavescens),它是伊利湖(Erie)一种在经济和生态方面都很重要的物种,伊利湖是一个淡水系统,其湖下层经常出现缺氧情况。与哺乳动物类似,鱼类拥有分子氧传感器——缺氧诱导因子-1(HIF-1),这是一种转录因子,可影响许多与动物生长和运动、蛋白质合成以及ATP和氨基酸代谢相关的下游基因的表达。HIF-1是一种异源二聚体,由两个亚基组成:分别是氧敏感亚基和氧不敏感亚基α和β。在本研究中,我们首次报告了黄鲈HIF-1α的分子克隆和测序。分离出全长互补DNA(cDNA)并提交至GenBank,登录号为KT783483。它由3529个碱基对(bp)组成,带有一个单一的开放阅读框,该开放阅读框包含2250 bp的编码区、247 bp的5'非翻译区(UTR)和1032 bp的3'UTR。还展示了由749个氨基酸组成的HIF-1α蛋白三维结构的“从头”预测。然后,我们利用一步法Taqman®实时逆转录聚合酶链反应(RT-PCR)技术来监测慢性缺氧应激下HIF-1α信使核糖核酸(mRNA)拷贝数的变化。使用游泳后14天、鳔未充气的黄鲈幼鱼进行了一项实验。该实验包括三个处理组:缺氧组、中度缺氧组和常氧组,每组四个重复(每个处理四个水箱),溶解氧水平分别为:3、4和>7 mg O/L。在实验结束时(2周)和中期(1周),从每个水箱中取样鱼进行体长测量和分子生物学分析。结果显示处理组之间的存活率(约90%)没有差异。氧浓度降至3.02±0.15(平均值±标准误)mg O/L,对鱼类存活没有不利影响。常氧组观察到最高的生长速率。鱼体长度也观察到类似趋势。鱼的生长速率随着水中溶解氧的降低而下降。缺氧、中度缺氧和常氧条件下HIF-1α mRNA拷贝数没有显著差异,实验中期和末期获得的鱼都是如此。图形摘要。

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