Antarctic Fish and MicroRNA Control of Development and Physiology

冰鱼生活在寒冷的南极海域，它们有一些独特的特征，比如没有红细胞，心脏增大，血管直径大，骨矿物质密度低，脂肪滴会破坏它们的肌肉。这些特征对其他动物是有害的。在生活在温暖水域的哺乳动物和鱼类中，参与这些特征的器官的发育是由编码特定蛋白质的基因调节的，但蛋白质生产的速度通常是由称为microRNAs （miRNAs）的短RNA分子调节的。编码蛋白质的基因必须首先复制RNA，而真正的蛋白质是由RNA复制的中间产物合成的。mirna通过结合RNA中间体并中断其蛋白质的产生来调节蛋白质的数量。mirna与RNA的结合在很大程度上取决于温度。南极鱼类生活在温度低于淡水冰点的海洋中，目前还没有研究过mirna对基因的调控。本项目将比较南极鱼和温水鱼的miRNA调控(1)，了解miRNA如何在持续寒冷条件下调控基因表达；2)南极冰鱼没有红细胞，心脏增大，骨密度降低，而与之密切相关的南极鱼类含有红细胞，心脏正常，骨骼致密。该项目将对国内和全球的科学和社会产生广泛的影响。首先，这将是对南极鱼类基因调控（miRNAs）重要因素的首次研究，这是南大洋整个生态的重要组成部分，并将阐明这些鱼类如何对南极水域变暖做出反应。其次，它将通过在南极实地考察季节举行视频会议和在研究实验室主持学生对南极鱼类的调查，将南极科学带给当地一所市中心高中的代表性不足的高中生。microRNAs （miRNAs）是调节温带动物发育和生理的关键基因表达转录后调节剂。尽管mirna通过与信使rna （mrna）结合而起作用，这是一个对温度非常敏感的过程，但尚未在南极动物中研究过mirna，包括主导南大洋的南极鱼。本项目将比较南极和温带鱼类的miRNA调节，以了解miRNA调节在适应持续寒冷中的作用；2)海底生活、密集骨骼、红血的南极鱼与高浮力、骨质减少、白血的冰鱼，以了解血红蛋白基因和红细胞丢失的进化过程中，特定器官中miRNA的调节，心脏和血管扩张的起源，以及浮力增加的进化过程，这是由骨矿化减少和脂质沉积增加引起的。目的1是验证南极鱼类进化出与mirna相关的基因组特化以应对持续寒冷的假设。该项目将把四种南极南极南极鱼与两种温带南极南极鱼和两种温带实验室物种进行比较，以验证以下假设：(a)南极鱼通过失去祖先基因和/或获得新基因而进化出miRNA基因组库，(b)表达与耐寒性有关的miRNA，以及(c)通过改变miRNA基因表达来应对温度变化。目的2是验证一种假设，即从红血底栖祖先进化而来的冰鱼伴随着miRNA基因组库、序列和/或表达的改变。该项目将测试以下假设：(a)冰鱼中的mirna在序列和/或在冰鱼特化的表达中进化，包括头肾（红细胞的起源）；心脏（血管系统改变）、头盖骨和胸带（骨密度降低）；和骨骼肌（脂质沉积），以及(b)在冰鱼特化中进化的mirna具有与其在冰鱼中衍生作用相关的祖先功能，这是通过在发育中的斑马鱼中对冰鱼mirna的斑马鱼同源物的功能测试确定的。该项目将使用高通量转录组学和新型软件分离、测序和确定mirna和mrna的表达。研究结果将揭示微rna系统是如何在被推向生理极端的脊椎动物中进化的，并为全球变暖最迅速地区的关键物种的前景提供洞见。

Icefish live in frigid Antarctic seas, and have unique traits such as the absence of red blood cells, enlarged hearts, large diameter blood vessels, low bone mineral densities, and fat droplets that disrupt their muscles. These features would be harmful in other animals. In mammals and fish inhabiting warm waters, development of organs involved in these traits is modulated by genes that encode specific proteins, but the rate of protein production is often regulated by short RNA molecules called microRNAs (miRNAs). Genes that code for proteins must first make an RNA copy, and the actual protein is made from this RNA copy intermediate. MiRNAs regulate the amount of protein that is made by binding to the RNA intermediate and interrupting its production of protein. Binding of miRNAs to RNA depends strongly on temperature. Regulation of genes by miRNAs has not been studied in Antarctic fish, which live in seas with temperatures below the freezing point of fresh water. This project will compare miRNA regulation 1) in Antarctic fish vs. warm-water fish to learn how miRNAs regulate gene expression in constant cold; and 2) in Antarctic icefish with no red blood cells, enlarged hearts, and reduced bone density vs. closely related Antarctic fish containing red blood cells, normal hearts, and dense bones. The project will have broad impacts to science and society nationally and globally. First, this will be the first study of important factors in gene regulation (miRNAs) in Antarctic fish, which are an essential component of the entire ecology of the Southern Ocean, and will shed light on how these fish might respond to the warming of Antarctic waters. Second, it will bring Antarctic science to under-represented high school students at a local alternative downtown high school by conducting video conferences during the Antarctic field seasons and hosting student investigations of Antarctic fish in the research laboratory. microRNAs (miRNAs) are key post-transcriptional regulators of gene expression that modulate development and physiology in temperate animals. Although miRNAs act by binding to messenger RNAs (mRNAs), a process that is strongly sensitive to temperature, miRNAs have yet not been studied in Antarctic animals, including Notothenioid fish, which dominate the Southern Ocean. This project will compare miRNA regulation in 1) Antarctic vs. temperate fish to learn the roles of miRNA regulation in adaptation to constant cold; and in 2) bottom-dwelling, dense-boned, red-blooded Nototheniods vs. high buoyancy, osteopenic, white-blooded icefish to understand miRNA regulation in specialized organs after the evolution of the loss of hemoglobin genes and red blood cells, the origin of enlarged heart and vasculature, and the evolution of increased buoyancy, which arose by decreased bone mineralization and increased lipid deposition. Aim 1 is to test the hypothesis that Antarctic fish evolved miRNA-related genome specializations in response to constant cold. The project will compare four Antarctic Notothenioid species to two temperate Notothenioids and two temperate laboratory species to test the hypotheses that (a) Antarctic fish evolved miRNA genome repertoires by loss of ancestral genes and/or gain of new genes, (b) express miRNAs that are involved in cold tolerance, and (c) respond to temperature change by changing miRNA gene expression. Aim 2 is to test the hypothesis that the evolution of icefish from red-blooded bottom-dwelling ancestors was accompanied by an altered miRNA genomic repertoire, sequence, and/or expression. The project will test the hypotheses that (a) miRNAs in icefish evolved in sequence and/or in expression in icefish specializations, including head kidney (origin of red blood cells); heart (changes in vascular system), cranium and pectoral girdle (reduced bone mineral density); and skeletal muscle (lipid deposition), and (b) miRNAs that evolved in icefish specializations had ancestral functions related to their derived roles in icefish, as determined by functional tests of zebrafish orthologs of icefish miRNAs in developing zebrafish. The program will isolate, sequence, and determine the expression of miRNAs and mRNAs using high-throughput transcriptomics and novel software. Results will show how the microRNA system evolves in vertebrate animals pushed to physiological extremes and provide insights into the prospects of key species in the most rapidly warming part of the globe.