Antarctic Fish and MicroRNA Control of Development and Physiology

冰鱼生活在寒冷的南极海域，具有独特的特征，如没有红细胞，心脏增大，血管直径大，骨矿物质密度低，脂肪滴会破坏肌肉。这些特征在其他动物身上是有害的。在生活在温暖沃茨的哺乳动物和鱼类中，与这些特征有关的器官的发育是由编码特定蛋白质的基因调节的，但蛋白质产生的速率通常由称为微RNA（miRNAs）的短RNA分子调节。编码蛋白质的基因必须首先复制RNA，而实际的蛋白质是由这个RNA复制中间体制成的。miRNAs通过与RNA中间体结合并中断其蛋白质的产生来调节蛋白质的量。miRNA与RNA的结合强烈依赖于温度。在南极鱼类中，miRNAs对基因的调控还没有研究过，南极鱼类生活在温度低于淡水冰点的海洋中。该项目将比较1）南极鱼与温水鱼的miRNA调控，以了解miRNA如何在恒定的寒冷中调节基因表达; 2）南极冰鱼没有红细胞，心脏增大，骨密度降低与含有红细胞，正常心脏和致密骨骼的密切相关的南极鱼。该项目将对国家和全球的科学和社会产生广泛影响。首先，这将是对南极鱼类基因调控（miRNAs）重要因素的首次研究，南极鱼类是南大洋整个生态的重要组成部分，并将揭示这些鱼类如何应对南极沃茨变暖。第二，它将在南极实地考察季节举行视频会议，并在研究实验室主持学生对南极鱼类的调查，从而将南极科学带到当地市中心替代高中的代表性不足的高中生中。microRNA（miRNAs）是调节温带动物发育和生理的基因表达的关键转录后调节因子。虽然miRNAs通过与信使RNA（mRNAs）结合而起作用，这是一个对温度非常敏感的过程，但miRNAs尚未在南极动物中进行研究，包括在南大洋占主导地位的Notothenioid鱼。本项目将比较1）南极和温带鱼类的miRNA调控，以了解miRNA调控在适应持续寒冷中的作用;以及2）在底栖的、骨密度高的、红血Nototheniods与高浮力的、骨质减少的、白血冰鱼中，以了解在血红蛋白基因和红细胞丧失的进化后，专门器官中的miRNA调控，心脏和脉管系统扩大的起源，以及浮力增加的演变，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miRNA相关基因组特化以应对持续寒冷的假设。该项目将比较四种南极鱼与两种温带南极鱼和两种温带实验室物种，以测试以下假设：（a）南极鱼通过丢失祖先基因和/或获得新基因进化出miRNA基因组库，（B）表达参与耐寒性的miRNA，以及（c）通过改变miRNA基因表达对温度变化作出反应。目的2是检验这一假设，即冰鱼从红血底栖祖先的进化伴随着改变的miRNA基因组库，序列和/或表达。该项目将测试以下假设：（a）冰鱼中的miRNAs在序列和/或在冰鱼专业化（包括头肾）中的表达中进化（红细胞的起源）;心脏（血管系统的变化），颅骨和肩带（骨密度降低）;和骨骼肌（脂质沉积），和（B）在冰鱼特化中进化的miRNA具有与它们在冰鱼中的衍生作用相关的祖先功能，如通过在发育中的斑马鱼中对冰鱼miRNA的斑马鱼直系同源物进行功能测试所确定的。该计划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。研究结果将展示microRNA系统如何在被推向生理极端的脊椎动物中进化，并为地球仪变暖最快的地区的关键物种的前景提供见解。

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.