Genomic Instability and the Evolution of the Antarctic Notothenioid Fishes

南极南极鱼的祖先很可能是在4000万至6000万年前在南极大陆温带大陆架水域进化而来的一种缓慢的海底生物。冰盖在大陆架上的搁浅和营养条件的变化可能消除了始新世晚期动物的分类多样性，并开始了南极鱼的快速多样化。在南极高大陆架上，南极鱼在鱼类动物群中占主导地位，被认为是海洋鱼类种群中为数不多的例子之一。这位研究者和其他研究者最近的工作强烈地表明，基因组水平上的进化变化渗透到南极鱼中，这可能是硬骨鱼类亚目快速多样化的主要驱动力。一个显著的例子是冰鱼（冰鱼科）丧失了产生功能性红细胞和血红蛋白的能力。这种红系的自然“敲除”使冰鱼分类群成为分析红系决定和分化的信号通路和转录机制的独特遗传资源。血红蛋白缺失和红细胞缺失的表型似乎至少部分是由重复元件和反转录转座子介导的基因组重排和/或缺失引起的。本提案的目的之一是评估基因组不稳定性在冰鱼造血程序进化中的作用。利用基于荧光原位杂交的细胞遗传学和细菌人工染色体（BAC）文库，将红细胞生成基因位点物理定位到两种南极鱼（红血岩鳕Notothenia coriiceps和白血冰鱼Chaenocephalus aceratus）的基因组中，以确定导致红细胞缺失的基因保留、失活和/或丢失模式。第二个目标将是利用新的比较基因组技术，利用冰鱼中红细胞的缺失来发现参与红细胞生成遗传程序的新基因。第三个目标将是通过斑马鱼胚胎中直系同源的功能分析来确定新发现的红系基因及其编码蛋白的功能。总之，本研究将评估南极冰鱼产生新的红细胞生成表型的进化机制，并利用这一新的分类单元对参与红细胞形成的基因进行公正的全基因组扫描。更广泛的影响包括基因组分子生物学的研究生培训，包括挖掘新红系基因潜在功能的数据集所需的生物信息学策略，以及使用斑马鱼模型系统来测试这些基因的假设功能。将建立一个网络托管数据库，以便其他实验室可以利用这项研究项目产生的基因组图谱数据，外行读者也可以了解基因组学在解决极地生物学重要问题方面的用途。此外，参与造血的新基因的发现可能为生物技术产业开发治疗贫血、白血病和其他血液疾病的疗法提供重要靶点。

The ancestral Notothenioid fish stock of Antarctica probably arose as a sluggish, bottom-dwelling species that evolved some 40-60 million years ago in the then temperate shelf waters of the Antarctic continent. The grounding of the ice sheet on the continental shelf and changing trophic conditions may have eliminated the taxonomically diverse late Eocene fauna and initiated the rapid diversification of the Notothenioids. On the High Antarctic shelf, Notothenioids today dominate the fish fauna and are recognized as one of the few examples of a species flock of marine fishes. Recent work conducted by this investigator and others suggests strongly that evolutionary change at the level of the genome permeates the Notothenioids and perhaps has been a major driver of the rapid diversification of this suborder of bony fishes. One striking example is the loss by the icefishes (Channichthyidae) of the ability to produce functional erythrocytes and hemoglobin. This natural "knockout" of the erythroid lineage makes the icefish taxon a unique genetic resource for analyzing the signaling pathways and transcriptional machinery involved in erythroid determination and differentiation. The hemoglobin-null and erythrocyte-null phenotypes appear to have been caused, at least in part, by genomic rearrangements and/or deletions mediated by repetitive elements and retrotransposons. One objective of this proposal will be to evaluate the role of genome instability in the evolution of the hematopoietic program of the icefishes. Using fluorescence in situ hybridization-based cytogenetics and bacterial artificial chromosome (BAC) libraries,erythropoietic gene loci will be mapped physically to the genomes of two notothenioids, the red-blooded rockcod Notothenia coriiceps and the white-blooded icefish Chaenocephalus aceratus, to determine the patterns of gene retention, inactivation, and/or loss that cause the erythrocyte-null condition. A second objective will be to exploit the absence of erythrocytes in icefishes to discover novel genes involved in the erythropoietic genetic program by use of new comparative genomic technologies. A third objective will be to determine the functions of newly discovered erythroid genes and their encoded proteins by functional analysis of zebrafish orthologs in zebrafish embryos. In summary, this research will evaluate the evolutionary mechanisms that generated the novel erythropoietic phenotype of the Antarctic icefishes and will exploit this novel taxon to conduct an unbiased, genome-wide scan for genes involved in red cell formation. The broader impacts include graduate student training in genomic molecular biology, including the bioinformatic strategies necessary to mine the data set for the potential functions of novel erythroid genes, and the use of a model system, the zebrafish, to test the hypothetical functions of these genes. A Web-hosted database will be created so that other laboratories can make use of the genome mapping data generated by this research project and lay audiences can learn about the uses of genomics for solving important questions in polar biology. In addition, the discovery of novel genes that participate in hematopoiesis may contribute important targets for the development by the biotechnology industry of therapeutics to treat anemias, leukemias, and other blood diseases.