Functional characterization of the Harbi1 and Naif1 transposon-derived genes in vertebrates

真核生物基因表达的时空协调依赖于转录和转录后调控网络。虽然这些网络的一些组成部分和功能已经被阐明，但这种复杂回路的起源和进化机制仍然知之甚少。真核生物基因组的很大一部分是由转座因子（te）组成的，这些转座因子能够在基因组中移动和复制。TEs最好被视为基因组寄生虫，对它们所居住的宿主细胞没有明显的适应价值。因此，在没有选择压力来维持迁移的情况下，绝大多数te在进化过程中已经转位失活，并且被视为没有任何明显细胞功能的“垃圾DNA”。然而，人们越来越认识到te在基因组中的扩散可能在新基因功能的进化中发挥了关键作用。例如，通过被称为“驯化”的进化过程，te衍生的转座酶蛋白作为调控元件被反复招募到细胞通路中。Harbi1和Naif1是脊椎动物中高度保守的基因，它们来自于大约5亿年前颌骨脊椎动物共同祖先的一个古老的PIF/Harbinger转座子。这些基因的保存表明它们在重要的细胞功能中受到了选择，它们的系统发育关系表明它们参与了相同的分子途径。Naif1与凋亡功能有关。我们的初步数据表明，Naif1是一种dna结合蛋白，可促进Harbi1的核输入，通过蛋白-蛋白相互作用将Harbi1招募到基因组位点。Harbi1催化结构域的序列保守表明它可能保留了核酸酶活性。然而，这两个基因在细胞稳态和有机体发育中的作用一直是谜。在这个项目中，我们利用体外生化分析，体外培养细胞全基因组ChIP和转录谱分析，以及斑马鱼模型中敲除（KO）动物的表型分析来理解和功能注释这些基因及其调控的遗传网络。该项目将极大地有助于我们理解te作为创造遗传新颖性的重要力量的机制和作用。

Spatio-temporal coordination of eukaryotic gene expression relies on transcriptional and post-transcriptional regulatory networks. Although some of the components and function of these networks have been elucidated, the mechanisms by which such intricate circuits originate and evolve remain poorly understood. Significant fractions of eukaryotic genomes are composed of transposable elements (TEs) that are able to move and replicate in the genome. TEs are best viewed as genomic parasites with no apparent adaptive value to the host cell in which they reside. Thus, in the absence of selective pressure to maintain mobility, the vast majority of TEs has become transpositionally inactivated over evolutionary time, and has been viewed as "junk DNA" without any apparent cellular function. However, it is being increasingly realized that the spread of TEs in the genome likely played a key role in the evolution of novel gene functions. For example, through an evolutionary process termed "domestication" TE-derived transposase proteins have been recurrently recruited into cellular pathways as regulatory elements. Harbi1 and Naif1 are highly conserved genes in vertebrates that have been derived from an ancient PIF/Harbinger transposon in a common ancestor of jawed vertebrates some 500 million years ago. Conservation of these genes implies that they have been under selection for important cellular functions, and their phylogenetic relationship suggests that both are involved in the same molecular pathway. Naif1 has been implicated in apoptotic functions. Our preliminary data suggest that Naif1 is a DNA-binding protein that promotes nuclear import of Harbi1, where it may recruit Harbi1 to genomic sites through protein-protein interactions. Sequence conservation of the catalytic domain of Harbi1 suggests that it might have retained nuclease activity. However, the role(s) of these two genes in cellular homeostasis and organismal development have been enigmatic. In this project we leverage biochemical assays in vitro, genome-wide ChIP and transcriptional profiling in cultured cells ex vivo, and phenotyping of knockout (KO) animals in the zebrafish model in vivo to understand and functionally annotate these genes and the genetic networks they regulate. The project will greatly contribute to our understanding of the mechanisms and roles of TEs as an important force in the creation of genetic novelty.