RNA Biology in Oxytricha: a Complex Unicellular Model

 描述（由申请人提供）： RNA分子显示一种新的行动模式，在池塘居住的纤毛虫原生生物，长和小的非编码RNA调节基因组重塑的复杂过程。Oxytricha中的母系遗传的皮尔纳分子标记了基因组的225，000个片段，并且长模板RNA在大规模基因组重排期间对这些DNA片段的顺序进行编程。这项研究的目标是了解这些长和小的非编码RNA的生物起源和调控，以及模式生物Oxytricha中RNA产生途径中其他步骤的调控。扩展我们用于研究和操纵这种新兴遗传系统的工具套件也是一个高度优先事项，其中一个目标是利用我们在这些生物体中的皮尔纳生物学知识来开发一种强大的基因敲除工具。这将反过来帮助我们更好地理解基因组重排的遗传学和发育生物学的其他方面，以及它对基因组完整性的维持和调节的影响。 拟议中的实验将检查参与这些复杂重排事件的分子的性质以及它们背后的机制。从拟议的研究中获得的最直接的见解是了解长和小非编码RNA分子的活动范围，特别是Oxytricha中的皮尔纳途径，并注意该系统与其他真核生物之间的差异和相似之处。具体目标包括：1.研究尖毛藻属中的piRNA及其生物发生，2.开发piRNA介导的基因敲除技术，作为尖毛藻属的第一个基因敲除工具，3.理解长的非编码模板RNA的产生及其相互作用的伙伴，4。鉴定和功能测试调控体细胞染色体基因表达的基序。

 DESCRIPTION (provided by applicant): RNA molecules display a novel mode of action in pond-dwelling ciliated protists, where both long and small noncoding RNAs regulate an intricate process of genome remodeling. Maternally-inherited piRNA molecules in Oxytricha mark which 225,000 segments of the genome to retain, and long, template RNAs program the order of those DNA segments during massive genome rearrangements. The proposed goal of this research is to understand the biogenesis and regulation of both these long and small noncoding RNAs, as well as the regulation of other steps in pathways of RNA production in the model organism Oxytricha. Expanding our suite of tools for studying and manipulating this emerging genetic system is also a high priority, and one aim will capitalize on our knowledge of piRNA biology in these organisms to develop a powerful tool for gene knockout. This will, in turn, help us better understand other aspects of the genetics and developmental biology of genome rearrangements, as well as its implications for the maintenance and regulation of genome integrity, more generally. The proposed experiments will examine the properties of both the molecules that participate in these complex rearrangement events and the machinery underlying them. The most immediate insight to be gained from the proposed research is an understanding of the range of activities accessible to both long and small noncoding RNA molecules, and the piRNA pathway in Oxytricha, in particular, with attention to both differences and similarities between this system and other eukaryotic organisms. Specific aims include: 1. Studying piRNAs and their biogenesis in Oxytricha, 2. Develop piRNA-mediated gene knockout as the first gene knockout tool in Oxytricha, 3. Understand long, noncoding template RNA production and their interacting partners, 4. Identify and functionally test motifs that regulate gene expression from somatic chromosomes.