Characterization of Small Silencing RNAs in Emiliania Huxleyi

描述(申请人提供)：球石藻类是海洋浮游植物，产生优雅雕刻的碳酸钙细胞覆盖物，称为球藻。虎纹伊蚊(E.huxleyi)是一种模式球石动物，在骨形成和疾病方面具有潜在的生物医学应用。小分子沉默RNA(SmRNAs)在从单细胞到多细胞动植物的许多真核生物转录后基因调控中起着关键作用。小RNA参与多种细胞过程，这些过程对基因组的稳定、发育和对环境胁迫的适应性反应至关重要。这些自然的smRNA沉默过程也被称为RNA干扰，通过触发mRNAs的酶促切割来抑制蛋白质的表达。单链RNA过程的特异性已被用于系统分析基因功能和潜在的治疗目的。MicroRNAs(MiRNAs)和siRNAs(SiRNAs)是植物中已知的两类主要的smRNAs。目前对SMRNAs及其在E.huxleyi中的调控作用知之甚少。支持这项研究的具体假设是，E.huxleyi含有小RNA，如miRNAs和siRNAs，它们通过RNA沉默来调节目标基因。最近联合基因组研究所(JGI)公布了E.huxleyi基因组和使用高通量Solexa测序产生的大量E.huxleyi小RNA数据集，使得系统地表征E.huxleyi中的smRNAs及其调控作用成为可能。初步研究表明，在E.huxleyi中存在miRNAs。在这里，我们建议使用计算方法结合实验验证来表征E.huxleyi中的小RNA。具体地说，我们建议1)通过对短RNA序列的计算分析来识别和表征湖北棘豆中的miRNA前体和成熟的miRNAs；2)计算预测和表征湖北棘豆miRNAs的靶标，并用实验证据验证预测；以及3)识别和表征湖北棘豆中的siRNAs和其他类型的小RNA。据我们所知，这项拟议的研究将是第一次系统分析E.huxleyi中的smRNAs，因此将为这种微藻的转录后基因调控提供重要的见解。 与公共卫生的相关性：食囊虫目前正在由 生物技术产业作为新药物、omega-3脂肪酸、 生物燃料和多糖。在以下方面，它们对人类健康也有重要影响 了解钙化的过程，因为它与骨病、肾结石和 心脏病。对E.huxleyi中smRNA控制的基因调控的洞察可能会带来更好的 了解这种重要的微钙化物质的基本生物学，这可能为促进健康的生物矿化和减轻与 疾病状态。

DESCRIPTION (provided by applicant): Coccolithophorids are marine phytoplanktons that produce elegantly sculptured calcium carbonate cell coverings known as coccoliths. Emiliania huxleyi (E. huxleyi) is a model coccolithophorid that has potential biomedical applications related to bone formation and diseases. Small silencing RNAs (smRNAs) are known to play a pivotal role in post transcriptional gene regulation in many eukaryotes, from unicellular to multicellular plants and animals. Small RNAs are involved in a variety of cellular processes that are essential to genome stability, development, and adaptive responses to environmental stress. These natural smRNA silencing processes otherwise known as RNA interference, suppress protein expression by triggering the enzymatic cleavage of mRNAs. The specificity of smRNA processes has been exploited for the systematic analysis of gene functions and potential therapeutic purposes. MicroRNAs (miRNAs) and small interfering RNAs (siRNAs) are two major classes of smRNAs known in plants. Very little is currently known about smRNAs and their regulatory roles in E. huxleyi. The specific hypothesis underlying this proposed research is that E. huxleyi contains small RNAs such as miRNAs and siRNAs, which regulate target genes through RNA silencing. The recent release of the E. huxleyi genome by the Joint Genome Institute (JGI) and a large dataset of small RNAs of E. huxleyi generated using high-throughput Solexa sequencing, make it feasible to systematically characterize smRNAs and their regulatory roles in E. huxleyi. Preliminary studies show strong evidence for the existence of miRNAs in E. huxleyi. We propose herein to characterize small RNAs in E. huxleyi using computational methods, combined with experimental validation. Specifically, we propose to 1) identify and characterize miRNA precursors and mature miRNAs in E. huxleyi by computationally analyzing the short RNA sequences; 2) computationally predict and characterize targets of E. huxleyi miRNAs and validate the predictions with experimental evidences; and 3) identify and characterize siRNAs and other classes of small RNAs in E. huxleyi. To our knowledge, this proposed research will be the first systemic analysis of smRNAs in E. huxleyi, and as such will provide important insights into post-transcriptional gene regulation in this microalga. PUBLIC HEALTH RELEVANCE: Coccolithophorids are presently being investigated by the biotechnology industry as a potential source for new pharmaceuticals, omega-3 fatty acids, biofuel, and polysaccharides. They also have important implications for human health in terms of understanding the process of calcification as it relates to bone disease, kidney stones, and heart disease. Insights into smRNA controlled gene regulation in E. huxleyi may lead to a better understanding of the fundamental biology of this important microcalcifier, which may offer new strategies to promote healthy biomineralization and mitigate calcification associated with disease states.