Genome-wide Analysis of miRNAs and siRNAs associated with Abiotic Stress in Arabidopsis

近年来分子生物学中最重要的发现之一是小RNA分子对调控基因非常重要。植物有两种主要的小RNA，microRNAs(MiRNAs)和小干扰RNAs(SiRNAs)，可以通过它们在细胞中的合成方式来区分。这些小RNA的大小从21-24个核苷酸不等，由于它们太小了，多年来一直没有找到。尽管发现较晚，但小RNA在植物和动物中都非常重要，因为它们的功能是关闭基因。小RNA通过碱基配对相互作用识别它们沉默的目标基因。该项目将在全基因组范围内研究模式植物拟南芥的小RNA分子，并调查那些与植物如何响应环境胁迫相关的功能意义。对来自关键的小RNA代谢突变体和遭受胁迫的植物的少量收集的深度测序将导致每个样本超过200,000个序列。对数据的分析将提供关于哪些小RNA是miRNA，哪些是siRNA的洞察力。组装包含许多小RNA序列的DNA微阵列将提供测试额外突变体和应激对各种小RNA水平的调节作用的手段。这些数据以及将miRNA/靶RNA信息整合到计算机模型中将导致关于小RNA在环境压力中的作用的新假设。许多检测这些角色的工具都是成熟的，例如miRNA传感器基因，以及过表达和敲除品系。如果时间允许，在资助期内开始测试其中一些假设的主要预测将是直截了当的。本项目将确定miRNA参与环境胁迫的关键实例，并为未来研究其机制细节奠定基础。由于许多拟南芥miRNAs在水稻和其他植物中是保守的，这项工作应该会导致发现具有普遍意义的新功能。

One of the most important discoveries in molecular biology in recent years was that small RNA molecules are very important for regulating genes. Plants have two main types of small RNA, the microRNAs (miRNAs) and the small interfering RNAs (siRNAs), that can be distinguished by how they are made in the cell. These small RNAs range in size from about 21-24 nucleotides, and were missed for many years because they are so tiny. Despite their late discovery, small RNAs are very important in both plants and animals because they function to shut off genes. Small RNAs recognize the target genes they silence by base-pairing interactions. This project will investigate small RNA molecules of the model plant Arabidopsis on a genome-wide scale and investigate the functional significance of those associated with how plants respond to environmental stress. Deep sequencing of a small number of collections made from key small RNA metabolism mutants and plants subjected to stress will result in more than 200,000 sequences per sample. Analysis of the data will provide insight about which small RNAs are miRNAs and which are siRNAs. Assembly of a DNA microarray containing many small RNA sequences will provide the means to test additional mutants and stresses for their regulatory effect on the levels of various small RNAs. These data as well as the incorporation of miRNA/target RNA information into a computer model will lead to new hypotheses about the roles of small RNAs in environmental stress. Many tools to examine these roles are well-established, such as miRNA sensor genes, and overexpression and knockout lines. If time permits, it will be straightforward to begin testing major predictions of some of these hypotheses within the funding period. This project will identify key examples of miRNA involvement in environmental stress and lay the foundation for studying the mechanistic details in the future. Because many Arabidopsis miRNAs are conserved in rice and other plants, this work should lead to the discovery of new functions of general significance.