Genetic Analysis of Conserved microRNAs in C. elegans

描述（由申请人提供）：微小RNA（miRNAs）是动物发育和生理所需的基因表达的不可或缺的调节因子。此外，miRNAs与多种人类疾病有关，特别是心血管疾病、神经退行性疾病、糖尿病和癌症。然而，仅描述了少数单个miRNA的生物学功能。识别由单个miRNA直接调控的途径和过程以及识别特定的miRNA靶标对于理解它们在动物发育以及人类疾病中的作用至关重要。作为一种遗传上非常温顺的动物，C。线虫提供了一个理想的模型系统，其中研究的功能，特别是因为许多miRNA显示完全或接近完全的保护蠕虫和人类。In C.在线虫中，大多数miRNA不是发育所必需的;携带miRNA基因突变的蠕虫基本上正常发育。miRNAs可能与其他miRNAs和其他调节因子一起发挥作用，以控制发育途径。为了揭示这种复杂的相互作用，我们研究了遗传致敏背景中单个miRNA突变的影响。我们已经在遗传背景中鉴定了miRNA依赖性表型，由于Argonaute编码基因alg-1的丢失，总体miRNA活性较低。在这个提议中，我们将分析一个单一的miRNA依赖性表型，它与五个miRNA的丢失有关。本研究的目的是确定这五种抑制性miRNA调控的生物学途径和直接mRNA靶点。该提案的具体目标是：1）定义抑制性miRNA发挥作用的遗传途径。为此，我们将进行遗传分析，以测试与发育定时基因，miRNA途径基因和单个miRNA基因的相互作用，其功能已被描述; 2）表征alg-1发育定时缺陷的miRNA依赖性抑制的分子机制。为了做到这一点，我们将确定抑制性miRNA是否影响miRNA的生物发生或活性，检查候选miRNA靶点，并进行转录组分析，以确定缺失抑制性miRNA活性的蠕虫中失调基因的概况。公共卫生相关性：该提案重点关注蠕虫和哺乳动物之间保守的miRNA。实现该项目的目标将提供有关发育过程中miRNA调控的途径的关键信息，这些途径可能在人类疾病中被错误调控，并进一步描述miRNA调控动物靶mRNA的生物学原理。 MicroRNA是动物发育和生理所需的基因表达的不可或缺的调节因子。此外，microRNA与多种人类疾病有关，特别是心血管疾病、神经退行性疾病、糖尿病和癌症。目前尚不清楚microRNA活性的变化是人类疾病的原因还是结果。鉴定单个microRNA的生物学功能对于了解它们在动物发育以及人类疾病中的作用至关重要。

DESCRIPTION (provided by applicant): MicroRNAs (miRNAs) are indispensable regulators of gene expression that are required for animal development and physiology. In addition, miRNAs have been implicated in a wide spectrum of human diseases, notably cardiovascular disease, neurodegenerative disease, diabetes, and cancer. However, the biological functions of only a small number of individual miRNAs have been described. The identification of pathways and processes directly regulated by individual miRNAs and the identification of specific miRNA targets is vital to understand their role in animal development as well as in human disease. As an eminently genetically tractable animal, C. elegans provides an ideal model system in which to study the functions of miRNAs, particularly as many miRNAs show complete or near-complete conservation between worms and humans. In C. elegans, most miRNAs are not individually required for development; worms carrying mutations in miRNA genes develop essentially normally. It is likely that miRNAs function with other miRNAs and with additional regulatory factors to control developmental pathways. To reveal such complex interactions, we have examined the effects of mutations in individual miRNAs in genetically sensitized backgrounds. We have identified miRNA- dependent phenotypes in a genetic background with lower overall miRNA activity due to loss of an Argonaute-encoding gene, alg-1. In this proposal, we will analyze a single miRNA-dependent phenotype that is associated with the loss of five miRNAs. The objective of this proposal is to identify the biological pathways and direct mRNA targets regulated by these five suppressor miRNAs. The specific aims in this proposal are to: 1) define the genetic pathway in which suppressor miRNAs function. To do this, we will perform genetic analysis to test for interactions with developmental timing genes, miRNA pathway genes, and individual miRNA genes, for which a function has been described and 2) characterize the molecular mechanism for miRNA-dependent suppression of alg-1 developmental timing defects. To do this, we will determine if suppressor miRNAs affect the biogenesis or activity of miRNAs, examine candidate miRNA targets, and perform transcriptome analysis to identify the profile of misregulated genes in worms missing suppressor miRNA activity. PUBLIC HEALTH RELEVANCE: This proposal focuses on miRNAs conserved between worms and mammals. Achieving the goals of this project will provide key information about pathways regulated by miRNAs during development that may be misregulated in human disease and to further describe the biological principles of miRNA regulation of target mRNAs in animals. PUBLIC HEALTH RELEVANCE: MicroRNAs are indispensable regulators of gene expression that are required for animal development and physiology. In addition, microRNAs have been implicated in a wide spectrum of human diseases, notably cardiovascular disease, neurodegenerative disease, diabetes, and cancer. It is not clear whether changes in microRNA activity are a cause or a consequence of human disease. The identification of biological functions of individual microRNAs is vital to understand their role in animal development as well as in human disease.