Regulation of microRNA Function and Cell Proliferation of Mouse Embryonic Stem Ce

MicroRNAs （miRNAs）是一类丰富的小rna，可抑制靶信使rna （mrna）的表达。mirna调节多种生物过程，对哺乳动物发育至关重要，并与许多疾病有关。尽管事实上mirna调控着哺乳动物基因组的50%，但mirna沉默途径的确切机制和组成部分仍然知之甚少。为了发挥其调节功能，miRNA被整合到一种称为miRNA诱导沉默复合物（miRISC）的核糖核蛋白复合物中，该复合物被引导到mRNA靶点。miRNA诱导的沉默效应是多种多样的；虽然一些靶标发生降解，但其他靶标的翻译受到抑制。越来越明显的是，其他蛋白质辅助因子可以与miRISC相互作用并调节miRISC活性的性质和功效。在这些蛋白辅助因子中，最近发现在各种生物体中控制发育和干细胞命运的TRIM-NHL蛋白家族成员调节miRNA功能。我们发现Trim71是trimm - nhl家族中受发育调节的成员，是一种新型miRISC相互作用蛋白，可增强mirna的功能，对小鼠胚胎干细胞（ES）的快速增殖至关重要。我们的目标是深入了解Trim71在小鼠胚胎干细胞中mirna介导的基因抑制和细胞增殖中的作用。我们的初步数据表明，Trim71是ES细胞中一个大的多亚基核糖核蛋白复合物的组成部分。我们将采用多种方法来鉴定含有trim71复合物的蛋白质和RNA成分。此外，我们将对Trim71进行详细的生化分析，以确定与miRISC相互作用、增强miRNA功能和控制ES细胞增殖所需的重要蛋白质结构域的功能。完成这一提议的目标将提供一个更好的理解干细胞是如何在分子水平上被调节的，并有助于开发新的治疗方法。

MicroRNAs (miRNAs) are an abundant class of small RNAs that repress the expression of target messenger RNAs (mRNAs). MiRNAs regulate a variety of biological processes, are essential for mammalian development and have been implicated in numerous diseases. Despite the fact that miRNAs regulate up to 50% of the mammalian genome, the exact mechanism and components of the miRNA-silencing pathway are still poorly understood. To exert their regulatory function, miRNAs are incorporated into a ribonucleoprotein complex termed miRNA- induced silencing complex (miRISC) that is guided to mRNA targets. The effects of miRNA- induced silencing are diverse; while some targets undergo degradation others are translationally repressed. It has become increasingly evident that additional protein cofactors can interact with miRISC and modulate the nature and efficacy of miRISC activity. Among these protein cofactors, members of the TRIM-NHL protein family that control development and stem cell fate in various organisms were recently found to modulate miRNA function. We have identified Trim71, a developmentally regulated member of the TRIM-NHL family, as a novel miRISC- interacting protein that enhances the function of miRNAs and is critical for rapid proliferation of mouse embryonic stem (ES) cells. Our goal is to gain mechanistic insight into the role of Trim71 in miRNA-mediated gene repression and cell proliferation in mouse ES cells. Our preliminary data indicates that Trim71 is a component of a large multi-subunit ribonucleoprotein complex in ES cells. We will employ a variety of approaches to identify the protein and RNA components of Trim71-containing complexes. In addition, we will perform a detailed biochemical analysis of Trim71 to functionally define important protein domains required for interaction with miRISC, enhancement of miRNA function and control of ES cell proliferation. Accomplishing the goals of this proposal will provide a better understanding of how stem cells are regulated at the molecular level and could aid in the development of novel therapeutics.