MicroRNA Function in Human Embryonic Stem Cells

MicroRNA 在人类胚胎干细胞中的功能

• 批准号: 7686802
• 负责人: Hannele RUOHOLA-BAKER
• 金额: $ 29.64万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-15 至 2012-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7686802
• 关键词: Adult; Binding; Biological Assay; Cell physiology; Cells; Dicer Enzyme; Drosophila genus; Functional RNA; Future; Gene Targeting; Genes; Genetic Translation; Goals; Informatics; Mediating; Messenger RNA; MicroRNAs; Mus; Phenotype; Plants; Process; RNA Interference; Regenerative Medicine; Regulation; Research; Role; Stem cells; Testing; Tissues; Transplantation; aptamer; base; embryonic stem cell; gain of function; genome-wide; human DICER1 protein; human embryonic stem cell; loss of function; public health relevance; research study; self-renewal; small molecule; tumor

DESCRIPTION (provided by applicant): The long-term goal of this research is to gain a better understanding and control over human embryonic stem cells (hESCs) by identifying and manipulating the key microRNAs required for hESC self-renewal and differentiation. MicroRNAs, small (21-23nt) non-coding RNA molecules that can regulate mRNA translation and stability by binding to the target mRNAs with incomplete complementarity, are required for normal stem cell function in mouse, Drosophila and plants. We have previously shown that microRNAs are required for germline stem cell (GSC) self-renewing divisions in Drosophila. Our preliminary results suggest that microRNAs are also critical for stem cell self-renewing division and differentiation in human ESCs, and the goal of this proposal is to identify the critical microRNAs and their mRNA targets in these cells. Towards this goal, we will analyze the phenotypes observed in hESCs lacking mature microRNAs due to gene knockdown (KD) of microRNA processing enzymes, Dicer or Drosha. We will identify up-regulated mRNAs in these KD-lines by genome wide mRNA profiling and determine whether they are responsible for the Dicer or Drosha phenotypes through loss-of-function (LOF), gain-of-function (GOF) and rescue experiments (Aim1). To identify the critical microRNAs that mediate this regulation we will carry out genome wide QPCR profiling of hESC microRNAs. The function of the hESC enriched microRNAs will be analyzed by LOF, GOF and rescue experiments (Aim2). The two aims will connect: The critical microRNAs should have the capacity to rescue dicer KD phenotypes (Aim 2.2) and may be sufficient to down-regulate the target genes identified in Aim 1.2 (Aim 2.3). This study should result in findings that allow better manipulation and more efficient differentiation of hESC which is of particular importance for regenerative medicine. PUBLIC HEALTH RELEVANCE: This study will be the first analysis of the functions of mature miRNAs in hESCs. Identification of the miRNA function in hESCs holds the future for better control and manipulation of the hESCs in regenerative medicine. This is important due to the troubling fact that embryonic stem cells tend to form tumors when transplanted into adult tissue. microRNAs carry hope as molecules which can control stem cell self-renewing division.

描述（由申请人提供）：本研究的长期目标是通过鉴定和操纵hESC自我更新和分化所需的关键microRNA，更好地了解和控制人胚胎干细胞（hESC）。MicroRNA是一种小的（21 - 23 nt）非编码RNA分子，可以通过与具有不完全互补性的靶mRNA结合来调节mRNA的翻译和稳定性，是小鼠、果蝇和植物正常干细胞功能所必需的。我们先前已经表明，microRNA是果蝇生殖干细胞（GSC）自我更新分裂所必需的。我们的初步研究结果表明，microRNA也是干细胞自我更新的分裂和分化在人类胚胎干细胞的关键，这个建议的目标是确定这些细胞中的关键microRNA及其mRNA的目标。为了实现这一目标，我们将分析由于microRNA加工酶Dicer或Drosha的基因敲低（KD）而缺乏成熟microRNA的hESC中观察到的表型。我们将通过全基因组mRNA分析鉴定这些KD系中上调的mRNA，并通过功能丧失（LOF）、功能获得（GOF）和拯救实验（Aim1）确定它们是否与Dicer或Drosha表型有关。为了鉴定介导这种调节的关键microRNA，我们将对hESC microRNA进行全基因组QPCR分析。将通过LOF、GOF和拯救实验（Aim2）分析hESC富集的microRNA的功能。这两个目标将相互联系：关键microRNA应具有拯救dicer KD表型的能力（目标2.2），并可能足以下调目标1.2中鉴定的靶基因（目标2.3）。这项研究的结果应该允许更好的操作和更有效的分化的hESC，这是再生医学特别重要的发现。公共卫生相关性：这项研究将是第一次分析成熟miRNAs在hESC中的功能。鉴定miRNA在hESC中的功能为在再生医学中更好地控制和操纵hESC提供了未来。这一点很重要，因为胚胎干细胞在移植到成人组织中时往往会形成肿瘤。microRNA作为能够控制干细胞自我更新分裂的分子带来了希望。