体细胞重编程中miRNA的鉴定及功能研究

程红

摘 要：该研究针对非编码RNA研究中的核心科学问题，研究体细胞重编程微RNA功能及调控网络。研究的5年预期目标是：利用分子生物学、细胞生物学、生物化学和信息生物学手段，完成重编程和体细胞重编程和多能性细胞分化多个阶段的微RNA的表达谱，鉴定出在以上多个阶段表达变化显著的微RNA。找到影响重编程或多能干细胞分化过程的几种微RNA及鉴定其靶基因，并揭示这些基因在上述过程的作用。鉴定到在转录后加工水平和核质转运过程受调控的微RNA，阐明这些特定微RNA的作用机理，揭示其核质转运变化与重编程和多能性细胞分化之间的关联，阐明其分子机制。阐明上述相关微RNA在胚胎发育、细胞分化和重编程过程中的作用及其自身生物合成规律，并揭示其在器官发育中的作用。近3年，已建立体细胞重编程和多能性细胞分化及体细胞转分化的研究系统。采用核移植技术，建立了可用于研究体细胞重编程中miRNA功能的新系统——来自孤雄囊胚的单倍体胚胎干细胞系。已经建立了研究miRNA核质转运和转录后加工调控的研究体系。制作了miRNA敲除小鼠。系统鉴定了体细胞重编程过程中差异表达的miRNA。系统鉴定了体细胞向肝细胞转分化过程中显著差异表达的miRNA。发现了RNA结合蛋白在转录后加工水平调控的特定miRNA的表达。发现在哺乳动物细胞核内存在一种读码机制，可能参与调控非编码RNA的核质转运。揭示了miR-126在小鼠胚胎血管新生和血管完整性维持中的作用，探究了其发挥作用的分子机制。

关键词：非编码RNA 细胞重编程 干细胞分化 微RNA 核质转运

Abstract： This proposal is focused on microRNAs that are involved in cell reprogramming. Using molecular biological， cell biological and biochemical methods， we plan to establish the expression profile of microRNAs at multiple stages of cell reprogramming and stem cell differentiation， and identify microRNAs whose expression level change significantly during these processes. Through these studies， we hope to identify microRNAs that play important regulatory roles in these processes and identify their target genes. We also plan to identify microRNAs whose expression levels are regulated at post-transcriptional levels and investigate the molecular mechanism. In the recent three years， we have established the systems that are suitable for study the processed of cell reprogramming and differentiation. Using nuclear transfer technique， we have established the systems for generation of mice by oocyte injection of androgenetic haploid embryonic stem cells that can be used for function of microRNA. We have also set up the research system for study of nucleocytoplasmic transportation and processing of microRNAs. We have obtained a line of microRNA knockout mouse. According to our plan， we have identified multiple microRNAs whose expression level changed significantly during cell reprogramming and trans-differentiation of somatic cells to hepatocellular cells. We have investigated the function and mechanism for mir-126 in regulating vescular integrity. In the past three years， we have published 10 research papers including one paper that was published in Cell.

Key Words： Non-coding RNA； Cell reprogramming； Stem cell differentiation； MicroRNA； Nucleocytoplasmic transportation

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