MicroRNA对小肠干细胞自我更新和分化的调控

小肠上皮是成人机体自我更新最快的组织，而隐窝基底的小肠干细胞是小肠上皮各类细胞的“源泉”。小肠干细胞的自我更新和分化受到多条信号通路的调控，如Wnt、Notch和BMP等信号通路。DGCR8是miRNA生物合成过程中的关键蛋白，其在小肠上皮中的功能还缺少系统性研究。有研究揭示miRNA在小肠干细胞中有重要功能，但尚缺少高通量的方法用于筛选小肠干细胞所必需的miRNA。本项目拟利用基因敲除小鼠、转基因小鼠、小肠类器官和miRNA测序等工具和方法，研究DGCR8在小鼠小肠上皮中的功能，分析其分子调控网络，并绘制小肠干细胞的miRNA表达谱。基于DGCR8非依赖的miRNA表达系统，在Dgcr8敲除的小肠类器官中筛选小肠干细胞必需的miRNA，进而研究其分子机制。本项目有望系统研究DGCR8在小鼠小肠上皮中的功能，筛选小肠干细胞自我更新和分化所必需的miRNA，并深入揭示其分子调控网络。

The intestinal epithelium is the fastest self-renewing tissue in the adult body and all mature cells within the epithelium are originated from intestinal stem cells residing at the bottom of crypts. Differentiation of intestinal stem cells is delicately regulated by multiple signaling pathways, including Wnt, Notch and BMP signaling. DGCR8 is essential for biogenesis of miRNA, but the function of DGCR8 in mouse small intestine epithelium has not been systematically studied. Few researches revealed functions of miRNAs in intestinal stem cells, but it lacks high throughout methods to screen essential miRNAs in the self-renewal and differentiation of intestinal stem cells. This project intends to reveal the function and regulatory network of DGCR8 in intestinal epithelium with Dgcr8 conditional knockout mice and intestinal organoids. And, it also draws miRNA expression profiles of intestinal stem cells with Lgr5-EGFP-IRES-creERT2 transgenic mice, fluorescence activated cell sorting and miRNA sequencing. Using DGCR8-independent stable miRNA expression strategy, we will perform screening essential miRNAs in intestinal stem cells with Dgcr8 conditional knockout intestinal organoids. Furthermore, we will study the functions of essential miRNAs in Dgcr8 knockout organoids. Finally, this project is expected to improve our understanding in the function of DGCR8 in mouse intestinal epithelium, screen essential miRNAs in the self-renewal and differentiation of intestinal stem cells with Dgcr8 knockout intestinal organoids, and reveals the regulatory networks of these miRNAs.