Department of Biomedical Sciences, College of Veterinary Medicine, Cornell University, Ithaca, New York.
Department of Immunology, University of Washington, Seattle, Washington.
Am J Physiol Gastrointest Liver Physiol. 2021 Dec 1;321(6):G668-G681. doi: 10.1152/ajpgi.00222.2021. Epub 2021 Oct 13.
MicroRNA-mediated regulation is critical for the proper development and function of the small intestinal (SI) epithelium. However, it is not known which microRNAs are expressed in each of the cell types of the SI epithelium. To bridge this important knowledge gap, we performed comprehensive microRNA profiling in all major cell types of the mouse SI epithelium. We used flow cytometry and fluorescence-activated cell sorting with multiple reporter mouse models to isolate intestinal stem cells, enterocytes, goblet cells, Paneth cells, enteroendocrine cells, tuft cells, and secretory progenitors. We then subjected these cell populations to small RNA-sequencing. The resulting atlas revealed highly enriched microRNA markers for almost every major cell type (https://sethupathy-lab.shinyapps.io/SI_miRNA/). Several of these lineage-enriched microRNAs (LEMs) were observed to be embedded in annotated host genes. We used chromatin-run-on sequencing to determine which of these LEMs are likely cotranscribed with their host genes. We then performed single-cell RNA-sequencing to define the cell type specificity of the host genes and embedded LEMs. We observed that the two most enriched microRNAs in secretory progenitors are miR-1224 and miR-672, the latter of which we found is deleted in hominin species. Finally, using several in vivo models, we established that miR-152 is a Paneth cell-specific microRNA. In this study, first, microRNA atlas (and searchable web server) across all major small intestinal epithelial cell types is presented. We have demonstrated microRNAs that uniquely mark several lineages, including enteroendocrine and tuft. Identification of a key marker of mouse secretory progenitor cells, miR-672, which we show is deleted in humans. We have used several in vivo models to establish miR-152 as a specific marker of Paneth cells, which are highly understudied in terms of microRNAs.
miRNA 介导的调控对于小肠 (SI) 上皮的正常发育和功能至关重要。然而，尚不清楚哪些 miRNA 表达于 SI 上皮的每种细胞类型中。为了填补这一重要的知识空白，我们对小鼠 SI 上皮的所有主要细胞类型进行了全面的 miRNA 谱分析。我们使用流式细胞术和荧光激活细胞分选技术，结合多个报告小鼠模型，分离出肠道干细胞、肠上皮细胞、杯状细胞、潘氏细胞、肠内分泌细胞、微绒毛细胞和分泌祖细胞。然后，我们对这些细胞群体进行了小 RNA 测序。由此产生的图谱揭示了几乎每种主要细胞类型都高度富集的 miRNA 标志物（https://sethupathy-lab.shinyapps.io/SI_miRNA/）。观察到其中一些谱系特异性 miRNA（LEMs）被嵌入到注释的宿主基因中。我们使用染色质转录跑速测序来确定这些 LEMs 中有哪些可能与其宿主基因共转录。然后，我们进行了单细胞 RNA 测序，以确定宿主基因和嵌入 LEM 的细胞类型特异性。我们观察到分泌祖细胞中最富集的两个 miRNA 是 miR-1224 和 miR-672，后者在人科物种中缺失。最后，使用几种体内模型，我们确定了 miR-152 是潘氏细胞特异性 miRNA。在这项研究中，首先呈现了 across all major small intestinal epithelial cell types 的 miRNA 图谱（和可搜索的网络服务器）。我们已经证明了一些独特标记几种谱系的 miRNA，包括肠内分泌细胞和微绒毛细胞。鉴定出了小鼠分泌祖细胞的关键标记物 miR-672，我们发现其在人类中缺失。我们使用几种体内模型确定了 miR-152 是潘氏细胞的特异性标记物，而潘氏细胞在 miRNA 方面的研究非常少。
