SLC7A5 is critical for Paneth cell integrity and function

The intestine is the critical organ not only for processing and resorbing nutrients from ingested food but also for defending the organism from external stresses such as pathogens. These functions are mainly carried out by the epithelium which is constantly being self-renewed throughout adult life. Intestinal epithelial homeostasis is maintained through well-controlled cell proliferation of the stem cells and transient amplifying cells and the apoptotic degeneration of epithelial cells, mostly at or near the tip of the villi. Many genes and pathways have been found to influence intestinal epithelial cell proliferation. Among them is the mTORC1 signaling pathway, whose activation is known to increase cell proliferation. Here, we report the first intestinal epithelial specific knockout (IEC-KO) of an amino acid transporter capable of activating mTORC1. We show that the transporter, slc7a5, is highly expressed in the intestinal crypt and slc7a5 IEC-KO leads to expected reduction in mTORC1 signal in the crypt or even in intestinal organoids in vitro. Surprisingly, slc7a5 IEC-KO leads to increased proliferation of both transit amplifying cells and crypt base stem cells but a reduction in the secretory cells, particularly mature Paneth cells in the crypt base. Our scRNA-seq and electron microscopic analyses reveals that slc7a5 IEC-KO causes dedifferentiation of the Paneth cells, leading to drastically reduced secretory granules and lysozyme expression without affecting the overall Paneth cell number. We further show that slc7a5 IEC-KO mice are prone to induced colitis due to this loss of Paneth cell differentiation. We propose a model where slc7a5 regulates secretory cell differentiation to affect stem cell niche and/or inflammatory response to regulate cell proliferation in the crypts. Comparison of slc7a5 fl/flwith slc7a5 IEC-KO enriched intestinal crypts

肠道不仅是消化并吸收摄入食物中营养物质的关键器官，同时也是机体抵御病原体等外界应激的重要防线。这些功能主要由上皮组织执行，而上皮组织在成年后仍会持续进行自我更新。肠道上皮稳态通过严格调控干细胞与瞬时扩增细胞的增殖，以及主要在绒毛顶端或其附近发生的上皮细胞凋亡退化得以维持。已有诸多基因及通路被发现可影响肠道上皮细胞的增殖，其中mTORC1信号通路（mTORC1 signaling pathway）的激活已被证实可促进细胞增殖。本研究首次报道了一种可激活mTORC1信号通路的氨基酸转运蛋白的肠道上皮特异性敲除（IEC-KO）模型。研究发现，该转运蛋白slc7a5在肠隐窝中高表达，且slc7a5 IEC-KO可导致隐窝甚至体外肠道类器官中的mTORC1信号通路活性出现预期的降低。令人意外的是，slc7a5 IEC-KO不仅会增加瞬时扩增细胞与隐窝基底干细胞的增殖，还会减少分泌细胞的数量，尤其是隐窝基底处的成熟潘氏细胞（Paneth cells）。通过单细胞RNA测序（scRNA-seq）与电镜分析，我们证实slc7a5 IEC-KO会引发潘氏细胞的去分化，使其分泌颗粒与溶菌酶的表达显著降低，但不会改变潘氏细胞的总数量。我们进一步发现，由于潘氏细胞分化功能的缺失，slc7a5 IEC-KO小鼠更易诱发结肠炎。据此我们提出模型：slc7a5通过调控分泌细胞分化，影响干细胞龛（stem cell niche）和/或炎症反应，进而调控肠隐窝内的细胞增殖。对slc7a5 fl/fl与slc7a5 IEC-KO富集的肠隐窝进行对比分析。