Deletion of Polycomb Repressive Complex 2 From Mouse Intestine Causes Loss of Stem Cells
Deletion of Polycomb Repressive Complex 2 From Mouse Intestine Causes Loss of Stem Cells
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DOI：
10.1053/j.gastro.2016.06.020
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发表时间：
2016-10-01
期刊：
影响因子：
29.4
通讯作者：
van Lohuizen, Maarten
中科院分区：
文献类型：
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作者：
Koppens, Martijn A. J.;Bounova, Gergana;van Lohuizen, Maarten
背景与目的：多梳抑制复合物 2 (PRC2) 通过抑制基因表达来调节分化，从而稳定干细胞及其后代的命运。 PRC2 有助于维持成体干细胞群，但对其在肠道干细胞中的功能知之甚少。我们研究了肠道特异性删除 PRC2 蛋白胚胎外胚层发育 (EED)（PRC2 功能所需的亚基）和 zeste 同源物 2 (EZH2)（一种组蛋白甲基转移酶）增强子的小鼠的表型。方法：我们对 AhCre 进行了研究； EedLoxP/LoxP（EED 敲除）小鼠和 AhCre； Ezh2LoxP/LoxP（EZH2 敲除）小鼠，其 EED 和 EZH2 分别具有肠道特异性破坏。分离小肠隐窝并随后培养以生长类器官。通过免疫组织化学、原位杂交、RNA 序列和染色质免疫沉淀方法对肠道和类器官进行分析。结果：与野生型对照小鼠相比，EED 基因敲除小鼠的肠道出现大量隐窝变性，增殖细胞数量减少。 Cdkn2a 的抑制被解除，我们检测到 P21 水平升高。我们没有观察到 EZH2 敲除小鼠和对照小鼠之间存在任何差异。 EED 敲除小鼠的肠隐窝有未定型隐窝细胞异常分化的迹象——这些细胞向分泌细胞谱系分化。此外，EED 敲除小鼠的隐窝 Wnt 信号传导受损，并伴随肠道干细胞损失，这种表型在类器官中异位刺激 Wnt 和 Notch 信号传导后不会逆转。对 EED 敲除小鼠肠道组织的基因表达模式分析显示，参与 Wnt 信号传导的多个基因失调。 Wnt 信号传导直接受 PRC2 调节。结论：在小鼠肠道组织中，PRC2 通过促进肠道干细胞区室增殖并防止分化来维持小肠干细胞。 PRC2 控制调节肠道稳态的多个信号通路中的基因表达。测序数据可在基因组数据存储库 GEO 中获取，参考系列 GSE81578； RNA 测序数据可在子系列 GSE81576 下获取； ChIP 测序数据可在子系列 GSE81577 下获取。
BACKGROUND & AIMS: The polycomb repressive complex 2 (PRC2) regulates differentiation by contributing to repression of gene expression and thereby stabilizing the fate of stem cells and their progeny. PRC2 helps to maintain adult stem cell populations, but little is known about its functions in intestinal stem cells. We studied phenotypes of mice with intestine-specific deletion of the PRC2 proteins embryonic ectoderm development (EED) (a subunit required for PRC2 function) and enhancer of zeste homolog 2 (EZH2) (a histone methyltransferase). METHODS: We performed studies of AhCre; EedLoxP/LoxP (EED knockout) mice and AhCre; Ezh2LoxP/LoxP (EZH2 knockout) mice, which have intestine-specific disruption in EED and EZH2, respectively. Small intestinal crypts were isolated and subsequently cultured to grow organoids. Intestines and organoids were analyzed by immunohistochemical, in situ hybridization, RNA sequence, and chromatin immunoprecipitation methods. RESULTS: Intestines of EED knockout mice had massive crypt degeneration and lower numbers of proliferating cells compared with wild-type control mice. Cdkn2a became derepressed and we detected increased levels of P21. We did not observe any differences between EZH2 knockout and control mice. Intestinal crypts from EED knockout mice had signs of aberrant differentiation of uncommitted crypt cells-these differentiated toward the secretory cell lineage. Furthermore, crypts from EED-knockout mice had impaired Wnt signaling and concomitant loss of intestinal stem cells, this phenotype was not reversed upon ectopic stimulation of Wnt and Notch signaling in organoids. Analysis of gene expression patterns from intestinal tissues of EED knockout mice showed dysregulation of several genes involved in Wnt signaling. Wnt signaling was regulated directly by PRC2. CONCLUSIONS: In intestinal tissues of mice, PRC2 maintains small intestinal stem cells by promoting proliferation and preventing differentiation in the intestinal stem cell compartment. PRC2 controls gene expression in multiple signaling pathways that regulate intestinal homeostasis. Sequencing data are available in the genomics data repository GEO under reference series GSE81578; RNA sequencing data are available under subseries GSE81576; and ChIP sequencing data are available under subseries GSE81577.