长链非编码 RNA Snhg3 对于小鼠胚胎干细胞自我更新和多能性至关重要。
The long non-coding RNA Snhg3 is essential for mouse embryonic stem cell self-renewal and pluripotency.
机构信息
The State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou, 510060, China.
MOE Key Laboratory of Gene Function and Regulation, Guangzhou Key Laboratory of Healthy Aging Research and State Key Laboratory of Biocontrol, SYSU-BCM Joint Research Center, School of Life Sciences, Sun Yat-sen University, Guangzhou, 510275, China.
出版信息
Stem Cell Res Ther. 2019 May 31;10(1):157. doi: 10.1186/s13287-019-1270-5.
BACKGROUND
Small nucleolar RNA host gene 3 (Snhg3) is a long non-coding RNA (lncRNA) that was shown to participate in the tumorigenesis of certain cancers. However, little is known about its role in embryonic stem cells (ESCs).
METHODS
Here, we investigated the role of Snhg3 in mouse ESCs (mESCs) through both loss-of-function (knockdown) and gain-of-function (overexpression) approaches. Alkaline phosphatase staining, secondary colony formation, propidium iodide staining, western blotting, and quantitative reverse transcription polymerase chain reaction (qRT-PCR) were used to access self-renewal capacity, whereas immunofluorescence, qRT-PCR, and embryoid body formation were performed to examine pluripotency. In addition, the effect of Snhg3 on mouse embryonic development was determined based on the morphological changes, blastocyst rate, and altered pluripotency marker (Nanog, Oct4) expression. Moreover, the relationship between Snhg3 and key pluripotency factors was evaluated by chromatin immunoprecipitation qPCR, qRT-PCR, subcellular fractionation, and RNA immunoprecipitation. Finally, RNA pull-down and mass spectrometry were applied to explore the potential interacting proteins of Snhg3 in mESCs.
RESULTS
We demonstrated that Snhg3 is essential for self-renewal and pluripotency maintenance in mESCs. In addition, Snhg3 knockdown disrupted mouse early embryo development. Mechanistically, Snhg3 formed a positive feedback network with Nanog and Oct4, and 126 Snhg3-interacting proteins were identified in mESCs.
CONCLUSIONS
Snhg3 is essential for mESC self-renewal and pluripotency, as well as mouse early embryo development.
背景
小核仁 RNA 宿主基因 3（Snhg3）是一种长链非编码 RNA（lncRNA），已被证明参与某些癌症的发生。然而，其在胚胎干细胞（ESCs）中的作用知之甚少。
方法
在这里，我们通过功能丧失（敲低）和功能获得（过表达）方法研究了 Snhg3 在小鼠胚胎干细胞（mESCs）中的作用。碱性磷酸酶染色、次级集落形成、碘化丙啶染色、Western blot 和实时定量聚合酶链反应（qRT-PCR）用于评估自我更新能力，而免疫荧光、qRT-PCR 和类胚体形成用于评估多能性。此外，基于形态变化、囊胚率和改变的多能性标记物（Nanog、Oct4）表达，确定了 Snhg3 对小鼠胚胎发育的影响。此外，通过染色质免疫沉淀 qPCR、qRT-PCR、亚细胞分离和 RNA 免疫沉淀评估了 Snhg3 与关键多能性因子之间的关系。最后，应用 RNA 下拉和质谱分析来探索 Snhg3 在 mESCs 中的潜在相互作用蛋白。
结果
我们证明了 Snhg3 对于 mESCs 的自我更新和多能性维持是必不可少的。此外，Snhg3 敲低破坏了小鼠早期胚胎发育。在机制上，Snhg3 与 Nanog 和 Oct4 形成正反馈网络，在 mESCs 中鉴定出 126 个 Snhg3 相互作用蛋白。
结论
Snhg3 对于 mESC 的自我更新和多能性以及小鼠早期胚胎发育是必不可少的。
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