哺乳類脳神経幹細胞の特性の経時変化を制御するエピゲノム修飾と細胞間シグナルの研究

在过去的一年里，吴继续分析了纤维蛋白在神经干细胞(NSCs)时间命运变化中的作用。结果，Wu发现，在增殖期，RNAi介导的Fibrillarin基因敲除~50%促进了NSCs的神经元提前分化。而对于神经干细胞自我更新起重要作用的Notch信号通路，在Fibrillarin被敲除后，其活性明显降低。相反，纤维蛋白的过度表达增加了神经干细胞的数量。这些结果表明，纤维素对神经干细胞的增殖是必需的。接下来，吴试图揭示Fibrillarin影响神经干细胞命运的机制。由于Fibrillarin是rRNA的甲基转移酶，rRNA的甲基化水平可能会影响翻译，吴问到，Fibrillarin的敲除是影响整体蛋白质翻译，还是只影响特定的靶点。利用化学探针追踪新合成的蛋白质，吴证实了RNAi介导的Fibrillarin基因敲除到~50%不会影响全球蛋白质的翻译。这些结果表明，Fibrillarin通过优先翻译靶mRNAs来调节细胞命运。然后，为了寻找Fibrillarin的靶标，吴与岩崎志野博士合作，进行了核糖体图谱分析，这是一种基于深度测序的检测翻译mRNAs的新方法。即使这项实验仍在进行中，我认为这项实验的结果将有助于我们更好地了解NSC命运是如何被精确控制的。在完成JSPS奖学金后，吴将继续在我的实验室完成这个项目。

In the last year, Wu continued to analyze the function of Fibrillarin in temporal fate change of neural stem cells (NSCs). As a result, Wu found that RNAi-mediated knockdown of Fibrillarin to ~50% at proliferative stage promoted premature neuronal differentiation of NSCs. Consistently, the activity of Notch signaling which is important for self-renewal of NSCs was strongly reduced after knockdown of Fibrillarin. In contrast, overexpression of Fibrillarin increased the population of NSCs. These results suggest that Fibrillarin is essential for proliferation of NSCs. Next, Wu tried to uncover the mechanism by which Fibrillarin affects the fate of NSCs. Because Fibrillarin is a methyltransferase of rRNA and methylation level of rRNA may affect translation, Wu asked that whether knockdown of Fibrillarin affects global protein translation or only impacts on specific targets. Using chemical probe to trace newly synthesized protein, Wu confirmed that RNAi-mediated knockdown of Fibrillarin to ~50% did not affect global protein translation. These results imply that Fibrillarin regulates cell fate through preferentially translating of target mRNAs. Then, to search the targets of Fibrillarin, Wu, cooperating with Dr. Iwasaki Shitano, performed ribosome profiling, a novel method to detect translating mRNAs based on deep-sequencing. Even this experiment is still ongoing, I think that results of this experiment will help us to better understand how NSC fate is precisely controlled. After finish of JSPS fellowship, Wu will continue to complete this project in my lab.