The neural RNA-binding protein Musashi1 translationally regulates mammalian numb gene expression by interacting with its mRNA
The neural RNA-binding protein Musashi1 translationally regulates mammalian numb gene expression by interacting with its mRNA
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DOI：
10.1128/mcb.21.12.3888-3900.2001
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发表时间：
2001-06-01
影响因子：
5.3
通讯作者：
Okano, H
中科院分区：
文献类型：
--
作者：
Imai, T;Tokunaga, A;Okano, H
Musashi1（Msi1）是一种RNA结合蛋白，在神经祖细胞（包括神经干细胞）中高度表达。在这项研究中，通过使用简并的50聚体序列库进行体外筛选，确定了Msi1的RNA结合序列。所有筛选出的RNA种类都包含（G/A）U(n)AGU（n = 1到3）序列的重复，这些序列对Msi1结合至关重要。在一些神经mRNA中鉴定出了这些共有元件。其中之一是哺乳动物的Numb（m - numb），它编码一种Notch信号的膜相关拮抗剂，很可能是Msi1的一个靶标。如通过亲和沉淀随后进行逆转录 - PCR所示，Msi1蛋白在体外转录的m - numb RNA的3' - 非翻译区（UTR）结合，并在体内结合内源性m - numb mRNA。此外，腺病毒诱导的Msi1表达导致内源性m - Numb蛋白表达下调。使用将荧光素酶和m - numb的3' - UTR组合的嵌合mRNA进行的报告基因分析表明，Msi1在不改变报告基因mRNA水平的情况下降低了报告基因活性。因此，我们的结果表明Msi1可以在翻译水平调节其靶基因的表达。此外，我们发现与m - numb基因的转录后下调相关，Msi1表达使Notch信号活性增加。
Musashi1 (Msi1) is an RNA-binding protein that is highly expressed in neural progenitor cells, including neural stem cells. In this study, the RNA-binding sequences for Msi1 were determined by in vitro selection using a pool of degenerate 50-mer sequences. All of the selected RNA species contained repeats of (G/A)U(n)AGU (n = 1 to 3) sequences which were essential for Msi1 binding. These consensus elements were identified in some neural mRNAs. One of these, mammalian numb (m-numb), which encodes a membrane-associated antagonist of Notch signaling, is a likely target of Msi1. Msi1 protein binds in vitro-transcribed m-numb RNA in its 3'-untranslated region (UTR) and binds endogenous m-numb mRNA in vivo, as shown by affinity precipitation followed by reverse transcription-PCR. Furthermore, adenovirus-induced Msi1 expression resulted in the down-regulation of endogenous m-Numb protein expression. Reporter assays using a chimeric mRNA that combined luciferase and the 3'-UTR of m-numb demonstrated that Msi1 decreased the reporter activity without altering the reporter mRNA level. Thus, our results suggested that Msi1 could regulate the expression of its target gene at the translational level. Furthermore, we found that Notch signaling activity was increased by Msi1 expression in connection with the posttranscriptional down-regulation of the m-numb gene.