MS2 Labeling of Endogenous Beta-Actin mRNA Does Not Result in Stabilization of Degradation Intermediates
MS2 Labeling of Endogenous Beta-Actin mRNA Does Not Result in Stabilization of Degradation Intermediates
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DOI：
10.14348/molcells.2019.2398
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发表时间：
2019-04-01
影响因子：
3.8
通讯作者：
Park, Hye Yoon
中科院分区：
文献类型：
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作者：
Kim, Songhee H.;Vieira, Melissa;Park, Hye Yoon
MS2噬菌体外壳蛋白(MCP)与MS2结合位点(MBS)RNA茎环序列的结合已被广泛用于单分子分辨率的活细胞成像的mRNA标记。然而，最近对萌芽酵母的研究表明，在MS2-GFP标记后出现了异常的mRNA代谢，这引起了人们的关注。为了研究MS2-GFP标记的信使核糖核酸在哺乳动物细胞和组织中的降解规律，我们用Northern印迹方法分析了从ActB-MBS基因敲入和MBSxMCP杂交小鼠模型中提取的ss-肌动蛋白信使核糖核酸。在永生化的小鼠胚胎细胞系和小鼠模型衍生的各种器官组织中，我们发现与野生型小鼠相比，没有明显的ss-肌动蛋白mRNA的衰变产物积累。我们的结果表明，MBS RNA衰变片段的积累并不总是发生的，这取决于mRNA的种类和所使用的模式生物。
The binding of MS2 bacteriophage coat protein (MCP) to MS2 binding site (MBS) RNA stem-loop sequences has been widely used to label mRNA for live-cell imaging at single-molecule resolution. However, concerns have been raised recently from studies with budding yeast showing aberrant mRNA metabolism following the MS2-GFP labeling. To investigate the degradation pattern of MS2-GFP-labeled mRNA in mammalian cells and tissues, we used Northern blot analysis of ss-actin mRNA extracted from the Actb-MBS knock-in and MBSxMCP hybrid mouse models. In the immortalized mouse embryonic cell lines and various organ tissues derived from the mouse models, we found no noticeable accumulation of decay products of ss-actin mRNA compared with the wild-type mice. Our results suggest that accumulation of MBS RNA decay fragments does not always happen depending on the mRNA species and the model organisms used.