Combining Spinach-tagged RNA and gene localization to image gene expression in live yeast.
Combining Spinach-tagged RNA and gene localization to image gene expression in live yeast.
复制标题
将菠菜标签的RNA和基因定位与活酵母中的基因表达相结合。
DOI：
10.1038/ncomms9882
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发表时间：
2015-11-19
影响因子：
16.6
通讯作者：
Dargemont C
中科院分区：
文献类型：
--
作者：
Guet D;Burns LT;Maji S;Boulanger J;Hersen P;Wente SR;Salamero J;Dargemont C
尽管已经描述了形成有输出能力的 mRNP 所需的许多因素，但由于技术限制，在单细胞水平上仍然部分缺少对 mRNP 从转录位点到核出口位点的时空协调级联的综合视图。在这里，我们报告说，RNA 菠菜适体是活体酿酒酵母 mRNA 成像的强大工具，具有高时空分辨率，并且不会干扰 mRNA 生物发生特性。开发了专用的图像处理工作流程，以允许检测非常低丰度的转录本、准确的定量动态研究，以及在一致的时间尺度下提供接近 100 nm 的定位精度。结合这些方法，通过定位诱导转录因子、靶基因位点和相应的转录本，提供了对活酵母渗透压休克反应的最先进的分析。 测量单细胞 mRNA 动态对于了解基因表达至关重要。 Guet 等人在这里使用 RNA Spinach 技术来检测非常低丰度的 mRNA。报告通过定位诱导转录因子、靶基因位点和相应转录本对活酵母中的渗透压休克反应进行分析。
Although many factors required for the formation of export-competent mRNPs have been described, an integrative view of the spatiotemporal coordinated cascade leading mRNPs from their site of transcription to their site of nuclear exit, at a single cell level, is still partially missing due to technological limitations. Here we report that the RNA Spinach aptamer is a powerful tool for mRNA imaging in live S. cerevisiae with high spatial-temporal resolution and no perturbation of the mRNA biogenesis properties. Dedicated image processing workflows are developed to allow detection of very low abundance of transcripts, accurate quantitative dynamic studies, as well as to provide a localization precision close to 100 nm at consistent time scales. Combining these approaches has provided a state-of-the-art analysis of the osmotic shock response in live yeast by localizing induced transcription factors, target gene loci and corresponding transcripts. Measuring single-cell mRNA dynamics is critical to understand gene expression. Here, using RNA Spinach technique to detect very low abundant mRNAs, Guet et al. report an analysis of the osmotic shock response in live yeast by localizing induced transcription factors, target gene loci and corresponding transcripts.