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Following the messenger: Recent innovations in live cell single molecule fluorescence imaging.
WIREs RNA ( IF 7.3 ) Pub Date : 2020-01-28 , DOI: 10.1002/wrna.1587
Andreas Schmidt 1 , Guoming Gao 1, 2 , Saffron R Little 1, 3 , Ameya P Jalihal 1, 4 , Nils G Walter 1
Affiliation  

Messenger RNAs (mRNAs) convey genetic information from the DNA genome to proteins and thus lie at the heart of gene expression and regulation of all cellular activities. Live cell single molecule tracking tools enable the investigation of mRNA trafficking, translation and degradation within the complex environment of the cell and in real time. Over the last 5 years, nearly all tools within the mRNA tracking toolbox have been improved to achieve high-quality multi-color tracking in live cells. For example, the bacteriophage-derived MS2-MCP system has been improved to facilitate cloning and achieve better signal-to-noise ratio, while the newer PP7-PCP system now allows for orthogonal tracking of a second mRNA or mRNA region. The coming of age of epitope-tagging technologies, such as the SunTag, MoonTag and Frankenbody, enables monitoring the translation of single mRNA molecules. Furthermore, the portfolio of fluorogenic RNA aptamers has been expanded to improve cellular stability and achieve a higher fluorescence "turn-on" signal upon fluorogen binding. Finally, microinjection-based tools have been shown to be able to track multiple RNAs with only small fluorescent appendages and to track mRNAs together with their interacting partners. We systematically review and compare the advantages, disadvantages and demonstrated applications in discovering new RNA biology of this refined, expanding toolbox. Finally, we discuss developments expected in the near future based on the limitations of the current methods. This article is categorized under: RNA Export and Localization > RNA Localization RNA Structure and Dynamics > RNA Structure, Dynamics, and Chemistry RNA Interactions with Proteins and Other Molecules > RNA-Protein Complexes.

中文翻译:

跟随信使:活细胞单分子荧光成像的最新创新。

信使 RNA (mRNA) 将遗传信息从 DNA 基因组传递到蛋白质,因此是基因表达和所有细胞活动调节的核心。活细胞单分子追踪工具能够实时研究细胞复杂环境中的 mRNA 运输、翻译和降解。在过去的 5 年里,mRNA 跟踪工具箱中的几乎所有工具都得到了改进,以在活细胞中实现高质量的多色跟踪。例如,噬菌体衍生的 MS2-MCP 系统已得到改进,以促进克隆并实现更好的信噪比,而较新的 PP7-PCP 系统现在允许正交跟踪第二个 mRNA 或 mRNA 区域。表位标记技术时代的到来,例如 SunTag、MoonTag 和 Frankenbody,能够监测单个 mRNA 分子的翻译。此外,荧光 RNA 适配体的产品组合已得到扩展,以提高细胞稳定性并在与荧光结合时获得更高的荧光“开启”信号。最后,基于显微注射的工具已被证明能够仅用小的荧光附属物跟踪多个 RNA,并与其相互作用的伙伴一起跟踪 mRNA。我们系统地回顾和比较了这个改进的、扩展的工具箱在发现新 RNA 生物学方面的优点、缺点和已证明的应用。最后,我们基于当前方法的局限性讨论了在不久的将来预期的发展。本文分类如下:RNA 导出和定位 > RNA 定位 RNA 结构和动力学 > RNA 结构、动力学、
更新日期:2020-01-28
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