Tethered Function Assays as Tools to Elucidate the Molecular Roles of RNA-Binding Proteins.
Tethered Function Assays as Tools to Elucidate the Molecular Roles of RNA-Binding Proteins.
复制标题
束缚功能分析是阐明RNA结合蛋白的分子作用的工具。
DOI：
10.1007/978-3-319-29073-7_3
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发表时间：
2016
影响因子：
--
通讯作者：
Yeo GW
中科院分区：
文献类型：
--
作者：
Bos TJ;Nussbacher JK;Aigner S;Yeo GW
RNA分子的动态调控对细胞的生存和发育至关重要。信使RNA作为内含子前mrna在细胞核中转录，并与RNA结合蛋白结合，通过调节RNA稳定性、剪接、聚腺苷化、翻译和细胞定位来控制信使RNA的命运。大多数rbp具有不同的mrna结合域和功能域；因此，通过人工将RBP招募到报告RNA上，然后测量RBP招募对报告RNA剪接、稳定性、翻译效率或细胞内运输的影响，可以独立于RNA结合来研究RBP的功能。因此，这些栓系功能测定不需要事先了解RBP的内源性RNA靶点或这些RNA中的结合位点。在这里，我们概述了实验策略以及常见系绳系统的优势和局限性。我们举例说明了在阐明各种rbp的功能分析应用的具体例子。我们还讨论了经典的栓系分析方法和从中获得的见解如何被最近的技术进步所增强，包括高效的基因组编辑和高通量rna测序。
Dynamic regulation of RNA molecules is critical to the survival and development of cells. Messenger RNAs are transcribed in the nucleus as intron-containing pre-mRNAs and bound by RNA-binding proteins, which control their fate by regulating RNA stability, splicing, polyadenylation, translation, and cellular localization. Most RBPs have distinct mRNA-binding and functional domains; thus, the function of an RBP can be studied independently of RNA-binding by artificially recruiting the RBP to a reporter RNA and then measuring the effect of RBP recruitment on reporter splicing, stability, translational efficiency, or intracellular trafficking. These tethered function assays therefore do not require prior knowledge of the RBP’s endogenous RNA targets or its binding sites within these RNAs. Here, we provide an overview of the experimental strategy and the strengths and limitations of common tethering systems. We illustrate specific examples of the application of the assay in elucidating the function of various classes of RBPs. We also discuss how classic tethering assay approaches and insights gained from them have been empowered by more recent technological advances, including efficient genome editing and high-throughput RNA-sequencing.
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DOI：
10.1016/j.bbrc.2006.06.177
发表时间：
2006-09-01
影响因子：
3.1
作者：
Barreau, Carine;Watrin, Tanguy;Paillard, Luc
通讯作者：
Paillard, Luc
影响因子：
4.5
作者：
Brodsky, AS;Silver, PA
通讯作者：
Silver, PA
影响因子：
2.9
作者：
CHEN, L;FRANKEL, AD
通讯作者：
FRANKEL, AD
影响因子：
4.5
作者：
De Gregorio, E;Baron, J;Hentze, MW
通讯作者：
Hentze, MW
DOI：
10.1038/nrm3918
发表时间：
2015-02
期刊：
Nature reviews. Molecular cell biology
影响因子：
--
作者：
通讯作者：
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