A dimerization-based fluorogenic dye-aptamer module for RNA imaging in live cells
A dimerization-based fluorogenic dye-aptamer module for RNA imaging in live cells
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DOI：
10.1038/s41589-019-0381-8
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发表时间：
2020-01-01
影响因子：
14.8
通讯作者：
Ryckelynck, Michael
中科院分区：
文献类型：
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作者：
Bouhedda, Farah;Fam, Kyong Tkhe;Ryckelynck, Michael
由于缺乏天然荧光RNA，RNA的活细胞成像仍然是一个挑战。最近开发的RNA适体可以点亮小的荧光染料，可以克服这一限制，但它们仍然受到亮度和光稳定性差的影响。在这里，我们提出了一个细胞可渗透的自淬灭磺酰罗丹明B染料（Gemini-561）和相应的二聚适体（o-珊瑚），可以大大提高目前的RNA成像方法的性能的荧光二聚体的概念。改进的亮度和光稳定性，以及该复合物的高亲和力，允许在活的哺乳动物细胞中的RNA聚合酶III转录产物以及用单拷贝的适体标记的信使RNA的直接荧光成像;即，没有标签多聚化。开发的荧光模块能够快速灵敏地检测活细胞内的RNA，而提出的设计概念开辟了新一代超亮RNA探针的路线。
Live-cell imaging of RNA has remained a challenge because of the lack of naturally fluorescent RNAs. Recently developed RNA aptamers that can light-up small fluorogenic dyes could overcome this limitation, but they still suffer from poor brightness and photostability. Here, we propose the concept of a cell-permeable fluorogenic dimer of self-quenched sulforhodamine B dyes (Gemini-561) and the corresponding dimerized aptamer (o-Coral) that can drastically enhance performance of the current RNA imaging method. The improved brightness and photostability, together with high affinity of this complex, allowed direct fluorescence imaging in live mammalian cells of RNA polymerase III transcription products as well as messenger RNAs labeled with a single copy of the aptamer; that is, without tag multimerization. The developed fluorogenic module enables fast and sensitive detection of RNA inside live cells, while the proposed design concept opens the route to new generation of ultrabright RNA probes.