RNA Infrastructure Profiling Illuminates Transcriptome Structure in Crowded Spaces.
RNA Infrastructure Profiling Illuminates Transcriptome Structure in Crowded Spaces.
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RNA 基础设施分析阐明了拥挤空间中的转录组结构。
DOI：
10.1101/2023.10.09.561413
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发表时间：
2023
期刊：
影响因子：
--
通讯作者：
Kool,EricT
中科院分区：
文献类型：
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作者：
Xiao,Lu;Fang,Linglan;Kool,EricT
RNA折叠成紧凑的结构，并在细胞中进行蛋白质相互作用。这些封闭的环境可以阻断探测底层RNA的试剂。可以在拥挤环境中分析结构的探针可以揭示RNA生物学。在这里，我们采用了2′-OH反应性探针，这些探针足够小，可以进入细胞内紧密分子接触的折叠RNA结构，为细胞内RNA结构分析提供了相当广泛的覆盖范围。首先用充分表征的人核糖体RNA分析数据，然后将其应用于全转录组的多腺苷酸化转录物。最小的探针乙酰咪唑（AcIm）产生80%以上的结构覆盖率比较大的常规试剂NAIN 3，提供增强的结构信息在数百个转录本。乙酰基探针还提供了用于鉴定转录物中m6 A修饰位点的上级信号，特别是在标准探针不可接近的位点中。我们的策略能够分析RNA基础结构，增强对转录组结构，修饰和细胞内相互作用的分析，特别是在空间拥挤的环境中。
RNAs fold into compact structures and undergo protein interactions in cells. These occluded environments can block reagents that probe the underlying RNAs. Probes that can analyze structure in crowded settings can shed light on RNA biology. Here, we employ 2′-OH-reactive probes that are small enough to access folded RNA structure underlying close molecular contacts within cells, providing considerably broader coverage for intracellular RNA structural analysis. The data are analyzed first with well-characterized human ribosomal RNAs and then applied transcriptome-wide to polyadenylated transcripts. The smallest probe acetylimidazole (AcIm) yields 80% greater structural coverage than larger conventional reagent NAIN3, providing enhanced structural information in hundreds of transcripts. The acetyl probe also provides superior signals for identifying m6A modification sites in transcripts, particularly in sites that are inaccessible to a standard probe. Our strategy enables profiling RNA infrastructure, enhancing analysis of transcriptome structure, modification, and intracellular interactions, especially in spatially crowded settings.