RNA structure probing dash seq.
RNA structure probing dash seq.
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RNA 结构探测短划线序列。
DOI：
10.1073/pnas.1107835108
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发表时间：
2011
影响因子：
11.1
通讯作者：
Weeks,KevinM
中科院分区：
文献类型：
--
作者：
Weeks,KevinM
RNA构成了生物系统中信息存储、传递和操作的中心管道（1，2）。RNA在两个水平上编码其关键信息。首先，一级序列指导蛋白质合成，并含有结合调节因子和其他RNA的简单顺式作用元件。其次，大多数RNA折叠以产生具有内在调节功能的复杂碱基配对和更高阶结构。直到最近，它一直很难或不可能询问大多数RNA的结构，特别是在复杂的生物环境中。核苷酸解析RNA结构探测的持续进展使得越来越严格的定量分析成为可能（3），最近的大规模和全基因组研究揭示了或更好地定义了RNA结构如何调节翻译起始、蛋白质折叠、剪接和进入蛋白质结合位点的规则（4-6）。下一代测序（NGS）改变基于核酸的分析（7，8）的明显力量已经激发了将化学和酶探测实验与NGS读数（称为seq实验）融合的重大努力（科学冲刺）。RNA结构探测实验与NGS读数的融合是转录组天堂的潜在婚姻。在RNA结构探测实验中，RNA最初与一种“试剂”一起孵育，这种试剂会稀疏地反应，并在RNA分子的整体上留下印记（图1，左）。每个探针的独特特征决定了所收集的结构信息的最终质量和有用性。RNA切割蛋白（RNA酶）和小的化学探针都被广泛使用。在反应完成时，探测过程中存在的RNA包含探测事件的完整和准确的印记（图1，左下）。挑战在于尽可能准确地提取这些信息。
RNA constitutes the central con-duit for information storage, conveyance, and manipulation in biological systems (1, 2). RNAs encode their critical information at two levels. First, the primary sequence directs protein synthesis and contains simple cis-acting elements that bind regulatory factors and other RNAs. Second, most RNAs fold to create complex basepaired and higher order structures with intrinsic regulatory functions. Until recently, it has been difficult or impossible to interrogate the structures of most RNAs, especially in complex biological environments. Ongoing advances in nucleotideresolution RNA structure probing have made possible increasingly rigorous and quantitative analyses (3), and recent largescale and whole-genome studies have revealed or better defined rules for how RNA structure regulates translation initiation, protein folding, splicing, and access to protein binding sites (4–6). The clear power of next-generation sequencing (NGS) to transform nucleic acid-based analyses (7, 8) has motivated significant efforts (a scientific dash) to meld chemical and enzymatic probing experiments with NGS readouts (termed seq experiments). The melding of RNA structure probing experiments with NGS readout is a potential marriage made in transcriptome heaven. Two papers in PNAS (9, 10) illustrate important progress toward this highly sought goal.In an RNA structure probing experiment, RNAs are initially incubated with a “reagent” that reacts sparsely and leaves an imprint on the ensemble of RNA molecules (Fig. 1, Left). Features unique to each probe govern the ultimate quality and usefulness of the structural information gleaned. Both RNA-cleaving proteins (RNases) and small chemical probes are widely used. On reaction completion, the RNAs present during probing contain a full and exact imprint of the probing event (Fig. 1, Lower Left). The challenge is to extract this information as accurately as possible.