RNA-Centric Approaches to Profile the RNA-Protein Interaction Landscape on Selected RNAs.
RNA-Centric Approaches to Profile the RNA-Protein Interaction Landscape on Selected RNAs.
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DOI：
10.3390/ncrna7010011
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发表时间：
2021-02-15
期刊：
影响因子：
4.3
通讯作者：
Gerber AP
中科院分区：
文献类型：
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作者：
Gerber AP
RNA-蛋白质相互作用构建转录后调控网络并调节转录和表观遗传学。虽然 RNA 测序技术的进步显着扩展了 RNA 的种类，但最近开发的生化方法与灵敏的质谱分析相结合，揭示了数百种以前未被识别的潜在新颖的 RNA 结合蛋白。然而，了解数千个 RNA 分子及其相互作用的蛋白质如何组装和控制细胞中每个单独 RNA 的命运仍然是一个重大挑战。在这里，我回顾了通过系统鉴定与活细胞中特定 RNA 相互作用的蛋白质来解决这个问题的最新方法学进展。因此，我们特别关注体内方法，包括通过紫外线照射或用化学物质处理细胞来交联 RNA-蛋白质相互作用，然后用反义寡核苷酸捕获所研究的 RNA，并用质谱法鉴定结合的蛋白质。重点介绍了最近几项定义长非编码 RNA、病毒 RNA 以及 mRNA 相互作用组的研究，并简要提及了最近的细胞内蛋白质标记技术。这些最近的实验改进可能为更广泛的应用打开大门，并研究 RNA-蛋白质复合物在不同环境因素和疾病中的重塑。
RNA–protein interactions frame post-transcriptional regulatory networks and modulate transcription and epigenetics. While the technological advances in RNA sequencing have significantly expanded the repertoire of RNAs, recently developed biochemical approaches combined with sensitive mass-spectrometry have revealed hundreds of previously unrecognized and potentially novel RNA-binding proteins. Nevertheless, a major challenge remains to understand how the thousands of RNA molecules and their interacting proteins assemble and control the fate of each individual RNA in a cell. Here, I review recent methodological advances to approach this problem through systematic identification of proteins that interact with particular RNAs in living cells. Thereby, a specific focus is given to in vivo approaches that involve crosslinking of RNA–protein interactions through ultraviolet irradiation or treatment of cells with chemicals, followed by capture of the RNA under study with antisense-oligonucleotides and identification of bound proteins with mass-spectrometry. Several recent studies defining interactomes of long non-coding RNAs, viral RNAs, as well as mRNAs are highlighted, and short reference is given to recent in-cell protein labeling techniques. These recent experimental improvements could open the door for broader applications and to study the remodeling of RNA–protein complexes upon different environmental cues and in disease.