Rapid and systematic analysis of the RNA recognition specificities of RNA-binding proteins
Rapid and systematic analysis of the RNA recognition specificities of RNA-binding proteins
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DOI：
10.1038/nbt.1550
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发表时间：
2009-07-01
影响因子：
46.9
通讯作者：
Hughes, Timothy R.
中科院分区：
文献类型：
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作者：
Ray, Debashish;Kazan, Hilal;Hughes, Timothy R.
后生动物基因组编码数百种RNA结合蛋白（RBP），但相对较少的RBP的RNA结合偏好已经得到很好的定义（1）。目前用于确定RNA靶标的技术，包括体外选择和RNA免疫共沉淀（2-5），需要大量的时间和劳动力投入。在这里，我们介绍RNAcompete，RNA结合特异性的系统分析方法，使用一个单一的结合反应，以确定相对偏好的RBP短RNA，包含一个完整的范围内的k-mer在结构化和非结构化的RNA上下文。我们通过分析9种不同的RBP（HuR、Vts 1、FUSIP 1、PTB、U1 A、SF 2/ASF、SLM 2、RBM 4和YB 1）来测试RNAcompete。RNAcompete确定了预期的和以前未知的RNA结合偏好。使用在体外和体内结合数据，我们证明，偏好的个人7-聚体确定的RNAcompete是一个更准确的代表性的结合活性比传统的基序模型。我们预计，RNAcompete将是一个有价值的工具，RNA-蛋白质相互作用的研究。
Metazoan genomes encode hundreds of RNA-binding proteins (RBPs) but RNA-binding preferences for relatively few RBPs have been well defined(1). Current techniques for determining RNA targets, including in vitro selection and RNA co-immunoprecipitation(2-5), require significant time and labor investment. Here we introduce RNAcompete, a method for the systematic analysis of RNA binding specificities that uses a single binding reaction to determine the relative preferences of RBPs for short RNAs that contain a complete range of k-mers in structured and unstructured RNA contexts. We tested RNAcompete by analyzing nine diverse RBPs (HuR, Vts1, FUSIP1, PTB, U1A, SF2/ASF, SLM2, RBM4 and YB1). RNAcompete identified expected and previously unknown RNA binding preferences. Using in vitro and in vivo binding data, we demonstrate that preferences for individual 7-mers identified by RNAcompete are a more accurate representation of binding activity than are conventional motif models. We anticipate that RNAcompete will be a valuable tool for the study of RNA-protein interactions.