核糖核蛋白复合体重塑与拆解的体外分析

In Vitro Analysis of Ribonucleoprotein Complex Remodeling and Disassembly.

作者信息

Zhou Hua-Lin, Lou Hua

机构信息

Department of Genetics and Genome Sciences, Case Western Reserve University, Cleveland, OH, USA.

出版信息

Methods Mol Biol. 2016;1421:69-78. doi: 10.1007/978-1-4939-3591-8_7.

DOI:10.1007/978-1-4939-3591-8_7

PMID:26965258

Abstract

Ribonucleoprotein (RNP) complexes play essential roles in gene expression. Their assembly and disassembly control the fate of mRNA molecules. Here, we describe a method that examines the remodeling and disassembly of RNPs. One unique aspect of this method is that the RNA-binding proteins (RBPs) of interest are produced in HeLa cells with or without the desired modification and the RNP is assembled in cellular extracts with synthetic RNA oligonucleotides. We use this method to investigate how ubiquitination of an RBP affects its ability to bind its RNA target.

摘要

核糖核蛋白（RNP）复合物在基因表达中发挥着至关重要的作用。它们的组装和解聚控制着mRNA分子的命运。在此，我们描述了一种检测RNP重塑和解聚的方法。该方法的一个独特之处在于，感兴趣的RNA结合蛋白（RBP）在有或没有所需修饰的HeLa细胞中产生，并且RNP在细胞提取物中与合成RNA寡核苷酸组装。我们使用这种方法来研究RBP的泛素化如何影响其结合RNA靶标的能力。