Höck Julia, Weinmann Lasse, Ender Christine, Rüdel Sabine, Kremmer Elisabeth, Raabe Monika, Urlaub Henning, Meister Gunter
Laboratory of RNA Biology, Max Planck Institute of Biochemistry, Am Klopferspitz 18, 82152 Martinsried, Germany.
EMBO Rep. 2007 Nov;8(11):1052-60. doi: 10.1038/sj.embor.7401088. Epub 2007 Oct 12.
Members of the Argonaute (Ago) protein family associate with small RNAs and have important roles in RNA silencing. Here, we analysed Ago1- and Ago2-containing protein complexes in human cells. Separation of Ago-associated messenger ribonucleoproteins (mRNPs) showed that Ago1 and Ago2 reside in three complexes with distinct Dicer and RNA-induced silencing complex activities. A comprehensive proteomic analysis of Ago-containing mRNPs identified a large number of proteins involved in RNA metabolism. By using co-immunoprecipitation experiments followed by RNase treatment, we biochemically mapped interactions within Ago mRNPs. Using reporter assays and knockdown experiments, we showed that the putative RNA-binding protein RBM4 is required for microRNA-guided gene regulation.
AGO(Argonaute)蛋白家族成员与小RNA结合,并在RNA沉默中发挥重要作用。在此,我们分析了人类细胞中含AGO1和AGO2的蛋白复合物。AGO相关信使核糖核蛋白(mRNP)的分离表明,AGO1和AGO2存在于三种具有不同Dicer和RNA诱导沉默复合体活性的复合物中。对含AGO的mRNP进行全面的蛋白质组学分析,鉴定出大量参与RNA代谢的蛋白质。通过免疫共沉淀实验并随后进行核糖核酸酶处理,我们对AGO mRNP内的相互作用进行了生化定位。利用报告基因检测和敲低实验,我们表明假定的RNA结合蛋白RBM4是microRNA引导的基因调控所必需的。