Laboratory of RNA Biology, Max-Planck-Institute of Biochemistry, Am Klopferspitz 18, 82152 Martinsried, Germany.
Mol Cell Proteomics. 2012 Nov;11(11):1442-56. doi: 10.1074/mcp.M112.017756. Epub 2012 Aug 23.
Argonaute (Ago) proteins interact with small regulatory RNAs such as microRNAs (miRNAs) and facilitate gene-silencing processes. miRNAs guide Ago proteins to specific mRNAs leading to translational silencing or mRNA decay. In order to understand the mechanistic details of miRNA function, it is important to characterize Ago protein interactors. Although several proteomic studies have been performed, it is not clear how the Ago interactome changes on miRNA or mRNA binding. Here, we report the analysis of Ago protein interactions in miRNA-containing and miRNA-depleted cells. Using stable isotope labeling in cell culture in conjunction with Dicer knock out mouse embryonic fibroblasts, we identify proteins that interact with Ago2 in the presence or the absence of Dicer. In contrast to our current view, we find that Ago-mRNA interactions can also take place in the absence of miRNAs. Our proteomics approach provides a rich resource for further functional studies on the cellular roles of Ago proteins.
Argonaute (Ago) 蛋白与小的调节 RNA 如 microRNAs (miRNAs) 相互作用，促进基因沉默过程。miRNAs 引导 Ago 蛋白到特定的 mRNAs，导致翻译沉默或 mRNA 降解。为了理解 miRNA 功能的机制细节，表征 Ago 蛋白相互作用体是很重要的。尽管已经进行了几项蛋白质组学研究，但尚不清楚 Ago 相互作用组在 miRNA 或 mRNA 结合时如何变化。在这里，我们报告了在含有 miRNA 和缺乏 miRNA 的细胞中 Ago 蛋白相互作用的分析。使用稳定同位素标记的细胞培养技术结合 Dicer 敲除的小鼠胚胎成纤维细胞，我们鉴定了在 Dicer 存在或不存在的情况下与 Ago2 相互作用的蛋白质。与我们目前的观点相反，我们发现 Ago-mRNA 相互作用也可以在没有 miRNAs 的情况下发生。我们的蛋白质组学方法为进一步研究 Ago 蛋白在细胞中的功能提供了丰富的资源。
