Argonaute HITS-CLIP decodes microRNA-mRNA interaction maps.

MicroRNAs (miRNAs) play critical roles in the regulation of gene expression. However, since miRNA activity requires base pairing with only 6-8 nucleotides of mRNA, predicting target mRNAs is a major challenge. Recently, high-throughput sequencing of RNAs isolated by crosslinking immunoprecipitation (HITS-CLIP) has identified functional protein-RNA interaction sites. Here we use HITS-CLIP to covalently crosslink native Argonaute (Ago) protein-RNA complexes in mouse brain. This produced two simultaneous datasets—Ago-miRNA and Ago-mRNA binding sites—that were combined with bioinformatic analysis to identify miRNA-target mRNA interaction sites. We validated genome-wide interaction maps for miR-124, and generated additional maps for the 20 most abundant miRNAs present in P13 mouse brain. Ago HITS-CLIP provides a general platform for exploring the specificity and range of miRNA action in vivo, and identifies precise sequences for targeting clinically relevant miRNA-mRNA interactions.

微小核糖核酸（miRNAs）在基因表达调控中起着关键作用。然而，由于miRNA的活性仅需要与信使核糖核酸（mRNA）的6 - 8个核苷酸进行碱基配对，预测目标mRNA是一项重大挑战。最近，通过交联免疫沉淀分离的RNA的高通量测序（HITS - CLIP）已经确定了功能性的蛋白质 - RNA相互作用位点。在此，我们利用HITS - CLIP使小鼠大脑中的天然Argonaute（Ago）蛋白 - RNA复合物共价交联。这产生了两个同时的数据集——Ago - miRNA和Ago - mRNA结合位点——将它们与生物信息学分析相结合，以确定miRNA - 目标mRNA相互作用位点。我们验证了miR - 124的全基因组相互作用图谱，并为出生后第13天（P13）小鼠大脑中存在的20种最丰富的miRNAs生成了其他图谱。Ago HITS - CLIP为探索体内miRNA作用的特异性和范围提供了一个通用平台，并确定了针对临床相关的miRNA - mRNA相互作用的精确序列。