Institute for Advanced Biosciences, Keio University, Tsuruoka 997-0017, Japan.
BMC Genomics. 2010 Feb 9;11:101. doi: 10.1186/1471-2164-11-101.
In many eukaryotes, microRNAs (miRNAs) bind to complementary sites in the 3'-untranslated regions (3'-UTRs) of target messenger RNAs (mRNAs) and regulate their expression at the stage of translation. Recent studies have revealed that many miRNAs are evolutionarily conserved; however, the evolution of their target genes has yet to be systematically characterized. We sought to elucidate a set of conserved miRNA/target-gene pairs and to analyse the mechanism underlying miRNA-mediated gene regulation in the early stage of bilaterian evolution.
Initially, we extracted five evolutionarily conserved miRNAs (let-7, miR-1, miR-124, miR-125/lin-4, and miR-34) among five diverse bilaterian animals. Subsequently, we designed a procedure to predict evolutionarily conserved miRNA/target-gene pairs by introducing orthologous gene information. As a result, we extracted 31 orthologous miRNA/target-gene pairs that were conserved among at least four diverse bilaterian animals; the prediction set showed prominent enrichment of orthologous miRNA/target-gene pairs that were verified experimentally. Approximately 84% of the target genes were regulated by three miRNAs (let-7, miR-1, and miR-124) and their function was classified mainly into the following categories: development, muscle formation, cell adhesion, and gene regulation. We used a reporter gene assay to experimentally verify the downregulation of six candidate pairs (out of six tested pairs) in HeLa cells.
The application of our new method enables the identification of 31 miRNA/target-gene pairs that were expected to have been regulated from the era of the common bilaterian ancestor. The downregulation of all six candidate pairs suggests that orthologous information contributed to the elucidation of the primordial set of genes that has been regulated by miRNAs; it was also an efficient tool for the elimination of false positives from the predicted candidates. In conclusion, our study identified potentially important miRNA-target pairs that were evolutionarily conserved throughout diverse bilaterian animals and that may provide new insights into early-stage miRNA functions.
在许多真核生物中，微小 RNA（miRNA）与靶信使 RNA（mRNA）的 3'非翻译区（3'-UTR）中的互补位点结合，并在翻译阶段调节其表达。最近的研究表明，许多 miRNA 在进化上是保守的；然而，它们的靶基因的进化尚未得到系统的描述。我们试图阐明一组保守的 miRNA/靶基因对，并分析 miRNA 介导的基因调控在两侧动物进化早期的机制。
最初，我们从五种不同的两侧动物中提取了五种进化上保守的 miRNA（let-7、miR-1、miR-124、miR-125/lin-4 和 miR-34）。随后，我们设计了一种通过引入同源基因信息来预测进化上保守的 miRNA/靶基因对的程序。结果，我们从至少四种不同的两侧动物中提取了 31 个保守的同源 miRNA/靶基因对；预测集显著富集了经过实验验证的同源 miRNA/靶基因对。约 84%的靶基因受到三种 miRNA（let-7、miR-1 和 miR-124）的调节，它们的功能主要分为以下几类：发育、肌肉形成、细胞粘附和基因调控。我们使用报告基因测定法在 HeLa 细胞中实验验证了六个候选对中的六个（六个测试对中的六个）的下调。
我们的新方法的应用能够识别出 31 个 miRNA/靶基因对，这些基因对有望从共同的两侧动物祖先时代就受到调节。所有六个候选对的下调表明，同源信息有助于阐明受 miRNAs 调节的原始基因集；它也是从预测的候选者中消除假阳性的有效工具。总之，我们的研究鉴定了在不同的两侧动物中进化保守的潜在重要 miRNA 靶对，这可能为早期 miRNA 功能提供新的见解。
