Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142, USA.
Genes Dev. 2010 May 15;24(10):992-1009. doi: 10.1101/gad.1884710. Epub 2010 Apr 22.
MicroRNAs (miRNAs) are small regulatory RNAs that derive from distinctive hairpin transcripts. To learn more about the miRNAs of mammals, we sequenced 60 million small RNAs from mouse brain, ovary, testes, embryonic stem cells, three embryonic stages, and whole newborns. Analysis of these sequences confirmed 398 annotated miRNA genes and identified 108 novel miRNA genes. More than 150 previously annotated miRNAs and hundreds of candidates failed to yield sequenced RNAs with miRNA-like features. Ectopically expressing these previously proposed miRNA hairpins also did not yield small RNAs, whereas ectopically expressing the confirmed and newly identified hairpins usually did yield small RNAs with the classical miRNA features, including dependence on the Drosha endonuclease for processing. These experiments, which suggest that previous estimates of conserved mammalian miRNAs were inflated, provide a substantially revised list of confidently identified murine miRNAs from which to infer the general features of mammalian miRNAs. Our analyses also revealed new aspects of miRNA biogenesis and modification, including tissue-specific strand preferences, sequential Dicer cleavage of a metazoan precursor miRNA (pre-miRNA), consequential 5' heterogeneity, newly identified instances of miRNA editing, and evidence for widespread pre-miRNA uridylation reminiscent of miRNA regulation by Lin28.
微小 RNA(miRNAs)是源自独特发夹转录本的小调控 RNA。为了更多地了解哺乳动物的 miRNAs,我们对来自小鼠大脑、卵巢、睾丸、胚胎干细胞、三个胚胎阶段和新生个体的 6000 万个小 RNA 进行了测序。对这些序列的分析证实了 398 个注释 miRNA 基因,并鉴定了 108 个新的 miRNA 基因。超过 150 个先前注释的 miRNAs 和数百个候选 miRNA 未能产生具有 miRNA 特征的测序 RNA。异位表达这些先前提出的 miRNA 发夹也未能产生小 RNA,而异位表达经证实和新鉴定的发夹通常会产生具有经典 miRNA 特征的小 RNA,包括依赖 Drosha 内切酶进行加工。这些实验表明,先前保守哺乳动物 miRNAs 的估计值过高,为推断哺乳动物 miRNAs 的一般特征提供了一份经过大幅修订的、有信心鉴定的鼠类 miRNAs 列表。我们的分析还揭示了 miRNA 生物发生和修饰的新方面,包括组织特异性链偏好、后生动物前体 miRNA(pre-miRNA)的连续 Dicer 切割、随后的 5'异质性、新发现的 miRNA 编辑实例,以及广泛存在的 pre-miRNA 尿嘧啶化证据,类似于 Lin28 对 miRNA 的调控。