Small RNA profile in fully grown mammalian oocytes

We aim to comprehensively characterize the small RNA population in oocytes Pseudogenes populate the mammalian genome as remnants of artifactual incorporation of coding mRNAs into transposon pathways 1. Here, we show that a subset of pseudogenes generates endogenous small interfering RNAs (endo-siRNAs) in mouse oocytes. In these cases, endo-siRNAs are often processed from double-stranded RNAs formed by hybridization of spliced transcripts from protein coding genes to antisense transcripts from homologous pseudogenes. In at least one case, an inverted repeat pseudogene gives rise to abundant small RNAs directly. A second class of endo-siRNAs may enforce repression of mobile genetic elements, acting in concert with piwi-interacting RNAs (piRNAs). Loss of Dicer increases expression of endo-siRNA targets, demonstrating the regulatory activity of these small RNAs. Our findings provide a function for pseudogenes in regulating gene expression via the RNAi pathway and may, in part, explain the evolutionary pressure to conserve Argonaute-mediated catalysis in mammals. Keywords: small RNAs profile Total RNA was isolated and size-fractionated by PAGE either into 19-24nt and 24-30nt, or 19-30nt. These were independently processed and sequenced on Illumina 1G platform.

本研究旨在全面解析卵母细胞中的小RNA群体。假基因（Pseudogenes）是编码mRNA被错误整合进转座子通路后的残留片段，广泛存在于哺乳动物基因组中[1]。本研究发现，部分假基因可在小鼠卵母细胞中生成内源性小干扰RNA（endo-siRNAs）。此类内源性小干扰RNA通常由双链RNA加工而来：双链RNA由蛋白编码基因的剪接转录本与同源假基因的反义转录本杂交形成。至少存在一例特殊情况：反向重复假基因可直接产生大量小RNA。第二类内源性小干扰RNA可与piwi互作RNA（piwi-interacting RNAs, piRNAs）协同作用，抑制移动遗传元件的表达。Dicer基因缺失会提升内源性小干扰RNA靶标的表达水平，证实了这类小RNA的调控活性。本研究结果揭示了假基因通过RNA干扰（RNAi）通路调控基因表达的功能，也在一定程度上解释了哺乳动物中保守Argonaute介导的催化作用的进化压力来源。关键词：小RNA谱。总RNA经分离后，通过聚丙烯酰胺凝胶电泳（Polyacrylamide Gel Electrophoresis, PAGE）按片段大小分级为19-24nt、24-30nt两组，或19-30nt单一组；分别对各组进行独立处理后，在Illumina 1G测序平台上完成测序。