Pachytene piRNAs target endogenous mRNAs for silencing during meiosis

During embryonic germ cell development in mice, transposon-enriched, piwi-interacting RNAs (piRNAs) guide MILI and MIWI2 to direct silencing of potentially active mobile element families. In contrast, we know much less about the function of the highly abundant and extremely diverse class of piRNAs, which partner with MIWI and MILI during meiosis. Both MIWI and its catalytic activity are required for successful spermatogenesis, strongly indicating that piRNA-guided cleavage is critical for germ cell development. To gain an understanding of meiotic piRNA targets, we augmented the mouse piRNA repertoire by introducing an entire human meiotic piRNA cluster. This triggered a spermatogenesis defect, presumably by inappropriately targeting the piRNA machinery to mouse RNAs essential for germ cell development. Through an analysis of such de novo targets, we derived a signature for pachytene piRNA target recognition. This enabled identification of both transposable elements and meiotically expressed protein coding genes as targets of native piRNAs. Cleavage of genic targets begins at the pachytene stage when meiotic piRNAs first appear. As such, target mRNA levels attenuate starting from the pachytene stage and are further repressed throughout meiosis. Target mRNA-piRNA pairs also show evidence of an ongoing cleavage-dependent amplification cycle, which is not normally a strong feature of meiotic piRNAs. Our data support the idea that meiotic piRNA populations must be strongly selected to enable successful spermatogenesis, both driving the response away from essential genes and directing the pathway toward mRNA targets that are regulated by small RNAs in meiotic cells. 48 samples在小鼠胚胎生殖细胞发育过程中，富集转座子的piwi相互作用RNA（piwi-interacting RNA，piRNA）可指导MILI与MIWI2，对潜在活跃的转座因子家族实施沉默调控。与之相对，人们对减数分裂过程中与MIWI、MILI结合的、含量极高且种类极为多样的piRNA类群的功能却知之甚少。MIWI及其催化活性均为正常精子发生所必需，这有力表明piRNA介导的切割过程对生殖细胞发育至关重要。为解析减数分裂piRNA的靶标，研究团队通过导入完整的人类减数分裂piRNA簇，扩增了小鼠的piRNA库。该操作引发了精子发生缺陷，推测其机制是将piRNA调控机器错误靶向了小鼠生殖细胞发育所必需的RNA。通过对这类全新靶标的分析，研究人员推导得到了粗线期piRNA靶标识别的特征序列。借此得以确认转座因子以及减数分裂期表达的蛋白质编码基因均为内源性piRNA的靶标。基因靶标的切割始于粗线期，而减数分裂piRNA正是在该时期首次出现。因此，靶标信使RNA（mRNA）的表达水平自粗线期开始下降，并在整个减数分裂过程中持续受到抑制。靶标mRNA-piRNA配对还显示出存在持续的切割依赖性扩增循环的证据，而这通常并非减数分裂piRNA的典型特征。本研究数据支持如下观点：为保障正常精子发生，减数分裂piRNA群体必须受到严格筛选，既要使调控通路远离必需基因，又要将其导向减数分裂细胞中小RNA调控的mRNA靶标。共包含48个样本