The role of the piRNA pathway in mouse neocorticogenesis

PIWI-interacting RNAs (piRNAs) and their associated PIWI proteins play an important role in repressing transposable elements in animal germlines. However, there have been conflicting reports about piRNA expression and function in the soma, particularly in the brain. Here we investigated the role of piRNAs and an important PIWI family member, Piwi-like protein 1 (Piwil1; also known as Miwi in mouse) in the developing mouse neocortex. Using a Piwil1 knock-out mouse strain, we found that Piwil1 is essential for neocorticogenesis, including neocortical cell cycle, migration and dendritogenesis. Piwil1 deletion resulted in increased cell cycle re-entry at embryonic day 17 (E17) when predominantly intracortically projecting neurons are being produced. Prenatal Piwil1 deletion increased the number of Pax6+ radial glia at postnatal day 0 (P0) and disrupted migration of Satb2+ neurons within deep layers at E17, P0 and P7. Satb2+ neurons showed increased co-localization with Bcl11b, marker of subcortically projecting neurons. In addition, Piwil1 knock-outs have disrupted neocortical circuitry represented by thinning of the corpus callosum and altered dendritogenesis. These results are consistent with the rodent phenotypes reported by a recent study that mainly used RNAi knockdown experiments. We identified a disrupted set of genes associated with cell cycle, cell adhesion, transcription and migration in developing neocortices. We further investigated the potential role of piRNAs by sequencing piRNAs in developing neocortices in the wildtype and Piwil1 knock-out strain. Unexpectedly, we did not find a strong signal of absolute piRNA abundance or differential expression in the knock-out, suggesting that Piwil1 may act independently of piRNAs in mouse neocorticogenesis. Our results may help clarify some outstanding questions in the piRNA field about the role of the piRNA pathway in the brain.

PIWI互作RNA（PIWI-interacting RNAs，piRNAs）及其结合的PIWI蛋白，在动物生殖系的转座子沉默过程中发挥关键作用。然而，关于piRNAs在体细胞尤其是大脑中的表达与功能，目前已有诸多相悖的研究报道。本研究针对发育中小鼠大脑新皮层，探究了piRNAs及重要的Piwi家族成员——Piwi样蛋白1（Piwi-like protein 1，Piwil1；小鼠中又称Miwi）的作用。通过使用Piwil1基因敲除小鼠品系，我们发现Piwil1对新皮层发生至关重要，其调控过程涵盖新皮层细胞周期、神经元迁移与树突发生。在主要产生皮层内投射神经元的胚胎第17天（E17），Piwil1缺失会导致细胞周期再进入事件增多。产前Piwil1敲除会使出生后第0天（P0）的Pax6阳性放射状胶质细胞数量增加，并在E17、P0及出生后第7天（P7）破坏Satb2阳性神经元在皮层深层的迁移。Satb2阳性神经元与皮层下投射神经元标志物Bcl11b的共定位现象显著增多。此外，Piwil1基因敲除小鼠的新皮层环路出现异常，表现为胼胝体变薄以及树突发生紊乱。上述结果与近期一项主要采用RNA干扰敲低实验的啮齿类动物研究报道的表型相符。我们在发育中的新皮层中鉴定到一组与细胞周期、细胞黏附、转录及神经元迁移相关的基因表达失调情况。我们进一步通过对野生型与Piwil1敲除小鼠发育新皮层中的piRNAs进行测序，探究了piRNAs的潜在功能。出乎意料的是，我们未在敲除样本中检测到piRNA绝对丰度或差异表达的显著信号，这提示Piwil1在小鼠新皮层发生过程中可能不依赖piRNAs发挥作用。本研究结果或有助于阐明piRNA领域中关于piRNA通路在大脑中作用的若干悬而未决的问题。