Derepression of transposable elements (TEs) in the course of epigenetic reprogramming of the mouse embryonic germline necessitates the existence of a robust defense that is comprised of PIWI/piRNA pathway and de novo DNA methylation machinery. To gain further insight into biogenesis and function of piRNAs, we studied the intracellular localization of piRNA pathway components and used the combination of genetic, molecular, and cell biological approaches to examine the performance of the piRNA pathway in germ cells of mice lacking Maelstrom (MAEL), an evolutionarily conserved protein implicated in transposon silencing in fruit flies and mice. Here we show that principal components of the fetal piRNA pathway, MILI and MIWI2 proteins, localize to two distinct types of germinal cytoplasmic granules and exhibit differential association with components of the mRNA degradation/translational repression machinery. The first type of granules, pi-bodies, contains the MILI-TDRD1 module of the piRNA pathway and is likely equivalent to the enigmatic “cementing material” first described in electron micrographs of rat gonocytes over 35 years ago. The second type of granules, piP-bodies, harbors the MIWI2-TDRD9-MAEL module of the piRNA pathway and signature components of P-bodies, GW182, DCP1a, DDX6/p54, and XRN1 proteins. piP-bodies are found predominantly in the proximity of pi-bodies and the two frequently share mouse VASA homolog (MVH) protein, an RNA helicase. In Mael-mutant gonocytes, MIWI2, TDRD9, and MVH are lost from piP-bodies, whereas no effects on pi-body composition are observed. Further analysis revealed that MAEL appears to specifically facilitate MIWI2-dependent aspects of the piRNA pathway including biogenesis of secondary piRNAs, de novo DNA methylation, and efficient downregulation of TEs. Cumulatively, our data reveal elaborate cytoplasmic compartmentalization of the fetal piRNA pathway that relies on MAEL function.
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机译：在小鼠胚胎种系的表观遗传重编程过程中，转移元素（TES）的DEREMENTION需要存在由PIWI / PiRNA途径和DE Novo DNA甲基化机械组成的强大防御。为了进一步了解PiRNA的生物发生和功能，我们研究了PiRNA途径成分的细胞内定位，并使用了遗传，分子和细胞生物方法的组合来检查缺乏Maelstrom的小鼠生殖细胞中PiRNA途径的性能（Mael ），一种涉及在果蝇和小鼠中的转座子沉默的进化保守的蛋白质。在这里，我们显示胎儿piRNA途径，MILI和MIWI2蛋白的主要成分，定位为两个不同类型的生发细胞质颗粒，与mRNA降解/翻译镇压机械的组分表现出差异关系。第一种类型的颗粒，Pi体含有PiRNA途径的MILI-TDRD1模块，可能是35年前大鼠淋巴织物的电子显微照片中首先描述的神秘的“硬质材料”。第二种类型的颗粒，桶体，竖琴的PiRNA途径的MiWI2-TDRD9-MAEL模块，P-体，GW182，DCP1A，DDX6 / P54和XRN1蛋白的特征组分。管体主要发现在Pi体的邻近，并且两种经常共享小鼠VASA同源物（MVH）蛋白，RNA螺旋酶。在Mael-突变腺细胞中，MIWI2，TDRD9和MVH从桶体中丧失，而观察到对PI-B体组合物的影响。进一步的分析表明，MAEL似乎具体促进了PiRNA途径的MiWI2依赖性方面，包括二次PiRNA的生物发生，De Novo DNA甲基化，以及TES的有效下调。累积地，我们的数据揭示了依靠Mael功能的胎儿piRNA途径的精细细胞质隔室化。
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