Global analysis of gene expression changes following LINE1 inhibition [III]

Transposable elements make up nearly half of mammalian genomes, yet are generally described as 'junk DNA' or genome parasites. The LINE1 retrotransposon is the most abundant class and is thought to be deleterious for cells, but it is paradoxically expressed at high levels during early development. Here, we report that LINE1 plays essential roles in mouse embryonic stem (ES) cells and pre-implantation embryos. In ES cells, LINE1 acts as a nuclear RNA scaffold that recruits Nucleolin and Kap1/Trim28 to repress Dux, the master activator of a gene expression program specific to the 2-cell stage. In parallel, LINE1 RNA mediates binding of Nucleolin and Kap1 to rDNA, thereby promoting rRNA synthesis and ES cell self-renewal. In embryos, LINE1 RNA is required for silencing of Dux, proper synthesis of rRNA and exit from the 2-cell stage. These results reveal an essential partnership between nuclear LINE1 RNA and chromatin factors in the regulation of transcription, developmental potency and ES cell self-renewal. Overall design: 3 replicates each of mouse embryos at early 2C or 4C stage, after zygote microinjection with 0.75mM RC control or LINE1 ASOs used for RNA extraction and library generation (12 samples total)转座元件（Transposable elements）几乎占哺乳动物基因组的一半，却长期被统称为“垃圾DNA”或基因组寄生虫。LINE1逆转座子是丰度最高的转座元件类别，既往认为其对细胞具有有害性，但矛盾的是，它在早期发育过程中却呈现高表达状态。本研究发现，LINE1在小鼠胚胎干细胞（embryonic stem cells, ES cells）及植入前胚胎中发挥不可或缺的关键作用。在胚胎干细胞中，LINE1作为核RNA支架，招募核仁素（Nucleolin）与Kap1/Trim28，以抑制双细胞期特异性基因表达程序的核心激活因子Dux。与此同时，LINE1 RNA介导核仁素与Kap1结合至核糖体DNA（rDNA），进而促进核糖体RNA（rRNA）合成与胚胎干细胞的自我更新。在胚胎中，LINE1 RNA对于沉默Dux、正常合成rRNA以及退出双细胞期均为必需。上述研究结果揭示了核LINE1 RNA与染色质调控因子之间的关键协同关系，其参与调控转录过程、发育潜能及胚胎干细胞自我更新。实验设计概述：本研究共纳入12个样本，分别为经受精卵显微注射0.75mM RC对照（RC control）或LINE1反义寡核苷酸（antisense oligonucleotides, ASOs）后的早期双细胞期（early 2-cell stage, 2C）或四细胞期（4-cell stage, 4C）小鼠胚胎，每组设置3次生物学重复，后续用于RNA提取与测序文库构建。