Evollutionally dynamic L1 regulation in embryonic stem cells. Evollutionally dynamic L1 regulation in embryonic stem cells

Mobile elements are important evolutionary forces that challenge genomic integrity. Long interspersed element-1 (L1, also known as LINE-1) is the only autonomous transposon still active in the human genome. It displays an unusual pattern of evolution, with at any given time a single active L1 lineage amplifying to thousands of copies before getting replaced by a new lineage likely under pressure of host restriction factors, which act notably by silencing L1 expression during early embryogenesis. Here, we demonstrate that in human embryonic stem cells (hESC) KAP1, the master co-factor of KRAB-containing zinc finger proteins (KRAB-ZFP) previously implicated in the restriction of endogenous retroviruses, represses a discrete subset of L1 lineages predicted to have entered the ancestral genome between 26.8 and 7.6 million years ago. In the mouse, we documented a similar chronologically conditioned pattern, albeit with a much contracted time scale. We could further identify an L1-binding KRAB-ZFP, suggesting that this rapidly evolving protein family is more globally responsible for L1 recognition. KAP1 knockdown in hESC induced the expression of KAP1-bound L1 elements, but their younger, human-specific counterparts (L1Hs) were unaffected. Instead, they were stimulated by depleting DNA methyltransferases, consistent with recent evidence demonstrating that the PIWI-piRNA pathway regulates L1Hs in hESC. Altogether, these data indicate that the early embryonic control of L1 is an evolutionary dynamic process, and support a model whereby newly emerged lineages are first suppressed by DNA methylation-inducing small RNA-based mechanisms, before KAP1-recruiting protein repressors are selected. Overall design: HA-tagged Gm6871 ChIP-seq in mES cells, RNA-seq in control and Gm6871 KD mES cells, KAP1 ChIP-seq in WT mES cells, RNA-seq in control and DNMTs KD hES cells.

转座因子是一类威胁基因组完整性的重要演化驱动因子。长散在核元件-1（Long interspersed element-1，L1，亦称LINE-1）是目前人类基因组中仅存的具有自主转座活性的转座子。其演化模式极为特殊：在任意时间节点，单一活性L1谱系可扩增至数千拷贝，随后在宿主限制因子的选择压力下被新的谱系取代——宿主限制因子主要通过在早期胚胎发生过程中沉默L1的表达发挥功能。本研究证实，在人胚胎干细胞（human embryonic stem cells，hESC）中，KAP1——此前被证实可限制内源性逆转录病毒（endogenous retroviruses）的含KRAB结构域锌指蛋白（KRAB-containing zinc finger proteins，KRAB-ZFP）的核心辅因子——可抑制特定亚类的L1谱系，这类谱系预估于2680万至760万年前进入人类祖先基因组。在小鼠中，我们同样观测到了类似的时间依赖性调控模式，仅其时间尺度显著缩短。我们进一步鉴定出一种可结合L1的KRAB-ZFP，提示这一快速演化的蛋白质家族在全局范围内参与L1的识别过程。
在hESC中敲低KAP1会诱导KAP1结合的L1元件的表达，但人类特异性的年轻L1亚类（L1Hs）并未受到影响。与之相反，敲低DNA甲基转移酶（DNA methyltransferases）可激活这类年轻亚类，这与近期研究证据相符——PIWI-piRNA通路（PIWI-piRNA pathway）可在hESC中调控L1Hs。
综上，本研究数据表明，L1的早期胚胎调控是一个动态演化的过程，并支持如下模型：新出现的L1谱系首先由诱导DNA甲基化的小RNA介导机制所抑制，后续才会招募KAP1依赖的蛋白阻遏因子。
实验设计：小鼠胚胎干细胞（mouse embryonic stem cells，mES细胞）中HA标签的Gm6871染色质免疫共沉淀测序（ChIP-seq）、对照组与Gm6871敲低小鼠ES细胞的RNA测序（RNA-seq）、野生型小鼠ES细胞中KAP1的ChIP-seq，以及对照组与DNA甲基转移酶敲低人ES细胞的RNA-seq。