KLF4 inhibits early neural differentiation of ESCs by coordinating specific 3D chromatin structure, a new mechanism of early neural differentiation. KLF4 inhibits early neural differentiation of ESCs by coordinating specific 3D chromatin structure, a new mechanism of early neural differentiation

Circular RNA has been reported to be dynamically expressed during embryonic development and regulates human embryonic stem cells (hESCs), but the identification and regulation of functional circular RNA in mouse embryonic stem cells (mESC) remains unclear. Neural differentiation of embryonic stem cells (ESCs) requires precisely orchestrated gene regulation, a process governed in part by changes in 3D chromatin structure. How these changes regulate gene expression in this context remains unclear. In this study, we observed enrichment of the transcription factor KLF4 at some poised or closed enhancers at TSS-linked regions of genes associated with neural differentiation. Combination analysis of ChIP, HiChIP and RNA-seq data indicated that KLF4 loss in ESCs induced changes in 3D chromatin structure, including increased chromatin interaction loops between neural differentiation-associated genes and active enhancers, leading to upregulated expression of neural differentiation-associated genes and therefore early neural differentiation. This study suggests KLF4 inhibits early neural differentiation by regulation of 3D chromatin structure, which is a new mechanism of early neural differentiation. Conclusions: Our study suggests KLF4 inhibits early neural differentiation by regulation of 3D chromatin structure, which is a new mechanism of early neural differentiation. Overall design: Two biological replicates were analyzed for each experimental condition. WT and KO cells were RNA-seq/HiChIP in ESCs or neuro induction at day 12.

已有研究证实，环状RNA（Circular RNA）在胚胎发育进程中呈动态表达模式，并可调控人类胚胎干细胞（hESCs），但目前小鼠胚胎干细胞（mESC）中功能性环状RNA的鉴定及其调控机制仍未明确。
胚胎干细胞的神经分化依赖于精密有序的基因调控程序，该过程在一定程度上由三维染色质结构的变化所调控。目前，此类结构变化在此背景下如何调控基因表达仍有待阐明。
本研究观察到，转录因子KLF4富集于与神经分化相关基因的转录起始位点（TSS）相连区域中的部分预激活（poised）或沉默增强子处。
对染色质免疫沉淀（ChIP）、HiChIP及RNA测序（RNA-seq）数据的联合分析显示，胚胎干细胞中KLF4的缺失会引发三维染色质结构改变，具体包括神经分化相关基因与活性增强子之间的染色质互作环增多，进而导致神经分化相关基因表达上调，最终促进早期神经分化。
本研究表明，KLF4可通过调控三维染色质结构抑制早期神经分化，这是早期神经分化的一种全新机制。
结论：本研究揭示KLF4通过调控三维染色质结构抑制早期神经分化，这是早期神经分化的一种新型机制。
整体实验设计：每个实验条件均设置两个生物学重复。野生型（WT）与敲除型（KO）细胞分别在胚胎干细胞状态及第12天神经诱导阶段进行RNA-seq与HiChIP测序。