KLF4 inhibits early neural differentiation of ESCs by coordinating specific 3D chromatin structure

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, such as Pax6. Combination analysis employing 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. And changes of chromatin structure upregulated expression of neural differentiation-associated genes and promoted early neural differentiation. This study reveals that KLF4 has a differentiation inhibitory effect in regulating 3D chromatin structure and suggests KLF4 inhibits early neural differentiation by regulation of 3D chromatin structure, which is a new mechanism of early neural differentiation. 2 biological replicates were analyzed for WT and KLF4-KO by RNA-seq and HiChIP analysis.

胚胎干细胞（ESCs）的神经分化依赖于精准调控的基因表达网络，该过程部分由三维染色质结构的动态变化所调控。目前，此类染色质结构变化在此背景下如何调控基因表达的机制仍不明确。本研究发现，在与神经分化相关基因（如Pax6）的转录起始位点（TSS）关联区域的部分预激活或封闭增强子处，转录因子KLF4存在富集现象。通过联合分析ChIP、HiChIP及RNA-seq数据，我们观察到胚胎干细胞中KLF4的缺失会引发三维染色质结构改变，具体表现为神经分化相关基因与活性增强子之间的染色质相互作用环增多。此类染色质结构变化上调了神经分化相关基因的表达水平，并促进了早期神经分化进程。本研究揭示了KLF4在三维染色质结构调控中发挥分化抑制作用，并表明KLF4可通过调控三维染色质结构抑制早期神经分化，这为早期神经分化的机制研究提供了全新视角。本研究针对野生型（WT）与KLF4基因敲除（KLF4-KO）样本，通过RNA-seq与HiChIP分析完成了2次生物学重复检测。