Chromatin activity of IκBα mediates the exit from naïve pluripotency (RNA-Seq). Chromatin activity of IκBα mediates the exit from naïve pluripotency (RNA-Seq)

Inflammatory signals are key in development and cell differentiation but their orchestration with pluripotency and stemness signals is poorly understood. Our previous work identified a chromatin function of IκBα, the NF-κB inhibitor, that is crucial for differentiation in different types of somatic stem cells. Here we demonstrate that deficiency of IκBα imposes a profound chromatin rewiring defect that impacts on DNA methylation, histone post-translational modifications and transcriptional regulation, stabilizing mouse embryonic stem cells (ESCs) in a ground state of pluripotency while preventing them from pluripotency exit and differentiation. By engineering separation-of-function mutants of IκBα with specific binding to either NF-κB or histones, we demonstrate that regulation of pluripotency state by IκBα is independent of NF-kB but requires the chromatin-related IκBα function. Overall design: We performed bulk RNAseq analysis in mESCs and two early differentiation stages (EBs 48h and EBs 96h). Per timepoint, 3xWT and IκBα 3xKO independent biological replicates were tested resulting in a total of 18 samples.

炎症信号在发育与细胞分化过程中发挥关键调控作用，但其与多能性（pluripotency）及干细胞干性（stemness）信号的协同调控机制仍未被充分阐明。本课题组前期研究发现，核因子κB（NF-κB）抑制剂IκBα具有染色质调控功能，该功能对多种成体干细胞的分化进程至关重要。本研究证实，IκBα缺失会引发严重的染色质重编程缺陷，该缺陷可影响DNA甲基化、组蛋白翻译后修饰及转录调控，能够将小鼠胚胎干细胞（ESCs）稳定维持在多能性基态，同时阻止其退出多能状态并启动分化程序。通过构建仅能特异性结合核因子κB（NF-κB）或组蛋白的IκBα功能分离突变体，本研究证实IκBα对多能状态的调控不依赖于核因子κB通路，而是依赖其染色质相关调控功能。实验设计：本研究对小鼠胚胎干细胞（mESCs）以及两个早期分化阶段的样本（培养48小时的胚状体（EBs 48h）与培养96小时的胚状体（EBs 96h））开展了批量RNA测序（bulk RNAseq）分析。每个时间点设置3份野生型（WT）与3份IκBα敲除（KO）独立生物学重复样本，最终总计获得18个测序样本。