Thrombopoietin signaling to chromatin elicits rapid and pervasive epigenome remodeling within poised chromatin architectures. Mus musculus

Thrombopoietin (TPO) is a critical cytokine regulating hematopoietic stem cell maintenance and differentiation into the megakaryocytic lineage. However, the transcriptional and chromatin dynamics elicited by TPO signaling are poorly understood. Here, we study the immediate early transcriptional and cis-regulatory responses to TPO in hematopoietic stem/progenitor cells (HSPCs) and use this paradigm of cytokine signaling to chromatin to dissect the relation between cis-regulatory activity and chromatin architecture. We show that TPO profoundly alters the transcriptome of HSPCs, with key hematopoietic regulators being transcriptionally repressed within 30 minutes of TPO. By examining cis-regulatory dynamics and chromatin architectures, we demonstrate that these changes are accompanied by rapid and extensive epigenome remodeling of cis-regulatory landscapes that is spatially coordinated within topologically associating domains (TADs). Moreover, homotypic TPO-responsive enhancers are spatially clustered and engage in preferential intra- and inter-TAD interactions that are only moderately perturbed by TPO signaling. By further examining the link between cis-regulatory dynamics and chromatin looping, we show that rapid modulation of cis-regulatory activity is largely independent of chromatin looping dynamics. Finally, we show that activated and repressed cis-regulatory elements share a remarkably similar DNA sequence composition and that in vivo transcription factor binding patterns accurately predict rapid cis-regulatory responses to TPO. Overall design: Subcellular RNA-seq, ChIP-seq, Hi-C and Promoter Capture Hi-C in HPC7 cells and two biological conditions.

血小板生成素（Thrombopoietin, TPO）是调控造血干细胞维持以及向巨核细胞系分化的关键细胞因子。然而，人们对TPO信号通路所引发的转录与染色质动态变化尚缺乏深入了解。本研究聚焦于造血干/祖细胞（hematopoietic stem/progenitor cells, HSPCs）对TPO的即刻早期转录与顺式调控应答，并以该细胞因子信号调控染色质的模型为基础，解析顺式调控活性与染色质三维结构之间的关联。研究结果显示，TPO可显著改变HSPCs的转录组，在TPO处理30分钟内，关键造血调控因子即发生转录抑制。通过分析顺式调控动态变化与染色质三维结构，我们发现这些转录改变伴随顺式调控区域的快速大规模表观基因组重塑，且该重塑在拓扑关联结构域（topologically associating domains, TADs）内呈现空间协同性。此外，同源的TPO应答增强子存在空间聚集，并倾向于发生偏好性的TAD内部及跨TAD相互作用，而此类相互作用仅被TPO信号通路轻度扰动。进一步探究顺式调控动态与染色质环之间的关联后，我们发现顺式调控活性的快速调控在很大程度上独立于染色质环的动态变化。最后，本研究证实，激活与抑制的顺式调控元件具有高度相似的DNA序列组成，且体内转录因子结合模式可精准预测细胞对TPO的快速顺式调控应答。整体实验设计：在HPC7细胞中设置两种生物学条件，分别进行亚细胞RNA测序（subcellular RNA-seq）、染色质免疫沉淀测序（ChIP-seq）、Hi-C测序及启动子捕获Hi-C（Promoter Capture Hi-C）检测。