System-wide Dissection of the Transcriptional Response to RUNX1 During Hematopoietic Specification [ChIP-seq]. System-wide Dissection of the Transcriptional Response to RUNX1 During Hematopoietic Specification [ChIP-seq]

The specification of hematopoietic cells in the developing embryo occurs in specific stages and is regulated by the successive establishment of specific transcriptional networks. However, the molecular mechanisms of how the different stages switch from one to another are still not well understood. Hematopoietic cells arise from endothelial cells within the dorsal aorta which transit into hematopoietic cells by a process called the endothelial-hematopoietic transition (EHT) which does not involve DNA replication. The transcription factor RUNX1 is essential for this process. Using the differentiation of mouse embryonic stem cells carrying an inducible version of RUNX1, we have previously shown that hematopoietic genes are primed prior to the EHT by the binding of transcription factors required to form both endothelial and hematopoietic cells (FLI-1 and SCL/TAL1). We demonstrated that after induction RUNX1 reshapes the transcription factor binding landscape by causing a relocation of these factors and pulling them towards its binding sites. In the study presented here, we employed the same system to globally dissect the transcriptional processes that underlay the EHT. We demonstrate that the RUNX1-mediated movement of FLI-1 involves the recruitment of the basal transcription components CDK9 and BRD4 to promoters. The looping factor LDB1 to binds to distal elements and after induction relocates towards RUNX1/FLI-1 to form a co-localizing complex in chromatin. This entire process is blocked by treatment with the BRD4 inhibitor JQ1. Our study constitutes a paradigm for transcriptional processes driving transitions in cellular shape and function which are widely observed in development and disease. Overall design: ChIP-seq expreiments have been used to study RUNX1 transcription factor during Hematopoietic specification

发育胚胎中的造血细胞（hematopoietic cells）特化具有特定阶段，并受特异性转录网络的相继建立所调控。然而，不同发育阶段间的转换分子机制目前仍未完全阐明。造血细胞源自背主动脉内的内皮细胞，后者通过一种名为内皮-造血转化（endothelial-hematopoietic transition, EHT）的过程转化为造血细胞，该过程不涉及DNA复制。转录因子RUNX1是该过程的必需因子。本团队此前利用携带可诱导型RUNX1的小鼠胚胎干细胞分化模型，证实造血基因在EHT发生前已被预启动，该过程由构建内皮与造血细胞所需的转录因子（FLI-1与SCL/TAL1）结合所介导。研究表明，RUNX1诱导后可重塑转录因子结合图谱：通过促使上述转录因子发生重定位，并将其招募至自身结合位点处。本研究采用同一模型，对EHT背后的全局转录调控过程进行解析。研究证实，RUNX1介导的FLI-1移动过程，涉及将基础转录组分CDK9与BRD4招募至基因启动子区域。环化因子LDB1可结合远端调控元件，诱导发生后可向RUNX1/FLI-1发生重定位，进而在染色质中形成共定位复合物。该完整过程可被BRD4抑制剂JQ1处理所阻断。本研究为驱动细胞形态与功能转变的转录调控过程提供了范式，这类转变在发育与疾病进程中广泛存在。实验设计：本研究采用染色质免疫共沉淀测序（ChIP-seq）技术，对造血细胞特化过程中的RUNX1转录因子进行研究。