Abstract
Hematopoietic stem cell (HSC) ontogeny is accompanied by dynamic changes in gene regulatory networks. We performed RNA-seq and histone mark ChIP-seq to define the transcriptomes and epigenomes of cells representing key developmental stages of HSC ontogeny in mice. The five populations analyzed were embryonic day 10.5 (E10.5) endothelium and hemogenic endothelium from the major arteries, an enriched population of prehematopoietic stem cells (pre-HSCs), fetal liver HSCs, and adult bone marrow HSCs. Using epigenetic signatures, we identified enhancers for each developmental stage. Only 12% of enhancers are primed, and 78% are active, suggesting the vast majority of enhancers are established de novo without prior priming in earlier stages. We constructed developmental stage-specific transcriptional regulatory networks by linking enhancers and predicted bound transcription factors to their target promoters using a novel computational algorithm, target inference via physical connection (TIPC). TIPC predicted known transcriptional regulators for the endothelial-to-hematopoietic transition, validating our overall approach, and identified putative novel transcription factors, including the broadly expressed transcription factors SP3 and MAZ. Finally, we validated a role for SP3 and MAZ in the formation of hemogenic endothelium. Our data and computational analyses provide a useful resource for uncovering regulators of HSC formation.
Highlights
Hematopoietic stem and progenitor cells (HSPCs) differentiate from a small population of endothelial cells in the embryo called hemogenic endothelium (HE) (Slukvin and Uenishi 2019)
The process of HSPC formation from HE involves an endothelial to hematopoietic transition (EHT), in which flat HE cells in a monolayer transition to rounded cells that detach from the endothelial layer and enter circulation (Slukvin and Uenishi 2019)
We predict that distinct requirements for HSPC formation from arterial endothelium are layered on top of a canonical EHT pathway that is required in all endothelial cells
Summary
Hematopoietic stem and progenitor cells (HSPCs) differentiate from a small population of endothelial cells in the embryo called hemogenic endothelium (HE) (Slukvin and Uenishi 2019). Chromatin analyses combined with ChIP-seq determination of transcription factor (TF) occupancy were used to define the chromatin landscape and predict transcriptional regulatory networks (TRNs) at each step of the differentiation process from ES cells to adherent macrophages (Goode et al 2016). This comprehensive study identified potential regulators of specific steps in the developmental trajectory that were validated in functional assays. We used a novel algorithm for predicting TRNs that we call target inference via physical connection (TIPC) to identify TFs operational at discrete stages of HSC formation, and demonstrate the importance of two broadly expressed TFs predicted by TIPC, SP3 and MAZ, in regulating the number of HE cells
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