Metazoan development involves the successive activation and silencing of specific gene expression programs and is driven by tissue-specific transcription factors programming the chromatin landscape. To understand how this process executes an entire developmental pathway, we generated global gene expression, chromatin accessibility, histone modification, and transcription factor binding data from purified embryonic stem cell-derived cells representing six sequential stages of hematopoietic specification and differentiation. Our data reveal the nature of regulatory elements driving differential gene expression and inform how transcription factor binding impacts on promoter activity. We present a dynamic core regulatory network model for hematopoietic specification and demonstrate its utility for the design of reprogramming experiments. Functional studies motivated by our genome-wide data uncovered a stage-specific role for TEAD/YAP factors in mammalian hematopoietic specification. Our study presents a powerful resource for studying hematopoiesis and demonstrates how such data advance our understanding of mammalian development.

Recent studies have established that during embryonic development, hematopoietic progenitors and stem cells are generated from hemogenic endothelium precursors through a process termed endothelial to hematopoietic transition (EHT). The transcription factor RUNX1 is essential for this process, but its main downstream effectors remain largely unknown. Here, we report the identification of Gfi1 and Gfi1b as direct targets of RUNX1 and critical regulators of EHT. GFI1 and GFI1B are able to trigger, in the absence of RUNX1, the down-regulation of endothelial markers and the formation of round cells, a morphologic change characteristic of EHT. Conversely, blood progenitors in Gfi1- and Gfi1b-deficient embryos maintain the expression of endothelial genes. Moreover, those cells are not released from the yolk sac and disseminated into embryonic tissues. Taken together, our findings demonstrate a critical and specific role of the GFI1 transcription factors in the first steps of the process leading to the generation of hematopoietic progenitors from hemogenic endothelium.

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The expression of eukaryotic genes is regulated by cis-regulatory elements such as promoters and enhancers, which bind sequence-specific DNA-binding proteins. One of the great challenges in the gene regulation field is to characterise these elements. This involves the identification of transcription factor (TF) binding sites within regulatory elements that are occupied in a defined regulatory context. Digestion with DNase and the subsequent analysis of regions protected from cleavage (DNase footprinting) has for many years been used to identify specific binding sites occupied by TFs at individual cis-elements with high resolution. This methodology has recently been adapted for high-throughput sequencing (DNase-seq). In this study, we describe an imbalance in the DNA strand-specific alignment information of DNase-seq data surrounding protein–DNA interactions that allows accurate prediction of occupied TF binding sites. Our study introduces a novel algorithm, Wellington, which considers the imbalance in this strand-specific information to efficiently identify DNA footprints. This algorithm significantly enhances specificity by reducing the proportion of false positives and requires significantly fewer predictions than previously reported methods to recapitulate an equal amount of ChIP-seq data. We also provide an open-source software package, pyDNase, which implements the Wellington algorithm to interface with DNase-seq data and expedite analyses.