Enhancer Landscape Dynamics and the Role of IRF8 in Mononuclear Phagocyte Development

Monocytes and dendritic cells (DCs) are critical mononuclear phagocytes that regulate innate and adaptive immune responses. Hematopoietic stem cells give rise to monocytes and DCs via intermediate myeloid progenitor populations, such as granulocyte-monocyte progenitors (GMPs), monocyte-DC progenitors (MDPs), and common monocyte progenitors (cMoPs) or common DC progenitors (CDPs). However, the molecular mechanism underlying their lineage determination is poorly understood. Recently, promoter-distal enhancers have been found to be key for cell fate decision. In this study, we performed chromatin immunoprecipitation-sequencing (ChIP-seq) analysis of three histone modifications (H3K4me1, H3K27ac, and H3K4me3, representing primed enhancers, active enhancers, and transcriptional intiation, respectively) in seven myeloid populations (GMPs, MDPs, cMoPs, CDPs, monocytes, DCs, and neutrophils), and identified approximately sixty thousand putative enhancer regions. We found that a majority of monocyte- and DC-specific active enhancers were gradually established at progenitor stages prior to gene expression and terminal differentiation. Computational DNA motif analysis implicated that these enhancers were regulated by combinations of lineage-determining transcription factors such as PU.1, RUNX, C/EBP, and IRF. Indeed, ChIP-seq of PU.1 confirmed that the myeloid master regulator PU.1 was the common transcription factors bound to enhancers at all the stages examined. IRF binding motifs were enriched at the enhancer regions in MDPs, cMoPs and CDPs, but not GMPs. Among IRFs, IRF8, a partner of PU.1, has been shown to be highly expressed in MDPs and their descendants, and to be required for MDP-to-CDP and cMoP-to-monocyte transitions. Indeed, ChIP-seq analysis in Irf8^-/- progenitors demonstrated that the enhancer landscapes of Irf8^-/- GMPs, MDPs, and cMoPs all remained similar to that of wild-type GMPs. Moreover, ChIP-seq for IRF8 binding revealed that IRF8 directly promoted the priming and activation of many enhancers in MDPs and cMoPs. These results contribute to a comprehensive understanding of how transcription factors govern the enhancer landscape dynamics during mononuclear phagocyte development.