Understanding the Transcriptional Mechanisms Underlying Dendritic Cell Development

Dendritic cells (DCs) comprise an important immune lineage that plays a critical role in initiating and sustaining the proper immune response. They can be divided into two distinct branches, classical/conventional DCs (cDCs) or plasmacytoid DCs (pDCs). cDCs can further be classified as cDC1 or cDC2. Each DC subset exerts unique functions in vivo and are necessary for a complete immune response. The precise transcriptional programs underlying DC specification and commitment remain unclear. cDC1, cDC2, and pDC all arise from the common DC progenitor (CDP) in the bone marrow. How the CDP gives rise to all three DC subsets in an important outstanding question in the field. Several transcription factors have been shown to be important for the development of certain subsets. The transcription factors Irf8, Batf3, Id2, Nfil3, and Bcl6 are required for the cDC1 lineage, while the transcription factors Klf4 and Notch2 are necessary for specific cDC2 subsets. pDCs rely on the transcription factors Tcf4 and Zeb2 for their development. Despite knowing that these factors influence DC development, the interactions between these factors and their timing of action are unclear. Recently, understanding of how the CDP specifies benefited from identifying cDC progenitors (pre-cDCs) that were found to include clonogenic populations separately committed to cDC1 or cDC2 lineages. Two Irf8 enhancers were found to affect cDC1 development in different stages: an E-protein dependent enhancer located 41 kilobases downstream of the transcription start site of IRF8 (+41 kb Irf8 enhancer) is required for the specification of the pre-cDC1, and a BATF-dependent +32 kb Irf8 enhancer required for the maturation to the cDC1. To understand the switch in Irf8 enhancer usage during cDC1 specification, we used single-cell RNA-sequencing of the CDP and identified a cluster of cells that expressed transcription factors that influence cDC1 development, such as Nfil3, Id2, and Zeb2. We then performed genetic epistasis to determine the functional hierarchy of transcription factors involved in cDC1 specification. We organized a transcriptional circuit that explains the switch in Irf8 expression from being Batf3-independent to being Batf3-dependent. The CDP originates in a Zeb2hi and Id2lo state in which Irf8 expression is maintained by the +41 kb Irf8 enhancer. Single-cell RNA-sequencing identified a fraction of the CDP that exclusively possesses cDC1 fate potential. This fraction’s development arises when Nfil3 induces a transition into a Zeb2lo and Id2hi state. A circuit of mutual Zeb2-Id2 repression serves to stabilize states before and after this transition. Id2 expression in the specified pre-cDC1 inhibits E proteins, blocking activity of the +41 kb Irf8 enhancer, and thereby imposing a new requirement for Batf3 for maintaining Irf8 expression via the +32 kb Irf8 enhancer

Indoleamine 2,3-dioxygenase 1 activation in mature cDC1 promotes tolerogenic education of inflammatory cDC2 via metabolic communication

Conventional dendritic cells (cDCs), cDC1 and cDC2, act both to initiate immunity and maintain self-tolerance. The tryptophan metabolic enzyme indoleamine 2,3-dioxygenase 1 (IDO1) is used by cDCs in maintaining tolerance, but its role in different subsets remains unclear. At homeostasis, only mature CCR

Cis interactions in the Irf8 locus regulate stage-dependent enhancer activation

Individual elements within a superenhancer can act in a cooperative or temporal manner, but the underlying mechanisms remain obscure. We recently identified a

Altered compensatory cytokine signaling underlies the discrepancy between Flt3–/– and Flt3l–/– mice

The receptor Flt3 and its ligand Flt3L are both critical for dendritic cell (DC) development, but DC deficiency is more severe i

Mafb lineage tracing to distinguish macrophages from other immune lineages reveals dual identity of Langerhans cells

Current systems for conditional gene deletion within mouse macrophage lineages are limited by ectopic activity or low efficiency. In this study, we generated a Mafb-driven Cre strain to determine whether any dendritic cells (DCs) identified by Zbtb46-GFP expression originate from a Mafb-expressing population. Lineage tracing distinguished macrophages from classical DCs, neutrophils, and B cells in all organs examined. At steady state, Langerhans cells (LCs) were lineage traced but also expressed Zbtb46-GFP, a phenotype not observed in any other population. After exposure to house dust mite antigen, Zbtb46-negative CD64(+) inflammatory cells infiltrating the lung were substantially lineage traced, but Zbtb46-positive CD64(−) cells were not. These results provide new evidence for the unique identity of LCs and challenge the notion that some inflammatory cells are a population of monocyte-derived DCs

Distinct Transcriptional Programs Control Cross-Priming in Classical and Monocyte-Derived Dendritic Cells

Both classical DCs (cDCs) and monocyte-derived DCs (Mo-DCs) are capable of cross-priming CD8+ T cells in response to cell-associated antigens. We found that Ly-6ChiTREML4− monocytes can differentiate into Zbtb46+ Mo-DCs in response to granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4) but that Ly-6ChiTREML4+ monocytes were committed to differentiate into Ly-6CloTREML4+ monocytes. Differentiation of Zbtb46+ Mo-DCs capable of efficient cross-priming required both GM-CSF and IL-4 and was accompanied by the induction of Batf3 and Irf4. However, monocytes require IRF4, but not BATF3, to differentiate into Zbtb46+ Mo-DCs capable of cross-priming CD8+ T cells. Instead, Irf4−/− monocytes differentiate into macrophages in response to GM-CSF and IL-4. Thus, cDCs and Mo-DCs require distinct transcriptional programs of differentiation in acquiring the capacity to prime CD8+ T cells. These differences may be of consideration in the use of therapeutic DC vaccines based on Mo-DCs

Oral Antibiotic Treatment of Mice Exacerbates the Disease Severity of Multiple Flavivirus Infections

Summary: Although the outcome of flavivirus infection can vary from asymptomatic to lethal, environmental factors modulating disease severity are poorly defined. Here, we observed increased susceptibility of mice to severe West Nile (WNV), Dengue, and Zika virus infections after treatment with oral antibiotics (Abx) that depleted the gut microbiota. Abx treatment impaired the development of optimal T cell responses, with decreased levels of WNV-specific CD8+ T cells associated with increased infection and immunopathology. Abx treatments that resulted in enhanced WNV susceptibility generated changes in the overall structure of the gut bacterial community and in the abundance of specific bacterial taxa. As little as 3 days of treatment with ampicillin was sufficient to alter host immunity and WNV outcome. Our results identify oral Abx therapy as a potential environmental determinant of systemic viral disease, and they raise the possibility that perturbation of the gut microbiota may have deleterious consequences for subsequent flavivirus infections. : Thackray et al. observed increased susceptibility to West Nile, Zika, and Dengue virus infections following oral antibiotic treatment in mice. Antibiotics altered the bacterial abundance and community structure and the development of optimal T cell immunity. These data suggest that antibiotics may have deleterious consequences for subsequent flavivirus infections. Keywords: West Nile virus, Dengue virus, Zika virus, flavivirus, oral antibiotics, gut microbiota, risk factors, pathogenesis determinants, immunit

Differential usage of transcriptional repressor Zeb2 enhancers distinguishes adult and embryonic hematopoiesis

The transcriptional repressor ZEB2 regulates development of many cell fates among somatic, neural, and hematopoietic lineages, but the basis for its requirement in these diverse lineages is unclear. Here, we identified a 400-basepair (bp) region located 165 kilobases (kb) upstream of the Zeb2 transcriptional start site (TSS) that binds the E proteins at several E-box motifs and was active in hematopoietic lineages. Germline deletion of this 400-bp region (Zeb2Δ-165mice) specifically prevented Zeb2 expression in hematopoietic stem cell (HSC)-derived lineages. Zeb2Δ-165 mice lacked development of plasmacytoid dendritic cells (pDCs), monocytes, and B cells. All macrophages in Zeb2Δ-165 mice were exclusively of embryonic origin. Using single-cell chromatin profiling, we identified a second Zeb2 enhancer located at +164-kb that was selectively active in embryonically derived lineages, but not HSC-derived ones. Thus, Zeb2 expression in adult, but not embryonic, hematopoiesis is selectively controlled by the -165-kb Zeb2 enhancer