Ablation of cDC2 specification by triple mutations in the Zeb2 enhancer [ChIP-Seq]
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA728352 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR14472212.fastq.gz | Fastqsanger.gz | |||
| SRR14472213.fastq.gz | Fastqsanger.gz |
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Project description:The divergence of the common dendritic cell progenitor (CDP) into specified progenitors for the cDC1 and cDC2 dendritic cells subsets is poorly understood. Some transcription factors (TFs) act in commitment of already specified progenitors, such as Batf3 which stabilizes Irf8 autoactivation at the +32 kb Irf8 enhancer, but the mechanism of CDP divergence remains unknown. Here, we report the transcriptional basis of CDP divergence and describe the first requirements for pre-cDC2 specification. Genetic epistasis analysis suggested that Nfil3 acts upstream of Id2, Batf3, and Zeb2 in cDC1 development but has not revealed its mechanism or targets. Analysis of newly generated NFIL3 reporter mice showed extremely transient NFIL3 expression during cDC1 specification. CUT&RUN and ChIP-seq analysis identified endogenous NFIL3 binding in the –165 kb Zeb2 enhancer at three sites that also bind CCAAT-enhancer-binding proteins C/EBPa and C/EBPb. In vivo mutational analysis using CRISPR/Cas9 targeting showed that these NFIL3/C/EBP sites are functionally redundant, with C/EBPs supporting and NFIL3 repressing Zeb2 expression at these sites, respectively. Mutation of all three NFIL3/C/EBP sites ablated Zeb2 expression in myeloid, but not lymphoid progenitors, causing complete loss of pre-cDC2 specification and mature cDC2 development in vivo. These mice failed to generate TH2 responses against H. polygyrus infection, consistent with cDC2 supporting TH2 responses to helminths. Thus, CDP divergence is controlled by competition between NFIL3 and C/EBPs at the –165 kb Zeb2 enhancer.
2022-04-19 | GSE174108 | GEO
Project description:The divergence of the common dendritic cell progenitor (CDP) into specified progenitors for the cDC1 and cDC2 dendritic cells subsets is poorly understood. Some transcription factors (TFs) act in commitment of already specified progenitors, such as Batf3 which stabilizes Irf8 autoactivation at the +32 kb Irf8 enhancer, but the mechanism of CDP divergence remains unknown. Here, we report the transcriptional basis of CDP divergence and describe the first requirements for pre-cDC2 specification. Genetic epistasis analysis suggested that Nfil3 acts upstream of Id2, Batf3, and Zeb2 in cDC1 development but has not revealed its mechanism or targets. Analysis of newly generated NFIL3 reporter mice showed extremely transient NFIL3 expression during cDC1 specification. CUT&RUN and ChIP-seq analysis identified endogenous NFIL3 binding in the –165 kb Zeb2 enhancer at three sites that also bind CCAAT-enhancer-binding proteins C/EBPa and C/EBPb. In vivo mutational analysis using CRISPR/Cas9 targeting showed that these NFIL3/C/EBP sites are functionally redundant, with C/EBPs supporting and NFIL3 repressing Zeb2 expression at these sites, respectively. Mutation of all three NFIL3/C/EBP sites ablated Zeb2 expression in myeloid, but not lymphoid progenitors, causing complete loss of pre-cDC2 specification and mature cDC2 development in vivo. These mice failed to generate TH2 responses against H. polygyrus infection, consistent with cDC2 supporting TH2 responses to helminths. Thus, CDP divergence is controlled by competition between NFIL3 and C/EBPs at the –165 kb Zeb2 enhancer.
2022-04-19 | GSE188564 | GEO
Project description:The divergence of the common dendritic cell progenitor (CDP) into specified progenitors for the cDC1 and cDC2 dendritic cells subsets is poorly understood. Some transcription factors (TFs) act in commitment of already specified progenitors, such as Batf3 which stabilizes Irf8 autoactivation at the +32 kb Irf8 enhancer, but the mechanism of CDP divergence remains unknown. Here, we report the transcriptional basis of CDP divergence and describe the first requirements for pre-cDC2 specification. Genetic epistasis analysis suggested that Nfil3 acts upstream of Id2, Batf3, and Zeb2 in cDC1 development but has not revealed its mechanism or targets. Analysis of newly generated NFIL3 reporter mice showed extremely transient NFIL3 expression during cDC1 specification. CUT&RUN and ChIP-seq analysis identified endogenous NFIL3 binding in the –165 kb Zeb2 enhancer at three sites that also bind CCAAT-enhancer-binding proteins C/EBPa and C/EBPb. In vivo mutational analysis using CRISPR/Cas9 targeting showed that these NFIL3/C/EBP sites are functionally redundant, with C/EBPs supporting and NFIL3 repressing Zeb2 expression at these sites, respectively. Mutation of all three NFIL3/C/EBP sites ablated Zeb2 expression in myeloid, but not lymphoid progenitors, causing complete loss of pre-cDC2 specification and mature cDC2 development in vivo. These mice failed to generate TH2 responses against H. polygyrus infection, consistent with cDC2 supporting TH2 responses to helminths. Thus, CDP divergence is controlled by competition between NFIL3 and C/EBPs at the –165 kb Zeb2 enhancer.
2022-04-19 | GSE188482 | GEO
Project description:Ablation of cDC2 specification by triple mutations in the Zeb2 enhancer [CUT&RUN]
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Project description:Ablation of cDC2 specification by triple mutations in the Zeb2 enhancer [RNA-seq]
| PRJNA779428 | ENA
Project description:The divergence of the common dendritic cell progenitor (CDP) into the conventional type 1 and type 2 dendritic cell (cDC1 and cDC2, respectively) lineages is poorly understood. Some transcription factors act in the commitment of already specified progenitors-such as BATF3, which stabilizes Irf8 autoactivation at the +32 kb Irf8 enhancer-but the mechanisms controlling the initial divergence of CDPs remain unknown. Here we report the transcriptional basis of CDP divergence and describe the first requirements for pre-cDC2 specification. Genetic epistasis analysis suggested that Nfil3 acts upstream of Id2, Batf3 and Zeb2 in cDC1 development but did not reveal its mechanism or targets. Analysis of newly generated NFIL3 reporter mice showed extremely transient NFIL3 expression during cDC1 specification. CUT&RUN and chromatin immunoprecipitation followed by sequencing identified endogenous NFIL3 binding in the -165 kb Zeb2 enhancer at three sites that also bind the CCAAT-enhancer-binding proteins C/EBPα and C/EBPβ. In vivo mutational analysis using CRISPR-Cas9 targeting showed that these NFIL3-C/EBP sites are functionally redundant, with C/EBPs supporting and NFIL3 repressing Zeb2 expression at these sites. A triple mutation of all three NFIL3-C/EBP sites ablated Zeb2 expression in myeloid, but not lymphoid progenitors, causing the complete loss of pre-cDC2 specification and mature cDC2 development in vivo. These mice did not generate T helper 2 (TH2) cell responses against Heligmosomoides polygyrus infection, consistent with cDC2 supporting TH2 responses to helminths. Thus, CDP divergence into cDC1 or cDC2 is controlled by competition between NFIL3 and C/EBPs at the -165 kb Zeb2 enhancer.
2022-04-19 | GSE188579 | GEO
Project description:Ablation of cDC2 development by triple mutations within the Zeb2 enhancer
| PRJNA779440 | ENA
Project description:The divergence of the common dendritic cell progenitor1-3 (CDP) into the conventional type 1 and type 2 dendritic cell (cDC1 and cDC2, respectively) lineages4,5 is poorly understood. Some transcription factors act in the commitment of already specified progenitors-such as BATF3, which stabilizes Irf8 autoactivation at the +32 kb Irf8 enhancer4,6-but the mechanisms controlling the initial divergence of CDPs remain unknown. Here we report the transcriptional basis of CDP divergence and describe the first requirements for pre-cDC2 specification. Genetic epistasis analysis7 suggested that Nfil3 acts upstream of Id2, Batf3 and Zeb2 in cDC1 development but did not reveal its mechanism or targets. Analysis of newly generated NFIL3 reporter mice showed extremely transient NFIL3 expression during cDC1 specification. CUT&RUN and chromatin immunoprecipitation followed by sequencing identified endogenous NFIL3 binding in the -165 kb Zeb2 enhancer8 at three sites that also bind the CCAAT-enhancer-binding proteins C/EBPα and C/EBPβ. In vivo mutational analysis using CRISPR-Cas9 targeting showed that these NFIL3-C/EBP sites are functionally redundant, with C/EBPs supporting and NFIL3 repressing Zeb2 expression at these sites. A triple mutation of all three NFIL3-C/EBP sites ablated Zeb2 expression in myeloid, but not lymphoid progenitors, causing the complete loss of pre-cDC2 specification and mature cDC2 development in vivo. These mice did not generate T helper 2 (TH2) cell responses against Heligmosomoides polygyrus infection, consistent with cDC2 supporting TH2 responses to helminths9-11. Thus, CDP divergence into cDC1 or cDC2 is controlled by competition between NFIL3 and C/EBPs at the -165 kb Zeb2 enhancer.
| S-EPMC10358283 | biostudies-literature
Project description:Plasmacytoid dendritic cells (pDCs) develop from pre-pDCs, while two lineages of conventional DCs (cDC1s and cDC2s) develop from lineage-committed pre-cDCs. A number of transcription factors (TFs) have been implicated in regulating the development of pDCs (E2-2, Id2) and cDC1s (IRF8, Id2 and Batf3) however, those required for the early commitment of pre-cDCs towards the cDC2 lineage are unknown. Here we identified the TF Zinc finger E box binding homeobox 2 (Zeb2), to play a crucial role in regulating DC development. Zeb2 was expressed from the pre-pDC and pre-cDC stage onwards, and highly expressed in mature pDCs and cDC2s. Mice conditionally lacking Zeb2 in CD11c+ cells had a cell intrinsic reduction in pDCs and cDC2s, coupled with an increase in cDC1s. Conversely, mice in which CD11c+ cells overexpressed Zeb2 displayed a reduction in cDC1s. This was accompanied by altered expression of Id2, which was upregulated in cDC2s and pDCs from conditional knock-out mice. Zeb2 ChIP analysis revealed Id2 to be a direct target of Zeb2. Thus, we conclude that Zeb2 regulates commitment to both the cDC2 and pDC lineages through repression of Id2.
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Project description:Type 1 conventional dendritic cells (cDC1) are required for effective CD8 T cell responses to many viruses and tumors and for effective checkpoint blockade immunotherapy. Recently, cytokines produced in association with certain tumors were reported to impair anti-tumor immune responses by reducing the abundance of cDC1. However, the reported mechanism of this reduction remains unclear, attributed either to reduced cDC1 development or decreased peripheral cDC1 survival. Here we show that tumor-derived IL-6 blocks cDC1 development from both in murine and human systems. We show that mechanism of this blockade is the IL-6-dependent increase in C/EBPβ expression in the common dendritic cell progenitor (CDP). C/EBPβ and NFIL3 compete for binding to sites in the -165 kb Zeb2 enhancer, and support or repress Zeb2 expression respectively. At homeostastis, pre-cDC1 specification occurs upon Nfil3 induction and consequent Zeb2 suppression. However, IL-6 strongly induces C/EBPβ expression in CDPs, thereby preventing normal NFIL3-dependent pre-cDC1 specification. Importantly, the ability of IL-6 to block cDC1 development is dependent on the presence of C/EBPβ binding sites in the -165 kb Zeb2 enhancer, as this effect is lost in Δ1+2+3 mutnat mice in which these binding sites are mutated. These results explain how tumor-associated IL-6 suppresses cDC1 development and suggest therapeutic approaches preventing abnormal C/EBPβ induction in CDPs may help reestablish cDC1 development to enhance anti-tumor immunity.
2023-06-22 | GSE215751 | GEO