ABSTRACT
While the role of transcription factors and coactivators in controlling enhancer activity and chromatin structure linked to gene expression is well established, the involvement of corepressors is not. Using inflammatory macrophage activation as a model, we investigate here a corepressor complex containing GPS2 and SMRT both genome-wide and at the Ccl2 locus, encoding the chemokine CCL2 (MCP-1). We report that corepressors co-occupy candidate enhancers along with the coactivators CBP (H3K27 acetylase) and MED1 (mediator) but act antagonistically by repressing eRNA transcription-coupled H3K27 acetylation. Genome editing, transcriptional interference, and cistrome analysis reveals that apparently related enhancer and silencer elements control Ccl2 transcription in opposite ways. 4C-seq indicates that corepressor depletion or inflammatory signaling functions mechanistically similarly to trigger enhancer activation. In ob/ob mice, adipose tissue macrophage-selective depletion of the Ccl2 enhancer-transcribed eRNA reduces metaflammation. Thus, the identified corepressor-eRNA-chemokine pathway operates in vivo and suggests therapeutic opportunities by targeting eRNAs in immuno-metabolic diseases.
Subject(s)
Chemokine CCL2/genetics , Co-Repressor Proteins/genetics , Enhancer Elements, Genetic , Intracellular Signaling Peptides and Proteins/genetics , Nuclear Receptor Co-Repressor 2/genetics , Obesity/genetics , Silencer Elements, Transcriptional , Adipose Tissue/immunology , Adipose Tissue/pathology , Animals , CRISPR-Cas Systems , Chemokine CCL2/immunology , Co-Repressor Proteins/immunology , Gene Editing , Gene Expression Regulation/drug effects , HEK293 Cells , Histone Acetyltransferases/genetics , Histone Acetyltransferases/immunology , Histones/genetics , Histones/immunology , Humans , Intracellular Signaling Peptides and Proteins/immunology , Lipopolysaccharides/pharmacology , Macrophage Activation/drug effects , Male , Mediator Complex Subunit 1/genetics , Mediator Complex Subunit 1/immunology , Mice , Mice, Obese , Nuclear Receptor Co-Repressor 2/immunology , Obesity/immunology , Obesity/pathology , RAW 264.7 Cells , RNA, Untranslated/genetics , RNA, Untranslated/immunology , Signal TransductionABSTRACT
Human in vitro generated monocyte-derived dendritic cells (moDCs) and macrophages are used clinically, e.g., to induce immunity against cancer. However, their physiological counterparts, ontogeny, transcriptional regulation, and heterogeneity remains largely unknown, hampering their clinical use. High-dimensional techniques were used to elucidate transcriptional, phenotypic, and functional differences between human in vivo and in vitro generated mononuclear phagocytes to facilitate their full potential in the clinic. We demonstrate that monocytes differentiated by macrophage colony-stimulating factor (M-CSF) or granulocyte macrophage colony-stimulating factor (GM-CSF) resembled in vivo inflammatory macrophages, while moDCs resembled in vivo inflammatory DCs. Moreover, differentiated monocytes presented with profound transcriptomic, phenotypic, and functional differences. Monocytes integrated GM-CSF and IL-4 stimulation combinatorically and temporally, resulting in a mode- and time-dependent differentiation relying on NCOR2. Finally, moDCs are phenotypically heterogeneous and therefore necessitate the use of high-dimensional phenotyping to open new possibilities for better clinical tailoring of these cellular therapies.
Subject(s)
Dendritic Cells/immunology , Interleukin-4/immunology , Macrophages/immunology , Monocytes/immunology , Nuclear Receptor Co-Repressor 2/immunology , Signal Transduction/immunology , Cell Differentiation , Cell Lineage , Dendritic Cells/cytology , Dendritic Cells/drug effects , Gene Expression Profiling , Gene Expression Regulation , Granulocyte-Macrophage Colony-Stimulating Factor/pharmacology , Humans , Immunophenotyping , Interleukin-4/genetics , Interleukin-4/pharmacology , Macrophage Activation , Macrophage Colony-Stimulating Factor/pharmacology , Macrophages/cytology , Macrophages/drug effects , Monocytes/cytology , Monocytes/drug effects , Nuclear Receptor Co-Repressor 2/genetics , Primary Cell Culture , Time Factors , Transcription, GeneticABSTRACT
Distinct CD4(+) T cell subsets are critical for host defense and immunoregulation. Although these subsets can act as terminally differentiated lineages, they have been increasingly noted to demonstrated plasticity. MicroRNAs are factors that control T cell stability and plasticity. Here we report that naturally occurring regulatory T cells (T(reg) cells) had high expression of the microRNA miR-10a and that miR-10a was induced by retinoic acid and transforming growth factor-ß (TGF-ß) in inducible T(reg) cells. By simultaneously targeting the transcriptional repressor Bcl-6 and the corepressor Ncor2, miR-10a attenuated the phenotypic conversion of inducible T(reg) cells into follicular helper T cells. We also found that miR-10a limited differentiation into the T(H)17 subset of helper T cells and therefore represents a factor that can fine-tune the plasticity and fate of helper T cells.