No response by an immune cell occurs in isolation but is instead the summation of multiple interacting signals. Ivashkiv and colleagues describe the epigenetic landscape that results from the integration of the inflammatory cytokines type I interferons and TNF. Cross-regulation of Toll-like receptor (TLR) responses by cytokines is essential for effective host defense, avoidance of toxicity and homeostasis, but the underlying mechanisms are not well understood. Our comprehensive epigenomics approach to the analysis of human macrophages showed that the proinflammatory cytokines TNF and type I interferons induced transcriptional cascades that altered chromatin states to broadly reprogram responses induced by TLR4. TNF tolerized genes encoding inflammatory molecules to prevent toxicity while preserving the induction of genes encoding antiviral and metabolic molecules. Type I interferons potentiated the inflammatory function of TNF by priming chromatin to prevent the silencing of target genes of the transcription factor NF-κB that encode inflammatory molecules. The priming of chromatin enabled robust transcriptional responses to weak upstream signals. Similar chromatin regulation occurred in human diseases. Our findings reveal that signaling crosstalk between interferons and TNF is integrated at the level of chromatin to reprogram inflammatory responses, and identify previously unknown functions and mechanisms of action of these cytokines.

Activation of macrophage proinflammatory and antimicrobial phenotypes is regulated by IFN-γ and LPS via synergistic induction of canonical, inflammatory NF-κB target genes. However, whether IFN-γ negatively regulates components of the LPS response, and how this may affect macrophage activation, is still unclear. Here we use combined transcriptomic and epigenomic approaches to find that IFN-γ selectively abrogates LPS-induced feedback and alters macrophage metabolic pathways by suppressing TLR4-mediated gene activation. In contrast to superinduction of inflammatory genes via enhancers that bind IRF1 and STAT1, IFN-γ represses target enhancers that bind STAT3. TLR4-activated but IFN-γ-suppressed enhancers comprise two subsets discernable by differential regulation of histone acetylation and recruitment of STAT3, CDK8 and cohesin. Our findings thus show that IFN-γ suppresses feedback inhibitory and metabolic components of TLR responses to enhance macrophage activation; they also provide insights for IFN-γ-mediated selective inhibition of TLR4-induced transcription. Such inhibition can contribute to severe and sustained inflammatory responses.

The immunoregulatory cytokine interleukin 10 (IL-10) modulates the function of diverse immune and non-immune cells. Here, we examine the chromatin structural changes associated with IL10 gene transcription by naïve and differentiated murine T cells. Naïve T cells lack DNase I hypersensitive (HS) sites in the vicinity of the IL10 gene, whereas differentiated T cells display a strong 3′ constitutive HS site as well as several inducible sites. The majority of HS sites map to regions that are strongly conserved in sequence between mouse and human genomes. In committed Th1 cells, the mechanism of IL10 gene silencing is associated with the development of repressive histone modifications near the IL10 promoter and also near intronic hypersensitive regions of the IL10 gene. Our results constitute the first report of chromatin structural differences within the IL10 gene in differentiated Th1 and Th2 cells and emphasize the surprising diversity of mechanisms used to regulate cytokine gene expression at the chromatin level.

The mechanisms by which IFN-γ activates expression of interferon-stimulated genes that have inflammatory and host defense functions are well understood. In contrast, little is known about how IFN-γ represses gene expression. By using transcriptomic and epigenomic analysis, we found that stable repression of a small group of genes by IFN-γ is associated with recruitment of the histone methyltransferase EZH2 and deposition of the negative mark histone 3 lysine 27 trimethylation (H3K27me3) at their promoters. Repressed genes included MERTK, PPARG, and RANK, which have anti-inflammatory functions and promote osteoclast differentiation. Gene repression and H3K27me3 persisted after IFN-γ signaling was terminated, and these silenced genes were no longer responsive to glucocorticoids, IL-4, and M-CSF. These results identify cytokine-induced H3K27 trimethylation as a mechanism that stabilizes gene silencing in macrophages. IFN-γ-induced macrophage activation is thus reinforced by a chromatin-based mechanism that blocks anti-inflammatory and opposing pathways.