Inflammation itself creates a microenvironment able to trigger epigenetic reprogramming that can lead to a distorted immune response. Epigenetic modifications lie at the heart of gene-specific control of the inflammatory response.34, 52, 53 Much of the capacity for selective regulation of inflammatory genes appears to be established at the chromatin level during development.54 Chromatin structure and histone modifications have been shown to be essential selective regulators of a large range of inflammatory genes,34, 52, 53 demonstrating the potential importance of these chromatin-based events in amplifying and perpetuating the inflammatory response (see below).
Regulation of Inflammation-induced Transcriptional Activation
Transcriptional activation involves a gene-specific multistep recruitment of distinct families of proteins to form an active transcription complex or enhanceosome.53, 55 In general, the first step requires a defined set of transcriptional activators to interact with specific DNA sequences present in the promoter and enhancers of genes. This leads to the recruitment of coactivator complexes containing chromatin-modifying enzymes and chromatin remodeling complexes, which in turn facilitate the recruitment and assembly of the preinitiation complex. This comprises RNA polymerase II and general transcription factors to render genes transcriptionally competent (Fig. 1). Then, in order to initiate RNA synthesis and productive elongation of RNA, the sequential phosphorylation of a number of serines at the carboxy-terminal domain of RNA polymerase II is required.55, 56 This creates binding sites for other transcription proteins that are important for mRNA processing and elongation.
A well-studied example of this model of transcriptional activation is given by human interferon-β (IFN-β) gene expression.57, 58 Upon viral infection, a multiprotein complex assembles on the regulatory region of the IFN-β promoter through the binding of inducible transcription factors including NF-κB, interferon regulatory factors, and ATF2/c-jun. This leads to the sequential recruitment of the p300 and GCN5 HATs and the SWI/SNF nucleosome remodeling complex, resulting in nucleosome displacement and recruitment of general transcription factors and RNA polymerase II to activate gene transcription.
Inflammatory gene transcription requires precise combinations of specific regulatory mechanisms based on cell type, stimulus, and individual genetic variation and is also associated with specific histone modifications.59 The induction of some primary response genes (such as IL6, IL8, MCP-1, and IL-12p40) by Toll-like receptor (TLR) signaling is marked by phosphorylation of histone 3 at serine 10 (H3S10), methylation at H3K4 and acetylation at H3K9/K14.60–62
Recent studies exploiting the lipopolysaccharide (LPS)-inducible transcriptional program as a model for understanding the regulation of inflammatory genes have shed light on the role of chromatin structure as a crucial regulator of the kinetics of NF-κB target gene activation.63, 64 Two classes of TLR-induced genes have been identified that are activated with differential kinetics. The first class are primary/immediate early genes that are rapidly induced and protein synthesis-independent, e.g., IL-1β and TNF-α in resting macrophages are packaged in a poised structure that facilitates rapid gene activation under physiological stimulatory conditions.60, 63, 64 These genes have a chromatin structure that resembles that of constitutively active genes and inducible transcription factors that enhance transcription initiation, elongation, and pre-mRNA processing. Evidence for the abnormal formation of a poised structure at cytokine promoters under disease conditions has been reported.65 The epigenetic variability underlying the variation in cytokine production in the human population may explain the differences in the inflammatory response between individuals with inflammatory/autoimmune diseases, including IBD.
Transcriptional activation of the second class of genes, the so-called secondary (delayed and protein synthesis dependent) genes, requires that a chromatin barrier is overcome, e.g., in the case of NF-κB target genes after LPS stimulation of macrophages.60, 63, 66 In other cases, the removal of corepressor complexes, such as NCoR and SMRT, is required, which contain HDACs and other subunits that serve to maintain a repressive chromatin structure.67 Hence, the assembly of promoters into nucleosomes provides a barrier to transcriptional activation and the requirement for nucleosome remodeling is gene-dependent. Another subset of LPS-inducible genes has been identified that undergoes demethylation of H3K9 and H3K27 as a prerequisite for induction.61, 68
Interestingly, some cytokines have been reported to induce changes in chromatin states. IL-6 can increase DNMT1 expression and total DNMT1 activity, suggesting that mediators of chronic inflammation can reprogram genes by altering chromatin components.69
Epigenetic Regulation of Tolerance
Endotoxin tolerance is a state of hyporesponsiveness to LPS (and other inflammatory stimuli) that is induced during conditions of excessive inflammation. Its purpose is to limit inflammation-induced associated pathology.70 Some cell types in specific compartments in the body are naturally tolerant: for instance, intestinal macrophages, although constantly exposed to LPS, make almost no proinflammatory cytokines due to downregulation by stromal cell-derived factors.71 Hence, LPS-tolerant cells are refractory to activation by TNF-α and IL-1β, and the available evidence suggests that this is, at least in part, mediated by epigenetic mechanisms. Studies of endotoxin-tolerant human monocytic cell lines have shown that Rel B directly interacts with the methylase G9a, impairs NF-κB p65 binding, and causes silencing of the IL-1β gene.72, 73 Silencing of TNF-α occurs by sustained binding of heterochromatin protein 1 alpha (HP1α) and methylation of H3K9, reduced phosphorylation of H3S10, and disrupted recruitment of p65 NF-κB to the promoter.74 Dynamic repositioning of nucleosomes from permissive to repressive locations has also been reported to be a contributory factor in silencing of TNF-α during sepsis.75 Similarly, association of HMGB1 and HP1 with the promoters of TNF-α and IL-1β genes promotes Rel B- mediated silencing of these cytokines in tolerant cells.76
Chromatin modifications can allow the selective control of distinct sets of genes after repeated exposure of murine macrophages to inflammatory stimuli.77 For example, LPS can induce the expression of both antimicrobial and proinflammatory genes, but restimulation with LPS induces only antimicrobial genes, suggesting that this is due to persistent gene-specific histone modifications that prime cells to respond to subsequent LPS stimulation. In contrast, the antiinflammatory genes do not retain their histone marks after initial LPS stimulation and so these genes are not expressed upon restimulation with LPS. Hence, defining how epigenetic/chromatin states regulate antimicrobial and inflammatory genes will provide important new insights into IBD pathogenesis as well as potential new targets for IBD therapy.
Aberrant epigenetic/chromatin changes have also been reported to mediate the breakdown of tolerance to specific self-antigens that can lead to a variety of autoimmune diseases such as systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA) (see below).
Epigenetic Regulation of T-cell Lineage Commitment
Recent advances have highlighted the importance of epigenetic mechanisms in determining the gene expression pattern and specification of T-cell lineage commitment.78 The proper differentiation and development of naïve T lymphocytes into effector T cells (Th1, Th2, Th17, or Treg) regulates immune responses by producing lineage-specific effector cytokines. The dysregulation of these T-cell subtypes can result in a variety of pathological conditions including IBD.1 The specification of T-cell subsets is controlled by epigenetic factors, chromatin modifications, and networks of lineage-specifying transcription factors. An example of this is the Th1/Th2 differentiation process that depends on chromatin modifiers such as histone methylation, histone acetylation, and DNA methylation at conserved noncoding regions in close proximity to specific cytokine loci. Th2 differentiation is dependent on epigenetic silencing of the Th1 Ifng cytokine locus, and differentiation of Th1 cells is determined by silencing of the Th2 cytokine locus that comprises the Il4, Il13, Rad5, and Il5 genes.79
Epigenetic regulation has also proven essential for Th17 cells and Tregs, even though extensive investigations are still needed to elucidate the role of epigenetic reprogramming and histone-modifying factors in the generation of these two T-cell subtypes. IL-17A and IL-17F are typical cytokines representative of differentiated Th17 cells in vivo and in vitro. Histone H3 acetylation and H3K4me3 at the Il17/Il17f promoter and intronic regions have been described during Th17 differentiation.80 The Forkhead box transcription factor Foxp3 is the major transcription factor that determines the fate and identity of Treg and its enhanced expression is required for regulatory effector function level.81, 82 Recent work indicates that both induction and stabilization of Foxp3 expression is under the control of DNA methylation at the promoter level.82