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Innate lymphoid cells (ILCs) are well-characterized immune cells that play key roles in host defense and tissue homeostasis. Yet, how the three-dimensional (3D) genome organization underlies the development and functions of ILCs is unknown. Herein, we carried out an integrative analysis of the 3D genome structure, chromatin accessibility and gene expression in mature ILCs. Our results revealed that the local 3D configuration of the genome is rewired specifically at loci associated with ILC biology to promote their development and functional differentiation. Importantly, we demonstrated that the ontogenesis of ILC2s and the progression of allergic airway inflammation are determined by a unique local 3D configuration of the region containing the ILC-lineage-defining factor Id2, which is characterized by multiple interactions between the Id2 promoter and distal regulatory elements bound by the transcription factors GATA-3 and RORα, unveiling the mechanism whereby the Id2 expression is specifically controlled in group 2 ILCs.

Exhausted CD8 T cells (TEX) are a distinct state of T cell differentiation associated with failure to clear chronic viruses and cancer. Immunotherapies such as PD-1 blockade can reinvigorate TEX cells, but reinvigoration is not durable. A major unanswered question is whether TEX cells differentiate into functional durable memory T cells (TMEM) upon antigen clearance. Here, using a mouse model, we found that upon eliminating chronic antigenic stimulation, TEX cells partially (re)acquire phenotypic and transcriptional features of TMEM cells. These ‘recovering’ TEX cells originated from the T cell factor (TCF-1+) TEX progenitor subset. Nevertheless, the recall capacity of these recovering TEX cells remained compromised as compared to TMEM cells. Chromatin-accessibility profiling revealed a failure to recover core memory epigenetic circuits and maintenance of a largely exhausted open chromatin landscape. Thus, despite some phenotypic and transcriptional recovery upon antigen clearance, exhaustion leaves durable epigenetic scars constraining future immune responses. These results support epigenetic remodeling interventions for TEX cell–targeted immunotherapies. Wherry and colleagues examine whether exhausted T cells (TEX) can differentiate into functional memory T cells (TMEM) when chronic antigen is withdrawn. Using the chronic LCMV infection mouse model, they show that ‘recovering’ TEX cells (REC-TEX) only partially recover immunophenotypic and functional characteristics of TMEM cells. The epigenomic status of REC-TEX cells more closely resembles that of TEX cells, and, upon rechallenge, the REC-TEX cells were still compromised in their ability to respond to virus.

There is paucity of information about DNA methylation dynamics in immune cells. Roy et al. mapped the DNA methylation status of several thousand differentially methylated CpGs in human immune cells. They reported that the extent of cell type-specific hypermethylation is intriguingly most prevalent in adaptive immune cells rather than innate cells.

An extraordinary degree of condensation is required to fit the eukaryotic genome inside the nucleus. This compaction is attained by first coiling the DNA around structures called nucleosomes. Mammalian genomes are further folded into sophisticated three-dimensional (3D) configurations, enabling the genetic code to dictate a diverse range of cell fates. Recent advances in molecular and computational technologies have enabled the query of higher-order chromatin architecture at an unprecedented resolution and scale. In T lymphocytes, similar to other developmental programs, the hierarchical genome organization is shaped by a highly coordinated division of labor among different classes of sequence-specific transcription factors. In this review, we will summarize the general principles of 1D and 3D genome organization, introduce the common experimental and computational techniques to measure the multilayer chromatin organization, and discuss the pervasive role of transcription factors on chromatin organization in T lymphocytes.

Summary Understanding the mechanisms that establish regulatory T (Treg) cell identity is central to understanding Treg cell function. van der Veeken et al. now show that the lineage-determining transcription factor Foxp3 establishes Treg-cell-specific chromatin architecture indirectly, mostly by decreasing the expression of other transcriptional regulators, including TCF1.

The high mobility group (HMG) transcription factor TCF-1 is essential for early T cell development. Although in vitro biochemical assays suggest that HMG proteins can serve as architectural elements in the assembly of higher-order nuclear organization, the contribution of TCF-1 on the control of three-dimensional (3D) genome structures during T cell development remains unknown. Here, we investigated the role of TCF-1 in 3D genome reconfiguration. Using gain and loss of function experiments, we discovered that the co-occupancy of TCF-1 and the architectural protein CTCF altered the structure of topologically associating domains in T cell progenitors, leading to interactions between previously insulated regulatory elements and target genes at late stages of T cell development. The TCF-1 dependent gain in long-range interactions was linked to deposition of active enhancer mark H3K27ac and recruitment of the cohesin-loading factor NIPBL at active enhancers. These data indicate that TCF-1 has a role in controlling the global genome organization during T cell development.

TCF-1 is a key transcription factor in progenitor exhausted CD8 T cells (Tex). Moreover, this Tex cell subset mediates responses to PD-1 checkpoint pathway blockade. However, the role of the transcription factor TCF-1 in early fate decisions and initial generation of Tex cells is unclear. Single cell RNA-sequencing (scRNA-seq) and lineage tracing identified a TCF-1(+)Ly108(+)PD-1(+) CD8 T cell population early during chronic infection that seeds development of mature Tex cells. TCF-1 mediated the bifurcation between divergent fates, repressing development of terminal KLRG1(Hi) effectors while fostering KLRG1(Lo) Tex precursor cells, and PD-1 stabilized this TCF-1(+) Tex precursor cell pool. TCF-1 mediated a T-bet to Eomes transcription factor transition in Tex precursors by promoting Eomes expression and drove c-Myb expression that controlled Bcl-2 and survival. These data define a role for TCF-1 in early fate bifurcation driving Tex precursor cells, and also identify PD-1 as a protector of this early TCF-1 subset.

Summary Genetics is a major determinant of susceptibility to autoimmune disorders. Here, we examined whether genome organization provides resilience or susceptibility to sequence variations, and how this would contribute to the molecular etiology of an autoimmune disease. We generated high-resolution maps of linear and 3D genome organization in thymocytes of NOD mice, a model of type 1 diabetes (T1D), and the diabetes-resistant C57BL/6 mice. Multi-enhancer interactions formed at genomic regions harboring genes with prominent roles in T cell development in both strains. However, diabetes risk-conferring loci coalesced enhancers and promoters in NOD, but not C57BL/6 thymocytes. 3D genome mapping of NODxC57BL/6 F1 thymocytes revealed that genomic misfolding in NOD mice is mediated in cis. Moreover, immune cells infiltrating the pancreas of humans with T1D exhibited increased expression of genes located on misfolded loci in mice. Thus, genetic variation leads to altered 3D chromatin architecture and associated changes in gene expression that may underlie autoimmune pathology.

Summary The generation of high-affinity neutralizing antibodies, the objective of most vaccine strategies, occurs in B cells within germinal centers (GCs) and requires rate-limiting “help” from follicular helper CD4+ T (Tfh) cells. Although Tfh differentiation is an attribute of MHC II-restricted CD4+ T cells, the transcription factors driving Tfh differentiation, notably Bcl6, are not restricted to CD4+ T cells. Here, we identified a requirement for the CD4+-specific transcription factor Thpok during Tfh cell differentiation, GC formation, and antibody maturation. Thpok promoted Bcl6 expression and bound to a Thpok-responsive region in the first intron of Bcl6. Thpok also promoted the expression of Bcl6-independent genes, including the transcription factor Maf, which cooperated with Bcl6 to mediate the effect of Thpok on Tfh cell differentiation. Our findings identify a transcriptional program that links the CD4+ lineage with Tfh differentiation, a limiting factor for efficient B cell responses, and suggest avenues to optimize vaccine generation.

Interleukin-23 (IL-23) is a pro-inflammatory cytokine required for the pathogenicity of T helper 17 (Th17) cells but the molecular mechanisms governing this process remain unclear. We identified the transcription factor Blimp-1 (Prdm1) as a key IL-23-induced factor that drove the inflammatory function of Th17 cells. In contrast to thymic deletion of Blimp-1, which causes T cell development defects and spontaneous autoimmunity, peripheral deletion of this transcription factor resulted in reduced Th17 activation and reduced severity of autoimmune encephalomyelitis. Furthermore, genome-wide occupancy and overexpression studies in Th17 cells revealed that Blimp-1 co-localized with transcription factors RORγt, STAT-3, and p300 at the Il23r, Il17a/f, and Csf2 cytokine loci to enhance their expression. Blimp-1 also directly bound to and repressed cytokine loci Il2 and Bcl6. Taken together, our results demonstrate that Blimp-1 is an essential transcription factor downstream of IL-23 that acts in concert with RORγt to activate the Th17 inflammatory program.

A long-standing question in immunology has been to understand how transcription factors (TF) determine cell-type-specific transcription programs. Traditionally, investigating TFs in immune cells has been limited to measuring the effect of a single TF on a limited number of gene targets. The advent of next generation sequencing methods makes it possible to measure the effects of multiple transcription factors on a genome-wide scale. This holistic approach gives us a better understanding of the influence that multiple TFs have on cell-specific programs. In this issue of European Journal of Immunology, Fang et al. [Eur. J. Immunol. 2015. 45: 3150-3157] show that genomic regions cooccupied with two or more TFs show increased epigenetic histone marks of active enhancers, which correspond to increased transcriptional activity.

CD4+ T follicular helper cells (TFH) are critical for the formation and function of B cell responses to infection or immunization, but also play an important role in autoimmunity. The factors that contribute to the differentiation of this helper cell subset are incompletely understood, although several cytokines including IL-6, IL-21, and IL-12 can promote TFH cell formation. Yet, none of these factors, nor their downstream cognate STATs, have emerged as nonredundant, essential drivers of TFH cells. This suggests a model in which multiple factors can contribute to the phenotypic characteristics of TFH cells. Because type I IFNs are often generated in immune responses, we set out to investigate whether these factors are relevant to TFH cell differentiation. Type I IFNs promote Th1 responses, thus one possibility was these factors antagonized TFH-expressed genes. However, we show that type I IFNs (IFN-α/β) induced B cell lymphoma 6 (Bcl6) expression, the master regulator transcription factor for TFH cells, and CXCR5 and programmed cell death-1 (encoded by Pdcd1), key surface molecules expressed by TFH cells. In contrast, type I IFNs failed to induce IL-21, the signature cytokine for TFH cells. The induction of Bcl6 was regulated directly by STAT1, which bound to the Bcl6, Cxcr5, and Pdcd1 loci. These data suggest that type I IFNs (IFN-α/β) and STAT1 can contribute to some features of TFH cells but are inadequate in inducing complete programming of this subset.

Summary Interleukin-27 (IL-27) is a key immunosuppressive cytokine that counters T helper 17 (Th17) cell-mediated pathology. To identify mechanisms by which IL-27 might exert its immunosuppressive effect, we analyzed genes in T cells rapidly induced by IL-27. We found that IL-27 priming of naive T cells upregulated expression of programmed death ligand 1 (PD-L1) in a signal transducer and activator of transcription 1 (STAT1)-dependent manner. When cocultured with naive CD4 + T cells, IL-27-primed T cells inhibited the differentiation of Th17 cells in trans through a PD-1-PD-L1 interaction. In vivo, coadministration of naive TCR transgenic T cells (2D2 T cells) with IL-27-primed T cells expressing PD-L1 inhibited the development of Th17 cells and protected from severe autoimmune encephalomyelitis. Thus, these data identify a suppressive activity of IL-27, by which CD4 + T cells can restrict differentiation of Th17 cells in trans .

T helper cell differentiation occurs in the context of the extracellular cytokine milieu evoked by diverse microbes and other pathogenic stimuli along with T cell receptor stimulation. The culmination of these signals results in specification of T helper lineages, which occurs through the combinatorial action of multiple transcription factors that establish distinctive transcriptomes. In this manner, inducible, but constitutively active, master regulators work in conjunction with factors such as the signal transducer and activator of transcriptions (STATs) that sense the extracellular environment. The acquisition of a distinctive transcriptome also depends on chromatin modifications that impact key cis elements as well as the changes in global genomic organization. Thus, signal transduction and epigenetics are linked in these processes of differentiation. In this review, recent advances in understanding T helper lineage specification and deciphering the action of transcription factors are summarized with emphasis on comprehensive views of the dynamic T cell epigenome.

The transcription factor T-bet drives the differentiation of NKp46-expressing IL-22–producing innate lymphoid cells, Interleukin (IL)-22–producing innate lymphoid cells (ILCs; ILC22) comprise a heterogeneous population of cells that are dependent on the transcription factor retinoid-related orphan γt (RORγt) and are critical for barrier function of the intestinal mucosa. A distinct ILC22 subset expresses the natural cytotoxicity receptor NKp46 (NKp46+ ILC22); however, the factors that contribute to the generation of this population versus other subsets are largely unknown. Herein, we show that T-bet (encoded by Tbx21) was highly expressed in NKp46+ ILC22, a feature shared by all NKp46+ cells present in the intestine but not by other IL-22–producing populations. Accordingly, the absence of T-bet resulted in loss of NKp46+ ILC22 in the intestinal lamina propria. The residual NKp46+ ILC22 present in Tbx21−/− mice showed a marked reduction of Rorγt expression and impairment in IL-22 production. Generation and functions of gut NK1.1+ cells were also altered. Bone marrow chimera experiments revealed a cell-intrinsic requirement for T-bet in these subsets and competitive reconstitution experiments revealed roles for T-bet in multiple ILC subsets. Thus, T-bet has a general importance for ILC in the gut and plays a selective and critical role in the generation of NKp46+ ILC22.

CD4(+) T cells have critical roles in orchestrating immune responses to diverse microbial pathogens. This is accomplished through the differentiation of CD4(+) T helper cells to specialized subsets in response to microbial pathogens, which evoke a distinct cytokine milieu. Signal transducer and activator of transcription family transcription factors sense these cytokines and they in turn regulate expression of lineage-defining master regulators that programme selective gene expression, resulting in distinctive phenotypes. However, phenotype and restricted gene expression are determined not only by the action of transcription factors; chromatin accessibility is required for these factors to exert their effect. Technical advances have greatly expanded our understanding of transcription factor action and dynamic changes in the epigenome that accompany cellular differentiation. In this review, we will discuss recent progress in the understanding of how cytokines influence gene expression and epigenetic modifications, and the impact of these findings on our views of helper cell lineage commitment and plasticity.

Interleukin 2 (IL-2), a cytokine linked to human autoimmune disease, limits IL-17 production. Here we found that deletion of the gene encoding the transcription factor STAT3 in T cells abrogated IL-17 production and attenuated autoimmunity associated with IL-2 deficiency. Whereas STAT3 induced IL-17 and the transcription factor RORγt and inhibited the transcription factor Foxp3, IL-2 inhibited IL-17 independently of Foxp3 and RORγt. STAT3 and STAT5 bound to multiple common sites across the locus encoding IL-17. The induction of STAT5 binding by IL-2 was associated with less binding of STAT3 at these sites and the inhibition of associated active epigenetic marks. 'Titration' of the relative activation of STAT3 and STAT5 modulated the specification of cells to the IL-17-producing helper T cell (T(H)17 cell) subset. Thus, the balance rather than the absolute magnitude of these signals determined the propensity of cells to make a key inflammatory cytokine.

A characteristic feature of asthma is the aberrant accumulation, differentiation or function of memory CD4+ T cells that produce type 2 cytokines (TH2 cells). By mapping genome-wide histone modification profiles for subsets of T cells isolated from peripheral blood of healthy and asthmatic individuals, we identified enhancers with known and potential roles in the normal differentiation of human TH1 cells and TH2 cells. We discovered disease-specific enhancers in T cells that differ between healthy and asthmatic individuals. Enhancers that gained the histone H3 Lys4 dimethyl (H3K4me2) mark during TH2 cell development showed the highest enrichment for asthma-associated single nucleotide polymorphisms (SNPs), which supported a pathogenic role for TH2 cells in asthma. In silico analysis of cell-specific enhancers revealed transcription factors, microRNAs and genes potentially linked to human TH2 cell differentiation. Our results establish the feasibility and utility of enhancer profiling in well-defined populations of specialized cell types involved in disease pathogenesis.

Super-enhancers (SEs) are regions of the genome consisting of clusters of regulatory elements bound with very high amounts of transcription factors, and this architecture appears to be the hallmark of genes and noncoding RNAs linked with cell identity. Recent studies have identified SEs in CD4(+) T cells and have further linked these regions to single nucleotide polymorphisms (SNPs) associated with immune-mediated disorders, pointing to an important role for these structures in the T cell differentiation and function. Here we review the features that define SEs, and discuss their function within the broader understanding of the mechanisms that define immune cell identity and function. We propose that SEs present crucial regulatory hubs, coordinating intrinsic and extrinsic differentiation signals, and argue that delineating these regions will provide important insight into the factors and mechanisms that define immune cell identity.

International audience; T cell development is orchestrated by transcription factors that regulate the expression of genes initially buried within inaccessible chromatin, but the transcription factors that establish the regulatory landscape of the T cell lineage remain unknown. Profiling chromatin accessibility at eight stages of T cell development revealed the selective enrichment of TCF-1 at genomic regions that became accessible at the earliest stages of development. TCF-1 was further required for the accessibility of these regulatory elements and at the single-cell level, it dictated a coordinate opening of chromatin in T cells. TCF-1 expression in fibroblasts generated de novo chromatin accessibility even at chromatin regions with repressive marks, inducing the expression of T cell-restricted genes. These results indicate that a mechanism by which TCF-1 controls T cell fate is through its widespread ability to target silent chromatin and establish the epigenetic identity of T cells.

SummaryInterleukin-6 (IL-6) and IL-27 signal through a shared receptor subunit and employ the same downstream STAT transcription proteins, but yet are ascribed unique and overlapping functions. To evaluate the specificity and redundancy for these cytokines, we quantified their global transcriptomic changes and determined the relative contributions of STAT1 and STAT3 using genetic models and chromatin immunoprecipitation-sequencing (ChIP-seq) approaches. We found an extensive overlap of the transcriptomes induced by IL-6 and IL-27 and few examples in which the cytokines acted in opposition. Using STAT-deficient cells and T cells from patients with gain-of-function STAT1 mutations, we demonstrated that STAT3 is responsible for the overall transcriptional output driven by both cytokines, whereas STAT1 is the principal driver of specificity. STAT1 cannot compensate in the absence of STAT3 and, in fact, much of STAT1 binding to chromatin is STAT3 dependent. Thus, STAT1 shapes the specific cytokine signature superimposed upon STAT3’s action.

The discovery of the specification of CD4(+) helper T cells to discrete effector 'lineages' represented a watershed event in conceptualizing mechanisms of host defense and immunoregulation. However, our appreciation for the actual complexity of helper T-cell subsets continues unabated. Just as the Sami language of Scandinavia has 1000 different words for reindeer, immunologists recognize the range of fates available for a CD4(+) T cell is numerous and may be underestimated. Added to the crowded scene for helper T-cell subsets is the continuously growing family of innate lymphoid cells (ILCs), endowed with common effector responses and the previously defined 'master regulators' for CD4(+) helper T-cell subsets are also shared by ILC subsets. Within the context of this extraordinary complexity are concomitant advances in the understanding of transcriptomes and epigenomes. So what do terms like 'lineage commitment' and helper T-cell 'specification' mean in the early 21st century? How do we put all of this together in a coherent conceptual framework? It would be arrogant to assume that we have a sophisticated enough understanding to seriously answer these questions. Instead, we review the current status of the flexibility of helper T-cell responses in relation to their genetic regulatory networks and epigenetic landscapes. Recent data have provided major surprises as to what master regulators can or cannot do, how they interact with other transcription factors and impact global genome-wide changes, and how all these factors come together to influence helper cell function.

CD4(+) T cells are critical for the elimination of an immense array of microbial pathogens. Among the ways they accomplish this task is to generate progeny with specialized, characteristic patterns of gene expression. From this perspective, helper cells can be viewed as pluripotent precursors that adopt distinct cell fates. Although there are aspects of helper cell differentiation that can be modeled as a classic cell fate commitment, CD4(+) T cells also maintain considerable flexibility in their transcriptional program. This makes sense in terms of host defense, but raises the question of how these remarkable cells balance both these requirements, a high degree of specific gene expression and the capacity for plasticity. In this review, we discuss recent advances in our understanding of CD4(+) T-cell specification, focusing on how genomic perspectives have influenced our views of these processes. The relative contributions of sensors of the cytokine milieu, especially the signal transducer and activator of transcription family transcription factors, 'master regulators', and other transcription factors are considered as they relate to the helper cell transcriptome and epigenome.

Naive T cells differentiate and become distinct subsets in response to changes in the cytokine milieu. Such specialization arises through a complex and dynamic utilization of cis-regulatory enhancer elements. In this brief essay, we review recent findings on the relative contributions of sensors of the cytokine milieu, especially the signal transducer and activator of transcription family transcription factors, 'master regulators', and other transcription factors in the enhancer architecture of T cells. These findings provide new insights into how signal transduction impinges upon the genome.

CD4 helper T cells are critical for proper immune cell homeostasis and host defense but are also major contributors to immune and inflammatory disease. Arising from a simple biphasic model of differentiation (ie, TH1 and TH2 cells). A bewildering number of fates seem possible for helper T cells. To what extent different helper cell subsets maintain their characteristic gene expression profiles or exhibit functional plasticity is a hotly debated topic. In this review we will discuss how the expression of "signature cytokines" and "master regulator" transcription factors do not neatly conform to a simple helper T-cell paradigm. Although this might seem confusing, the good news is that the newly recognized complexity fits better with our understanding of immunopathogenesis. Finally, we will discuss factors, including epigenetic regulation and metabolic alterations, that contribute to helper cell specificity and plasticity.

Through their functional diversification, distinct lineages of CD4 + T cells play key roles in either driving or constraining immune-mediated pathology. Transcription factors are critical in the generation of cellular diversity, and negative regulators antagonistic to alternate fates often act in conjunction with positive regulators to stabilize lineage commitment. Genetic polymorphisms within a single locus encoding the transcription factor BACH2 are associated with numerous autoimmune and allergic diseases including asthma, Crohn’s disease, coeliac disease, vitiligo, multiple sclerosis and type 1 diabetes. While these associations point to a shared mechanism underlying susceptibility to diverse immune-mediated diseases, a function for BACH2in the maintenance of immune homeostasis had not been established. We have found that BACH2 plays a broad role in maintaining immune homeostasis, by stabilizing T reg -mediated immunoregulatory capacity while repressing the differentiation programmes of multiple effector lineages in CD4 + T cells. BACH2 was required for efficient formation of regulatory ( T reg ) cells and consequently for suppression of lethal inflammation in a manner that was T reg cell dependent. Assessment of the genome-wide function of BACH2, however, revealed that it represses genes associated with effector cell differentiation. Consequently, its absence during T reg polarization resulted in inappropriate diversion to effector lineages and aberrant expression of effector cytokines. Remarkably, blockade of effector cytokines IFN-γ and IL-4 during iTreg cell polarization could restore FoxP3 expression amongst BACH2-deficient cells. In addition, BACH2constrained full effector differentiation and production of IFN-γ, IL-13 and IL17-A in Th1, Th2 and Th17 cell lineages, respectively. These findings identify BACH2 as a key regulator of CD4 + T-cell differentiation that prevents inflammatory disease by controlling the balance between tolerance and immunity.

Interleukin (IL)-27 is a key immunosuppressive cytokine that counters Th17-mediated pathology. To identify mechanisms by which IL-27 might exert its immunosuppressive effect, we analyzed genes in naïve CD4+ T cells rapidly induced by IL-27 stimulation for 3 hours. We discovered that IL-27 priming of naïve T cells upregulated expression of the ligand of PD-1 (PD-L1) in a STAT1-dependent manner. To assess the relevance of IL-27 in controlling PD-L1 expression in vivo, we employed a T. gondii infection model in which IL-27 is important for controlling immunopathology. We found that expression of PD-L1 on CD4+ T cells was significantly lower in EBI3-/- mice. When added to naïve CD4+ T cells, IL-27-primed T cells inhibited the differentiation of Th17 cells in trans in vitro. In vivo, co-administration of IL-27-primed T cells expressing PD-L1 inhibited the development of Th17 cells and protected from severe autoimmune encephalomyelitis. Moreover this inhibitory effect of IL-27 in vivo was largely reversed by elimination of PD-L1 on T cells. Thus, these data identify a novel suppressive activity of IL-27, by which CD4+ T cells can restrict differentiation of Th17 cells in trans. It is conceivable that the strategy of IL-27-pretreatment of naïve T cells ex vivo with subsequent adoptive transfer might be used therapeutically to attenuate autoimmune disease.

SummarySTAT3, an essential transcription factor with pleiotropic functions, plays critical roles in the pathogenesis of autoimmunity. Despite recent data linking STAT3 with inflammatory bowel disease, exactly how it contributes to chronic intestinal inflammation is not known. Using a T cell transfer model of colitis, we found that STAT3 expression in T cells was essential for the induction of both colitis and systemic inflammation. STAT3 was critical in modulating the balance of T helper 17 (Th17) and regulatory T (Treg) cells, as well as in promoting CD4+ T cell proliferation. We used chromatin immunoprecipitation and massive parallel sequencing (ChIP-Seq) to define the genome-wide targets of STAT3 in CD4+ T cells. We found that STAT3 bound to multiple genes involved in Th17 cell differentiation, cell activation, proliferation, and survival, regulating both expression and epigenetic modifications. Thus, STAT3 orchestrates multiple critical aspects of T cell function in inflammation and homeostasis.

SummarySignal transducer and activator of transcription 4 (STAT4) and STAT6 are key factors in the specification of helper T cells; however, their direct roles in driving differentiation are not well understood. Using chromatin immunoprecipitation and massive parallel sequencing, we quantitated the full complement of STAT-bound genes, concurrently assessing global STAT-dependent epigenetic modifications and gene transcription by using cells from cognate STAT-deficient mice. STAT4 and STAT6 each bound over 4000 genes with distinct binding motifs. Both played critical roles in maintaining chromatin configuration and transcription of a core subset of genes through the combination of different epigenetic patterns. Globally, STAT4 had a more dominant role in promoting active epigenetic marks, whereas STAT6 had a more prominent role in antagonizing repressive marks. Clusters of genes negatively regulated by STATs were also identified, highlighting previously unappreciated repressive roles of STATs. Therefore, STAT4 and STAT6 play wide regulatory roles in T helper cell specification.