Regulated transcription controls the diversity, developmental pathways and spatial organization of the hundreds of cell types that make up a mammal. Using single-molecule cDNA sequencing, we mapped transcription start sites (TSSs) and their usage in human and mouse primary cells, cell lines and tissues to produce a comprehensive overview of mammalian gene expression across the human body. We find that few genes are truly ‘housekeeping’, whereas many mammalian promoters are composite entities composed of several closely separated TSSs, with independent cell-type-specific expression profiles. TSSs specific to different cell types evolve at different rates, whereas promoters of broadly expressed genes are the most conserved. Promoter-based expression analysis reveals key transcription factors defining cell states and links them to binding-site motifs. The functions of identified novel transcripts can be predicted by coexpression and sample ontology enrichment analyses. The functional annotation of the mammalian genome 5 (FANTOM5) project provides comprehensive expression profiles and functional annotation of mammalian cell-type-specific transcriptomes with wide applications in biomedical research.

The transcription factor Foxp3 is essential for the development of regulatory T (Treg) cells, yet its expression is insufficient for establishing the Treg cell lineage. Here we showed that Treg cell development was achieved by the combination of two independent processes, i.e., the expression of Foxp3 and the establishment of Treg cell-specific CpG hypomethylation pattern. Both events were induced by T cell receptor stimulation. The Treg cell-type CpG hypomethylation began in the thymus and continued to proceed in the periphery and could be fully established without Foxp3. The hypomethylation was required for Foxp3(+) T cells to acquire Treg cell-type gene expression, lineage stability, and full suppressive activity. Thus, those T cells in which the two events have concurrently occurred are developmentally set into the Treg cell lineage. This model explains how Treg cell fate and plasticity is controlled and can be exploited to generate functionally stable Treg cells.

CD4(+) T cells that express the forkhead box P3 (FOXP3) transcription factor function as regulatory T (Treg) cells and hinder effective immune responses against cancer cells. Abundant Treg cell infiltration into tumors is associated with poor clinical outcomes in various types of cancers. However, the role of Treg cells is controversial in colorectal cancers (CRCs), in which FOXP3(+) T cell infiltration indicated better prognosis in some studies. Here we show that CRCs, which are commonly infiltrated by suppression-competent FOXP3(hi) Treg cells, can be classified into two types by the degree of additional infiltration of FOXP3(lo) nonsuppressive T cells. The latter, which are distinguished from FOXP3(+) Treg cells by non-expression of the naive T cell marker CD45RA and instability of FOXP3, secreted inflammatory cytokines. Indeed, CRCs with abundant infiltration of FOXP3(lo) T cells showed significantly better prognosis than those with predominantly FOXP3(hi) Treg cell infiltration. Development of such inflammatory FOXP3(lo) non-Treg cells may depend on secretion of interleukin (IL)-12 and transforming growth factor (TGF)-β by tissues and their presence was correlated with tumor invasion by intestinal bacteria, especially Fusobacterium nucleatum. Thus, functionally distinct subpopulations of tumor-infiltrating FOXP3(+) T cells contribute in opposing ways to determining CRC prognosis. Depletion of FOXP3(hi) Treg cells from tumor tissues, which would augment antitumor immunity, could thus be used as an effective treatment strategy for CRCs and other cancers, whereas strategies that locally increase the population of FOXP3(lo) non-Treg cells could be used to suppress or prevent tumor formation.

Naturally arising CD25+CD4+ regulatory T cells (T(R) cells) are engaged in the maintenance of immunological self-tolerance and immune homeostasis by suppressing aberrant or excessive immune responses, such as autoimmune disease and allergy. T(R) cells specifically express the transcription factor Foxp3, a key regulator of T(R)-cell development and function. Ectopic expression of Foxp3 in conventional T cells is indeed sufficient to confer suppressive activity, repress the production of cytokines such as interleukin-2 (IL-2) and interferon-gamma (IFN-gamma), and upregulate T(R)-cell-associated molecules such as CD25, cytotoxic T-lymphocyte-associated antigen-4, and glucocorticoid-induced TNF-receptor-family-related protein. However, the method by which Foxp3 controls these molecular events has yet to be explained. Here we show that the transcription factor AML1 (acute myeloid leukaemia 1)/Runx1 (Runt-related transcription factor 1), which is crucially required for normal haematopoiesis including thymic T-cell development, activates IL-2 and IFN-gamma gene expression in conventional CD4+ T cells through binding to their respective promoters. In natural T(R) cells, Foxp3 interacts physically with AML1. Several lines of evidence support a model in which the interaction suppresses IL-2 and IFN-gamma production, upregulates T(R)-cell-associated molecules, and exerts suppressive activity. This transcriptional control of T(R)-cell function by an interaction between Foxp3 and AML1 can be exploited to control physiological and pathological T-cell-mediated immune responses.

Regulatory T (Treg) cells are a developmentally and functionally distinct T cell subpopulation that is engaged in sustaining immunological self-tolerance and homeostasis. The transcription factor Foxp3 plays a key role in Treg cell development and function. However, expression of Foxp3 alone is not sufficient for conferring and maintaining Treg cell function and phenotype. Complementing the insufficiency, Treg-cell-specific epigenetic changes are also critical in the process of Treg cell specification, in regulating its potential plasticity, and hence in establishing a stable lineage. Understanding how epigenetic alterations and Foxp3 expression coordinately control Treg-cell-specific gene regulation will enable better control of immune responses by targeting the generation and maintenance of Treg cells.

Naturally occurring CD4+ regulatory T cells (Tregs), which specifically express the transcription factor FoxP3 in the nucleus and CD25 and CTLA-4 on the cell surface, are a functionally distinct T cell subpopulation actively engaged in the maintenance of immunological self-tolerance and homeostasis. Recent studies have facilitated our understanding of the cellular and molecular basis of their generation, function, phenotypic and functional stability, and adaptability. It is under investigation in humans how functional or numerical Treg anomalies, whether genetically determined or environmentally induced, contribute to immunological diseases such as autoimmune diseases. Also being addressed is how Tregs can be targeted to control physiological and pathological immune responses, for example, by depleting them to enhance tumor immunity or by expanding them to treat immunological diseases. This review discusses our current understanding of Treg immunobiology in normal and disease states, with a perspective on the realization of Treg-targeting therapies in the clinic.

Most Foxp3+ regulatory T (Treg) cells develop in the thymus as a functionally mature T cell subpopulation specialized for immune suppression. Their cell fate appears to be determined before Foxp3 expression; yet molecular events that prime Foxp3− Treg precursor cells are largely obscure. We found that Treg cell–specific super-enhancers (Treg-SEs), which were associated with Foxp3 and other Treg cell signature genes, began to be activated in Treg precursor cells. T cell–specific deficiency of the genome organizer Satb1 impaired Treg-SE activation and the subsequent expression of Treg signature genes, causing severe autoimmunity due to Treg cell deficiency. These results suggest that Satb1-dependent Treg-SE activation is crucial for Treg cell lineage specification in the thymus and that its perturbation is causative of autoimmune and other immunological diseases.

Human T-cell leukemia virus type 1 (HTLV-1) is the causal agent of a neoplastic disease of CD4+ T cells, adult T-cell leukemia (ATL), and inflammatory diseases including HTLV-1 associated myelopathy/tropical spastic paraparesis, dermatitis, and inflammatory lung diseases. ATL cells, which constitutively express CD25, resemble CD25+CD4+ regulatory T cells (Treg). Approximately 60% of ATL cases indeed harbor leukemic cells that express FoxP3, a key transcription factor for Treg cells. HTLV-1 encodes an antisense transcript, HTLV-1 bZIP factor (HBZ), which is expressed in all ATL cases. In this study, we show that transgenic expression of HBZ in CD4+ T cells induced T-cell lymphomas and systemic inflammation in mice, resembling diseases observed in HTLV-1 infected individuals. In HBZ-transgenic mice, CD4+Foxp3+ Treg cells and effector/memory CD4+ T cells increased in vivo. As a mechanism of increased Treg cells, HBZ expression directly induced Foxp3 gene transcription in T cells. The increased CD4+Foxp3+ Treg cells in HBZ transgenic mice were functionally impaired while their proliferation was enhanced. HBZ could physically interact with Foxp3 and NFAT, thereby impairing the suppressive function of Treg cells. Thus, the expression of HBZ in CD4+ T cells is a key mechanism of HTLV-1-induced neoplastic and inflammatory diseases.