Transcriptional Regulation of Immune Cell Development, Activation and Functions

National Institute Of Allergy And Infectious Diseases

Investigators

Linked publications, trials & patents

Abstract

CD4+ T lymphocytes play a central role in orchestrating adaptive immune responses. After activation through their T cell receptor (TCR) in a particular cytokine milieu, naive CD4+ T cells differentiate into distinct T helper (Th) lineages, including Th1, Th2 and Th17 cells that produce interferon (IFN)-g, interleukin (IL)-4 and IL-17, respectively, as their signature effector cytokines. Through the production of these distinct effector cytokines, specific Th subsets mediate crucial functions during different types of protective immune responses to various microorganisms. Th1 cells are important for host defense against intracellular bacteria and viruses; Th2 cells for expelling extracellular parasites such as helminths; and Th17 cells for controlling extracellular bacteria and fungi. Inappropriate Th responses to pathogens may lead to chronic infection and/or tissue damage to the host, whereas aberrant Th cell differentiation may result in many inflammatory allergic or autoimmune diseases including asthma, inflammatory bowel diseases (IBD), rheumatoid arthritis (RA) and multiple sclerosis (MS). Innate lymphoid cells (ILCs), which lack expression of antigen receptors, require signaling through the IL-2 receptor (IL-2R) common gamma chain and IL-7Ra, for their development, maturation or homeostasis. Distinct ILC subsets mirror different Th cell subsets in their cytokine production. Therefore, ILCs are classified into type 1 innate lymphoid cells (ILC1s) that produce IFNg, type 2 innate lymphoid cells (ILC2s) that produce IL-5 and IL-13, and type 3 innate lymphoid cells (ILC3s) that produce IL-17 and IL-22. Although some ILCs, such as lymphoid tissue inducer (LTi) cells, are specifically critical for lymphoid organogenesis, most ILCs, like Th cells, are important for protective immune responses to infections and contribute to the pathogenesis of many inflammatory diseases. The activation, differentiation and expansion of Th cells are tightly regulated by specific transcription factors that are induced and/or activated by a combination of cytokines and TCR-mediate signaling. Our major research goal is to better understand the transcriptional regulatory networks and mechanisms that control differentiation processes leading to the distinct Th and ILC lineages. We have chosen to focus on the master regulators (also known as lineage-determining transcription factors, LDTFs) including T-bet, GATA3, and RORgt (for type 1, type 2 and type 3 lymphocytes, respectively), because we hypothesize that they are the major nodes in these networks. By comparing the regulation and actions of T-bet, GATA3 and RORgt in distinct Th and ILC lineages, we aim to identify new components and/or connections of these complex networks controlling Th cell differentiation and ILC development. Comparing gene regulation at the transcriptomic and epigenomic level between these two cell types will allow us to identify the core elements that determine their shared functionality and unique molecules/pathways that control their specialized functions. During the past fiscal year, we have reported a representative study focusing on the regulation of T-bet expression in type 1 lymphocytes to elucidate the similarities and differences in the induction of LDTFs in innate and adaptive lymphocytes (Immunity 55:639-655, 2022). In this manuscript, we showed that Th1 cells and NK cells displayed distinct epigenomes at the Tbx21 locus, which encodes T-bet, the LDTF for type 1 lymphocytes. The initial induction of T-bet in NK precursors was dependent on the NK-specific DNase I hypersensitive site Tbx21-CNS-3 and the expression of IL-18 receptor; IL-18 induced T-bet expression through RUNX3, which bound to Tbx21-CNS-3. By contrast, STAT-binding motifs within Tbx21-CNS-12 were critical for IL-12-induced T-bet expression during Th1 cell differentiation both in vitro and in vivo. Thus, type 1 innate and adaptive lymphocytes utilize distinct enhancer elements for their development and differentiation. In previous years, we have used T-bet-ZsGreen (or AmCyan)-RORgt-E2Crimson-Foxp3-RFP triple reporter mice to investigate the dynamic expression of T-bet and RORgt in EAE, a mouse model for MS. We found that RORgt-expressing cells can generate T-bet/RORgt dual-expressors as well as cells only expressing T-bet after adoptive transfer. RNA-Seq analysis of these subsets harvested from the spinal cord of the disease mice indicates that T-bet and RORgt each regulate (induce or inhibit) distinct and shared sets of genes, however, we did not observe any synergistic effect between T-bet and RORgt in gene regulation. Nevertheless, mice with conditional deletion of Rorc gene by T-bet-driven Cre were completely resistant to the disease induction. By generating other novel mouse models, we also demonstrated that a transition from expressing only RORgt to expressing both RORgt and T-bet, and finally to expressing only T-bet is a natural process in autoimmunity. In the past fiscal year, we have further demonstrated that the cells expressing only RORgt contain a stem-like population and have a better capacity in populating T-bet-expressing effector cells than T-bet-expressing cells themselves upon transfer. Thus, all three cell subsets (RORgt and T-bet single and dual expressors) play unique roles in the disease induction in EAE. We are now also testing this in IBD. We have previously reported that GATA3 serves as a switch in determining the development of LTi cells versus other ILC lineages (Immunity. 52: 83-95, 2020). While GATA3 is absolutely required for the generation of PLZF-expressing non-LTi progenitors, which express high level of GATA3, it is not necessary for the generation of RORt-expressing LTi progenitors consistent with low levels of GATA3 expression in these progenitors. In the past fiscal year, we further found that the transcription factor TCF-1, just as GATA3, was indispensable for the development of non-LTi ILC subsets. While LTi cells were still present in the TCF-1-deficient mice, the organogenesis of Peyers patches (PPs) but not of lymph nodes was impaired in these mice. LTi cells from different tissues had distinct gene expression patterns and TCF-1 regulated the expression of lymphotoxin specifically in PP LTi cells. Mechanistically, TCF-1 indirectly regulated Lta through promoting the expression of GATA3. Thus, the TCF-1-GATA3 axis, which plays an important role during T cell development, also critically regulates the development of non-LTi cells and tissue-specific functions of LTi cells. We continued our investigation on the relative importance and crosstalk between ILC2s and Th2 cells during type 2 immune responses by utilizing unique mouse strains that are specifically deficient in either ILC2s or Th2 cells. In previous years, we have found that IL-33-mediated ILC2 activation promoted Th2 cell differentiation in the papain model, however, Th2 cell differentiation was completely independent of ILC2s in the OVA/alum immunization model. On the other hand, Th2 cells induced the expression of IL-25, IL-33, and TSLP, which played a redundant role in promoting ILC2 expansion. During a helminth infection, ILC2s and Th2 cells participated in different phases of host defense and collaborated in promoting each others expansion. During the past fiscal year, we have further demonstrated that IL-4 produced by Th2 cells played a critical role in inducing type 2 alarmin expression. While IL-25 and IL-33 played a redundant role in activating ILC2s, TSLP was important for ILC2-independent Th2 cell response induced by OVA/alum immunization. In addition, pre-activation of ILC2s by either IL-33 or IL-25 in the ILC2-independent OVA model further promoted Th2 cell differentiation. Thus, alarmin- and IL-4-mediated crosstalk between ILC2s and Th2 cells varies in different type 2 immune responses in mice