CD4 T cells play a central role in orchestrating adaptive immune responses. After being activated 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. These cells are indispensable for different types of immunity to various microorganisms. Inappropriate Th responses to pathogens may lead to chronic infection and/or tissue damage to the host. Similarly, unnecessary activation of Th1, Th17 or Th2 cells by harmless environmental- or self-antigens can cause organ-specific autoimmune diseases or allergic inflammatory diseases. There are innate counterparts of Th cells. A class of innate lymphoid cells (ILCs), whose development requires signaling through the IL-2 receptor (IL-2R) common g chain and IL-7Ra, has been recently discovered. Distinct subsets of ILCs are capable of producing similar sets of characteristic effector cytokines as produced by Th cells. Therefore, they 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. Within the ILC3s all of which express RORgt, there are two subsets CCR6+ (mainly lymphoid tissue inducers, LTis) and CCR6- ILC3s - with the latter having the potential to develop into NKp46+ ILC3s that express both RORgt and T-bet. CCR6+ and NKp46+ ILC3s seem to have distinct biological functions and develop from different precursors. Like Th cells, ILCs are important for protective immune responses to infections and are responsible for the pathogenesis of many inflammatory diseases. Some ILCs such as LTis are critical for lymphoid organ development. The activation, differentiation and expansion of Th cells are tightly regulated by specific transcription factors. Among the lineage-specific transcription factors, T-bet, GATA3, RORgt and Foxp3 are deterministic for the differentiation of Th1, Th2, Th17 and Treg cells, respectively. These transcription factors have been referred as to master regulators. The differentiation of Th lineages is usually mutually exclusive, possibly due to the cross-regulation of the key transcription factors expressed by each lineage. However, many reports have indicated that the master regulator of one lineage may be expressed in other lineages. For example, among the Treg population, there are T-bet-expressing and GATA3-expressing Foxp3+ Treg cells. In addition, RORgt and T-bet co-expressing Th cells have been identified both in mice and in humans. How these master regulators function in a same cell is an intriguing question. The ILCs also express one or two or even three of the master regulators T-bet, GATA3 and RORgt, in a single cell level, and these factors are critical for the development and functions of ILC subsets. During the past year, we reported that T-bet and GATA3 are dynamically expressed by regulatory T cells and such dynamic expression is critical for maintaining immune tolerance (Nature Immunology, 16: 197-206, 2015). By using a T-bet-AmCyan/GATA3-GFP/Foxp3-RFP triple reporter mouse strain, we found that T-bet and GATA3 were dynamically expressed by Treg cells under the influence of cytokine environment. T-bet fate-mapping reporter mice (T-bet-ZsGreen-T2A-CreERT2 X ROSA26-loxp-STOP-loxp-tdTomato) confirmed dynamic expression of T-bet by Treg cells in the steady state. Single deletion of either Tbx21 or Gata3 gene specifically in Treg cells did not result in an obvious phenotype, however, combinatorial deletion of both Tbx21 and Gata3 in Treg cells allowed the development of autoimmune-like diseases in mice in the steady state. Loss of suppressive functions in T-bet-GATA3 double-deficient Treg cells was associated with the upregulation of RORgt expression and IL-17 production in a cell-intrinsic manner. T-bet-GATA3 double-deficient Treg cells also become unstable in maintaining Foxp3 expression in transfer models. Overall, our results demonstrate that T-bet and GATA3-expressing Treg cells do not represent stable Treg subsets. In the steady state, Treg cells can transiently upregulate either T-bet or GATA3 to antagonize RORgt; such mechanism is critical for the maintenance of Treg function and stability, which controls peripheral tolerance. We have previously reported that GATA3 plays an essential role in the development of all IL-7Ra-expressing ILCs but not conventional NK cells (Immunity, 40: 378-88, 2014). This mirrors the essential function of GATA3 during CD4 but not CD8 T cell development indicating that NK cells may represent innate version of CD8 T cells and that GATA3 plays parallel roles in establishing and regulating both adaptive and innate lymphocyte subsets. Mice lacking GATA3 in all hematopoietic cells do not develop lymph node structures and Peyers patches and are also susceptible to Citrobacter rodentium infection due to the failure of ILC3 development. We have also reported that GATA3 is indispensable for maintaining the functions and survival of ILC2s similar to its functions in Th2 cells. Hundreds of GATA3-regulated genes in ILC2s including many critical genes that are involved in type 2 immune responses were identified through RNA-Seq. We have recently performed anti-GATA3 ChIP-Seq with ILC2s and found that GATA3 directly binds to many of its target genes as it does in Th2 cells. GATA3 is expressed at low level in mature ILC3s, but the functional role of low GATA3 expression in committed ILC3s has not been identified. Using a novel GATA3 conditional-deficient mouse strain (Gata3fl/fl-Rorc-Cre) in which Gata3 is only deleted in cells that have expressed and/or are expressing RORgt, we specifically assessed the function of GATA3 in committed ILC3s. Strikingly, we found that GATA3 has different but important functions in regulating homeostasis, development and functions of distinct ILC3 subsets. By comparing gene expression patterns in wild type (WT) and GATA3 deficient ILC3s, we identified three critical genes that are regulated by GATA3 in ILC3s Il7r, Rorc and Il22. Anti-GATA3 ChIP-Seq analysis indicates that all three genes are GATA3 direct targets in ILC3s. We further show that by promoting IL-7Rα expression, GATA3 regulates homeostasis of both CCR6+ and CCR6- ILC3 subsets. In addition, we demonstrate that through its regulation of Il22 expression, GATA3 promotes ILC3-dependent protective immunity to Citrobacter rodentium infection. Most importantly, we show that by fine-tuning RORgt expression and thus functional balance between T-bet and RORgt expression, GATA3 dictates the development of NKp46+ T-bet/RORgt co-expressing ILC3 subsets. To compare global gene expression patterns between the CCR6+ and NKp46+ ILC3 subsets, we generated a T-bet-ZsGreen/RORgt-E2-Crimson dual reporter mouse strain. Through RNA-Seq analysis of ILC3 subsets isolated from the dual reporter mice, we identified hundreds of CCR6+ and NKp46+ ILC3 lineage specific genes. Also through RNA-Seq analysis of ILC3 subsets that are deficient in GATA3, we found that GATA3 positively regulates NKp46+ ILC3 lineage specific genes but negatively regulates CCR6+ ILC3 lineage specific genes in NKp46+ ILC3s. These results suggest that GATA3 may serve as a switch in determining the CCR6+ and NKp46+ ILC3 lineages. Consequently, unlike the Gata3fl/fl-VavCre mice that do not have lymph nodes and fail to deal with C. rodentium infection due to lack of both CCR6+ LTi cells and NKp46+ ILC3s, Gata3fl/fl-Rorc-Cre mice develop lymph node structure normally yet still succumb to C. rodentium infection pointing to a differential function of ILC3 subsets regulated by GATA3.

Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Niaid Extramural Activities
Zip Code
Zhu, Jinfang (2018) Seventeen-Year Journey Working With a Master. Front Immunol 9:960
Zhong, Chao; Zheng, Mingzhu; Zhu, Jinfang (2018) Lymphoid tissue inducer-A divergent member of the ILC family. Cytokine Growth Factor Rev 42:5-12
Zhu, Jinfang (2018) T Helper Cell Differentiation, Heterogeneity, and Plasticity. Cold Spring Harb Perspect Biol 10:
Fang, Difeng; Cui, Kairong; Mao, Kairui et al. (2018) Transient T-bet expression functionally specifies a distinct T follicular helper subset. J Exp Med 215:2705-2714
Fang, Difeng; Cui, Kairong; Hu, Gangqing et al. (2018) Bcl11b, a novel GATA3-interacting protein, suppresses Th1 while limiting Th2 cell differentiation. J Exp Med 215:1449-1462
Hu, Gangqing; Cui, Kairong; Fang, Difeng et al. (2018) Transformation of Accessible Chromatin and 3D Nucleome Underlies Lineage Commitment of Early T Cells. Immunity 48:227-242.e8
Yu, Fang; Sharma, Suveena; Jankovic, Dragana et al. (2018) The transcription factor Bhlhe40 is a switch of inflammatory versus antiinflammatory Th1 cell fate determination. J Exp Med 215:1813-1821
Kawabe, Takeshi; Zhu, Jinfang; Sher, Alan (2018) Foreign antigen-independent memory-phenotype CD4+ T cells: a new player in innate immunity? Nat Rev Immunol 18:1
Huang, Yuefeng; Mao, Kairui; Chen, Xi et al. (2018) S1P-dependent interorgan trafficking of group 2 innate lymphoid cells supports host defense. Science 359:114-119
Zhu, Jinfang (2017) GATA3 Regulates the Development and Functions of Innate Lymphoid Cell Subsets at Multiple Stages. Front Immunol 8:1571

Showing the most recent 10 out of 57 publications