Bcl11b, a novel GATA3-interacting protein, suppresses Th1 while limiting Th2 cell differentiation

GATA-binding protein 3 (GATA3) acts as the master transcription factor for type 2 T helper (Th2) cell differentiation and function. However, it is still elusive how GATA3 function is precisely regulated in Th2 cells. Here, we show that the transcription factor B cell lymphoma 11b (Bcl11b), a previously unknown component of GATA3 transcriptional complex, is involved in GATA3-mediated gene regulation. Bcl11b binds to GATA3 through protein–protein interaction, and they colocalize at many important cis-regulatory elements in Th2 cells. The expression of type 2 cytokines, including IL-4, IL-5, and IL-13, is up-regulated in Bcl11b-deficient Th2 cells both in vitro and in vivo; such up-regulation is completely GATA3 dependent. Genome-wide analyses of Bcl11b- and GATA3-regulated genes (from RNA sequencing), cobinding patterns (from chromatin immunoprecipitation sequencing), and Bcl11b-modulated epigenetic modification and gene accessibility suggest that GATA3/Bcl11b complex is involved in limiting Th2 gene expression, as well as in inhibiting non-Th2 gene expression. Thus, Bcl11b controls both GATA3-mediated gene activation and repression in Th2 cells.

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Signal Transducer and Activator of Transcription 6 Controls Chemokine Production and T Helper Cell Type 2 Cell Trafficking in Allergic Pulmonary Inflammation

Journal of Experimental Medicine ◽

10.1084/jem.193.9.1087 ◽

2001 ◽

Vol 193 (9) ◽

pp. 1087-1096 ◽

Cited By ~ 130

Author(s):

Anuja Mathew ◽

James A. MacLean ◽

Elliot DeHaan ◽

Andrew M. Tager ◽

Francis H.Y. Green ◽

...

Keyword(s):

Transcription Factor ◽

Cell Differentiation ◽

Effector Function ◽

Th2 Cells ◽

Cell Trafficking ◽

Signal Transducer ◽

T Helper ◽

Chemokine Production ◽

Th2 Cell

Antigen-specific CD4 T helper type 2 (Th2) cells play a pivotal role in the induction of allergic asthma, but the mechanisms regulating their recruitment into the airways are unknown. Signal transducer and activator of transcription factor (Stat)6 is a transcription factor essential for Th2 cell differentiation. Here we show that Stat6 also controls Th2 cell recruitment and effector function in allergic inflammation in vivo. To isolate the role of Stat6 in regulating Th2 cell trafficking and effector function from its role in Th2 cell differentiation, we used a murine model of asthma in which in vitro–differentiated Stat6+/+ antigen-specific Th2 cells were adoptively transferred into naive Stat6−/− and Stat6+/+ mice followed by aerosol antigen challenge. We found that all of the features of asthma, including Th2 cell accumulation, Th2 and eosinophil-active chemokine production, and airway eosinophilia, mucus production, and hyperresponsiveness seen in Stat6+/+ mice, were dramatically absent in Stat6−/− mice that received Stat6+/+ antigen-specific Th2 cells. Our findings establish Stat6 as essential for Th2 cell trafficking and effector function and suggest that interruption of Stat6 signaling in resident cells of the lung is a novel approach to asthma therapy.

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ILC2 Cells Promote Th2 Cell Differentiation in AECOPD Through Activated Notch-GATA3 Signaling Pathway

Frontiers in Immunology ◽

10.3389/fimmu.2021.685400 ◽

2021 ◽

Vol 12 ◽

Author(s):

Min Jiang ◽

Ren Cai ◽

Jing Wang ◽

Zheng Li ◽

Dan Xu ◽

...

Keyword(s):

Chronic Obstructive Pulmonary Disease ◽

Peripheral Blood ◽

Culture Supernatant ◽

Th2 Cells ◽

Chronic Obstructive ◽

Obstructive Pulmonary Disease ◽

Protein Levels ◽

Th2 Cell

This study is to investigate the capacity of type 2 innate lymphoid cells (ILC2s) in regulating the Th2 type adaptive immune response of acute exacerbation of chronic obstructive pulmonary disease (AECOPD). The study enrolled healthy people, stable chronic obstructive pulmonary disease (COPD) patients, and AECOPD patients. Flow cytometry was used to detect Th2 and ILC2 cells in the peripheral blood. In addition, ILC2s from the peripheral blood of AECOPD patients were stimulated with PBS, IL-33, Jagged1, DAPT, IL-33+Jagged1, IL-33+DAPT, and IL-33+Jagged-1+DAP in vitro. The levels of cytokines in the culture supernatant were detected by ELISA and the culture supernatant was used to culture CD4 + T cells. The mRNA and protein levels of Notch1, hes1, GATA3, RORα, and NF-κB of ILC2s were detected by real-time PCR and Western blot. The proportion of Th2 and ILC2s was significantly increased in the peripheral blood of AECOPD patients, alone with the increased Notch1, hes1, and GATA3 mRNA levels. In vitro results showed that the mRNA and protein levels of Notch1, hes1, GATA3 and NF-κB were significantly increased after stimulation with Notch agonist, meanwhile, the level of type 2 cytokines were increased in the supernatant of cells stimulated with Notch agonist, and significantly promoted differentiation of Th2 cells in vitro. Disruption of Notch pathway weakened GATA3 expression and cytokine production, and ultimately affected the differentiation of Th2 cells. In conclusion, our results suggest that ILC2s can promote Th2 cell differentiation in AECOPD via activated Notch-GATA3 signal pathway.

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Downregulation of miRNA-451a Promotes the Differentiation of CD4+ T Cells towards Th2 Cells by Upregulating ETS1 in Childhood Asthma

Journal of Innate Immunity ◽

10.1159/000509714 ◽

2020 ◽

pp. 1-11

Author(s):

Tianyue Wang ◽

Qianlan Zhou ◽

Yunxiao Shang

Keyword(s):

T Cells ◽

Cell Differentiation ◽

Cd4 T Cells ◽

Peripheral Blood Lymphocytes ◽

Cell Polarization ◽

Luciferase Assay ◽

Th2 Cells ◽

Th2 Cell ◽

Novel Target

Children exposed to common aeroallergens may develop asthma that progresses into adulthood. Inflammation regulated by T helper 2 (Th2) cells, a specific subpopulation of CD4+ T lymphocytes, is involved in asthmatic injury. Herein, our microarray data indicated that microRNA-451a-5p (miRNA-451a) expression decreased by 4.6-fold and ETS proto-oncogene 1 (ETS1) increased by 2.2-fold in the peripheral blood lymphocytes isolated from asthmatic children (n = 4) as compared to control individuals (n = 4). The negative correlation between miRNA-451a and ETS1 was further validated in 40 CD4+ T cell samples (10 healthy vs. 30 asthmatic samples). In vitro, naïve CD4+ T cells isolated from control individuals were cultured under Th2 cell polarizing condition. miRNA-451a expression decreased while ETS1 increased in CD4+ T cells in the setting of Th2 cell polarization. Moreover, miRNA-451a knockdown enhanced Th2 cell polarization – cells positive for both GATA3 (GATA binding protein 3, a Th2-transcription factor) and CD4 increased, and the generation of Th2 cell cytokines, interleukin (IL)5 and IL13, increased. In contrast, miRNA-451a overexpression inhibited Th2 cell differentiation. Interestingly, dual-Luciferase assay proved ETS1 as a novel target of miRNA-451a. Moreover, enforced expression of ETS1 partially restored miRNA-451a-induced inhibition of IL5 and IL13, and increased the GATA3+CD4+ cell population. Collectively, our work demonstrates that downregulation of miRNA-451a upregulates ETS1 expression in CD4+ T cells, which may contribute to Th2 cell differentiation in pediatric asthma.

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The Wnt signaling pathway effector TCF7L2 is upregulated by insulin and represses hepatic gluconeogenesis

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00249.2012 ◽

2012 ◽

Vol 303 (9) ◽

pp. E1166-E1176 ◽

Cited By ~ 44

Author(s):

Wilfred Ip ◽

Weijuan Shao ◽

Yu-ting Alex Chiang ◽

Tianru Jin

Keyword(s):

Gene Expression ◽

Type 2 Diabetes ◽

Transcription Factor ◽

Wnt Signaling ◽

Wnt Signaling Pathway ◽

Glucose Production ◽

Hepatic Gluconeogenesis

Certain single nucleotide polymorphisms (SNPs) in transcription factor 7-like 2 (TCF7L2) are strongly associated with the risk of type 2 diabetes. TCF7L2 and β-catenin (β-cat) form the bipartite transcription factor cat/TCF in stimulating Wnt target gene expression. cat/TCF may also mediate the effect of other signaling cascades, including that of cAMP and insulin in cell-type specific manners. As carriers of TCF7L2 type 2 diabetes risk SNPs demonstrated increased hepatic glucose production, we aimed to determine whether TCF7L2 expression is regulated by nutrient availability and whether TCF7L2 and Wnt regulate hepatic gluconeogenesis. We examined hepatic Wnt activity in the TOPGAL transgenic mouse, assessed hepatic TCF7L2 expression in mice upon feeding, determined the effect of insulin on TCF7L2 expression and β-cat Ser675 phosphorylation, and investigated the effect of Wnt activation and TCF7L2 knockdown on gluconeogenic gene expression and glucose production in hepatocytes. Wnt activity was observed in pericentral hepatocytes in the TOPGAL mouse, whereas TCF7L2 expression was detected in human and mouse hepatocytes. Insulin and feeding stimulated hepatic TCF7L2 expression in vitro and in vivo, respectively. In addition, insulin activated β-cat Ser675 phosphorylation. Wnt activation by intraperitoneal lithium injection repressed hepatic gluconeogenic gene expression in vivo, whereas lithium or Wnt-3a reduced gluconeogenic gene expression and glucose production in hepatic cells in vitro. Small interfering RNA-mediated TCF7L2 knockdown increased glucose production and gluconeogenic gene expression in cultured hepatocytes. These observations suggest that Wnt signaling and TCF7L2 are negative regulators of hepatic gluconeogenesis, and TCF7L2 is among the downstream effectors of insulin in hepatocytes.

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Pyrrothiogatain acts as an inhibitor of GATA family proteins and inhibits Th2 cell differentiation in vitro

Scientific Reports ◽

10.1038/s41598-019-53856-1 ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 5

Author(s):

Shunsuke Nomura ◽

Hirotaka Takahashi ◽

Junpei Suzuki ◽

Makoto Kuwahara ◽

Masakatsu Yamashita ◽

...

Keyword(s):

Transcription Factor ◽

Cell Differentiation ◽

Dna Binding ◽

Binding Activity ◽

Th2 Cells ◽

Th2 Cytokines ◽

Screening System ◽

Dna Binding Activity ◽

Th2 Cell ◽

Gata Family

AbstractThe transcription factor GATA3 is a master regulator that modulates T helper 2 (Th2) cell differentiation and induces expression of Th2 cytokines, such as IL-4, IL-5, and IL-13. Th2 cytokines are involved in the protective immune response against foreign pathogens, such as parasites. However, excessive production of Th2 cytokines results in type-2 allergic inflammation. Therefore, the application of a GATA3 inhibitor provides a new therapeutic strategy to regulate Th2 cytokine production. Here, we established a novel high-throughput screening system for an inhibitor of a DNA-binding protein, such as a transcription factor, and identified pyrrothiogatain as a novel inhibitor of GATA3 DNA-binding activity. Pyrrothiogatain inhibited the DNA-binding activity of GATA3 and other members of the GATA family. Pyrrothiogatain also inhibited the interaction between GATA3 and SOX4, suggesting that it interacts with the DNA-binding region of GATA3. Furthermore, pyrrothiogatain significantly suppressed Th2 cell differentiation, without impairing Th1 cell differentiation, and inhibited the expression and production of Th2 cytokines. Our results suggest that pyrrothiogatain regulates the differentiation and function of Th2 cells via inhibition of GATA3 DNA binding activity, which demonstrates the efficiency of our drug screening system for the development of novel small compounds that inhibit the DNA-binding activity of transcription factors.

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T Cell Receptor–Induced Calcineurin Activation Regulates T Helper Type 2 Cell Development by Modifying the Interleukin 4 Receptor Signaling Complex

Journal of Experimental Medicine ◽

10.1084/jem.191.11.1869 ◽

2000 ◽

Vol 191 (11) ◽

pp. 1869-1880 ◽

Cited By ~ 82

Author(s):

Masakatsu Yamashita ◽

Makoto Katsumata ◽

Makio Iwashima ◽

Motoko Kimura ◽

Chiori Shimizu ◽

...

Keyword(s):

Cell Receptor ◽

Cell Development ◽

Interleukin 4 ◽

Th2 Cells ◽

T Helper ◽

Signaling Complex ◽

Th2 Cell ◽

Interleukin 4 Receptor

The activation of downstream signaling pathways of both T cell receptor (TCR) and interleukin 4 receptor (IL-4R) is essential for T helper type 2 (Th2) cell development, which is central to understanding immune responses against helminthic parasites and in allergic and autoimmune diseases. However, little is known about how these two distinct signaling pathways cooperate with each other to induce Th2 cells. Here, we show that successful Th2 cell development depends on the effectiveness of TCR-induced activation of calcineurin. An inhibitor of calcineurin activation, FK506, inhibited the in vitro anti-TCR–induced Th2 cell generation in a dose-dependent manner. Furthermore, the development of Th2 cells was significantly impaired in naive T cells from dominant-negative calcineurin Aα transgenic mice, whereas that of Th1 cells was less affected. Efficient calcineurin activation in naive T cells upregulated Janus kinase (Jak)3 transcription and the amount of protein. The generation of Th2 cells induced in vitro by anti-TCR stimulation was inhibited significantly by the presence of Jak3 antisense oligonucleotides, suggesting that the Jak3 upregulation is an important event for the Th2 cell development. Interestingly, signal transducer and activator of transcription (STAT)5 became physically and functionally associated with the IL-4R in the anti-TCR–activated developing Th2 cells that received efficient calcineurin activation, and also in established cloned Th2 cells. In either cell population, the inhibition of STAT5 activation resulted in a diminished IL-4–induced proliferation. Moreover, our results suggest that IL-4–induced STAT5 activation is required for the expansion process of developing Th2 cells. Thus, Th2 cell development is controlled by TCR-mediated activation of the Ca2+/calcineurin pathway, at least in part, by modifying the functional structure of the IL-4R signaling complex.

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IL-2 Modulates Th2 cell Responses to Glucocorticoid: A Cause of Persistent Type 2 Inflammation?

10.1101/365783 ◽

2018 ◽

Author(s):

Tharsan Kanagalingam ◽

Meerah Vijeyakumaran ◽

Nami Shrestha Palikhe ◽

Lauren Solomon ◽

Harissios Vliagoftis ◽

...

Keyword(s):

Peripheral Blood ◽

Blood Cells ◽

Asthma Severity ◽

Th2 Cells ◽

Peripheral Blood Cells ◽

Cell Responses ◽

Th2 Cell ◽

Type 2 Inflammation

ABSTRACTBackgroundInhaled glucocorticosteroids (GCs) are the main treatment for asthma as they reduce type 2 cytokine (IL-4, IL-5 and IL-13) expression and induce apoptosis. Asthma severity is associated with GC insensitivity, increased type 2 inflammation and circulating Th2 cells. Since IL-2 is a T cell survival factor, we assessed whether IL-2 levels associate with the proportion of Th2 cells and/or correlate with clinical features of asthma severity.MethodsPeripheral blood from asthma patients (n=18) was obtained and Th2 cell numbers determined by flow cytometry. Peripheral blood cells were activated with mitogen (24hrs) and supernatant levels of IL-2 and IL-13 measured by ELISA. In vitro differentiated Th2 cells were treated with dexamethasone and IL-2 and assessed for apoptosis by flow cytometry staining of Annexin V. Level of mRNA for anti-apoptotic (BCL-2) and pro-apoptotic (BIM) genes as well as IL-13 were determined by qRT-PCR.ResultsIL-2 produced by activated peripheral blood cells correlated negatively with lung function (FEV1) and positively with daily dose of inhaled GC. When patients were stratified based on IL-2 level, high IL-2 producers made more IL-13 and had more circulating Th2 cells. In vitro, increasing the level of IL-2 in the culture media was associated with resistance to DEX-induced apoptosis, more BCL-2 and less BIM mRNA. Th2 cells cultured with higher IL-2 also had more IL-13 mRNA and required higher concentrations of DEX for cytokine suppression.Conclusions and Clinical RelevanceIL-2 modulates Th2 cell responses to GC, supporting both their survival and pro-inflammatory capacity, suggesting that a patient’s potential to produce IL-2 may be a determinant in asthma severity.

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Loss of the Pioneer Factor FOXA1 Results in Genome-wide Epigenetic Reprogramming and activation of Interferon-Response Genes including CD274/PD-L1

10.1101/2020.09.11.294041 ◽

2020 ◽

Author(s):

Wenhuo Hu ◽

Hironobu Yamashita ◽

Jenna Craig ◽

Vonn Walter ◽

Joshua I. Warrick ◽

...

Keyword(s):

Gene Expression ◽

Bladder Cancer ◽

Transcription Factor ◽

Epigenetic Modification ◽

Regulatory Elements ◽

Epigenetic Reprogramming ◽

Gene Set Enrichment Analysis ◽

Interferon Response ◽

Integrated Analysis ◽

The Impact

AbstractForkhead Box A1 (FOXA1) is a pioneer transcription factor critical in epigenetic regulation of chromatin and cell fate determination. Reduced FOXA1 expression is an independent predictor of poor overall survival in bladder cancer patients. However, the impact of FOXA1 loss on chromatin epigenetics in bladder cancer is unknown. Therefore, we determined the impact of FOXA1 knock out (KO) on epigenetic modification of chromatin and associated gene expression. We identified 8,230 differentially expressed genes following FOXA1 KO. Surprisingly, Gene Set Enrichment Analysis (GSEA) identified IFNɑ/ɣ gene expression signatures as enriched following FOXA1 KO. FOXA1 KO induced both increased and decreased numbers of histone 3 lysine 27 acetylation (H3K27ac) sites throughout the genome. As expected, the majority of differences in H3K27ac across genomic areas in FOXA1 KO cells is mapped to intergenic and intronic regions where enhancers reside. In addition, a subset of differential H3K27ac levels were also mapped to proximal promoters and within gene bodies. Integrated analysis of RNA/ChIP-seq data shows changes in gene expression that are mirrored by differences in H3K27ac. Motif analysis of DNA sequence enriched for H3K27ac identified significant increases in transcription factor binding motifs including the interferon sensitive response element (ISRE) and interferon response factors such as IRF1. Moreover, we identified increased H3K27ac of regulatory elements as being associated with several upregulated interferon sensitive genes (ISGs) in FOXA1 KO cells, including CD274/PD-L1. Western blotting and Q-RT-PCR confirmed upregulation of CD274/PD-L1 following FOXA1 KO. Analysis of TCGA data confirmed an inverse relationship between FOXA1 and CD274 in bladder cancer, as well as in other cancers. In summary, we provide evidence of widespread epigenetic reprogramming after FOXA1 KO in bladder cancer cells. Additionally, we provide evidence that FOXA1 KO-induced epigenetic changes contribute to activation of a global interferon-dominant expression signature, including the immune checkpoint target CD274/PD-L1 in a cancer cell-intrinsic manner.

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Lack of WDFY4 Aggravates Ovalbumin-Induced Asthma via Enhanced Th2 Cell Differentiation

International Archives of Allergy and Immunology ◽

10.1159/000516970 ◽

2021 ◽

pp. 1-8

Author(s):

Yan Li ◽

Anran Wang ◽

Feng Long ◽

Fei Gao ◽

Shang Gao ◽

...

Keyword(s):

Cell Differentiation ◽

Susceptibility Gene ◽

Inflammatory Cells ◽

Lavage Fluid ◽

Th2 Cells ◽

Th2 Cytokines ◽

Wild Type ◽

Th2 Cell ◽

Study Results

Background: Asthma is a chronic inflammatory airway disease, and Th2 cells play an important role in asthma. WDFY4 (WDFY family member 4) is a susceptibility gene in several autoimmune diseases. Objective: In this study, the roles of WDFY4 in Th2 cell differentiation and Th2-dependent asthma were investigated. Methods: Naïve CD4+ T cells were isolated from wild-type and WDFY4-deficient mice and induced to differentiate in vitro. Subsequently, a mouse model of asthma was established by sensitization with ovalbumin. Results: Study results showed that WDFY4 deficiency could promote the differentiation of Th2 cells and the production of Th2 cytokines. WDFY4-deficient asthmatic mice showed higher levels of Th2 cytokines in the lungs and bronchoalveolar lavage fluid than wild-type mice. Moreover, infiltration of inflammatory cells, hyperplasia of goblet cells, production of mucus, and deposition of collagen were enhanced in WDFY4-deficient asthmatic mice. Conclusions: Our study demonstrates the pivotal role of WDFY4 in the pathogenesis of asthma and in Th2 cell differentiation.

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HNF1alpha is involved in tissue-specific regulation of CFTR gene expression

Biochemical Journal ◽

10.1042/bj20031157 ◽

2004 ◽

Vol 378 (3) ◽

pp. 909-918 ◽

Cited By ~ 44

Author(s):

Nathalie MOUCHEL ◽

Sytse A. HENSTRA ◽

Victoria A. McCARTHY ◽

Sarah H. WILLIAMS ◽

Marios PHYLACTIDES ◽

...

Keyword(s):

Gene Expression ◽

Transcription Factor ◽

Tissue Specificity ◽

Regulatory Elements ◽

Upstream Region ◽

Mrna Levels ◽

Tissue Specific ◽

Specific Regulation

The CFTR (cystic fibrosis transmembrane conductance regulator) gene shows a complex pattern of expression with tissue-specific and temporal regulation. However, the genetic elements and transcription factors that control CFTR expression are largely unidentified. The CFTR promoter does not confer tissue specificity on gene expression, suggesting that there are regulatory elements outside the upstream region. Analysis of potential regulatory elements defined as DNase 1-hypersensitive sites within introns of the gene revealed multiple predicted binding sites for the HNF1α (hepatocyte nuclear factor 1α) transcription factor. HNF1α, which is expressed in many of the same epithelial cell types as CFTR and shows similar differentiation-dependent changes in gene expression, bound to these sites in vitro. Overexpression of heterologous HNF1α augmented CFTR transcription in vivo. In contrast, antisense inhibition of HNF1α transcription decreased the CFTR mRNA levels. Hnf1α knockout mice showed lower levels of CFTR mRNA in their small intestine in comparison with wild-type mice. This is the first report of a transcription factor, which confers tissue specificity on the expression of this important disease-associated gene.

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