Eun Sook Hwang

T-bet represses T H 17 differentiation by preventing Runx1-mediated activation of the gene encoding RORγt

Overactive TH17 responses are tightly linked to the development of autoimmunity, yet the factors that negatively regulate differentiation of this lineage remain unknown. Here, we report that T-bet suppresses the development of the TH17 cell lineage by inhibiting the transcription of Rorc. T-bet interacts with the transcription factor Runx1 and this interaction blocks Runx1-mediated transactivation of Rorc. T-bet residue Tyr304 is required for T-bet-Runx1 complex formation, for blocking Runx1 activity and for inhibiting the TH17 differentiation program. These data reinforce the concept of master regulators that shape immune responses by simultaneously activating one genetic program while silencing the activity of competing regulators in a common progenitor cell.Overactive responses by interleukin 17 (IL-17)-producing helper T cells (TH17 cells) are tightly linked to the development of autoimmunity, yet the factors that negatively regulate the differentiation of this lineage remain unknown. Here we report that the transcription factor T-bet suppressed development of the TH17 cell lineage by inhibiting transcription of Rorc (which encodes the transcription factor RORγt). T-bet interacted with the transcription factor Runx1, and this interaction blocked Runx1-mediated transactivation of Rorc. T-bet Tyr304 was required for formation of the T-bet–Runx1 complex, for blockade of Runx1 activity and for inhibition of the TH17 differentiation program. Our data reinforce the idea of master regulators that shape immune responses by simultaneously activating one genetic program while silencing the activity of competing regulators in a common progenitor cell.

TBX21: A functional variant predicts improvement in asthma with the use of inhaled corticosteroids

TBX21 encodes for the transcription factor T-bet (T-box expressed in T cells), which influences naïve T lymphocyte development and has been implicated in asthma pathogenesis. Specifically, the T-bet knockout mouse spontaneously develops airway hyperresponsiveness and other changes consistent with asthma. Because airway responsiveness is moderated by the use of inhaled corticosteroids in asthma, it is conceivable that genetic variation in TBX21 may alter asthma phenotypes in a treatment-specific fashion. Here we demonstrate that the nonsynonymous variation in TBX21 coding for replacement of histidine 33 with glutamine is associated with significant improvement in the PC20 (a measure of airway responsiveness) of asthmatic children in a large clinical trial spanning 4 years. We note that this increase occurs only in the children randomized to inhaled corticosteroids and that it dramatically enhances the overall improvement in PC20 associated with inhaled corticosteroid usage. The average PC20 at trial end for subjects on inhaled corticosteroids possessing a variant allele was in the normal range for nonasthmatics. In cellular models, we show that the TBX21 variant increases T helper 1 and decreases T helper 2 cytokine expression comparably with wild type. TBX21 may thus be an important determinant pharmacogenetic response to the therapy of asthma with inhaled corticosteroids.

IL-2 production in developing Th1 cells is regulated by heterodimerization of RelA and T-bet and requires T-bet serine residue 508

Interleukin (IL)-2 is the predominant cytokine that is produced by naive Th cells in a primary response. It is required for proliferation and differentiation of Th precursor cells into effector cells. Initial high-level IL-2 production is followed by its decline, and the concomitant induction of cytokines that are typical of the differentiated state. Although the factors that are responsible for the early induction of IL-2 are well defined, the mechanisms that are responsible for its down-regulation in later stages of Th development have not been studied as much. Previous work from our laboratory revealed a repressor function for the T-box transcription factor, T-bet, in IL-2 gene transcription. Here, we report that T-betS508 is required for the optimal repression of IL-2 production in developing Th1 cells. Phosphorylation of T-betS508 by casein kinase I and glycogen synthase kinase-3 kinases accompanies T-bets interaction with the RelA nuclear factor–κB transcription factor. Heterodimerization of T-bet and RelA interferes with the binding of RelA to the IL-2 promoter, and hence, transcriptional activation of the IL-2 gene by RelA.

Spontaneous and aging-dependent development of arthritis in NADPH oxidase 2 deficiency through altered differentiation of CD11b+ and Th/Treg cells

Emerging evidence indicates that NADPH oxidase (NOX) and its reactive oxygen species (ROS) products modulate a variety of cellular events, including proliferation, differentiation, and apoptosis. In this study, we investigated the functions of NOX2 and ROS in immune modulation using NOX2 knockout (KO) mice. Interestingly, NOX2 KO mice spontaneously developed arthritis with onset at 6–7 wk of age and high incidence (60%) at 15–18 wk of age. Arthritis severity in NOX2 KO mice was proportionally increased with age and higher in females than in males. Bone destruction was confirmed by microcomputed tomography scanning and histological analyses of joints. Inflammatory factors, including TNF-α, IL-1β, and RANKL, and serum level of anti–type II collagen IgG were significantly increased in NOX2 KO mice. In addition, NOX2 deficiency perturbed the immune system upon aging. NOX2 KO mice demonstrated preferred development of CD11b+Gr-1+ myeloid cells with profound production of proinflammatory cytokines and augmented expression of IL-17 through the activation of STAT3 and RORγt in vivo. NOX2 deficiency increased differentiation of effector Th cells in vitro and decreased CD25+FoxP3+ Treg cells both in vitro and in vivo. Furthermore, adoptive transfer of NOX2-deficient CD4+ T cells into RAG KO mice increased arthritic inflammation compared with WT cells. These results demonstrated that NOX2 deficiency affected the development of CD11b+ myeloid cells and Th17/Treg cells, and thus promoted inflammatory cytokine production and inflammatory arthritis development, strongly supporting a crucial role for ROS generation in the modulation of Th17/Treg cell development and its related inflammatory immune response upon aging.

TAZ as a novel enhancer of MyoD-mediated myogenic differentiation

Myoblast differentiation is indispensable for skeletal muscle formation and is governed by the precisely coordinated regulation of a series of transcription factors, including MyoD and myogenin, and transcriptional coregulators. TAZ (transcriptional coactivator with PDZ‐binding motif) has been characterized as a modulator of mesenchymal stem cell differentiation into osteoblasts and adipocytes through its regulation of lineage‐specific master transcription factors. In this study, we investigated whether TAZ affects myoblast differentiation, which is one of the differentiated lineages of mesenchymal stem cells. Ectopic overexpression of TAZ in myoblasts increases myogenic gene expression in a MyoD‐dependent manner and hastens myofiber formation, whereas TAZ knockdown delays myogenic differentiation. In addition, enforced coexpression of TAZ and MyoD in fibroblasts accelerates MyoD‐induced myogenic differentiation. TAZ physically interacts with MyoD through the WW domain and activates MyoD‐dependent gene transcription. TAZ additionally enhances the interaction of MyoD with the myogenin gene promoter. These results strongly suggest that TAZ functions as a novel transcriptional modulator of myogenic differentiation by promoting MyoD‐mediated myogenic gene expression.—Jeong, H., Bae, S., An, S. Y., Byun, M. R., Hwang, J.‐H., Yaffe, M. B., Hong, J.‐H., Hwang, E. S. TAZ as a novel enhancer of MyoD‐mediated myogenic differentiation. FASEB J. 24, 3310–3320 (2010). www.fasebj.org

Shear stress induced by an interstitial level of slow flow increases the osteogenic differentiation of mesenchymal stem cells through TAZ activation.

Shear stress activates cellular signaling involved in cellular proliferation, differentiation, and migration. However, the mechanisms of mesenchymal stem cell (MSC) differentiation under interstitial flow are not fully understood. Here, we show the increased osteogenic differentiation of MSCs under exposure to constant, extremely low shear stress created by osmotic pressure-induced flow in a microfluidic chip. The interstitial level of shear stress in the proposed microfluidic system stimulated nuclear localization of TAZ (transcriptional coactivator with PDZ-binding motif), a transcriptional modulator of MSCs, activated TAZ target genes such as CTGF and Cyr61, and induced osteogenic differentiation. TAZ-depleted cells showed defects in shear stress-induced osteogenic differentiation. In shear stress induced cellular signaling, Rho signaling pathway was important forthe nuclear localization of TAZ. Taken together, these results suggest that TAZ is an important mediator of interstitial flow-driven shear stress signaling in osteoblast differentiation of MSCs.

An IL-4-independent and CD25-mediated function of c-maf in promoting the production of Th2 cytokines

c-maf is a T helper (Th)2 cell-specific transcription factor, which promotes the differentiation of Th2 cells mainly by an IL-4-dependent mechanism. It remains unclear whether c-maf possesses any IL-4-independent function in regulating the production of Th2 cytokines. Here, we provide evidence demonstrating that c-maf, independent of IL-4, is essential for normal induction of CD25 in developing Th2 cells. The levels of CD25 are significantly higher in developing Th2 cells than in developing Th1 cells during in vitro differentiation. In addition, timely blockade of IL-2 receptor signaling selectively inhibits the production of Th2 cytokines, but not IFN-γ or IL-2. Taken together, our results uncover an IL-4-independent and CD25-mediated function of c-maf in promoting the production of Th2 cytokines.

T-Bet Plays a Key Role in NK-Mediated Control of Melanoma Metastatic Disease

Antitumor responses depend on type 1 immunity, which is severely impaired in mice deficient for the T-box expressed in T cells (T-bet) transcription factor. Both T-bet-deficient (T-bet−/−) NK and CTL show defective function, which can be overcome by strong stimuli due to the expression of eomesodermin, another member of the T-box family. The effective response from T-bet−/− mice to viral infection and tumor initiation corroborates with these findings. However, T-bet−/− animals fail to control cancer metastasis and are, therefore, highly susceptible to tumor spread. The mechanism of T-bet-dependent resistance to metastatic disease is not known. In this study, we show that T-bet plays a role in inhibiting cancer metastasis by regulating the longevity and function of NK cells. Our data demonstrate that the absence of a proper innate immune response driven by NK cells in T-bet−/− mice precludes the initiation of a potent adaptive response to tumors. Adoptive transfer of wild-type activated NK cells protects T-bet−/− animals after melanoma challenge showing that reconstitution of the NK compartment in these mice is sufficient to mediate a significant reduction in tumor burden. Transfer of T-bet−/− A-NK cells fails to do so, due to their reduced in vivo survival, inefficient lysis of cancer cells, and poor IFN-γ production. Taken together, these results show for the first time an irreplaceable role for T-bet in the NK-mediated cross-talk between innate and adaptive immune responses to metastatic disease.

T-bet, a T-Cell–Associated Transcription Factor, Is Expressed in a Subset of B-Cell Lymphoproliferative Disorders

T-bet, a T-box transcription factor, is expressed in CD4+ T lymphocytes committed to Th1 T-cell development and in a subset of T-cell non-Hodgkin lymphomas. Recent evidence indicates that T-bet also is expressed in B lymphocytes and might participate in immunoglobulin class switching. We examined T-bet expression in 116 cases of B-cell lymphoproliferative disorders by immunostaining and found that T-bet was expressed consistently in precursor B-cell lymphoblastic leukemia/lymphoblastic lymphoma (14/14 cases) in contrast with precursor T-cell lymphoblastic leukemia/lymphoblastic lymphoma, which is consistently T-bet- (13 cases), as previously reported. T-bet is expressed in memory B cell-derived neoplasms (chronic lymphocytic leukemia, marginal zone lymphoma, hairy cell leukemia; 35/42 cases) but not in cases of mantle cell, follicular, and large cell lymphoma (43 cases). Expression of T-bet in precursor B-cell lymphoblastic leukemia/lymphoblastic lymphoma was confirmed by Western blot analysis. The expression of T-bet in a significant subset of B-cell lymphoproliferative disorders but not in the vast majority of reactive B cells suggests it might have a role in the oncogenesis of T-bet+ B-cell neoplasms. In addition, T-bet should serve as a useful new marker for the diagnosis and subtyping of B-cell lymphoproliferative disorders.

Transcriptional regulation of T helper 17 cell differentiation.

The third lineage of T helper subsets, Th17, has recently been identified as an IL-17-producing CD4+ Th cell, and its functions and regulatory mechanisms have been extensively characterized in immune responses. Functional studies have provided evidence that Th17 cells are important for the modulation of autoimmune responses, such as chronic asthma, rheumatoid arthritis, inflammatory bowel diseases, and multiple sclerosis. Murine Th17 cell differentiation is enhanced by the coordinated functions of distinct cytokines including TGFβ, IL-6, IL-21, and IL-23, whereas IL-2, IL-4, IFNγ, and IL-27 inhibit its differentiation. In addition, Th17 cells are controlled by several transcription factors such as RORγ t, IRF4, BATF, FoxP3, T-bet, PPARγ, E-FABP, and SOCSs. This review focuses on the functions and regulatory mechanisms of several transcription factors in the control of Th17 cell differentiation.