BACKGROUND

The transcription factor GATA3 has recently been shown to be necessary for mammary gland morphogenesis and luminal cell differentiation. There is also an increasing body of data linking GATA3 to the estrogen receptor alpha (ERalpha) pathway. Among these it was shown that GATA3 associates with the promoter of the ERalpha gene and ERalpha can reciprocally associate with the GATA3 gene. GATA3 has also been directly implicated in a differentiated phenotype in mouse models of mammary tumourigenesis. The purpose of our study was to compare coexpressed genes, by meta-analysis, of GATA3 and relate these to a similar analysis for ERalpha to determine the depth of overlap.

RESULTS

We have used a newly described method of meta-analysis of multiple cancer studies within the Oncomine database, focusing here predominantly upon breast cancer studies. We demonstrate that ERalpha and GATA3 reciprocally have the highest overlap with one another. Furthermore, we show that when both coexpression meta-analysis lists for ERalpha and GATA3 are compared there is a significant overlap between both and, like ERalpha, GATA3 coexpresses with ERalpha pathway partners such as pS2 (TFF1), TFF3, FOXA1, BCL2, ERBB4, XBP1, NRIP1, IL6ST, keratin 18(KRT18) and cyclin D1 (CCND1). Moreover, as these data are derived from human tumour samples this adds credence to previous cell-culture or murine based studies.

CONCLUSIONS

GATA3 is hypothesized to be integral to the ERalpha pathway given the following: (1) The large overlap of coexpressed genes as seen by meta-analysis, between GATA3 and ERalpha, (2) The highest coexpressing gene for GATA3 was ERalpha and vice-versa, (3) GATA3, like ERalpha, coexpresses with many well-known ERalpha pathway partners such as pS2.

How loss-of-function of GATA3 contributes to the development of breast cancer is poorly understood. Here, we report that GATA3 nucleates a transcription repression program composed of G9A and MTA3-, but not MTA1- or MTA2-, constituted NuRD complex. Genome-wide analysis of the GATA3/G9A/NuRD(MTA3) targets identified a cohort of genes including ZEB2 that are critically involved in epithelial-to-mesenchymal transition and cell invasion. We demonstrate that the GATA3/G9A/NuRD(MTA3) complex inhibits the invasive potential of breast cancer cells in vitro and suppresses breast cancer metastasis in vivo. Strikingly, the expression of GATA3, G9A, and MTA3 is concurrently downregulated during breast cancer progression, leading to an elevated expression of ZEB2, which, in turn, represses the expression of G9A and MTA3 through the recruitment of G9A/NuRD(MTA1).

IgG4-related disease (IgG4-RD) is a systemic condition of unknown cause characterized by highly fibrotic lesions with dense lymphoplasmacytic infiltrates. CD4(+) T cells constitute the major inflammatory cell population in IgG4-RD lesions.

We used an unbiased approach to characterize CD4(+) T-cell subsets in patients with IgG4-RD based on their clonal expansion and ability to infiltrate affected tissue sites.

We used flow cytometry to identify CD4(+) effector/memory T cells in a cohort of 101 patients with IgG4-RD. These expanded cells were characterized by means of gene expression analysis and flow cytometry. Next-generation sequencing of the T-cell receptor β chain gene was performed on CD4(+)SLAMF7(+) cytotoxic T lymphocytes (CTLs) and CD4(+)GATA3(+) TH2 cells in a subset of patients to identify their clonality. Tissue infiltration by specific T cells was examined by using quantitative multicolor imaging.

CD4(+) effector/memory T cells with a cytolytic phenotype were expanded in patients with IgG4-RD. Next-generation sequencing revealed prominent clonal expansions of these CD4(+) CTLs but not CD4(+)GATA3(+) memory TH2 cells in patients with IgG4-RD. The dominant T cells infiltrating a range of inflamed IgG4-RD tissue sites were clonally expanded CD4(+) CTLs that expressed SLAMF7, granzyme A, IL-1β, and TGF-β1. Clinical remission induced by rituximab-mediated B-cell depletion was associated with a reduction in numbers of disease-associated CD4(+) CTLs.

IgG4-RD is prominently linked to clonally expanded IL-1β- and TGF-β1-secreting CD4(+) CTLs in both peripheral blood and inflammatory tissue lesions. These active, terminally differentiated, cytokine-secreting effector CD4(+) T cells are now linked to a human disease characterized by chronic inflammation and fibrosis.

Metanephric kidney induction critically depends on mesenchymal-epithelial interactions in the caudal region of the nephric (or Wolffian) duct. Central to this process, GDNF secreted from the metanephric mesenchyme induces ureter budding by activating the Ret receptor expressed in the nephric duct epithelium. A failure to regulate this pathway is believed to be responsible for a large proportion of the developmental anomalies affecting the urogenital system. Here, we show that the nephric duct-specific inactivation of the transcription factor gene Gata3 leads to massive ectopic ureter budding. This results in a spectrum of urogenital malformations including kidney adysplasia, duplex systems, and hydroureter, as well as vas deferens hyperplasia and uterine agenesis. The variability of developmental defects is reminiscent of the congenital anomalies of the kidney and urinary tract (CAKUT) observed in human. We show that Gata3 inactivation causes premature nephric duct cell differentiation and loss of Ret receptor gene expression. These changes ultimately affect nephric duct epithelium homeostasis, leading to ectopic budding of interspersed cells still expressing the Ret receptor. Importantly, the formation of these ectopic buds requires both GDNF/Ret and Fgf signaling activities. We further identify Gata3 as a central mediator of beta-catenin function in the nephric duct and demonstrate that the beta-catenin/Gata3 pathway prevents premature cell differentiation independently of its role in regulating Ret expression. Together, these results establish a genetic cascade in which Gata3 acts downstream of beta-catenin, but upstream of Ret, to prevent ectopic ureter budding and premature cell differentiation in the nephric duct.

Differentiation of naive CD4+ T cells into helper T (Th) cells is controlled by a combination of several transcriptional factors. In this study, we examined the functional role of the Runx1 transcription factor in Th cell differentiation. Naive T cells from transgenic mice expressing a dominant interfering form of Runx1 exhibited enhanced interleukin 4 production and efficient Th2 differentiation. In contrast, transduction of Runx1 into wild-type T cells caused a complete attenuation of Th2 differentiation and was accompanied by the cessation of GATA3 expression. Furthermore, endogenous expression of Runx1 in naive T cells declined after T cell receptor stimulation, at the same time that expression of GATA3 increased. We conclude that Runx1 plays a novel role as a negative regulator of GATA3 expression, thereby inhibiting the Th2 cell differentiation.

The oncogenic transcription factor TAL1/SCL is aberrantly expressed in 60% of cases of human T cell acute lymphoblastic leukemia (T-ALL) and initiates T-ALL in mouse models. By performing global microRNA (miRNA) expression profiling after depletion of TAL1, together with genome-wide analysis of TAL1 occupancy by chromatin immunoprecipitation coupled to massively parallel DNA sequencing, we identified the miRNA genes directly controlled by TAL1 and its regulatory partners HEB, E2A, LMO1/2, GATA3, and RUNX1. The most dynamically regulated miRNA was miR-223, which is bound at its promoter and up-regulated by the TAL1 complex. miR-223 expression mirrors TAL1 levels during thymic development, with high expression in early thymocytes and marked down-regulation after the double-negative-2 stage of maturation. We demonstrate that aberrant miR-223 up-regulation by TAL1 is important for optimal growth of TAL1-positive T-ALL cells and that sustained expression of miR-223 partially rescues T-ALL cells after TAL1 knockdown. Overexpression of miR-223 also leads to marked down-regulation of FBXW7 protein expression, whereas knockdown of TAL1 leads to up-regulation of FBXW7 protein levels, with a marked reduction of its substrates MYC, MYB, NOTCH1, and CYCLIN E. We conclude that TAL1-mediated up-regulation of miR-223 promotes the malignant phenotype in T-ALL through repression of the FBXW7 tumor suppressor.

During the development of immune responses to pathogens, self-antigens, or environmental allergens, naive CD4(+) T cells differentiate into subsets of effector cells including Th1, Th2, and Th17 cells. The differentiation into these subsets is controlled by specific transcription factors. The activity of these effector cells is limited by nTregs and iTregs, whose differentiation and maintenance are dependent on the transcription factor Foxp3. The regulation of autoimmune diseases mediated by Th1 and Th17 cells by Tregs has been studied and reviewed extensively. However, much less has been presented about the interplay between Tregs and Th2 cells and their contribution to allergic disease. In this perspective, we discuss the regulation of Th2 cells by Tregs and vice versa, focusing on the interplay between the IL-4-activated STAT6/GATA3 pathway and Foxp3.

The clinical symptomatology in the X-linked Wiskott-Aldrich syndrome (WAS), a combined immunodeficiency and autoimmune disease resulting from WAS protein (WASp) deficiency, reflects the underlying coexistence of an impaired T helper 1 (TH1) immunity alongside intact TH2 immunity. This suggests a role for WASp in patterning T(H) subtype immunity, yet the molecular basis for the TH1-TH2 imbalance in human WAS is unknown. We have discovered a nuclear role for WASp in the transcriptional regulation of the TH1 regulator gene TBX21 at the chromatin level. In primary TH1-differentiating cells, a fraction of WASp is found in the nucleus, where it is recruited to the proximal promoter locus of the TBX21 gene, but not to the core promoter of GATA3 (a TH2 regulator gene) or RORc (a TH17 regulator gene). Genome-wide mapping demonstrates association of WASp in vivo with the gene-regulatory network that orchestrates TH1 cell fate choice in the human TH cell genome. Functionally, nuclear WASp associates with H3K4 trimethyltransferase [RBBP5 (retinoblastoma-binding protein 5)] and H3K9/H3K36 tridemethylase [JMJD2A (Jumonji domain-containing protein 2A)] proteins, and their enzymatic activity in vitro and in vivo is required for achieving transcription-permissive chromatin dynamics at the TBX21 proximal promoter in primary differentiating TH1 cells. During TH1 differentiation, the loss of WASp accompanies decreased enrichment of RBBP5 and, in a subset of WAS patients, also of filamentous actin at the TBX21 proximal promoter locus. Accordingly, human WASp-deficient TH cells, from natural mutation or RNA interference-mediated depletion, demonstrate repressed TBX21 promoter dynamics when driven under TH1-differentiating conditions. These chromatin derangements accompany deficient T-BET messenger RNA and protein expression and impaired TH1 function, defects that are ameliorated by reintroducing WASp. Our findings reveal a previously unappreciated role of WASp in the epigenetic control of T-BET transcription and provide a new mechanism for the pathogenesis of WAS by linking aberrant histone methylation at the TBX21 promoter to dysregulated adaptive immunity.

Breast cancer is characterized by an important histoclinical heterogeneity that currently hampers the selection of the most appropriate treatment for each case. This problem could be solved by the identification of new parameters that better predict the natural history of the disease and its sensitivity to treatment. A large-scale molecular characterization of breast cancer could help in this context. Using cDNA arrays, we studied the quantitative mRNA expression levels of 176 candidate genes in 34 primary breast carcinomas along three directions: comparison of tumor samples, correlations of molecular data with conventional histoclinical prognostic features and gene correlations. The study evidenced extensive heterogeneity of breast tumors at the transcriptional level. A hierarchical clustering algorithm identified two molecularly distinct subgroups of tumors characterized by a different clinical outcome after chemotherapy. This outcome could not have been predicted by the commonly used histoclinical parameters. No correlation was found with the age of patients, tumor size, histological type and grade. However, expression of genes was differential in tumors with lymph node metastasis and according to the estrogen receptor status; ERBB2 expression was strongly correlated with the lymph node status (P < 0.0001) and that of GATA3 with the presence of estrogen receptors (P < 0.001). Thus, our results identified new ways to group tumors according to outcome and new potential targets of carcinogenesis. They show that the systematic use of cDNA array testing holds great promise to improve the classification of breast cancer in terms of prognosis and chemosensitivity and to provide new potential therapeutic targets.

The adult zebrafish brain, unlike mammalian counterparts, can regenerate after injury owing to the neurogenic capacity of stem cells with radial glial character. We hypothesized that injury-induced regenerative programs might be turned on after injury in zebrafish brain and enable regenerative neurogenesis. Here we identify one such gene-the transcription factor gata3-which is expressed only after injury in different zebrafish organs. Gata3 is required for reactive proliferation of radial glia cells, subsequent regenerative neurogenesis, and migration of the newborn neurons. We found that these regeneration-specific roles of Gata3 are dependent on the injury because Gata3 overexpression in the unlesioned adult zebrafish brain is not sufficient to induce neurogenesis. Thus, gata3 acts as a specific injury-induced proregenerative factor that is essential for the regenerative capacity in vertebrates.

BACKGROUND

It has been suggested that bladder cancer can be divided into two molecular subtypes referred to as luminal and basal with distinct clinical behaviors and sensitivities to chemotherapy. We aimed to validate these subtypes in several clinical cohorts and identify signature immunohistochemical markers that would permit simple and cost-effective classification of the disease in primary care centers.

METHODS

We analyzed genomic expression profiles of bladder cancer in three cohorts of fresh frozen tumor samples: MD Anderson (n=132), Lund (n=308), and The Cancer Genome Atlas (TCGA) (n=408) to validate the expression signatures of luminal and basal subtypes and relate them to clinical follow-up data. We also used an MD Anderson cohort of archival bladder tumor samples (n=89) and a parallel tissue microarray to identify immunohistochemical markers that permitted the molecular classification of bladder cancer.

RESULTS

Bladder cancers could be assigned to two candidate intrinsic molecular subtypes referred to here as luminal and basal in all of the datasets analyzed. Luminal tumors were characterized by the expression signature similar to the intermediate/superficial layers of normal urothelium. They showed the upregulation of PPARγ target genes and the enrichment for FGFR3, ELF3, CDKN1A, and TSC1 mutations. In addition, luminal tumors were characterized by the overexpression of E-Cadherin, HER2/3, Rab-25, and Src. Basal tumors showed the expression signature similar to the basal layer of normal urothelium. They showed the upregulation of p63 target genes, the enrichment for TP53 and RB1 mutations, and overexpression of CD49, Cyclin B1, and EGFR. Survival analyses showed that the muscle-invasive basal bladder cancers were more aggressive when compared to luminal cancers. The immunohistochemical expressions of only two markers, luminal (GATA3) and basal (KRT5/6), were sufficient to identify the molecular subtypes of bladder cancer with over 90% accuracy.

CONCLUSIONS

The molecular subtypes of bladder cancer have distinct clinical behaviors and sensitivities to chemotherapy, and a simple two-marker immunohistochemical classifier can be used for prognostic and therapeutic stratification.

BACKGROUND

U.S. National Cancer Institute and National Institute of Health.

The 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors or statins are newly identified immunomodulators. In vivo treatment of SJL/J mice with lovastatin reduced the duration and clinical severity of active and passive experimental autoimmune encephalomyelitis (EAE), an animal model for multiple sclerosis. Lovastatin induced the expression of GATA3 and the phosphorylation of STAT6, whereas it inhibited tyrosine phosphorylation of Janus kinase 2, tyrosine kinase 2, and STAT4. Inhibition of the Janus kinase-STAT4 pathway by lovastatin modulated T0 to Th1 differentiation and reduced cytokine (IFN-gamma and TNF-alpha) production, thus inducing Th2 cytokines (IL-4, IL-5, and IL-10). It inhibited T-bet (T box transcription factor) and NF-kappaB in activated T cells and significantly reduced infiltration of CD4- and MHC class II-positive cells to CNS. Further, it stabilized IL-4 production and GATA-3 expression in differentiated Th2 cells, whereas in differentiated Th1 cells it inhibited the expression of T-bet and reduced the production of IFN-gamma. Moreover, lovastatin-exposed macrophage and BV2 (microglia) in allogeneic MLRs induced the production of the anti-inflammatory cytokine IL-10. These observations indicate that the anti-inflammatory effects of lovastatin are mediated via T cells as well as APCs, because it modulates the polarization patterns of naive T cell activation in an APC-independent system. Together, these findings reveal that lovastatin may have possible therapeutic value involving new targets (in both APCs and T cells) for the treatment of multiple sclerosis and other inflammatory diseases.

T helper 2 cells regulate inflammatory responses to helminth infections while also mediating pathological processes of asthma and allergy. IL-4 promotes Th2 development by inducing the expression of the GATA3 transcription factor, and the Th2 phenotype is stabilized by a GATA3-dependent autoregulatory loop. In this study, we found that type I IFN (IFN-alpha/beta) blocked human Th2 development and inhibited cytokine secretion from committed Th2 cells. This negative regulatory pathway was operative in human but not mouse CD4(+) T cells and was selective to type I IFN, as neither IFN-gamma nor IL-12 mediated such inhibition. IFN-alpha/beta blocked Th2 cytokine secretion through the inhibition of GATA3 during Th2 development and in fully committed Th2 cells. Ectopic expression of GATA3 via retrovirus did not overcome IFN-alpha/beta-mediated inhibition of Th2 commitment. Thus, we demonstrate a novel role for IFN-alpha/beta in blocking Th2 cells, suggesting its potential as a promising therapy for atopy and asthma.

The GATA family members are zinc finger transcription factors involved in cell differentiation and proliferation. GATA3 in particular is necessary for mammary gland maturation, and its loss has been implicated in breast cancer development. Our goal was to validate the ability of GATA3 expression to predict survival in breast cancer patients. Protein expression of GATA3 was analyzed on a high-density tissue microarray consisting of 242 cases of breast cancer. We associated GATA3 expression with patient outcomes and clinicopathologic variables. Expression of GATA3 was significantly increased in breast cancer, in situ lesions, and hyperplastic tissue compared with normal breast tissue. GATA3 expression decreased with increasing tumor grade. Low GATA3 expression was a significant predictor of disease-related death in all patients, as well as in subgroups of estrogen receptor-positive or low-grade patients. In addition, low GATA3 expression correlated with increased tumor size and estrogen and progesterone receptor negativity. GATA3 is an important predictor of disease outcome in breast cancer patients. This finding has been validated in a diverse set of populations. Thus, GATA3 expression has utility as a prognostic indicator in breast cancer.

Interleukin-4 (IL-4) is crucial for the differentiation of naive T helper (T(H)) cells into the T(H)2 effector cells that promote humoral (antibody) immunity and provide protection against intestinal helminths. IL-4 also has a central role in the pathogenesis of allergic inflammation. Many transcription factors are involved in the regulation of expression of the gene encoding IL-4. Initiation of transcription of the gene encoding IL-4 in naive T(H) cells is regulated by the T(H)2-specific transcription factor GATA3, whereas acute expression of the gene encoding IL-4 in T(H)2 cells is mediated by inducible, ubiquitous transcription factors after antigen encounter. This review focuses on acute activation of the gene encoding IL-4 in T cells and discusses therapeutic perspectives at the transcriptional level.

The double-positive (DP) to single-positive (SP) transition during T cell development is initiated by downregulation of the E protein transcription factors HEB and E2A. Here, we have demonstrated that in addition to regulating the onset of this transition, HEB and E2A also play a separate role in CD4(+) lineage choice. Deletion of HEB and E2A in DP thymocytes specifically blocked the development of CD4(+) lineage T cells. Furthermore, deletion of the E protein inhibitors Id2 and Id3 allowed CD4(+) T cell development but blocked CD8(+) lineage development. Analysis of the CD4(+) lineage transcriptional regulators ThPOK and Gata3 placed HEB and E2A upstream of CD4(+) lineage specification. These studies identify an important role for E proteins in the activation of CD4(+) lineage differentiation as thymocytes undergo the DP to SP transition.

Cross-talk between TGF-beta and IL-6 has been shown to direct the differentiation of CD4(+) cells into special IL-17-secreting cells, which are termed Th17 cells. In this study, we demonstrated that TGF-beta and IL-6 could stimulate CD8(+) cells to differentiate into noncytotoxic, IL-17-producing cells in MLC. These IL-17-producing CD8(+) cells exhibit a unique granzyme B(-)IFN-gamma(-)IL-10(-) phenotype. The mRNA level of Th2/T cytotoxic 2 (Tc2) transcription factors GATA3 and Th1/Tc1 transcription factors T-box expressed in T cell (T-bet) as well as its target H2.O-like homeobox (Hlx) is decreased in CD8(+) cells from TGF-beta- and IL-6-treated MLC. In addition, these CD8(+) cells display a marked up-regulation of retinoic acid-related orphan receptor-gammat, a key IL-17 transcription factor. These results demonstrate that the existence of an IL-17-producing CD8(+) subset belongs to neither the Tc1 nor the Tc2 subset and can be categorized as a T noncytotoxic 17 (Tnc17) subset.

The transcription factor Gata3 is crucially involved in epidermis and hair follicle differentiation. Yet, little is known about how Gata3 co-ordinates stem cell lineage determination in skin, what pathways are involved and how Gata3 differentially regulates distinct cell populations within the hair follicle. Here, we describe a conditional Gata3-/- mouse (K14-Gata3-/-) in which Gata3 is specifically deleted in epidermis and hair follicles. K14-Gata3-/- mice show aberrant postnatal growth and development, delayed hair growth and maintenance, abnormal hair follicle organization and irregular pigmentation. After the first hair cycle, the germinative layer surrounding the dermal papilla was not restored; instead, proliferation was pronounced in basal epidermal cells. Transcriptome analysis of laser-dissected K14-Gata3-/- hair follicles revealed mitosis, epithelial differentiation and the Notch, Wnt and BMP signaling pathways to be significantly overrepresented. Elucidation of these pathways at the RNA and protein levels and physiologic endpoints suggests that Gata3 integrates diverse signaling networks to regulate the balance between hair follicle and epidermal cell fates.

CD4+ T cells with a block in the NF-kappaB signaling pathway exhibit decreases in Th1 responses and diminished nuclear levels of multiple transactivating NF-kappaB/Rel/IkappaB proteins. To determine the lineage-intrinsic contributions of these transactivators to Th differentiation, T cells from mice deficient in specific subunits were cultured in exogenous cytokines promoting either Th1 or Th2 differentiation. RelB-deficient cells exhibited dramatic defects in Th1 differentiation and IFN-gamma production, whereas no consistent defect in either Th1 or Th2 responses was observed with c-Rel-deficient cells. In sharp contrast, Bcl-3-null T cells displayed no defect in IFN-gamma production, but their Th2 differentiation and IL-4, IL-5, and IL-13 production were significantly impaired. The absence of RelB led to a dramatic decrease in the expression of T-box expressed in T cells and Stat4. In contrast, Bcl-3-deficient cells exhibited decreased GATA-3, consistent with evidence that Bcl-3 can transactivate a gata3 promoter. These data indicate that Bcl-3 and RelB exert distinct and opposing effects on the expression of subset-determining transcription factors, suggesting that the characteristics of Th cell responses may be regulated by titrating the stoichiometry of transactivating NF-kappaB/Rel/IkappaB complexes in the nuclei of developing helper effector cells.

Insm1 (IA-1) encodes a Zn-finger factor that is expressed in the developing nervous system. We demonstrate here that the development of the sympatho-adrenal lineage is severely impaired in Insm1 mutant mice. Differentiation of sympatho-adrenal precursors, as assessed by the expression of neuronal subtype-specific genes such as Th and Dbh, is delayed in a pronounced manner, which is accompanied by a reduced proliferation. Sympathetic neurons eventually overcome the differentiation blockade and mature correctly, but sympathetic ganglia remain small. By contrast, terminal differentiation of adrenal chromaffin cells does not occur. The transcription factors Mash1 (Ascl1), Phox2a, Gata3 and Hand2 (previously dHand) control the differentiation of sympatho-adrenal precursor cells, and their deregulated expression in Insm1 mutant mice demonstrates that Insm1 acts in the transcriptional network that controls differentiation of this lineage. Pronounced similarities between Mash1 and Insm1 phenotypes are apparent, which suggests that Insm1 might mediate aspects of Mash1 function in the subtype-specific differentiation of sympatho-adrenal precursors. Noradrenaline is the major catecholamine produced by developing sympatho-adrenal cells and is required for fetal survival. We demonstrate that the fetal lethality of Insm1 mutant mice is caused by catecholamine deficiency, which highlights the importance of Insm1 in the development of the sympatho-adrenal lineage.

Palmitoylation is a key post-translational modification mediated by a family of DHHC-containing palmitoyl acyl-transferases (PATs). Unlike other lipid modifications, palmitoylation is reversible and thus often regulates dynamic protein interactions. We find that the mouse hair loss mutant, depilated, (dep) is due to a single amino acid deletion in the PAT, Zdhhc21, resulting in protein mislocalization and loss of palmitoylation activity. We examined expression of Zdhhc21 protein in skin and find it restricted to specific hair lineages. Loss of Zdhhc21 function results in delayed hair shaft differentiation, at the site of expression of the gene, but also leads to hyperplasia of the interfollicular epidermis (IFE) and sebaceous glands, distant from the expression site. The specific delay in follicle differentiation is associated with attenuated anagen propagation and is reflected by decreased levels of Lef1, nuclear beta-catenin, and Foxn1 in hair shaft progenitors. In the thickened basal compartment of mutant IFE, phospho-ERK and cell proliferation are increased, suggesting increased signaling through EGFR or integrin-related receptors, with a parallel reduction in expression of the key differentiation factor Gata3. We show that the Src-family kinase, Fyn, involved in keratinocyte differentiation, is a direct palmitoylation target of Zdhhc21 and is mislocalized in mutant follicles. This study is the first to demonstrate a key role for palmitoylation in regulating developmental signals in mammalian tissue homeostasis.

OBJECTIVE

To investigate the potential therapeutic effects of mesenchymal stem cells (MSCs) in inflammatory bowel disease (IBD), we transplanted MSCs into an experimental model of IBD.

METHODS

A rectal enema of trinitrobenzene sulfonic acid (TNBS) (100 mg/kg body weight) was administered to female BALB/c mice. Bone marrow mesenchymal stem cells (BMSCs) were derived from male green fluorescent protein (GFP) transgenic mice and were transplanted intravenously into the experimental animals after disease onset. Clinical activity scores and histological changes were evaluated. GFP and Sex determining region Y gene (SRY) expression were used for cell tracking. Ki67 positive cells and Lgr5-expressing cells were determined to measure proliferative activity. Inflammatory response was determined by measuring the levels of different inflammatory mediators in the colon and serum. The inflammatory cytokines included tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), interleukin-2 (IL-2), IL-6, IL-17, IL-4, IL-10, and transforming growth factor (TGF-β). Master regulators of Th1 cells (T-box expressed in T cells, T-bet), Th17 cells (retinoid related orphan receptor gamma(t), RORγt), Th2 cells (GATA family of transcription factors 3, GATA3) and regulatory T cells (forkhead box P3, Foxp3) were also determined.

RESULTS

Systemic infusion of GFP-BMSCs ameliorated the clinical and histopathologic severity of colitis, including body weight loss, diarrhea and inflammation, and increased survival (P < 0.05). The cell tracking study showed that MSCs homed to the injured colon. MSCs promoted proliferation of intestinal epithelial cells and differentiation of intestinal stem cells (P < 0.01). This therapeutic effect was mainly mediated by down-regulation of both Th1-Th17-driven autoimmune and inflammatory responses (IL-2, TNF-α, IFN-γ, T-bet; IL-6, IL-17, RORγt), and by up-regulation of Th2 activities (IL-4, IL-10, GATA-3) (P < 0.05). MSCs also induced activated CD4(+)CD25(+)Foxp3(+) regulatory T cells (TGF-β, IL-10, Foxp3) with a suppressive capacity on Th1-Th17 effecter responses and promoted Th2 differentiation in vivo (P < 0.05).

CONCLUSIONS

MSCs are key regulators of immune and inflammatory responses and may be an attractive candidate for cell-based therapy of IBD.

Over the past years, increasing numbers of distinct subsets have been discovered and identified for a T lymphocytes' entity. Differentiation and function of each T cell subset are controlled by a specific master transcription factor. Importantly, Runt-related transcription factors, particularly Runx1 and Runx3, interplay with these master regulators in various aspects of T cells' immunity. In this review article, we first explain roles of Th-Pok and Runx3 in differentiation of CD4 versus CD8 single positive cells, and later focus on cross-regulation of Th-Pok and Runx3 and their relationship with other factors such as TCR strength. Next, we provide evidences for the direct interplay of Runx1/3 with T-bet and GATA3 during Th1 versus Th2 commitment to activate or silence transcription of signature cytokine genes, IFNγ and IL4. Lastly, we explain feed-forward relationship between Runx1 and Foxp3 and discuss roles of Runx1 in regulatory T cells' suppressive activity. This review highlights an essential importance of Runx molecules in controlling various T cell subsets' differentiation and functions through molecular interplay with the master transcription factors in terms of protein-protein interaction as well as regulation of gene expression.

The TCR signals for the release of CD4 effector function are poorly understood. Itk plays an essential role in Th2, but not Th1, responses. However, when Itk is required during Th2 development is unclear. We followed the fate of Itk-deficient T cells during Th2 development in vitro and in vivo using an IL-4/GFP reporter. Surprisingly, a similar frequency of itk(-/-) CD4(+) cells differentiated and committed to the Th2 lineage as wild-type cells. However, Itk-deficient Th2 cells failed to exert effector function upon TCR triggering. Loss of function was marked by defective transcriptional enhancement of Th2 cytokines and GATA3. IL-4 production in itk(-/-) Th2s could be rescued by the expression of kinase-active Itk. Thus, Itk is necessary for the release, but not gain, of Th2 function. We suggest that the liberation of effector function is tightly controlled through qualitative changes in TCR signals, facilitating postdifferentiation regulation of cytokine responses.