GATA binding protein 3 (GATA3) is a transcription factor, which belongs to a distinct family of tumor suppressor genes. It is involved in human cancer cell growth and differentiation, and plays an important role in cell proliferation and apoptosis. Although, its expression has been reported in various cancers, there are limited data in genitourinary malignancies. Recent studies found GATA3 to be a sensitive marker for urothelial carcinoma (UC) and associated with prognostic pathologic features. Its level of expression was found to be an independent factor predicting cancer recurrence.

In this article, immunohistochemical evaluation of GATA3 expression in genitourinary malignancies (invasive UC, renal cell carcinoma, and prostatic adenocarcinomas) was performed.

GATA3 was positive in 56/79 (70.8%) of invasive UC, and was negative in all renal cell carcinoma and prostatic adenocarcinomas. The pattern of GATA3 staining, when positive, was intensely nuclear within the clusters of malignant cells. No cytoplasmic staining was noted. Negative controls were all negative. High GATA3 expression was associated with larger tumor size in invasive UC (3.19cm vs. 1.65cm, P = 0.01). GATA3 expression did not correlate with other clinicopathologic parameters in UC.

This data suggest that GATA3 is a sensitive marker in confirming invasive UC, and may be helpful in differentiating it from metastatic tumors of renal and prostatic origin. Furthermore, strong GATA3 expression was noted to have an effect on tumor size in patients with UC.

Helminths and their products can shape immune responses by modulating immune cells, which are dysfunctional in inflammatory diseases such as asthma. We previously identified SJMHE1, a small molecule peptide from the HSP60 protein of Schistosoma japonicum. SJMHE1 can inhibit delayed-type hypersensitivity and collagen-induced arthritis in mice. In the present study, we evaluated this peptide's potential intervention effect and mechanism on ovalbumin-induced asthma in mice. SJMHE1 treatment suppressed airway inflammation in allergic mice, decreased the infiltrating inflammatory cells in the lungs and bronchoalveolar lavage fluid, modulated the production of pro-inflammatory and anti-inflammatory cytokines in the splenocytes and lungs of allergic mice, reduced the percentage of Th2 cells and increased the proportion of Th1 and regulatory T cells (Tregs). At the same time, Foxp3 and T-bet expression increased, and GATA3 and RORγt decreased in the lungs of allergic mice. We proved that SJMHE1 can interrupt the development of asthma by diminishing airway inflammation in mice. The down-regulation of Th2 response and the up-regulation of Th1 and Tregs response may contribute to the protection induced by SJMHE1 in allergic mice. SJMHE1 can serve as a novel therapy for asthma and other allergic or inflammatory diseases.

Trophectoderm (TE) lineage development is pivotal for proper implantation, placentation, and healthy pregnancy. However, only a few TE-specific transcription factors (TFs) have been systematically characterized, hindering our understanding of the process. To elucidate regulatory mechanisms underlying TE development, here we map super-enhancers (SEs) in trophoblast stem cells (TSCs) as a model. We find both prominent TE-specific master TFs (Cdx2, Gata3, and Tead4), and >150 TFs that had not been previously implicated in TE lineage, that are SE-associated. Mapping targets of 27 SE-predicted TFs reveals a highly intertwined transcriptional regulatory circuitry. Intriguingly, SE-predicted TFs show 4 distinct expression patterns with dynamic alterations of their targets during TSC differentiation. Furthermore, depletion of a subset of TFs results in dysregulation of the markers for specialized cell types in placenta, suggesting a role during TE differentiation. Collectively, we characterize an expanded TE-specific regulatory network, providing a framework for understanding TE lineage development and placentation.

Neuroendocrine breast carcinomas represent a rare subtype of breast cancer. Their definition, prevalence, and prognosis remain controversial in the literature. The 2012 WHO classification of breast cancer categorizes neuroendocrine carcinomas into three morphologically distinct subtypes: well-differentiated neuroendocrine tumors, poorly differentiated neuroendocrine carcinomas, and invasive breast carcinomas with neuroendocrine differentiation. We aimed to gain insight into the clinical, morphologic, phenotypic, and molecular features of 47 neuroendocrine breast carcinomas. Targeted next-generation sequencing by an AmpliSeq 22 cancer gene hotspot panel and the Prosigna assay were performed on 42/47 and 35/47 cases, respectively. Average age at diagnosis was 69 years. All tumors were estrogen receptor-positive and the large majority expressed progesterone receptor (89%), GATA3 (98%), FOXA1 (96%), and CK8/18 (98%). There was an almost equal distribution of luminal A (52%) and B (48%) carcinomas. Almost half of the cohort (49%) displayed a high risk of recurrence score with the Prosigna test. Patients with a neuroendocrine carcinoma had a shorter disease-free survival compared with those affected by carcinomas of no special type matched for age, size, grade, and estrogen receptor status. No significant differences were observed in terms of overall survival. Stratification of neuroendocrine carcinomas using the 2012 WHO criteria did not reveal statistically significant differences among the distinct categories (well-differentiated neuroendocrine tumors, poorly differentiated neuroendocrine carcinomas, and invasive breast carcinomas with neuroendocrine differentiation), in terms of either progression-free or overall survival. Our targeted sequencing analysis found three cases (7%) harboring a PIK3CA mutation, and in three other cases (7%) TP53 mutations were detected. This study showed that neuroendocrine breast carcinoma is a distinct subtype of luminal carcinoma with a low rate of PIK3CA mutations and with an aggressive clinical behavior. An accurate identification of neuroendocrine differentiation may be useful to better tailor patient adjuvant therapy within luminal carcinomas.

Allergic asthma is a chronic inflammatory disease induced by the inhalation of allergens, which trigger the activation of T helper type 2 (Th2) cells that release Th2 cytokines. Recently, herbal medicines are being considered a major source of novel agents to treat various diseases. In the present study, we evaluated the anti-asthmatic effects of a Codonopsis lanceolata extract (CLE) and the mechanisms involved in its anti-inflammatory effects. Treatment with CLE reduced infiltration of inflammatory cells, especially eosinophils, and the production of mucus in lung tissues. Levels of Th2 cytokines, such as IL-4, IL-5, and IL-13, and chemokines were also decreased following treatment with CLE. Moreover, Th2 cell proportion in vivo and differentiation in vitro were reduced as evidenced by the decreased expression of GATA3⁺. Furthermore, the expression of superoxide dismutase (SOD)2, a mitochondrial ROS (mROS) scavenger, was increased, which was related to Th2 cell regulation. Interestingly, treatment with CLE increased the number of macrophages in the lungs and enhanced the immune-suppressive property of macrophages. Our findings indicate that CLE has potential as a novel therapeutic agent to inhibit Th2 cell differentiation by regulating mROS scavenging.

Estrogen receptor (ER)-positive progesterone receptor (PR)-negative breast cancers are infrequent but clinically challenging. Despite the volume of genomic data available on these tumors, their biology remains poorly understood. Here, we aimed to identify clinically relevant subclasses of ER+/PR- breast cancers based on their mutational landscape. The Cancer Genomics Data Server was interrogated for mutational and clinical data of all ER+ breast cancers with information on PR status from The Cancer Genome Atlas (TCGA), Memorial Sloan Kettering (MSK), and Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) projects. Clustering analysis was performed using gplots, ggplot2, and ComplexHeatmap packages. Comparisons between groups were performed using the Student's t-test and the test of Equal or Given Proportions. Survival curves were built according to the Kaplan⁻Meier method; differences in survival were assessed with the log-rank test. A total of 3570 ER+ breast cancers (PR- n = 959, 27%; PR+ n = 2611, 73%) were analyzed. Mutations in well-known cancer genes such as TP53, GATA3, CDH1, HER2, CDH1, and BRAF were private to or enriched for in PR- tumors. Mutual exclusivity analysis revealed the presence of four molecular clusters with significantly different prognosis on the basis of PIK3CA and TP53 status. ER+/PR- breast cancers are genetically heterogeneous and encompass a variety of distinct entities in terms of prognostic and predictive information.

Much research effort has been focused on investigating new compounds derived from low-cost sources, such as natural products, for treating leishmaniasis. Oleuropein derived from numerous plants, particularly from the olive tree, Olea europaea L. (Oleaceae), is a biophenol with many biological activities. Our previous findings showed that oleuropein exhibits leishmanicidal effects against three Leishmania spp. in vitro, and minimizes the parasite burden in L. donovani-infected BALB/c mice. The aim of the present study is to investigate the possible mechanism(s) that mediate this leishmanicidal activity.

We determined the efficacy of oleuropein in elevating ROS and NO production in L. donovani-infected J774A.1 macrophages and in explanted splenocytes and hepatocytes obtained from L. donovani-infected BALB/c mice. We also assessed the expression of genes that are related to inflammation, T-cell polarization and antioxidant defense, in splenocytes. Finally, we determined the ratios of specific IgG2a/IgG1 antibodies and DTH reactions in L. donovani-infected BALB/c mice treated with oleuropein.

Oleuropein was able to elevate ROS production in both in vitro and in vivo models of visceral leishmaniasis and raised NO production in ex vivo cultures of splenocytes and hepatocytes. The extensive oxidative stress found in oleuropein-treated mice was obviated by the upregulation of the host's antioxidant enzyme (mGCLC) and the simultaneous downregulation of the corresponding enzyme of the parasite (LdGCLC). Moreover, oleuropein was able to mount a significant Th1 polarization characterized by the expression of immune genes (IL-12β, IL-10, TGF-β1, IFN-γ) and transcription factors (Tbx21 and GATA3). Moreover, this immunomodulatory effect was also correlated with an inhibitory effect on IL-1β gene expression, rather than with the expression of IL-1α, IL-1rn and TNF-α. Furthermore, oleuropein-treated BALB/c mice mounted a delayed-type hypersensitivity (DTH) response and an elevated Leishmania-specific IgG2a/IgG1 ratio that clearly demonstrated an in vivo protective mechanism.

The ability of Oleuropein to promote a Th1 type immune response in L. donovani-infected BALB/c mice points towards the candidacy of this bioactive compound as an immunomodulatory agent that may complement therapeutic approaches to leishmaniasis.

Estrogen receptor-positive, progesterone receptor-negative, and human epidermal growth factor receptor 2 (HER2)-negative (ER+PR-HER2-) breast cancer comprise a special type of breast cancer that constitutes ~10% of all breast cancer patients. ER+PR-HER2- tumor benefits less from endocrine therapy, while its genomic features remain elusive. In this study, we systematically assessed the multiomic landscape and endocrine responsiveness of ER+PR-HER2- breast cancer. Methods: This study incorporated five cohorts. The first and second cohorts were from the Surveillance, Epidemiology, and End Results database (n=130,856) and Molecular Taxonomy of Breast Cancer International Consortium (n=1,055) for analyzing survival outcomes and endocrine responsiveness. The third cohort was from The Cancer Genome Atlas (n=630) for multiomic analysis and endocrine-resistant subgroup exploration. The fourth cohort, from the MD Anderson database (n=92), was employed to assist gene selection. The fifth cohort was a prospective observational cohort from Fudan University Shanghai Cancer Center (n=245) that was utilized to validate the gene-defined subgroup by immunohistochemistry (IHC). Results: Clinically, ER+PR-HER2- tumors showed lower endocrine responsiveness than did ER+PR+HER2- tumors. Genomically, copy number loss or promoter methylation of PR genes occurred in 75% of ER+PR-HER2- tumors, collectively explaining PR loss. ER+PR-HER2- tumors had higher TP53 (30.3% vs. 17.0%) and lower PIK3CA mutation rates (25.8% vs. 42.7%) and exhibited more ZNF703 (21.5% vs. 13.6%) and RPS6KB1 (18.5% vs. 7.8%) amplification events than ER+PR+HER2- tumors. Among ER+PR-HER2- tumors, nearly 20% were of the PAM50-defined non-luminal-like subgroup and manifested lower endocrine sensitivity scores and enriched biosynthesis, metabolism and DNA replication pathways. We further identified the non-luminal-like subgroup using three IHC markers, GATA3, CK5, and EGFR. These IHC-defined non-luminal-like (GATA3-negative, CK5-positive and/or EGFR-positive) tumors received limited benefit from adjuvant endocrine therapy. Conclusion: ER+PR-HER2- breast cancer consists of clinically and genomically distinct groups that may require different treatment strategies. The non-luminal-like subgroup was associated with reduced benefit from endocrine therapy.

Recent work indicates that the nuclear envelope is a major signaling node for the cell that can influence tissue differentiation processes. Here we present two nuclear envelope trans-membrane proteins TMEM120A and TMEM120B that are paralogs encoded by the Tmem120A and Tmem120B genes. The TMEM120 proteins are expressed preferentially in fat and both are induced during 3T3-L1 adipocyte differentiation. Knockdown of one or the other protein altered expression of several genes required for adipocyte differentiation, Gata3, Fasn, Glut4, while knockdown of both together additionally affected Pparg and Adipoq. The double knockdown also increased the strength of effects, reducing for example Glut4 levels by 95% compared to control 3T3-L1 cells upon pharmacologically induced differentiation. Accordingly, TMEM120A and B knockdown individually and together impacted on adipocyte differentiation/metabolism as measured by lipid accumulation through binding of Oil Red O and coherent anti-Stokes Raman scattering microscopy (CARS). The nuclear envelope is linked to several lipodystrophies through mutations in lamin A; however, lamin A is widely expressed. Thus it is possible that the TMEM120A and B fat-specific nuclear envelope transmembrane proteins may play a contributory role in the tissue-specific pathology of this disorder or in the wider problem of obesity.

Immunohistochemistry of a transcription factor, GATA3, has been widely used as a promising urothelial marker in diagnostic surgical pathology practice. However, the expression status of GATA3 in upper urinary tract urothelial carcinomas (UUTUCs) and its prognostic significance have not been fully investigated. We immunohistochemically stained for GATA3 in 99 UUTUC samples and paired nonneoplastic urothelial tissues. GATA3 was positive in 51 (51.5%; 32 [32.3%] weak, 11 [11.1%] moderate, 8 [8.1%] strong) of 99 UUTUCs, which was significantly lower than in benign urothelium (79 [96.3%] of 82; 33 [40.2%] weak, 35 [42.7%] moderate, 11 [13.4%] strong; P<.001). However, there were no statistically significant associations between GATA3 expression and tumor grade, pT stage, lymph node involvement, or distant metastasis. Meanwhile, the rate of GATA3 positivity was significantly higher (P=.004) in ureteral tumors (66.0%) than in renal pelvic tumors (35.6%). Kaplan-Meier and log-rank tests revealed that GATA3 negativity was significantly associated with lower recurrence-free survival (P=.037 for all cases, P=.026 for muscle-invasive tumors) and cancer-specific survival (P=.007 for all cases, P=.012 for muscle-invasive tumors, P=.035 for cases with adjuvant chemotherapy) rates. Multivariate analysis further identified strong correlations of GATA3 expression with tumor progression (all cases: hazard ratio [HR], 0.479 [95% confidence interval {CI},0.229-1.003; P=.051]; muscle-invasive tumors: HR, 0.387 [95% CI, 0.166-0.903; P=.028) or cancer-specific mortality (all cases: HR, 0.354 [95% CI, 0.135-0.925; P=.034]; muscle-invasive tumors: HR, 0.402 [95% CI, 0.149-1.086; P=.072]). Thus, compared with nonneoplastic urothelium, a significant decrease in the expression of GATA3 in UUTUC was seen. Moreover, loss of GATA3 expression was found to be an independent predictor of poor patient outcomes. Of note was that only roughly half of high-grade and/or muscle-invasive UUTUCs were immunoreactive for GATA3.

In metastatic breast cancer (MBC), it can be difficult to establish the origin if the primary tumor is triple negative or if there is a loss of biomarker expression. SOX10 expression has been reported in primary triple negative breast cancer but is poorly studied in metastatic lesions. In this study, the diagnostic utility of a panel of SOX10, GATA3, and androgen receptor (AR) in MBC negative for estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) was evaluated and compared to the expression of these markers in the matched primary breast cancer (PBC). In a series of 57 triple negative MBC, 82% were positive for GATA3, 58% for SOX10, and 25% for AR. Nearly all MBC (95%) were positive for either GATA3 or SOX10, with 46% dual positive and 5% of cases negative for both markers. Most GATA3 negative MBC cases were SOX10 positive (70%). AR expression was only seen in GATA3 positive MBC (25%), and was significantly more frequent in SOX10 negative MBC (48%) vs. SOX10 positive MBC (9%, P=.001). Concordance for GATA3, SOX10, and AR between the primary and metastasis was 89%, 88%, and 80% respectively. While GATA3 is a more sensitive lineage marker than SOX10 in MBC, SOX10 is a useful adjunct because it is positive in the majority of GATA3 negative breast metastases. Utilizing both GATA3 and SOX10 is recommended for confirming breast as the site of origin in metastases that lack ER, PR, and HER2 expression whereas the addition of AR is not helpful.

Male breast cancer (MBC) is rare and poorly characterized. Like the female counterpart, most MBCs are hormonally driven, but relapse after hormonal treatment is also noted. The pan-hormonal action of steroid hormonal receptors, including estrogen receptor alpha (ERα), androgen receptor (AR), progesterone receptor (PR), and glucocorticoid receptor (GR) in this understudied tumor type remains wholly unexamined. This study reveals genomic cross-talk of steroid hormone receptor action and interplay in human tumors, here in the context of MBC, in relation to the female disease and patient outcome. Here we report the characterization of human breast tumors of both genders for cistromic make-up of hormonal regulation in human tumors, revealing genome-wide chromatin binding landscapes of ERα, AR, PR, GR, FOXA1, and GATA3 and enhancer-enriched histone mark H3K4me1. We integrate these data with transcriptomics to reveal gender-selective and genomic location-specific hormone receptor actions, which associate with survival in MBC patients.

In breast cancer, GATA3 mutations have been associated with a favorable prognosis and the response to neoadjuvant aromatase inhibitor treatment. Therefore, we investigated whether GATA3 mutations predict the outcome of tamoxifen treatment in the advanced setting. In a retrospective study consisting of 235 hormone-naive patients with ER-positive breast cancer who received tamoxifen as first-line treatment for recurrent disease, GATA3 mutations (in 14.0% of patients) did not significantly associate with either the overall response rate (ORR) or with the length of progression-free survival (PFS) after the start of tamoxifen therapy. Interestingly, among 148 patients for whom both mutation and mRNA expression data were available, GATA3 mutations associated with an increased expression of GATA3. However, only 23.7% of GATA3 high tumors had a mutation. Evaluation of the clinical significance of GATA3 mRNA revealed that it was associated with prolonged PFS, but not with the ORR, also in multivariate analysis. Thus, GATA3 mRNA expression, but not GATA3 mutation, is an independent predictor of prolonged PFS in ER-positive breast cancer patients who received first-line tamoxifen for recurrent disease. Besides GATA3 mutation, other mechanisms must exist that underlie increased GATA3 levels.

We have previously shown that the in vitro embryonic development and the yield of viable calves were increased by using a two-step chemically defined medium for post-fertilization culture of bovine embryos. In this study, we explored the embryonic development and the temporal behavioral interaction of the genes involved in IFNτ gene expression and how they behave in an orchestrated manner to increase the developmental competence of IVF produced embryos by culturing in the chemically defined medium. Behavior of genes included ETS2, CDX2, GATA2, GATA3, OCT4 and NANOG was analyzed in early bovine IVF produced embryos, (from compact morulae to the blastocyst hatching stages), by semi- and relative quantitative PCR and compared between two in vitro culture (IVC) systems, two-step chemically defined medium and modified synthetic oviductal fluid (mSOF) containing 8 mg/mL, BSA. Early embryonic development was found to be better in two-step chemically defined culture system than that of mSOF as indicated by the increment of blastocyst yield, 33.1% in two-step culture system vs 18.8% in mSOF medium, and the blastocyst hatching, 52.3% in two-step culture system vs 33.5% in mSOF medium. Relative quantitative gene expression showed harmonic behavior in the two-step culture system rather than the culture in mSOF, IFNτ showed even increase throughout the embryonic development in the two-step culture medium while it decreased with blastocyst hatching in mSOF culture condition. Temporal dominance of OCT4 over all the transcription factors was found in regulation of IFNτ expression (the major factor of expression regulation but in inverse manner). However, ETS2, CDX2, GATA2 and GATA3 are potent IFNτ stimulator in cumulative manner but in case of OCT4 decrement. CDX2 directly related with IFNτ, but still under OCT4 dominance and also regulated by the subservient of OCT4 which is NANOG. In conclusion, this study confirmed our previous results about the usefulness of using the two-step chemically defined culture medium for increasing the developmental competence of IVF produced embryos and elucidated the dominance of OCT4 over the other genes implicated in regulation of IFNτ expression.

BACKGROUND

Th2 cells are crucially important in allergic disease and the possible involvement of Treg and Th17 cells has not been clearly identified.

OBJECTIVE

To identify the mRNA expression of T cell transcription factors in nasal mucosa in patients with allergic rhinitis (AR) and to reveal their correlations with clinical features.

METHODS

Eighteen patients with AR and 12 controls with turbinate hypertrophy were included. mRNA expression of the following transcriptional factors in nasal mucosa were measured by quantitative polymerase chain reaction; T-bet (Th1), GATA3 (Th2), retinoic acid-related orphan receptor C (RORC; Th17), and forkhead box P3 (Foxp3; Treg). mRNA expression was compared among groups and correlation between mRNA expression level and clinical features (rhinitis symptoms, eosinophil count, and IgE) were also investigated.

RESULTS

GATA3 and RORC were significantly increased and Foxp3 was significantly decreased in the AR group. Moderate-to-severe AR group also had increased expression of GATA3 and RORC than mild AR group, suggesting severity of AR influence expression of transcription factors. Correlation analysis showed that none of these transcription factors were associated with severity of clinical symptoms, eosinophil counts and skin prick test severity and that IgE level was significantly correlated with expression level of GATA3 and RORC, suggesting an association of IgE production with Th2 and Th17 cells.

CONCLUSIONS

Increased mRNA expression of GATA3 (Th2), increased expression of RORC and decreased expression of Foxp3 may be important in pathogenesis of AR. GATA3 and RORC may be closed related with IgE level.

GABAergic neurons in the ventral mesodiencephalic region are highly important for the function of dopaminergic pathways that regulate multiple aspects of behavior. However, development of these neurons is poorly understood. We recently showed that molecular regulation of differentiation of the GABAergic neurons associated with the dopaminergic nuclei in the ventral midbrain (VTA and SNpr) is distinct from the rest of midbrain, but the reason for this difference remained elusive. Here, we have analyzed the developmental origin of the VTA and SNpr GABAergic neurons by genetic fate mapping. We demonstrate that the majority of these GABAergic neurons originate outside the midbrain, from rhombomere 1, and move into the ventral midbrain only as postmitotic neuronal precursors. We further show that Gata2, Gata3 and Tal1 define a subpopulation of GABAergic precursors in ventral rhombomere 1. A failure in GABAergic neuron differentiation in this region correlates with loss of VTA and SNpr GABAergic neurons in Tal1 mutant mice. In contrast to midbrain, GABAergic neurons of the anterior SNpr in the diencephalon are not derived from the rhombomere 1. These results suggest unique migratory pathways for the precursors of important GABAergic neuron subpopulations, and provide the basis for understanding diversity within midbrain GABAergic neurons.

BACKGROUND

Conventional karyotyping (550 bands resolution) is able to identify chromosomal aberrations >5-10 Mb, which represent a known cause of intellectual disability/developmental delay (ID/DD) and/or multiple congenital anomalies (MCA). Array-Comparative Genomic Hybridization (array-CGH) has increased the diagnostic yield of 15-20%.

RESULTS

In a cohort of 700 ID/DD cases with or without MCA, including 15 prenatal diagnoses, we identified a subgroup of seven patients with a normal karyotype and a large complex rearrangement detected by array-CGH (at least 6, and up to 18 Mb). FISH analysis could be performed on six cases and showed that rearrangements were translocation derivatives, indistinguishable from a normal karyotype as they involved a similar band pattern and size. Five were inherited from a parent with a balanced translocation, whereas two were apparently de novo. Genes spanning the rearrangements could be associated with some phenotypic features in three cases (case 3: DOCK8; case 4: GATA3, AKR1C4; case 6: AS/PWS deletion, CHRNA7), and in two, likely disease genes were present (case 5: NR2F2, TP63, IGF1R; case 7: CDON). Three of our cases were prenatal diagnoses with an apparently normal karyotype.

CONCLUSIONS

Large complex rearrangements of up to 18 Mb, involving chromosomal regions with similar size and band appearance may be overlooked by conventional karyotyping. Array-CGH allows a precise chromosomal diagnosis and recurrence risk definition, further confirming this analysis as a first tier approach to clarify molecular bases of ID/DD and/or MCA. In prenatal tests, array-CGH is confirmed as an important tool to avoid false negative results due to karyotype intrinsic limit of detection.

The Sonic hedgehog (Shh) signaling pathway is crucial for pattern formation in early central nervous system development. By systematically analyzing high-throughput in situ hybridization data of E11.5 mouse brain, we found that Shh and its receptor Ptch1 define two adjacent mutually exclusive gene expression domains: Shh+Ptch1- and Shh-Ptch1+. These two domains are associated respectively with Foxa2 and Gata3, two transcription factors that play key roles in specifying them. Gata3 ChIP-seq experiments and RNA-seq assays on Gata3-knockdown cells revealed that Gata3 up-regulates the genes that are enriched in the Shh-Ptch1+ domain. Important Gata3 targets include Slit2 and Slit3, which are involved in the process of axon guidance, as well as Slc18a1, Th and Qdpr, which are associated with neurotransmitter synthesis and release. By contrast, Foxa2 both up-regulates the genes expressed in the Shh+Ptch1- domain and down-regulates the genes characteristic of the Shh-Ptch1+ domain. From these and other data, we were able to reconstruct a gene regulatory network governing both domains. Our work provides the first genome-wide characterization of the gene regulatory network involved in the Shh pathway that underlies pattern formation in the early mouse brain.

Recent reports have revealed that CD4(+) T(h) cell subsets have the ability to alter their gene expression pattern in response to extracellular stimuli. We previously highlighted the plasticity of T(h)1 cells by demonstrating that T(h)1 cells gain the capacity to produce IL-3, IL-9, IL-13 and granulocyte macrophage colony-stimulating factor in response to antigen, IL-2 and IL-18, and based on their unique function, we designated these activated T(h)1 cells as 'super T(h)1 cells'. However, the precise molecular mechanism underlying IL-13 production by super T(h)1 cells has not been elucidated. Here, we show that the GATA-binding protein 3 (Gata3) is essentially required for II13 gene expression in super T(h)1 cells. Gata3 is synergistically induced in T-box expressed in T-cells (T-bet)-expressing T(h)1 cells when co-stimulated with anti-CD3, IL-18 and IL-4 through the activation of nuclear factor of activated T cells, nuclear factor kappa-light-chain-enhancer of activated B cells and signal transducer and activator of transcription 6, respectively. However, Gata3 induction is not satisfactory, and additional TCR or anti-CD3 signaling is prerequisite for triggering IL-13 production by Gata3 plus T-bet-expressing T(h)1 cells. These findings suggest that Gata3, which is not originally expressed in T(h)1 cells, alters the cytokine production profile by T(h)1 cells.

The inner ear is a complex sensory organ with hearing and balance functions. Gata3 and Gata2 are expressed in the inner ear, and to gain more insight into their roles in otic development, we made a detailed expression analysis in chicken embryos. At early stages, their expression was highly overlapping. At later stages, Gata2 expression became prominent in vestibular and cochlear nonsensory epithelia. In contrast to Gata2, Gata3 was mainly expressed in the developing sensory epithelia, reflecting the importance of this factor in the sensory-neural development of the inner ear. While the later expression patterns of both Gata3 and Gata2 were highly conserved between chicken and mouse, important differences were observed especially with Gata3 during early otic development, providing indications of divergent molecular control during placode invagination in mice and chickens. We also found indications that the regulatory hierarchy observed in mouse, where Gata3 is upstream of Gata2 and Fgf10, could be conserved in chicken.

Differential diagnosis of collecting duct carcinoma (CDC) from invasive upper tract urothelial carcinoma (UTUC) can be challenging. PAX8 and p63 are 2 markers often used in this setting. GATA binding protein 3 (GATA3) is a marker of urothelial differentiation. We investigated GATA3 expression in CDC and UTUC and its use in this differential. Eighteen CDC and 25 UTUC cases were used to build 2 tissue microarrays. GATA3, p63, and PAX8 nuclear expression was evaluated using standard immunohistochemistry. Staining intensity and percentage of positive cells were assessed. Sensitivity, specificity, and positive and negative predictive values of the markers and their combination were also evaluated. We found GATA3 positivity in 22 (88%) of 25 UTUCs and 1 (6%) of 18 CDCs. The median GATA3 extent of expression was higher in UTUC than in CDC (74% versus 0%, P = .00). We found p63 positivity in 23 (92%) of 25 UTUCs and 2 (11%) of 18 CDCs. PAX8 was positive in 3 (12%) of 25 UTUCs and all (100%) CDCs. GATA3 sensitivity and specificity for UTUC were 88% and 94%, respectively. p63 sensitivity and specificity for UTUC were 92% and 89%, respectively. The p63+/PAX8- profile showed higher sensitivity for UTUC than did the GATA3+/PAX8- profile (80% versus 76%). Both showed a specificity of 100% for UTUC. GATA3+ or p63+/PAX8- sensitivity and specificity for UTUC were 84% and 100%, respectively. Immunohistochemical expression of GATA3 was higher in UTUC, suggesting a potential role for distinguishing UTUC from CDC. Adding this marker to the combination panel of p63 and PAX8 might improve its performance in the diagnosis of epithelial neoplasms involving the renal sinus.

BACKGROUND

Major transcription factors controlling T(H)1 and T(H)2 development, such as T-box transcription factor and GATA3, might be centrally involved in asthma and atopic diseases. Only recently, the homeobox transcription factor H.20-like homeobox 1 (HLX1), interacting closely with T-box transcription factor, has been identified as an important regulator of T(H)1 differentiation and suppressor of T(H)2 commitment.

OBJECTIVE

We investigated whether genetic variations in the HLX1 gene exist and whether these could affect the development of childhood asthma.

METHODS

The HLX1 gene was resequenced in 80 chromosomes. Associations between identified polymorphisms, asthma, and atopic diseases were investigated in German children (total n = 3099) from the cross-sectional International Study of Asthma and Allergy in Childhood phase II. Functional properties of polymorphisms were studied by using luciferase reporter gene assays and electrophoretic mobility shift assays in T cells. All statistical analyses were performed with SAS/Genetics software (SAS Institute, Inc, Cary, NC).

RESULTS

Nineteen polymorphisms were identified in the HLX1 gene, and 2 tagging single nucleotide polymorphisms representing 7 polymorphisms were associated with childhood asthma in our study population. Two promoter polymorphisms, C-1407T and C-742G, contained in 1 tagging block were associated with asthma (odds ratio, 1.44; 95% CI, 1.11-1.86; P = .0061), significantly decrease promoter transactivation, and disrupt specificity protein-transcription factor binding in in vitro experiments.

CONCLUSIONS

Our data suggest that polymorphisms in the HLX1 gene increase the risk for childhood asthma. On the cellular level, altered binding of specificity protein-transcription factors to the HLX1 promoter and subsequent changes in HLX1 gene expression might contribute to these effects.