GATA-binding Factor Research Articles

Integrated analyses of copy number variations and gene differential expression in lung squamous-cell carcinoma.

BackgroundAlthough numerous efforts have been made, the pathogenesis underlying lung squamous-cell carcinoma (SCC) remains unclear. This study aimed to identify the CNV-driven genes by an integrated analysis of both the gene differential expression and copy number variation (CNV).ResultsA higher burden of the CNVs was found in 10–50 kb length. The 16 CNV-driven genes mainly located in chr 1 and chr 3 were enriched in immune response [e.g. complement factor H (CFH) and Fc fragment of IgG, low affinity IIIa, receptor (FCGR3A)], starch and sucrose metabolism [e.g. amylase alpha 2A (AMY2A)]. Furthermore, 38 TFs were screened for the 9 CNV-driven genes and then the regulatory network was constructed, in which the GATA-binding factor 1, 2, and 3 (GATA1, GATA2, GATA3) jointly regulated the expression of TP63.ConclusionsThe above CNV-driven genes might be potential contributors to the development of lung SCC.

GATA-Binding Factor 6 Contributes to Atrioventricular Node Development and Function.

Several transcription factors regulate cardiac conduction system (CCS) development and function but the role of each in specifying distinct CCS components remains unclear. GATA-binding factor 6 (GATA6) is a zinc-finger transcription factor that is critical for patterning the cardiovascular system. However, the role of GATA6 in the embryonic heart and CCS has never been shown. We report that Gata6 is expressed abundantly in the proximal CCS during midgestation in mice. Myocardial-specific deletion of the carboxyl zinc-finger of Gata6 induces loss of hyperpolarizing cyclic nucleotide-gated channel, subtype 4 staining in the compact atrioventricular node with some retention of hyperpolarizing cyclic nucleotide-gated channel, subtype 4 staining in the atrioventricular bundle, but has no significant effect on the connexin-40-positive bundle branches. Furthermore, myocardial-specific deletion of the carboxyl zinc-finger of Gata6 alters atrioventricular conduction in postnatal life as assessed by surface and invasive electrophysiological evaluation, as well as decreasing the number of ventricular myocytes and inducing compensatory myocyte hypertrophy. Myocardial-specific deletion of the carboxyl zinc-finger of Gata6 is also associated with downregulation of the transcriptional repressor ID2 and the cardiac sodium-calcium exchanger NCX1 in the proximal CCS, where GATA6 transactivates both of these factors. Finally, carboxyl zinc-finger deletion of Gata6 reduces cell-cycle exit of TBX3+ myocytes in the developing atrioventricular bundle during the period of atrioventricular node specification, which results in fewer TBX3+ cells in the proximal CCS of mature mutant mice. GATA6 contributes to the development and postnatal function of the murine atrioventricular node by promoting cell-cycle exit of specified cardiomyocytes toward a conduction system lineage.

Identification of differentially expressed microRNAs in the ovary of polycystic ovary syndrome with hyperandrogenism and insulin resistance.

Background:Polycystic ovary syndrome (PCOS) is the commonest endocrinopathy in women of reproductive age. The patients often develop insulin resistance (IR) or hyperinsulinemia despite manifesting anovulation and signs of hyperandrogenism. The cause and effect relationship of hyperinsulinemia and hyperandrogenemia (HA) is still debated. Micro-ribonucleic acids (miRNAs) have recently been shown to play a role in regulation of ovarian function. Our current study focused on the altered expression of miRNAs with PCOS.Methods:Ovarian theca interna tissues were obtained from 10 PCOS patients and 8 controls that were non-PCOS and had normal insulin sensitivity undergoing laparoscopy and/or ovarian wedge resection. Total RNA of all samples was extracted. We studied the repertoire of miRNAs in both PCOS and non-PCOS women by microarray hybridization. Bioinformatic analysis was performed for predicting targets of the differentially expressed miRNAs. Furthermore, selected miRNAs were validated by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR).Results:A total of 27 miRNAs were differentially expressed in PCOS patients with respect to the controls in our discovery evaluationand two (miR-92a and miR-92b) of them were significantly downregulated in PCOS women in followed validation (P < 0.05). Targets prediction revealed that miR-92a targeted both GATA family of zinc finger transcription factor GATA-binding factor 6 (GATA6) and insulin receptor substrate proteins 2 (IRS-2).Conclusions:MiRNAs are differentially expressed between PCOS patients and controls. We identified and validated two miRNAs-miR-92a and miR-92b. They are significantly downregulated and may be involved in the pathogenesis of PCOS.

Functional analysis of Fenneropenaeus chinensis anti-lipopolysaccharide factor promoter regulated by lipopolysaccharide and (1,3)-β-D-glucan

Current knowledge on cis-regulatory elements of immune genes of shrimp is poor. In this study, the genomic sequence of the Fenneropenaeus chinensis anti-lipopolysaccharide factor (ALFFc) gene was obtained by using PCR and genome walking techniques, and the promoter was identified. The ALFFc gene contained three exons interrupted by two introns. Immune-related transcription factor binding sites recognized by nuclear factor-kappa B, octamer binding protein 1, GATA binding factor 1 and specificity protein 1 were identified in the regin from +1 to −702. The activity of ALFFc promoter was analyzed in insect sf9 cell lines. The putative promoter sequence of pALF-702 drive the expression of reporter EGFP gene successfully by adding lipopolysaccharide or (1,3)-β-D-glucan, but the shorter promoter sequence pALF-318 is only by (1,3)-β-D-glucan. The results pointed out that these transcription elements might contribute to the differences in promoter of ALFFc. Our results would provide supports for future studies to identify the functional transcription elements in the ALF promoter and to expand our knowledge on regulation of innate immune genes in Chinese shrimp.

Piceatannol attenuates cardiac hypertrophy in an animal model through regulation of the expression and binding of the transcription factor GATA binding factor 6

Piceatannol is found in grapes, passion fruit, and Japanese knotweed. Piceatannol pretreatment suppresses cardiac hypertrophy induced by isoproterenol as assessed by heart weight/body weight ratio, cross-sectional area, and expression of hypertrophic markers. The anti-hypertrophic effect of piceatannol in rat neonatal cardiomyocytes is the same as that in vivo. Piceatannol inhibits lentiviral-GATA6-induced cardiomyocyte hypertrophy. Furthermore, piceatannol reduces the interaction between GATA4 and GATA6 as well as the DNA-binding activity of endogenous GATA6 in the ANP promoter. Our results suggest that piceatannol may be a novel therapeutic agent for the prevention of cardiac hypertrophy.

Global analysis of induced transcription factors and cofactors identifies Tfdp2 as an essential coregulator during terminal erythropoiesis

Key transcriptional regulators of terminal erythropoiesis, such as GATA-binding factor 1 (GATA1) and T-cell acute lymphocytic leukemia protein 1 (TAL1), have been well characterized, but transcription factors and cofactors and their expression modulations have not yet been explored on a global scale. Here, we use global gene expression analysis to identify 28 transcription factors and 19 transcriptional cofactors induced during terminal erythroid differentiation whose promoters are enriched for binding by GATA1 and TAL1. Utilizing protein-protein interaction databases to identify cofactors for each transcription factor, we pinpoint several co-induced pairs, of which E2f2 and its cofactor transcription factor Dp-2 (Tfdp2) were the most highly induced. TFDP2 is a critical cofactor required for proper cell cycle control and gene expression. GATA1 and TAL1 are bound to the regulatory regions of Tfdp2 and upregulate its expression and knockdown of Tfdp2 results in significantly reduced rates of proliferation as well as reduced upregulation of many erythroid-important genes. Loss of Tfdp2 also globally inhibits the normal downregulation of many E2F2 target genes, including those that regulate the cell cycle, causing cells to accumulate in S phase and resulting in increased erythrocyte size. Our findings highlight the importance of TFDP2 in coupling the erythroid cell cycle with terminal differentiation and validate this study as a resource for future work on elucidating the role of diverse transcription factors and coregulators in erythropoiesis.

Gene set enrichment analysis of the NF-κB/Snail/YY1/RKIP circuitry in multiple myeloma

The presence of a dysregulated NF-κB/Snail/YY1/RKIP loop was recently established in metastatic prostate cancer cells and non-Hodgkin's lymphoma; however, its involvement in multiple myeloma (MM) has yet to be investigated. Aim of the study was to investigate the role of the NF-κB/Snail/YY1/RKIP circuitry in MM and how each gene is correlated with the remaining genes of the loop. Using gene set enrichment analysis and gene neighbours analysis in data received from four datasets included in the Multiple Myeloma Genomics Portal of the Multiple Myeloma Research Consortium, we identified various enriched gene sets associated with each member of the NF-κB/Snail/YY1/RKIP circuitry. In each dataset, the 20 most co-expressed genes with the circuitry genes were isolated subjected to Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment. Among many, we highlighted on FNDC3B, TPD52, BBX, MBNL1 and MFAP2. Many co-expressed genes participated in the regulation of metabolic processes and nucleic acid binding, or were transcription factor binding genes and genes with metallopeptidase activity. The transcription factors FOXO4, GATA binding factor, Sp1 and AP4 most likely affect the expression of the NF-κB/Snail/YY1/RKIP circuitry genes. Computational analysis of various GEO datasets revealed elevated YY1 and RKIP levels in MM vs. the normal plasma cells, as well as elevated RKIP levels in MM vs. normal B lymphocytes. The present study highlights the relationships of the NF-κB/Snail/YY1/RKIP circuitry genes with specific cancer-related gene sets in multiple myeloma.

The role of PKCε-dependent signaling for cardiac differentiation

Protein kinase Cepsilon (PKCε) exerts a well-known cardio-protective activity in ischemia-reperfusion injury and plays a pivotal role in stem cell proliferation and differentiation. Although many studies have been performed on physiological and morphological effects of PKCε mis-expression in cardiomyocytes, molecular information on the role of PKCε on early cardiac gene expression are still lacking. We addressed the molecular role of PKCε in cardiac cells using mouse cardiomyocytes and rat bone marrow mesenchymal stem cells. We show that PKCε is modulated in cardiac differentiation producing an opposite regulation of the cardiac genes NK2 transcription factor related, locus 5 (nkx2.5) and GATA binding protein 4 (gata4) both in vivo and in vitro. Phospho-extracellular regulated mitogen-activated protein kinase 1/2 (p-ERK1/2) levels increase in PKCε over-expressing cells, while pkcε siRNAs produce a decrease in p-ERK1/2. Indeed, pharmacological inhibition of ERK1/2 rescues the expression levels of both nkx2.5 and gata4, suggesting that a reinforced (mitogen-activated protein kinase) MAPK signaling is at the basis of the observed inhibition of cardiac gene expression in the PKCε over-expressing hearts. We demonstrate that PKCε is critical for cardiac cell early gene expression evidencing that this protein is a regulator that has to be fine tuned in precursor cardiac cells.

Analysis of multiple polymorphisms in the bovine neuropeptide Y5 receptor gene and structural modelling of the encoded protein

The neuropeptide Y 5 receptor (NPY5R) plays an important role in the regulation of appetite and feeding behaviour in mammals by modulating the effect of the neurotransmitter neuropeptide Y. As single nucleotide polymorphism (SNP) variation in the bovine NPY5R gene is likely to influence the expression and/or function of this gene, the objectives of this study were to identify SNPs in the bovine NPY5R gene and to predict their functional role in the expression and physico-chemical characteristics of the protein product. Nineteen novel SNPs were identified in a 2.1kb genomic region of the NPY5R gene in a total of 419 beef cattle from 13 Bostaurus breeds and eight Bosindicus animals. Four of these SNPs were non-synonymous (Met→Ile, Leu→Phe, Pro→Leu, Arg→Stop codon), while 10 were synonymous. Of particular interest was one non-synonymous SNP (c.1090C>T) that introduced a stop codon in the third intracellular loop of the NPY5R molecule. This stop codon is predicted to create a truncated NPY5R molecule with different physico-chemical properties compared to the native NPY5R protein. A further four SNPs were located in the 5' untranslated region (UTR) and one in the 3'UTR. Two of the 5'UTR SNPs affected putative transcription factor binding sites (GATA binding factor and snRNA-activating protein complex). In conclusion, regulatory and functional SNPs were identified in the bovine NPY5R gene. These include SNPs which potentially modify transcription factor binding sites as well as SNPs that cause amino acid changes and premature termination of the NPY5R protein. Such polymorphisms are likely to play vital physiological roles in the neuropeptide Y mediated appetite, feed intake and energy homeostasis in cattle.

The Notch ligand Jagged2 promotes lung adenocarcinoma metastasis through a miR-200–dependent pathway in mice

Epithelial tumor cells transit to a mesenchymal state in response to extracellular cues, in a process known as epithelial-to-mesenchymal transition (EMT). The precise nature of these cues has not been fully defined, an important issue given that EMT is an early event in tumor metastasis. Here, we have found that a population of metastasis-prone mouse lung adenocarcinoma cells expresses Notch and Notch ligands and that the Notch ligand Jagged2 promotes metastasis. Mechanistically, Jagged2 was found to promote metastasis by increasing the expression of GATA-binding (Gata) factors, which suppressed expression of the microRNA-200 (miR-200) family of microRNAs that target the transcriptional repressors that drive EMT and thereby induced EMT. Reciprocally, miR-200 inhibited expression of Gata3, which reversed EMT and abrogated metastasis, suggesting that Gata3 and miR-200 are mutually inhibitory and have opposing effects on EMT and metastasis. Consistent with this, high levels of Gata3 expression correlated with EMT in primary tumors from 2 cohorts of lung adenocarcinoma patients. These findings reveal what we believe to be a novel Jagged2/miR-200-dependent pathway that mediates lung adenocarcinoma EMT and metastasis in mice and may have implications for the treatment of human epithelial tumors.

Repair of senescent myocardium by mesenchymal stem cells is dependent on the age of donor mice

Myocardial infarction is one of the leading causes of mortality in aged people. Whether age of donors of mesenchymal stem cells (MSCs) affects its ability to repair the senescent heart tissue is unknown. In the present study, MSCs from young (2 months) and aged (18 months) green fluorescent protein expressing C57BL/6 mice were characterized with p16INK4a and β-gal associated senescence. Myocardial infarction was produced in 18-month-old wild-type C57BL/6 mice transplanted with MSCs from young and aged animals in the border of the infarct region. Expression of p16INK4a in MSCs from aged animals was significantly higher (21.5%± 1.2, P < 0.05) as compared to those from young animals (9.2%± 2.8). A decline in the tube-forming ability on Matrigel was also observed in aged MSCs as well as down-regulation of insulin-like growth factor-1, fibroblast growth factor (FGF-2), vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) compared to young cells. Mice transplanted with young MSCs exhibited significant improvement in their left ventricle (LV) systolic and diastolic function as demonstrated by dp/dtmax, dp/dtmin, Pmax. Reduction in the LV fibrotic area was concomitant with neovascularization as demonstrated by CD31 and smooth muscle actin (SMA) expression. Real-time RT-PCR analysis for VEGF, stromal cell derived factor (SDF-1α) and GATA binding factor 4 (GATA-4) genes further confirmed the effect of age on MSC differentiation towards cardiac lineages and enhanced angiogenesis. These studies lead to the conclusion that repair potential of MSCs is dependent on the age of donors and the repair of senescent infarcted myocardium requires young healthy MSCs.

Altered gene expression in the brain and ovaries of zebrafish (Danio Rerio) exposed to the aromatase inhibitor fadrozole: Microarray analysis and hypothesis generation

As part of a research effort examining system-wide responses of the hypothalamic-pituitary-gonadal (HPG) axis in fish to endocrine-active chemicals (EACs) with different modes of action, zebrafish (Danio rerio) were exposed to 25 or 100 microg/L of the aromatase inhibitor fadrozole for 24, 48, or 96 h. Global transcriptional response in brain and ovarian tissue of fish exposed to 25 microg/L of fadrozole was compared to that in control fish using a commercially available, 22,000-gene oligonucleotide microarray. Transcripts altered in brain were functionally linked to differentiation, development, DNA replication, and cell cycle. Additionally, multiple genes associated with the one-carbon pool by folate pathway (KEGG 00670) were significantly up-regulated. Transcripts altered in ovary were functionally linked to cell-cell adhesion, extracellular matrix, vasculogenesis, and development. Promoter motif analysis identified GATA-binding factor 2, Ikaros 2, alcohol dehydrogenase gene regulator 1, myoblast-determining factor, and several heat shock factors as being associated with coexpressed gene clusters that were differentially expressed following exposure to fadrozole. Based on the transcriptional changes observed, it was hypothesized that fadrozole elicits neurodegenerative stress in brain tissue and that fish cope with this stress through proliferation of radial glial cells. Additionally, it was hypothesized that changes of gene expression in the ovary of fadrozole-exposed zebrafish reflect disruption of oocyte maturation and ovulation because of impaired vitellogenesis. These hypotheses and others derived from the microarray results provide a foundation for future studies aimed at understanding responses of the HPG axis to EACs and other chemical stressors.

NFAT directly regulates Nkx2‐5 transcription during cardiac cell differentiation

The transcription factor NFAT (nuclear factor of activated T-cell) family comprises important regulators in immuno-responses and mouse embryonic development, including early cardiovascular and heart valve development. The mechanism involved, however, is not fully understood. Nkx2-5 (NK2 transcription factor related, locus 5) is one of the earliest genes expressed in early cardiac progenitor cells and is essential for heart tube development by control of a subset of cardiac muscle-specific genes. Previously we found that downregulation of mitochondrial respiratory chain complex I caused severe cardiac deficiencies during heart tube development in Xenopus embryos associated with compromised Nkx2-5 expression. However, the heart defects and Nkx2-5 expression could be rescued by a constitutively activated NFAT, suggesting a possible link between NFAT and Nkx2-5 during early heart development. In the present study, we demonstrate that NFAT regulates Nkx2-5 expression in both mouse ES (embryonic stem) cells and P19 cells, a mouse model for embryonic differentiation. We found that there are six core NFAT-binding elements in the 5' regulatory region of the Nkx2-5 gene. Although NFAT is able to bind directly to all but one of these elements, it activates Nkx2-5 transcription only via a specific binding site in the distal enhancer region. Interestingly, the transcriptional activity of NFAT is largely dependent on the co-factor GATA (GATA-binding transcription factor), which binds to an element adjacent to this key NFAT-binding site. Furthermore, binding of the endogenous NFAT to this particular site was observed during cardiac differentiation in mouse ES and P19 cells. The results suggest that Nkx2-5 is a direct target of NFAT that co-ordinates with other transcription factors such as GATA4 to regulate Nkx2-5 during cardiogenesis.

Putative transcription factor binding sites and polymorphisms in the proximal promoter of the PRL‐A gene in percomorphs and European sea bass (Dicentrarchus labrax)

AbstractThere is an increasing interest in exploring how the mechanisms that regulate gene expression might generate diversity in phenotypes as a function of habitat utilisation (at both inter‐ and intra‐specific levels). Prolactin (PRL) is the main hormone involved in adaptation to low salinity in teleosts. Several mechanisms regulating PRL‐A gene expression have been described. The main mechanisms involve Pit‐1 binding sites and length polymorphism of a short sequence repeat (SSR) located in the proximal promoter (PP) of the gene. Together, these processes may generate phenotypes with potentially distinct physiological responses to salinity. To gain insight into their relative roles and importance, we sequenced the PP of the PRL‐A gene in the euryhaline European sea bass (Dicentrarchus labrax) and in the stenohaline sister species, Dicentrarchus punctatus. Then, we assessed the presence of both SSR and binding elements in PP among several teleost lineages, by checking available data from the literature and GenBank. The sequence of PRL‐A PP was completely distinct between the percomorphs and other lineages such as salmoniform, siluriform and cypriniform, except for short stretches of nucleotides that were highly conserved across all species, also including mammals. Most of them were identified as putative cis‐regulatory elements, including Pit‐1, but also δEF1, c‐ETS, CEBP, and GATA binding factor 2, previously unreported as regulating PRL‐A transcription in teleosts, but involved in gene regulation in mammals. This result suggests that there is greater conservation of PRL‐A regulatory mechanisms across vertebrate lineages than previously thought, and offers new perspectives for future studies. The presence and homology of an SSR in the PP of PRL‐A gene of percomorphs was demonstrated, but no SSR polymorphisms were found amongst the two Dicentrarchus species, or within natural D. labrax populations covering the known range of natural genetic variation, including marine, brackish water and freshwater populations.

Cloning, characterization and computational analysis of the 5′ regulatory region of ovine glucose 6-phosphate dehydrogenase gene

To better understand the structure and the function of ovine glucose 6-phosphate dehydrogenase (G6PD) promoter region, a genome-walking procedure was followed to isolate and sequence a 1628 bp fragment, containing the 5′ regulatory region of the G6PD gene. In silico analysis of the sequence showed many conserved blocks and features with other known mammalian G6PD promoter regions. The analysis also revealed the presence of one TATA box, three GC boxes, two E-boxes and several binding sites for Stimulating Protein 1 (Sp1) and Activator Protein 2 (AP2). Moreover, elements involved in the regulation of lipogenesis like USF (Upstream stimulating factor), HSF (Heat Shock Factor), F2F (Prolactin receptor), RAR (Retinoid Acid Receptor), STRE (STress Response Element), RORa (Retinoid related Orphan Receptor alpha), GATA (GATA binding factor), RFX (Regulatory Factor X), SREBP (Sterol Regulatory Element Binding Protein), MEP (Metal Element Protein), CREB (insulin receptor), PRE (Progesterone receptor), and HNF4 (Hepatic Nuclear Factor 4) were detected. The most important regulatory motifs were found to be conserved as compared to those in human and mouse counterparts. However, some differences were noted, likely indicating differences in the transcription regulation of G6PD gene between ruminant and non-ruminant species.

Cytochrome P450 aromatase in grey mullet: cDNA and promoter isolation; brain, pituitary and ovarian expression during puberty

In a study towards elucidating the role of aromatases during puberty in female grey mullet, the cDNAs of the brain (muCyp19b) and ovarian (muCyp19a) aromatase were isolated by RT-PCR and their relative expression levels were determined by quantitative real-time RT-PCR. The muCyp19a ORF of 1515 bp encoded 505 predicted amino acid residues, while that of muCyp19b was 1485 bp and encoded 495 predicted amino acid residues. The expression level of muCyp19b significantly increased in the brain as puberty advanced; however, its expression level in the pituitary increased only slightly with pubertal development. In the ovary, the muCyp19a expression level markedly increased as puberty progressed. The promoter regions of the two genes were also isolated and their functionality evaluated in vitro using luciferase as the reporter gene. The muCyp19a promoter sequence (650 bp) contained a consensus TATA box and putative transcription factor binding sites, including two half EREs, an SF-1, an AhR/Arnt, a PR and two GATA-3s. The muCyp19b promoter sequence (2500 bp) showed consensus TATA and CCAAT boxes and putative transcription binding sites, namely: a PR, an ERE, a half ERE, a SP-1, two GATA-binding factor, one half GATA-1, two C/EBPs, a GRE, a NFkappaB, three STATs, a PPAR/RXR, an Ahr/Arnt and a CRE. Basal activity of serially deleted promoter constructs transiently transfected into COS-7, αT3 and TE671 cells demonstrated the enhancing and silencing roles of the putative transcription factor binding sites. Quinpirole, a dopamine agonist, significantly reduced the promoter activity of muCyp19b in TE671. The results suggest tissue-specific regulation of the muCyp19 genes and a putative alternative promoter for muCyp19b.

Transcriptional regulation of the NADPH oxidase isoform, Nox1, in colon epithelial cells: Role of GATA-binding factor(s)

Nonphagocytic NADPH oxidases (Noxs) are major sources of reactive oxygen species (ROS) and exist as a family of isoenzymes with tissue-restricted expression and functions. Nox1, expressed in colon epithelium and vascular smooth muscle, is suggested to be involved in innate immune defense and cell growth or proliferation. The transcriptional regulation of Nox1 appears to be particularly important in the modulation of its activity but the underlying mechanisms are unknown. Here we have identified the functional Nox1 promoter in human colon epithelial Caco-2 cells, and show that a 520-bp genomic fragment encompassing the CAP site is sufficient to direct high levels of expression of a linked reporter gene in these cells. Deletion analyses together with electrophoretic mobility-shift assays (EMSAs) suggest that maximal promoter activity is dependent on a GATA-binding site, conserved between human and mouse, within the proximal promoter region. The ability of mouse GATA factors to transactivate the Nox1 promoter was demonstrated in Cos-7 cells and site-directed mutagenesis of the conserved GATA-binding site further demonstrates that the regulation of Nox1 transcription is mediated by the direct binding of a GATA factor to the Nox1 proximal promoter. We also identified more distal, upstream regions which act to repress significantly expression from the Nox1 promoter.

Identification ofcis-element regulating expression of the mouseFgf10 gene during inner ear development

Fibroblast growth factor (FGF) signaling is crucial for the induction and growth of the ear, a sensory organ that involves intimate tissue interactions. Here, we report the abnormality of Fgf10 null ear and the identification of a cis-regulatory element directing otic expression of Fgf10. In Fgf10 null inner ears, we found that the initial development of semicircular, vestibular, and cochlear divisions is roughly normal, after which there are abnormalities of semicircular canal/cristae and vestibular development. The mutant semicircular disks remain without canal formation by the perinatal stage. To elucidate regulation of the Fgf10 expression during inner ear development, we isolated a 6.6-kb fragment of its 5'-upstream region and examined its transcriptional activity with transgenic mice, using a lacZ-reporter system. From comparison of the mouse sequences of the 6.6-kb fragment with corresponding sequences of the human and chicken Fgf10, we identified a 0.4-kb enhancer sequence that drives Fgf10 expression in the developing inner ear. The enhancer sequences have motifs for many homeodomain-containing proteins (e.g., Prx, Hox, Nkx), in addition to POU-domain factors (e.g., Brn3), zinc-finger transcription factors (e.g., GATA-binding factors), TCF/LEF-1, and a SMAD-interacting protein. Thus, FGF10 signaling is dispensable for specification of otic compartment identity but is required for hollowing the semicircular disk. Furthermore, the analysis of a putative inner ear enhancer of Fgf10 has disclosed a complicated regulation of Fgf10 during inner ear development by numerous transcription factors and signaling pathways.

A paradoxical mutant GATA factor.

The niiA (nitrite reductase) and niaD (nitrate reductase) genes of Aspergillus nidulans are subject to both induction by nitrate and repression by ammonium or glutamine. The intergenic region between these genes functions as a bidirectional promoter. In this region, nucleosomes are positioned under nonexpression conditions. On nitrate induction under derepressing conditions, total loss of positioning occurs. This is independent of transcription and of the NirA-specific transcription factor but absolutely dependent on the wide-domain GATA-binding AreA factor. We show here that a 3-amino-acid deletion in the basic carboxy-terminal sequence of the DNA-binding domain results in a protein with paradoxical properties. Its weak DNA binding is consistent with its loss-of-function phenotype on most nitrogen sources. However, it results in constitutive expression and superinducibility of niiA and niaD. Nucleosome loss of positioning is also constitutive. The mutation partially suppresses null mutations in the transcription factor NirA. AreA binds NirA in vitro, and the mutation does not affect this interaction. The in vivo methylation pattern of the promoter is drastically altered, suggesting the recruitment of one or more unknown transcription factors and/or a local distortion on the DNA double helix.

959 Comparison of histological and clinical changes between before and after spontaneous loss of hepatitis B surface antigen

We have isolated a genomic clone encoding human selenoprotein P including the putative promoter region. The gene spans 12 kb and consists of five exons with a start codon in the second exon. A typical TATA sequence, the recognition motifs for a GATA-binding factor and the liver-specific factors, HNF-1 and HNF-3, were detected upstream from the transcription start point.