SEARCH · PubMed Health

Results for “regulatory element transcripts”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,495 records · Page 83Linked to original sources

PubMed2002

DNA methylation and the regulation of gene transcription.

The regulation of gene transcription is not simply dependent on the presence or absence of DNA-binding transcription factors that turn genes on or off, but also involves processes determining the ability of transcription factors to gain access to and bind their target DNA. Methylation of DNA cytosine bases leads to the inaccessibility of DNA regulatory elements to their transcription factors by a number of mechanisms. Our understanding of DNA methylation has advanced rapidly in recent years with the identification of an increasingly large number of novel proteins involved in this process. These include methylcytosine-binding proteins as well as additional members of the DNA methyltransferase family. The creation of mice with targeted deletions in a number of genes involved in DNA methylation has further elucidated the functions of many of these proteins. The characterization of complexes that contain proteins known to be involved in DNA methylation has led to the identification of additional proteins, especially those involved in histone deacetylation, indicating that DNA methylation and histone deacetylation very likely act in a synergistic fashion to regulate gene transcription. Finally, the implication of DNA methylation in tumorigenesis and the realization that some congenital diseases are caused by deficiency of proteins involved in DNA methylation has confirmed the importance of this process in regulating gene expression.

5-Methylcytosine↗

PubMed2004

Estrogens and progesterone promote persistent CCND1 gene activation during G1 by inducing transcriptional derepression via c-Jun/c-Fos/estrogen receptor (progesterone receptor) complex assembly to a distal regulatory element and recruitment of cyclin D1 to its own gene promoter.

Transcriptional activation of the cyclin D1 gene (CCND1) plays a pivotal role in G(1)-phase progression, which is thereby controlled by multiple regulatory factors, including nuclear receptors (NRs). Appropriate CCND1 gene activity is essential for normal development and physiology of the mammary gland, where it is regulated by ovarian steroids through a mechanism(s) that is not fully elucidated. We report here that CCND1 promoter activation by estrogens in human breast cancer cells is mediated by recruitment of a c-Jun/c-Fos/estrogen receptor alpha complex to the tetradecanoyl phorbol acetate-responsive element of the gene, together with Oct-1 to a site immediately adjacent. This process coincides with the release from the same DNA region of a transcriptional repressor complex including Yin-Yang 1 (YY1) and histone deacetylase 1 and is sufficient to induce the assembly of the basal transcription machinery on the promoter and to lead to initial cyclin D1 accumulation in the cell. Later on in estrogen stimulation, the cyclin D1/Cdk4 holoenzyme associates with the CCND1 promoter, where E2F and pRb can also be found, contributing to the long-lasting gene enhancement required to drive G(1)-phase completion. Interestingly, progesterone triggers similar regulatory events through its own NRs, suggesting that the gene regulation cascade described here represents a crossroad for the transcriptional control of G(1)-phase progression by different classes of NRs.

Base Sequence↗

PubMed1999

Neural restrictive silencer factor recruits mSin3 and histone deacetylase complex to repress neuron-specific target genes.

Accumulative evidence suggests that more than 20 neuron-specific genes are regulated by a transcriptional cis-regulatory element known as the neural restrictive silencer (NRS). A trans-acting repressor that binds the NRS, NRSF [also designated RE1-silencing transcription factor (REST)] has been cloned, but the mechanism by which it represses transcription is unknown. Here we show evidence that NRSF represses transcription of its target genes by recruiting mSin3 and histone deacetylase. Transfection experiments using a series of NRSF deletion constructs revealed the presence of two repression domains, RD-1 and RD-2, within the N- and C-terminal regions, respectively. A yeast two-hybrid screen using the RD-1 region as a bait identified a short form of mSin3B. In vitro pull-down assays and in vivo immunoprecipitation-Western analyses revealed a specific interaction between NRSF-RD1 and mSin3 PAH1-PAH2 domains. Furthermore, NRSF and mSin3 formed a complex with histone deacetylase 1, suggesting that NRSF-mediated repression involves histone deacetylation. When the deacetylation of histones was inhibited by tricostatin A in non-neuronal cells, mRNAs encoding several neuronal-specific genes such as SCG10, NMDAR1, and choline acetyltransferase became detectable. These results indicate that NRSF recruits mSin3 and histone deacetylase 1 to silence neural-specific genes and suggest further that repression of histone deacetylation is crucial for transcriptional activation of neural-specific genes during neuronal terminal differentiation.

3T3 Cells↗

PubMed1998

Stat6 inhibits human interleukin-4 promoter activity in T cells.

The differentiation of naive T-helper (Th) cells into cytokine-secreting effector Th cells requires exposure to multiple signals, including exogenous cytokines. Interleukin-4 (IL-4) plays a major role in this process by promoting the differentiation of IL-4-secreting Th2 cells. In Th2 cells, IL-4 gene expression is tightly controlled at the level of transcription by the coordinated binding of multiple transcription factors to regulatory elements in the proximal promoter region. Nuclear factor of activated T cell (NFAT) family members play a critical role in regulating IL-4 transcription and interact with up to five sequences (termed P0 through P4) in the IL-4 promoter. The molecular mechanisms by which IL-4 induces expression of the IL-4 gene are not known, although the IL-4-activated transcription factor signal transducer and activator of transcription 6 (Stat6) is required for this effect. We report here that Stat6 interacts with three binding sites in the human IL-4 promoter by electrophoretic mobility shift assays. These sites overlap the P1, P2, and P4 NFAT elements. To investigate the role of Stat6 in regulating IL-4 transcription, we used Stat6-deficient Jurkat T cells with different intact IL-4 promoter constructs in cotransfection assays. We show that, whereas a multimerized response element from the germline IgE promoter was highly induced by IL-4 in Stat6-expressing Jurkat cells, the intact human IL-4 promoter was repressed under similar conditions. We conclude that the function of Stat6 is highly dependent on promoter context and that this factor promotes IL-4 gene expression in an indirect manner.

Binding Sites↗

PubMed1998

The mating-type proteins of fission yeast induce meiosis by directly activating mei3 transcription.

Cell type control of meiotic gene regulation in the budding yeast Saccharomyces cerevisiae is mediated by a cascade of transcriptional repressors, a1-alpha2 and Rme1. Here, we investigate the analogous regulatory pathway in the fission yeast Schizosaccharomyces pombe by analyzing the promoter of mei3, the single gene whose expression is sufficient to trigger meiosis. The mei3 promoter does not appear to contain a negative regulatory element that represses transcription in haploid cells. Instead, correct regulation of mei3 transcription depends on a complex promoter that contains at least five positive elements upstream of the TATA sequence. These elements synergistically activate mei3 transcription, thereby constituting an on-off switch for the meiosis pathway. Element C is a large region containing multiple sequences that resemble binding sites for Mc, an HMG domain protein encoded by the mating-type locus. The function of element C is extremely sensitive to spacing changes but not to linker-scanning mutations, suggesting the possibility that Mc functions as an architectural transcription factor. Altered-specificity experiments indicate that element D interacts with Pm, a homeodomain protein encoded by the mating-type locus. This indicates that Pm functions as a direct activator of the meiosis pathway, whereas the homologous mating-type protein in S. cerevisiae (alpha2) functions as a repressor. Thus, despite the strong similarities between the mating-type loci of S. cerevisiae and S. pombe, the regulatory logic that governs the tight control of the key meiosis-inducing genes in these organisms is completely different.

Base Sequence↗

PubMed2004

Long-range control of gene expression: emerging mechanisms and disruption in disease.

Transcriptional control is a major mechanism for regulating gene expression. The complex machinery required to effect this control is still emerging from functional and evolutionary analysis of genomic architecture. In addition to the promoter, many other regulatory elements are required for spatiotemporally and quantitatively correct gene expression. Enhancer and repressor elements may reside in introns or up- and downstream of the transcription unit. For some genes with highly complex expression patterns--often those that function as key developmental control genes--the cis-regulatory domain can extend long distances outside the transcription unit. Some of the earliest hints of this came from disease-associated chromosomal breaks positioned well outside the relevant gene. With the availability of wide-ranging genome sequence comparisons, strong conservation of many noncoding regions became obvious. Functional studies have shown many of these conserved sites to be transcriptional regulatory elements that sometimes reside inside unrelated neighboring genes. Such sequence-conserved elements generally harbor sites for tissue-specific DNA-binding proteins. Developmentally variable chromatin conformation can control protein access to these sites and can regulate transcription. Disruption of these finely tuned mechanisms can cause disease. Some regulatory element mutations will be associated with phenotypes distinct from any identified for coding-region mutations.

Animals↗

PubMed2006

Structures, regulatory regions, and inductive expression patterns of antimicrobial peptide genes in the silkworm Bombyx mori.

Antimicrobial peptides (AMPs) are a group of immune proteins that protect the host from infection. In Drosophila, seven groups of inducible AMPs have been identified, with activities against fungi and gram-positive and gram-negative bacteria. On the basis of the silkworm genome sequence and expressed sequence tags, we identified 35 AMP genes, mostly belonging to the cecropin, moricin, and gloverin gene families. We predicted the core promoters required for gene transcription and the cis-regulatory elements for NF-kappaB/Rel and GATA transcription factors. The expression profiles of these genes after an immune challenge with lipopolysaccharide were examined by reverse transcription PCR. Members of the cecropin B and gloverin A subfamilies were intensely expressed in the fat body after induction. In contrast, those of the moricin B subfamily were not expressed under the same conditions. Such results suggest that these regulatory elements and their positions in the upstream regions play an important role in regulating the transcription of these defense genes.

Amino Acid Sequence↗

PubMed2002

Peroxisome-proliferator-activated receptor-alpha (PPARalpha) deficiency leads to dysregulation of hepatic lipid and carbohydrate metabolism by fatty acids and insulin.

The aim of the present study was to determine whether peroxisome-proliferator-activated receptor-alpha (PPARalpha) deficiency disrupts the normal regulation of triacylglycerol (TAG) accumulation, hepatic lipogenesis and glycogenesis by fatty acids and insulin using PPARalpha-null mice. In wild-type mice, hepatic TAG concentrations increased (P<0.01) with fasting (24 h), with substantial reversal after refeeding (6 h). Hepatic TAG levels in fed PPARalpha-null mice were 2.4-fold higher than in the wild-type (P<0.05), increased with fasting, but remained elevated after refeeding. PPARalpha deficiency also impaired hepatic glycogen repletion (P<0.001), despite normal insulin and glucose levels after refeeding. Higher levels of plasma insulin were required to support similar levels of hepatic lipogenesis de novo ((3)H(2)O incorporation) in the PPARalpha-null mice compared with the wild-type. This difference was reflected by corresponding changes in the relationship between plasma insulin and the mRNA expression of the lipogenic transcription factor sterol-regulatory-element-binding protein-1c, and that of one of its known targets, fatty acid synthase. In wild-type mice, hepatic pyruvate dehydrogenase kinase (PDK) 4 protein expression (a downstream marker of altered fatty acid catabolism) increased (P<0.01) in response to fasting, with suppression (P<0.001) by refeeding. Although PDK4 up-regulation after fasting was halved by PPARalpha deficiency, PDK4 suppression after refeeding was attenuated. In summary, PPARalpha deficiency leads to accumulation of hepatic TAG and elicits dysregulation of hepatic lipid and carbohydrate metabolism, emphasizing the importance of precise control of lipid oxidation for hepatic fuel homoeostasis.

Animals↗

PubMed1999

Co-stimulation of promoter for low density lipoprotein receptor gene by sterol regulatory element-binding protein and Sp1 is specifically disrupted by the yin yang 1 protein.

Sterol regulation of gene expression in mammalian cells is mediated by an interaction between the cholesterol-sensitive sterol regulatory element-binding proteins (SREBPs) and promoter-specific but generic co-regulatory transcription factors such as Sp1 and NF-Y/CBF. Thus, sterol-regulated promoters that require different co-regulatory factors could be regulated independently through targeting the specific interaction between the SREBPs and the individual co-regulatory proteins. In the present studies we demonstrate that transiently expressed yin yang 1 protein (YY1) inhibits the SREBP-mediated activation of the low density lipoprotein (LDL) receptor in a sensitive and dose-dependent manner. The inhibition is independent of YY1 binding directly to the LDL receptor promoter, and we show that the same region of YY1 that interacts in solution with Sp1 also interacts with SREBP. Furthermore, other SREBP-regulated genes that are not co-regulated by Sp1 are either not affected at all or are not as sensitive to the repression. Thus, the specific interaction that occurs between SREBPs and Sp1 to stimulate the LDL receptor promoter is a specific target for inhibition by the YY1 protein, and we provide evidence that the mechanism can be at least partially explained by the ability of YY1 to inhibit the interaction between SREBP and Sp1 in solution in vitro. The LDL receptor is the key gene of cholesterol uptake, and the rate-controlling genes of cholesterol synthesis are stimulated by the concerted action of SREBPs along with coregulators that are distinct from Sp1. Therefore, repression of gene expression through specifically targeting the interaction between SREBP and Sp1 would provide a molecular mechanism to explain how cholesterol uptake can be regulated independently from cholesterol biosynthesis in mammalian cells.

CCAAT-Enhancer-Binding Proteins↗

PubMed1997

Stabilization of atherosclerotic plaque during lipid lowering.

Lipid lowering therapy leads to a great reduction of cardiovascular complications, but has almost no effect on the degree of stenosis of coronary arteries. These findings have lead to a new paradigm of coronary artery disease, i.e. clinical prognosis is not only determined by the extent of a single stenosis, but mainly by the number and structure of atherosclerotic plaque. Rupture of an instable or vulnerable plaque, characterized by a large lipid-rich central core, inflammatory cells, and a thin fibrous cap, causes sudden thrombus formation and thereby acute coronary syndromes. There is accumulating evidence that cholesterol lowering can result in plaque stabilization and improvement of endothelial dysfunction, reviewed recently in this journal. Accordingly, this review focuses on new molecular mechanisms which provide evidence that reduction of cellular cholesterol activates novel transcription factors, called sterol regulatory element binding proteins, which can regulate not only the LDL receptor, thereby reducing plasma cholesterol levels, but also a diverse number of other genes. These gene regulatory events might link cholesterol lowering not only to cellular cholesterol metabolism, but also to triglyceride metabolism, cell differentiation and other events potentially stabilizing vulnerable plaque.

Anticholesteremic Agents↗

PubMed2005

Polyunsaturated fatty acid regulation of genes of lipid metabolism.

Apart from being an important macronutrient, dietary fat has recently gained much prominence for its role in regulating gene expression. Polyunsaturated fatty acids (PUFAs) affect gene expression through various mechanisms including, but not limited to, changes in membrane composition, intracellular calcium levels, and eicosanoid production. Furthermore, PUFAs and their various metabolites can act at the level of the nucleus, in conjunction with nuclear receptors and transcription factors, to affect the transcription of a variety of genes. Several of these transcription mediators have been identified and include the nuclear receptors peroxisome proliferator-activated receptor (PPAR), hepatocyte nuclear factor (HNF)-4alpha, and liver X receptor (LXR) and the transcription factors sterol-regulatory element binding protein (SREBP) and nuclear factor-kappaB (NFkappaB). Their interaction with PUFAs has been shown to be critical to the regulation of several key genes of lipid metabolism. Working out the mechanisms by which these interactions and consequent effects occur is proving to be complicated but is invaluable to our understanding of the role that dietary fat can play in disease management and prevention.

Animals↗

PubMed2005

Synthesis of long-chain polyunsaturated fatty acids in lactating mammary gland: role of Delta5 and Delta6 desaturases, SREBP-1, PPARalpha, and PGC-1.

The purpose of this work was to study whether rat lactating mammary gland can synthesize PUFAs through the expression of Delta5 and Delta6 desaturases (Delta5D and Delta6D), whether these enzymes are regulated by the transcription factors sterol-regulatory element binding protein 1 (SREBP-1) and peroxisome proliferator-activated receptor alpha (PPARalpha) and the coactivator peroxisome proliferator-activated receptor gamma coactivator 1beta (PGC-1beta), and whether these desaturases are regulated by the lipid concentration in the diet. The results showed that on day 12 of lactation, approximately 35% of the linoleic acid in the diet, which is the precursor of PUFAs, is transferred to the mammary gland. There was expression of Delta5D and Delta6D in mammary gland, and it was regulated by the corn oil content in the diet. The higher the corn oil content in the diet, the lower the expression of both desaturases. Induction of Delta5D and Delta6D was associated positively with similar changes in SREBP-1 and PGC-1beta. Expression of PPARalpha was barely detected and was not affected by the corn oil content in the diet, whereas PGC-1beta expression increased as the corn oil in the diet increased. These results indicate that the lactating mammary gland has the capacity to synthesize PUFAs and can be regulated by the lipid content in the diet.

Adipose Tissue↗

PubMed1995

Identification and characterization of a novel transcriptional silencer in the human collagen type IV gene COL4A2.

Collagen type IV [alpha 1(IV)2 alpha 2(IV)] is the basic structural component of all basement membranes. The two subunit genes COL4A1 and COL4A2 are found closely linked in the human and murine genomes and are transcribed divergently from a common promoter. Previously, activating elements had been detected within both genes which are indispensable for efficient transcription. An additional negative regulatory element has now been identified within the third intron of the COL4A2 gene which is able to inhibit transcription of both COL4 genes from their shared promoter, as well as the nonrelated herpes simplex virus thymidine kinase promoter. The element exerts its inhibitory effect largely independently from its relative orientation and distance from the initiation site of transcription. Therefore, the element represents a silencer which is named the "COL4 silencer." The minimal functional silencer could be narrowed down by deletion mapping to a sequence element located within intron 3 of the COL4A2 gene. This motif is specifically recognized by a nuclear protein, named "SILBF," and the binding site of which was determined by footprinting assays. Mutation studies and deletion analysis proved that the presence of this sequence element and its interaction with SILBF is not only essential but also sufficient for the silencing function. We assume that the COL4 silencer plays an important role in the control of overall expression and the balance of divergent transcription of both COL4 genes.

Animals↗

PubMed1995

Transcriptional regulation of the carcinoembryonic antigen gene. Identification of regulatory elements and multiple nuclear factors.

Human carcinoembryonic antigen (CEA) belongs to a family of membrane glycoproteins that are overexpressed in many carcinomas; CEA functions in vitro as a homotypic intercellular adhesion molecule and can inhibit differentiation when expressed ectopically in myoblasts. The regulation of expression of CEA is therefore of considerable interest. The CEA gene promoter region between -403 and -124 base pairs upstream of the translation initiation site directed high levels of expression in CEA-expression SW403 cells and was 3 times more active in differentiated than in undifferentiated Caco-2 cells, correlating exactly with the 3-fold increase in CEA mRNA seen in differentiated Caco-2 cells. Inclusion of additional upstream sequences between -1098 and -403 base pairs repressed all activity. By in vitro footprinting and deletion analyses, four cis-acting elements were mapped within the positive regulatory region, and one element within the silencing region. Several nuclear factors binding to these domains were identified: USF, Sp1, and an Sp1-like factor. By co-transfection, USF directly activated the CEA gene promoter in vivo in both SW403 and Caco-2 cells. In addition, the levels of factors binding to each positively acting element increased dramatically with differentiation in Caco-2 cells. Thus the transcriptional control of the CEA gene depends on the interaction of several regulatory elements that bind multiple specific factors.

Base Sequence↗

PubMed2005

The Prader-Willi syndrome murine imprinting center is not involved in the spatio-temporal transcriptional regulation of the Necdin gene.

BACKGROUND: The human Prader-Willi syndrome (PWS) domain and its mouse orthologue include a cluster of paternally expressed genes which imprinted expression is co-ordinately regulated by an imprinting center (IC) closely associated to the Snurf-Snrpn gene. Besides their co-regulated imprinted expression, two observations suggest that the spatio-temporal expression of these genes could also be co-regulated. First, the PWS genes have all been reported to be expressed in the mouse nervous system. Second, Snurf-Snrpn and its associated IC are the most ancient elements of the domain which later acquired additional functional genes by retrotransposition. Although located at least 1.5 megabases from the IC, these retroposons acquired the same imprinted regulation as Snurf-Snrpn. In this study, we ask whether the IC, in addition to its function in imprinting, could also be involved in the spatio-temporal regulation of genes in the PWS domain. RESULTS: We compared the expression pattern of Snurf-Snrpn and C/D-box small nucleolar RNAs (snoRNAs) MBII-85 and MBII-52 to the expression pattern of the two evolutionary related retroposons Ndn and Magel2, in the developing mouse embryo. We show that these genes have highly similar expression patterns in the central nervous system, suggesting that they share a common central nervous system-specific regulatory element. Among these genes, Ndn and Magel2 display the most similar expression patterns. Using transgenic mice containing the Ndn and Magel2 genes, we show that the transgenic Ndn gene whereas not imprinted is correctly expressed. Search for DNase I hypersensitive sites in the Ndn-Magel2 genomic region and comparative genomic analyses were performed in order to identify potential transcriptional cis-regulatory elements. CONCLUSIONS: These results strongly suggest that paternally expressed genes of the PWS domain share a common central nervous system-specific regulatory element. We proposed that this regulatory element could co-localize with the IC. However, we demonstrate that the IC, if required for imprinted regulation, is not involved in the spatio-temporal regulation of distantly located retrotransposed genes such as the Ndn gene in the PWS domain.

Animals↗

PubMed2001

Peroxisome proliferator-activated receptor gamma-mediated transcriptional up-regulation of the hepatocyte growth factor gene promoter via a novel composite cis-acting element.

Hepatocyte growth factor (HGF) is a pleotropic polypeptide that can function as a morphogen, motogen, mitogen, angiogen, carcinogen, and tumor suppressor, depending on the target cell and tissue. Previous studies from our laboratory using transgenic mice have shown that HGF gene expression is tightly regulated at the transcriptional level and that the upstream regulatory elements are crucial for the control of HGF gene transcription. In the present study, we have identified and characterized one of these elements as a peroxisome proliferator-activated receptor gamma (PPARgamma)-responsive element. This regulatory element was localized at -246 to -233 base pairs upstream from the transcription start site of the HGF gene promoter having the sequence GGGCCAGGTGACCT. Gel mobility shift and supershift assays demonstrated that this cis-acting element strongly binds to the PPARgamma isoforms as well as to chicken ovalbumin upstream promoter-transcription factor, a member of the orphan nuclear receptor subfamily. Mutational analysis and gel mobility band shift assays indicated that the binding site is an inverted repeat of the AGGTCA motif with two spacers (inverted repeat 2 configuration) and that the two spacers are important for PPARgamma binding. This binding site overlaps with functional binding sites for activating protein-2, nuclear factor 1, and upstream stimulatory factor, and together, they constitute a multifunctional composite binding site through which these different transcription factors exert their regulatory effects on HGF promoter activity. Functional assays revealed that PPARgamma, with its ligand, 15-deoxy-prostaglandin J2, strongly stimulates HGF promoter activity. On the other hand, nuclear factor 1, activating protein-2, and chicken ovalbumin upstream promoter-transcription factor transcription factors repress the stimulatory action of PPARgamma by competing with PPARgamma for their overlapping binding sites. Furthermore, for the first time, our studies demonstrate that the PPARgamma ligand, 15-deoxy-prostaglandin J2, induces endogenous HGF mRNA and protein expression in fibroblasts in culture.

3T3 Cells↗

PubMed2026

Transcriptional perturbation of LINE-1 elements reveals their cis-regulatory potential.

Long interspersed element-1 (LINE-1 or L1) retrotransposons constitute the largest transposable element family in mammalian genomes and contribute prominently to inter- and intra-individual genetic variation. Although most L1 elements are inactive, some evolutionary younger elements remain intact and genetically competent for transcription and occasionally retrotransposition. Despite being generally more abundant in gene-poor regions, intact or full-length L1s (FL-L1) are also enriched around specific classes of genes and on the eutherian X chromosome. How proximal FL-L1 may affect nearby gene expression remains unclear. Here, we examine this systematically using engineered mouse embryonic stem cells (ESCs) in which expression of one active L1 subfamily is perturbed. We find that FL-L1 activation leads to the misregulation of ~1024 genes, whereas FL-L1 repression affects ~81 genes. In most cases (68%), misexpressed genes contain an intronic FL-L1 or lie near a FL-L1 (< 260 kb). Gene ontology analysis shows that upon L1 activation, upregulated genes are enriched for neuronal function-related terms, suggesting that some L1 elements may have evolved to control neuronal gene networks. These results illustrate the cis-regulatory potential of FL-L1 elements and suggest a broader role for L1s than originally anticipated.

Animals↗

PubMed2002

Transcriptional regulation of the rat NHE3 gene. Functional interactions between GATA-5 and Sp family transcription factors.

Expression of sodium-hydrogen exchanger isoform 3 (NHE3) in the intestinal and renal epithelium plays a critical role in sodium absorption and acid/base homeostasis. To decipher rat NHE3 gene regulation, its cis-acting regulatory elements and associated transcription factors were characterized by transient transfection of Caco-2, IEC-6, Qt6, and Drosophila SL2 cells. Deletion and mutational analyses demonstrated that the atypical TATA box located at bp -26/-31 was not necessary for promoter activity, and that a -20/+8-bp fragment represents a functional initiator. Within the 81-bp upstream region, three Sp transcription factor binding sites were critical because their mutation drastically reduced promoter activity. The roles of Sp1 and Sp3 were further demonstrated by electromobility shift assay and by transactivation of the NHE3 promoter in SL2 cells by forced expression of Sp1 and Sp3. Both of these transcription factors were found to act synergistically with GATA-5 bound to a GATA box in exon 1 (+20/+23 bp). These studies demonstrate that rat NHE3 promoter is initiator-driven and controlled mainly by Sp1 and Sp3, which functionally interact with GATA-5. This interaction represents a novel regulatory mechanism, which is likely to participate in a gradient of intestinal gene expression along the crypt-villus axis.

Adenocarcinoma↗