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PubMed1994

Transcriptional regulation of the apolipoprotein A-IV gene involves synergism between a proximal orphan receptor response element and a distant enhancer located in the upstream promoter region of the apolipoprotein C-III gene.

Apolipoprotein A-IV expression is limited to intestinal and hepatic cells, suggesting a tissue specific transcriptional regulation of its gene. To investigate the mechanism controlling apo A-IV transcription we have analysed its promoter region by in vitro DNA binding and transient transfection experiments. DNase I footprinting analysis of the proximal promoter with rat liver nuclear extracts revealed four protected regions: AIVA (-32 to -22), AIVB (-84 to -42), AIVC (-148 to -92) and AIVD (-274 to -250). Element AIVC which is necessary for maximal promoter activity, binds HNF-4, Arp-1 and Ear-3 with similar affinity in a mutually exclusive manner. HNF-4 transactivated chimeric constructs containing intact AIVC site in the context of either the apo A-IV promoter or the heterologous thymidine kinase minimal promoter, while Arp-1 and Ear-3 repressed this activation. Increasing amounts of HNF-4 alleviated Arp-1 or Ear-3 mediated repression, suggesting that the observed opposing effects is a result of direct competition of these factors for the same recognition site. In transient transfection assays the apo A-IV promoter region (-700 to +10) had a very low activity in cells of hepatic (HepG2) and intestinal (CaCo2) origin. This activity was increased 13 to 18-fold when the upstream elements of the distantly linked apo C-III gene were fused to the proximal promoter. Results obtained with different 5' and 3' deletion constructs indicated that the cis-acting elements F to J between the nucleotides -500 and -890 of the apo C-III promoter were absolutely necessary to drive maximal enhancement in HepG2 and CaCo2 cells. The apo C-III upstream elements enhanced the activity of the minimal AdML promoter or the apo A-IV site C mutant less efficiently than the intact apo A-IV or AdML promoter constructs containing single HNF-4 sites. The findings suggest that the enhancer effect is mediated by synergistic interactions between the trans-acting factors which recognize the apo C-III regulatory elements and HNF-4 which binds to the proximal apo A-IV promoter.

Animals↗
PubMed2001

Conserved regulatory elements of the promoter sequence of the gene rpoH of enteric bacteria.

The rpoH regulatory region of different members of the enteric bacteria family was sequenced or downloaded from GenBank and compared. In addition, the transcriptional start sites of rpoH of Yersinia frederiksenii and Proteus mirabilis, two distant members of this family, were determined. Sequences similar to the sigma(70) promoters P1, P4 and P5, to the sigma(E) promoter P3 and to boxes DnaA1, DnaA2, cAMP receptor protein (CRP) boxes CRP1, CRP2 and box CytR present in Escherichia coli K12, were identified in sequences of closely related bacteria such as: E.coli, Shigella flexneri, Salmonella enterica serovar Typhimurium, Citrobacter freundii, Enterobacter cloacae and Klebsiella pneumoniae. In more distant bacteria, Y.frederiksenii and P.mirabilis, the rpoH regulatory region has a distal P1-like sigma(70) promoter and two proximal promoters: a heat-induced sigma(E)-like promoter and a sigma(70) promoter. Sequences similar to the regulatory boxes were not identified in these bacteria. This study suggests that the general pattern of transcription of the rpoH gene in enteric bacteria includes a distal sigma(70) promoter, >200 nt upstream of the initiation codon, and two proximal promoters: a heat-induced sigma(E)-like promoter and a sigma(70) promoter. A second proximal sigma(70) promoter under catabolite-regulation is probably present only in bacteria closely related to E.coli.

Base Composition↗
PubMed2003

A single-nucleotide natural variation (U4 to C4) in an influenza A virus promoter exhibits a large structural change: implications for differential viral RNA synthesis by RNA-dependent RNA polymerase.

The influenza A virus promoter is recognized by the influenza A virus RNA-dependent RNA polymerase, and directs both transcription and replication of the viral RNA genome. Within the sequence of this promoter, flu strains exhibit a natural, unique variation, either a U or a C, at the fourth position from the 3' end. Promoters that contain a C residue (C4 promoter), which are invariably found in genome segments that encode the three RNA polymerase subunits (PB1, PB2 and PA), down-regulate transcription but activate genome replication. Here, we have determined the structure of the C4 promoter by NMR spectroscopy and compared it with the structure of the U4 promoter, which was determined previously. The structure of the internal loop in the C4 promoter is similar to that of the U4 promoter. However, the terminal stem of the C4 promoter is strikingly different from that of the U4 promoter. These structural data suggest that the internal loop is important for polymerase binding to the promoter, and the terminal stem is crucial for differential regulation of transcription and replication.

Base Sequence↗
PubMed2003

Synergistic activation of the human orphan nuclear receptor SHP gene promoter by basic helix-loop-helix protein E2A and orphan nuclear receptor SF-1.

The orphan nuclear receptor small heterodimer partner (SHP; NR0B2) is an unusual orphan nuclear receptor that lacks a conventional DNA-binding domain and acts as a modulator of transcriptional activities of a number of nuclear receptors. We have previously reported that the orphan nuclear receptor ERRgamma activates the SHP promoter. In this study, we have found that basic helix-loop-helix (bHLH) transcription factors, the E2A proteins (E47, E12 and E2/5), activated the human but not the mouse SHP promoter. In contrast, the tissue-specific E47 heterodimer partner BETA2 repressed the E47- mediated transactivation of the human SHP (hSHP) promoter. Using serial deletions and E-box mutant constructs of the hSHP promoter, we identified two E-boxes (E6 and E7) as E47-responsive E-boxes, which are not conserved in the mouse SHP promoter. Moreover, gel shift, chromatin immunoprecipitation (ChIP) and northern blot assays demonstrated that E47 directly binds to the hSHP promoter in vivo and in vitro and that Id proteins inhibited E47 binding to the hSHP promoter. Finally, we found that E47 and steroidogenic factor 1 (SF-1), a regulator of the SHP promoter, synergistically activate the human but not the mouse SHP promoter. Our findings suggest that the E2A proteins differentially regulate the human and mouse SHP promoters and cooperate with orphan nuclear receptor SF-1 for transcriptional activation of the hSHP promoter.

Animals↗
PubMed2005

Multiple promoters are a common feature of mitochondrial genes in Arabidopsis.

Mitochondrial genes in the plant Arabidopsis thaliana are transcribed by two phage-type RNA polymerases encoded in the nucleus. Little is known about cis-elements that are recognized by these enzymes and mediate the transcription of the Arabidopsis mitochondrial genome. Here, 30 transcription initiation sites of 12 mitochondrial genes and gene clusters have been determined using 5'-RACE and ribonuclease protection analysis of primary transcripts labelled in vitro by guanylyltransferase. A total of 9 out of 12 genes were found to possess multiple promoters, revealing for the first time that multiple promoters are a common feature of mitochondrial genes in a dicotyledonous plant. No differences in promoter utilization were observed between leaves and flowers, suggesting that promoter multiplicity reflects a relaxed promoter specificity rather than a regulatory role of promoter selection. Nearly half the identified transcription initiation sites displayed immediately upstream a CRTA core sequence, which was mostly seen within the previously described CRTAAGAGA promoter motif or a novel CGTATATAA promoter element. About as many promoters possessed an ATTA or RGTA core. Our data indicate that the majority of mitochondrial promoters in Arabidopsis deviate significantly from the nonanucleotide consensus derived earlier for dicot mitochondrial promoters.

Arabidopsis↗
PubMed1997

Efficiency of recombination by Cre transient expression in embryonic stem cells: comparison of various promoters.

The Cre-loxP recombination system of bacteriophage P1 is frequently utilized in genetic manipulation in embryonic stem (ES) cells. The level of Cre expression is critical to induce loxP site-specific recombination in ES cells. To compare the efficiency of recombination, we constructed four Cre expression vectors driven by different promoters: cytomegarovirus/chicken beta-actin (CAG) promoter, human polypeptide chain elongation factor 1alpha (hEF-1alpha) promoter, mouse phosphoglycerate kinase-1 (mPGK) promoter, and polyoma enhancer/herpes simplex virus thymidine kinase (MC1) promoter. We introduced these Cre expression vectors by electroporation into three ES cell lines carrying a single copy of CAG-loxP-chloramphenicol acetyltransferase (CAT) gene-loxP-beta-galactosidase (beta-gal) gene construct. Since the Cre-mediated recombination leads to excision of the CAT gene, the efficiency of recombination can be monitored as beta-gal expression. No selection system was used in the experiments. The maximum recombination frequency was obtained when the CAG promoter was used, followed by the hEF-1alpha promoter, the mPGK promoter and the MC1 promoter in order. These results indicate that the efficiency of recombination in transient expression system correlates with the promoter activity of Cre expression vector. Thus, it is important to choose the promoter for effective recombination by Cre.

Actins↗
PubMed2002

Consensus promoter identification in the human genome utilizing expressed gene markers and gene modeling.

Deciphering the human genome includes locating the promoters that initiate transcription and identifying the exons of genes. Many promoter prediction programs have been proposed, but when they are applied to extended regions of the genome, most of their predictions are false-positives. The extensive collection of gene transcript sequences is an important new source of information, which has not been used previously in promoter predictions. Our approach is to enhance the specificity of predictions by restricting the genomic regions that are searched using gene transcript alignments as anchors in the genome for gene modeling. We developed a consensus promoter prediction method combining previously developed algorithms with the GENSCAN gene modeling program. Our method, CONPRO (CONsensus PROmoter), identifies promoters with very high confidence, and the predicted promoters are guaranteed to be associated with genes. On our test data set, the method correctly detects promoters for approximately half of all human genes (37%-71%), and most predictions are true promoters (85%-90%). Applying our method to the human genome and human genes from the Unigene data set, we find the promoters for 13,744 genes. Of these, 6440 are genes with a functionally cloned mRNA, and 7304 are novel genes for which only expressed sequence tags (ESTs) are available. Candidate promoters for many novel genes will be a useful resource in elucidating complex biological response mechanisms.

5' Untranslated Regions↗
PubMed2003

Identification of promoter motifs involved in the network of phytochrome A-regulated gene expression by combined analysis of genomic sequence and microarray data.

Several hundred Arabidopsis genes, transcriptionally regulated by phytochrome A (phyA), were previously identified using an oligonucleotide microarray. We have now identified, in silico, conserved sequence motifs in the promoters of these genes by comparing the promoter sequences to those of all the genes present on the microarray from which they were sampled. This was done using a Perl script (called Sift) that identifies over-represented motifs using an enumerative approach. The utility of Sift was verified by analysis of circadian-regulated promoters known to contain a biologically significant motif. Several elements were then identified in phyA-responsive promoters by their over-representation. Five previously undescribed motifs were detected in the promoters of phyA-induced genes. Four novel motifs were found in phyA-repressed promoters, plus a motif that strongly resembles the DE1 element. The G-box, CACGTG, was a prominent hit in both induced and repressed phyA-responsive promoters. Intriguingly, two distinct flanking consensus sequences were observed adjacent to the G-box core sequence: one predominating in phyA-induced promoters, the other in phyA-repressed promoters. Such different conserved flanking nucleotides around the core motif in these two sets of promoters may indicate that different members of the same family of DNA-binding proteins mediate phyA induction and repression. An increased abundance of G-box sequences was observed in the most rapidly phyA-responsive genes and in the promoters of phyA-regulated transcription factors, indicating that G-box-binding transcription factors are upstream components in a transcriptional cascade that mediates phyA-regulated development.

Amino Acid Sequence↗
PubMed1994

Site-specific mutagenesis of the nodule-infected cell expression (NICE) element and the AT-rich element ATRE-BS2* of the Sesbania rostrata leghemoglobin glb3 promoter.

Sesbania rostrata leghemoglobin glb3 (Srglb3) promoter sequences responsible for expression in infected cells of transgenic Lotus corniculatus nodules were delimited to a 78-bp Dral-Hinfl fragment. This region, which is located between coordinates -194 to -116 relative to the start codon of the Srglb3 gene, was named the nodule-infected cell expression (NICE) element. Insertion of the NICE element into the truncated nopaline synthase promoter was found to confer a nodule-specific expression pattern on this normally root-enhanced promoter. Within the NICE element, three distinct motifs ([A]AAAGAT, TTGTCTCTT, and CACCC[T]) were identified; they are highly conserved in the promoter regions of a variety of plant (leg)hemoglobin genes. The NICE element and the adjacent AT-rich element (ATRE-BS2*) were subjected to site-directed mutagenesis. The expression patterns of nine selected Srglb3 promoter fragments carrying mutations in ATRE-BS2* and 19 with mutations in the NICE element were examined. Mutations in ATRE-BS2* had varying effects on Srglb3 promoter activity, ranging from a two- to threefold reduction to a slight stimulation of activity. Mutations in the highly conserved (A)AAAGAT motif of the NICE element reduced Srglb3 promoter activity two- to fourfold, whereas mutations in the TCTT portion of the TTGTCTCTT motif virtually abolished promoter activity, demonstrating the essential nature of these motifs for Srglb3 gene expression. An A-to-T substitution in the CACCC(T) motif of the NICE element also abolished Srglb3 promoter activity, while a C-to-T mutation at position 4 resulted in a threefold reduction of promoter strength. The latter phenotypes resemble the effect of similar mutations in the conserved CACCC motif located in the promoter region of mammalian beta-globin genes. The possible analogies between these two systems will be discussed.

Base Sequence↗
PubMed2004

Promoting and protecting the health of children and young people.

BACKGROUND: The health-related behaviours adopted by children and young people can have both immediate and long-term health effects. Health promotion interventions that target children and young people can lay the foundations of a healthy lifestyle that may be sustained into adulthood. This paper is based on a selective review of evidence relating to health promotion in childhood, carried out to support the external working group on the 'Healthy Child' module of the Children's National Service Framework. METHODS: This is a selective review of mainly secondary research. It focuses on injury prevention, support for parenting and the promotion of good mental health, and promoting a healthy diet and physical activity amongst children and young people. FINDINGS: In many areas, the quality of primary research into health promotion interventions aimed at children and young people is poor. Interventions are heterogeneous and not described in sufficient detail. Sample sizes tend to be small, and there are commonly problems of bias. Despite these difficulties, there is good evidence for a range of interventions, including (1) area road safety schemes; (2) combining a variety of approaches to the promotion of the use of safety equipment, including legislation and enforcement, loan/assisted purchase/giveaway schemes, education, fitting and maintenance of safety equipment; (3) school-based mental health promotion; (4) parenting support; (5) interventions that promote and facilitate 'lifestyle' activity for children, such as walking and cycling to school, and those that aim to reduce sedentary behaviours such as parent education to reduce the time children spend watching TV and using computers; and (6) controlling advertising of unhealthy food that is aimed at children. CONCLUSIONS: There are effective interventions to promote and protect the health of children and young people that require action across the five areas described in the Ottawa Charter. Health, social care and education services have a direct role in the delivery of many of these interventions and, in other areas, a role in collaborative work with other agencies, in lobbying for policy change and in raising the profile of child health promotion. Further research is needed using larger study populations, and closely defined interventions, both targeted and universal, in order to fill some of the current gaps in the evidence base for health promotion in children and young people.

Accident Prevention↗
PubMed2006

Evaluation of the activities of the medfly and Drosophila hsp70 promoters in vivo in germ-line transformed medflies.

The promoter of the hsp70 gene of Drosophila melanogaster has been widely used for the expression of foreign genes in other insects. It has been generally assumed that because this gene is highly conserved, its promoter will function efficiently in other species. We report the results of a quantitative comparison of the activities of the medfly and D. melanogaster hsp70 promoters in vivo in transformed medflies. We constructed transformed lines containing the lacZ reporter gene under the control of the two promoters by using Minos-mediated germ-line transformation. The activity of each promoter was evaluated in 15 transformed lines by beta-galactosidase quantitative assays. The heat-inducible activity of the medfly promoter was found several times higher than the respective activity of the heterologous D. melanogaster promoter. These results were confirmed by northern blot analysis and indicate that the D. melanogaster promoter does not work efficiently in medfly. The -263/+105 medfly promoter region that was used in this study was found able to drive heat shock expression of the lacZ reporter gene in all stages of medfly, except early embryonic stages, in a similar fashion to the endogenous hsp70 genes. However the heat inducible RNA levels driven from this promoter region were significantly lower than the endogenous hsp70 RNA levels, suggesting that additional upstream and/or downstream sequences to the -263/+105 region may be necessary for optimum function of the medfly hsp70 promoter in vivo.

Animals↗
PubMed1989

Functional characterization of a putative internal promoter sequence between the 16S and the 23S RNA genes within the Escherichia coli rrnB operon.

Transcription of ribosomal RNAs in Escherichia coli is started from two strong tandem promoters, P1 and P2. It is known, however, that internal promoter-like structures occur and in a recent report (Mankin et al., 1987) a promoter sequence Pi within the 16S and 23S RNA spacer region showing good homology to the prokaryotic consensus promoter structure was identified. It was proposed that this putative promoter has a possible function in the transcription of ribosomal RNAs in E. coli. Fusion of various DNA fragments containing the putative promoter sequence and different parts of the 16S/23S spacer region as well as the 23S RNA to the galactokinase gene allowed us to assess the functional activity of the promoter in vivo. To determine any growth rate dependent function of the putative promoter, the measurements were performed under different growth conditions. The promoter activity did not exceed 7% of the lac promoter under in vivo assay conditions. In addition, transcription starting at the promoter Pi did not proceed through the entire 23S RNA gene. We conclude, therefore, that transcription from Pi does not contribute significantly to the synthesis of ribosomal RNAs. Thus its functional significance, if any, remains elusive.

DNA, Bacterial↗
PubMed1994

Structural and functional analysis of the promoter region involved in full expression of the cryIIIA toxin gene of Bacillus thuringiensis.

The promoter region of the cryIIIA toxin gene of Bacillus thuringiensis is composed of at least three domains: an upstream region extending from nucleotide positions -635 to -553 (with reference to the translational start codon of cryIIIA), an internal region extending from nucleotide positions -553 to -367, and a downstream region extending from nucleotide position -367 to +18. Deletion analysis and transcriptional fusions to the lacZ gene indicate that full expression of cryIIIA requires the association of the upstream and the downstream region. Primer extension experiments reveal a major cryIIIA transcript (designated T-129) starting at nucleotide position -129 and another transcript (designated T-558) starting at nucleotide position -558. Mutation in the -35 region of the promoter responsible for the initiation of T-558 indicates that the upstream promoter is essential for full expression of cryIIIA, although not sufficient. Deletion of the DNA region carrying the previously described cryIIIA promoter does not affect full expression of cryIIIA and does not modify the 5' end of T-129. Taken together, these results indicate that the 5' end of T-129 is not a trnascriptional start site. Therefore, we propose that T-129 results from the processing of the mRNA initiated at the upstream promoter (T-558), generating a stable mRNA with a 5' extremity at nucleotide position -129. From primer extension analysis and transcriptional fusions to lacZ, it appears that the upstream promoter is weakly but significantly expressed during the vegetative phase of growth, is activated at the onset of sporulation and remains active at least until t5. However, unlike the promoters of other cry genes, this promoter is similar to sigma A-dependent promoters rather than sporulation-specific promoters. This promoter may therefore be transcribed by the E sigma A form of RNA polymerase. Activation at the onset of sporulation could result from the disappearance of a repressor, or the appearance of a stationary-phase-specific activator.

Bacillus thuringiensis↗
PubMed1996

Differential binding of BvgA to two classes of virulence genes of Bordetella pertussis directs promoter selectivity by RNA polymerase.

Transcription of virulence genes of Bordetella pertussis is co-ordinately regulated by the BvgA and BvgS proteins, which are members of the two-component family of bacterial signal-transduction proteins. BvgS is the transmembrane sensor and BvgA the transcriptional regulator. By gel mobility shift assays we demonstrate that phosphorylated BvgA (BvgA approximately P) forms distinct complexes with the filamentous haemagglutinin (PFHA) promoter DNA at different BvgA approximately P: DNA ratios. DNase I protection analyses show that phosphorylation of BvgA not only enhances affinity of the protein for the binding sites of the PFHA and bvgP1 promoters, but it extends significantly the bound region towards position -35 of these promoters. Conversely, a 10-fold higher amount of BvgA approximately P is required for binding to a large DNA region, from -168 to -60, of the pertussis toxin (Ptox) promoter sequence. These findings suggest that the molecular interaction of BvgA approximately P with the Ptox promoter is different from its interaction with the PFHA and bvgP1 promoters. The sigma 70 Escherichia coli RNA polymerase (RNP) does not bind to the bvg-regulated promoters. However, following the formation of a BvgA approximately P-promoter complex, the E. coli RNP specifically recognizes and binds to the bvg-regulated promoters. Thus, BvgA approximately P exerts its action at the level of promoter recognition by directing promoter selectivity by RNP.

Adhesins, Bacterial↗
PubMed2005

Usage of an intronic promoter for stable gene expression in Saccharomyces cerevisiae.

AIMS: To construct expression vectors capable of switching promoters under different metabolic circumstances to obtain stable gene expression. METHODS AND RESULTS: In this study, we designed a series of constructs for the expression of the chicken lactate dehydrogenase (cldh) gene under the control of galactose-inducible GAL1 promoter and the high glucose-inducible HXT1 promoter in Saccharomyces cerevisiae. In one construct, the HXT1 promoter was placed between artificial splicing sequences to function as an intronic promoter. We checked all constructs for the usage of promoters by reverse transcriptional polymerase chain reaction and assayed the expression level of the reporter gene under different culturing conditions. In the presence of galactose, when the GAL1 promoter was linked with the intronic HXT1 promoter, the cldh gene showed 1.5-fold activity compared with single GAL1 promoter, while in the presence of glucose, the construct showed over twofold activity compared with that without splicing sequences. CONCLUSION: The intronic HXT1 promoter could be induced by the presence of high glucose concentration. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report detailing the use of an intronic promoter in the construction of stable expression vectors and the novel system could serve as a model of expression vectors for fermentation or other purposes.

Animals↗
PubMed2004

Significance of the tissue kallikrein promoter and transforming growth factor-beta1 polymorphisms with renal progression in children with vesicoureteral reflux.

BACKGROUND: Tissue kallikrein regulates blood circulation. Low urinary kallikrein excretion was associated with hypertension and renal disease in blacks. The polymorphic KLK1 promoter includes -130 GN coupled with multiple single base substitutions. The -130 G12 allele in the KLK1 promoter was associated with lower transcriptional activity and hypertensive end-stage renal disease (ESRD) in blacks. Transforming growth factor-beta1 (TGF-beta1) regulates matrix production, and induces fibrosis in a variety of tissues. High circulating TGF-beta1 levels mediating renal fibrosis and loss of function in transgenic mice. The -509 T allele in the TGF-beta1 promoter showed marginally higher transcriptional activity, and was associated with increased TGF-beta1 production in humans. The aim of this study was to investigate whether the tissue KLK1 promoter and TGF-beta1 polymorphism are involved in primary vesicoureteric reflux (VUR) with renal progression in children. METHODS: Seventy-four primary VUR children were studied with regular annual follow-up for more than 18 years, all of them more than grade II (diagnosed by voiding cystourethroradiography). All of them were born before 1984. Patients were classified into two groups according to the renal function with progressive deterioration or not. Patients with baseline creatinine clearance (CCr) less than 25 mL/min were defined as having chronic renal insufficiency (CRI). The TGF-beta1 -509 T-C polymorphism was analyzed by Bsu36I restriction fragment length polymorphism (RFLP)-polymerase chain reaction (PCR). In KLK1 promoter, the -130 GN length polymorphism and multiple single base substitutions were analyzed by electrophoresis of fluoresced PCR products in sequencing gels, single strand conformation polymorphism (SSCP), allele-specific PCR, and DNA sequencing. Patients' TGF-beta1 and KLK1 promoter polymorphisms were evaluated for association with VUR susceptibility and progression in Taiwanese children. Annual echocardiography study was used to evaluate left ventricular mass index (LVMI). RESULTS: Four alleles were identified in the complex KLK1 promoter: A (-130 G10), B (-130 G2CG7), H (-130 G11), and K (-130 G12). The polymorphic KLK1 promoter showed no association with VUR susceptibility. However, the frequency distribution of KLK1 promoter among VUR patients with or without CRI (A, 50.0% and 67.5%; B, 17.9% and 8.3%; H, 14.3% and 18.3%; K, 17.9% and 5.8%, respectively) was statistically different (P = 0.008). Significantly higher K allele frequency was present in primary VUR with CRI children, as it was in the renal survival curve study. A significant increase of LVMI was also found in the A allele group compared with the non-A allele group of KLK1 promoter gene at the age of 18 years old with renal progression. The TGF-beta1 gene polymorphism was determined, and we found significant over-representation of the TT genotype in primary VUR patients with CRI compared with normal renal function (P= 0.0035). CONCLUSION: The K allele of KLK1 promoter and TT genotype of TGF-beta1 may be a genetic KLK1 -130 GN and -128 G-C, and the susceptibility factor contributing to progressive renal deterioration in Taiwanese primary VUR children.

Alleles↗
PubMed2001

Xanthomonas oryzae pv. oryzae recA is transcribed and regulated from multiple promoters.

Transcription regulation of Xanthomonas oryzae pv. oryzae recA was characterized. Primer extension experiments showed that recA is transcribed from three promoters designated P1, P2 and P3. The sequences of -10 and -35 regions of these promoters have moderate homology to the proposed consensus sequence for a Xanthomonas promoter. Putative SOS boxes were identified in the vicinity of P1 and P2 promoters. Deletion analysis and in vivo monitoring of promoter activity of these promoters revealed that the three promoters have different characteristics. P1 and P2 show stress-inducible high and low promoter strengths respectively. P3 is a non-inducible moderate promoter strength. These promoters are regulated by two SOS boxes. The multiplicity of promoters and SOS boxes provides back-up systems to ensure proper regulation of recA.

Base Sequence↗
PubMed1990

Function of transfected globin promoters and the globin locus activator in K562 erythroleukemia cells.

We have examined the importance of cis-acting regulatory elements within the human gamma-globin gene promoter and the globin locus activating region in K562 cells. A gamma-globin or beta-globin promoter fragments were fused with the neomycin phosphotransferase gene in a plasmid-based vector (gamma-neo or beta-neo) and transiently transfected by electroporation into K562 cells. Correctly initiated gamma-neo or beta-neo transcripts were detected with an S1 nuclease protection assay that was internally controlled for transfection efficiency and RNA content. We first optimized the conditions for electroporation and then determined that a gamma-globin promoter fragment extending from -299 and +36 was active in the assay but that a beta-globin promoter extending from -375 to +46 was inactive. Deletion of the gamma-globin promoter to -199 did not affect promoter function, but deletion to -160 reduced promoter strength to 70% of that of control. Additional deletion to position -130 reduced promoter strength to 19% of the control value, and to position -61, 8.7% of the control value. Three gamma-globin promoters containing mutations associated with hereditary persistence of fetal hemoglobin (HPFH), -202 C----G, -196 C----T and -117 G----A, were not overexpressed in K562 cells, consistent with the hypothesis that these promoters are not overexpressed in fetal erythroblasts, only in adult red cells. When the beta-globin locus activating region (LAR) was added to a wild-type or an HPFH gamma-neo plasmid, the abundance of correctly initiated gamma-neo transcripts increased dramatically. However, beta-neo expression could not be activated by the LAR in K562 cells. These studies should allow us to further dissect the interactive roles of globin promoters and enhancers in K562 cells.

Cell Line↗