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At least 595 records · Page 33Linked to original sources
PubMed2005

Promoting physical activity in Australian general practices: a randomised trial of health promotion advice versus hypertension management.

A randomised controlled trial was conducted to determine if physicians' advice to promote physical activity to patients was more effective if the advice was tailored to the management of hypertension, compared with more general health promotion advice. Participants included inactive 40- to 70-year-old patients visiting the physicians' during study recruitment period. Physicians provided verbal physical activity advice and written materials, both tailored to either general health promotion messages or specifically as a means for treating or managing hypertension. Seventy-five physicians and 98% (767/780) of screened eligible patients participated in the study. Differences between intervention and control groups self-reported physical activity were assessed over 6 months. Follow-up response rates were 92 and 84% at the 2- and 6-month assessments. There were no consistent, significant differences between groups at the 2- or 6-month assessments. Thus, neither intervention strategy resulted in significant changes in patients self-reported physical activity, regardless of the whether the advice was tailored to hypertension management or general health promotion advice.

Adult↗
PubMed2000

The molecular basis for genetic polymorphism of human deoxyribonuclease II (DNase II): a single nucleotide substitution in the promoter region of human DNase II changes the promoter activity.

Deoxyribonuclease II (DNase II) levels in human vary depending on whether the individual has the DNASE2*H (high) allele or the DNASE2*L (low) allele. We examined the promoter activity of the 5'-flanking region of each of these alleles by transient transfection luciferase assay. DNASE2*H had 5-fold higher promoter activity than DNASE2*L in human hepatoma HepG2 cell. Comparison of the nucleotide sequences of the proximal promoter regions revealed a G to A transition at position -75; G and A residues were assigned to DNASE2*H and *L, respectively. Since no differences were found between the open reading frame sequences of these alleles, it is likely that the A-75G transition causes the allelic difference in the promoter activity of the gene, underlying the genetic polymorphism.

Alleles↗
PubMed1996

E. coli SSB activates N4 virion RNA polymerase promoters by stabilizing a DNA hairpin required for promoter recognition.

Bacteriophage N4 virion RNA polymerase transcription of double-stranded promoter-containing DNAs requires supercoiled template and E. coli single-stranded DNA-binding protein (EcoSSB); other single-stranded DNA-binding proteins cannot substitute. The DNA determinants of virion RNA polymerase binding at the promoter comprise a small template-strand hairpin. The requirement for EcoSSB is surprising, since single-stranded DNA-binding proteins destabilize hairpin structures. DNA footprinting of EcoSSB on wild-type and mutant promoters indicates that EcoSSB stabilizes the template-strand hairpin owing to the hairpin-loop sequences. Other single-stranded DNA-binding proteins destabilize the promoter hairpin, explaining the specificity of EcoSSB activation. We conclude that EcoSSB activates transcription by providing the appropriate DNA structure for polymerase binding. The existence of small hairpins stable to single-stranded protein binding suggests a novel mechanism that provides structural determinants for specific recognition in single-stranded DNA transactions by an otherwise nonspecific DNA-binding protein.

Base Sequence↗
PubMed1997

Promoting in tandem: the promoter for telomere transposon HeT-A and implications for the evolution of retroviral LTRs.

HeT-A elements are non-long terminal repeat (non-LTR) retrotransposons found in head-to-tail arrays on Drosophila chromosome ends, where they form telomeres. We report that HeT-A promoter activity is located in the 3' end of the element, unlike the 5' location seen for other non-LTR retrotransposons. In HeT-A arrays the 3' sequence of one element directs transcription of its downstream neighbor. Because the upstream promoter has the same sequence as the 3' end of the transcribed element, the HeT-A promoter is effectively equivalent to a 5' LTR in both structure and function. Retroviruses and LTR retrotransposons have their promoters and transcription initiation sites in their 5' LTRs. Thus HeT-A appears to have the structure of an evolutionary intermediate between non-LTR and LTR retrotransposons.

Animals↗
PubMed2003

Structure and promoter activity of the human glia maturation factor-gamma gene: a TATA-less, GC-rich and bidirectional promoter.

Human glia maturation factor-gamma (hGMFG) was recently identified as a gene that is homologous to glia maturation factor-beta (GMFB). In this study, we determined the organization of the 9.5-kb hGMFG gene and characterized its promoter activity. The 5'-flanking region of the first exon has putative elements for binding transcription factors Sp-1, GATA-1, AML-1a, Lyf-1 and Ets-1, but there were no TATA or CAAT boxes within a 226-bp sequence upstream from the initiation codon. Primer extension analysis and 5'RACE (rapid amplification of cDNA 5' ends) identified multiple transcription initiation sites within the region -84 to -70 nucleotides from the first ATG codon in a Kozak consensus sequence. A core promoter region was determined by transfecting a series of deletion constructs with a dual luciferase reporter system into rat astrocyte-derived ACT-57 cells. We found that 226 bp of the core promoter region exhibited bidirectional promoter activity.

Base Sequence↗
PubMed1998

Deletional and mutational analyses of the human CD4 gene promoter: characterization of a minimal tissue-specific promoter.

In recent years, considerable interest has arisen in understanding the mechanisms of the CD4 gene transcriptional control which resulted in the identification of a promoter, enhancers and a silencer. While the murine CD4 gene promoter has been well studied, little is known about its human counterpart that we previously identified as a 1.1 kb region. Here, we show that the -170/+20 region represents a minimal tissue-specific promoter with a size compatible with its inclusion in viral vectors for gene therapy. In addition, mutational analyses provided evidence that this fragment contains three regions critical for transcriptional activity: an initiator-like sequence, an Ets consensus site and an ATF consensus site. The latter site is absent in the murine promoter, suggesting that some differences exist between the transcriptional control of the human and murine CD4 genes.

Animals↗
PubMed1997

Analysis of genes for human snRNP Sm-D1 protein and identification of the promoter sequence which shows segmental homology to the promoters of Sm-E and U1 snRNA genes.

The Sm core proteins of U1, U2, U4/U6 and U5 snRNPs include B(B1), B'(B2), N(B3), D1, D2, D3, E, F and G polypeptides. We have isolated genomic clones encoding the Sm-D1 protein using the Sm-D1 cDNA as probe. Southern blotting and DNA sequencing analysis of these clones revealed the presence of an Sm-D1 multigene family in the human genome. Three gene members have been identified. Two of the genes are without introns and contain mutations compared to the cDNA sequence. They appear to be processed pseudogenes. The third gene, termed SNRPD1, shares 100% identity to the cDNA sequence including both 5'- and 3'-untranslated regions (UTR); it contains three introns. Analysis of the 5'-flanking region of the SNRPD1 gene revealed promoter activity, suggesting this is the functional gene that encodes the Sm-D1 protein. The promoter activity was localized in a 0.38 kb PstI fragment using CAT reporter gene fusion assays. Addition of an SV40 enhancer element did not enhance the transcription directed by that fragment. Sequence comparison of the 0.38 kb promoter sequence with the promoters of the Sm-E gene and U1 snRNA genes revealed several homologous motifs, suggesting that genes encoding the snRNP components may be coordinately regulated.

Amino Acid Sequence↗
PubMed2000

Distinct promoter elements mediate endodermal and mesodermal expression of the HNF1alpha promoter in transgenic Xenopus.

The gene encoding the tissue specific transcription factor HNF1alpha is expressed in vertebrates in tissues of endodermal origin such as the liver and the gut as well as in the kidney, a mesoderm derived organ. Using a 6 kb HNF1alpha promoter fragment linked to GFP we observed green fluorescence in transgenic embryos restricted to the liver and gut as well as to the pronephros, the embryonic kidney. By deletion and mutation analysis of the HNF1alpha promoter we succeeded in dissecting the HNF1alpha promoter into two entities that are either active in the endoderm or the mesoderm. In conclusion, our data establish that the generation of transgenic Xenopus allows the functional dissection of promoters in the context of the entire organism.

Animals↗
PubMed2003

Identification of the human HEX1/hExo1 gene promoter and characterization of elements responsible for promoter activity.

HEX1/hExo1 is a Class III nuclease of the RAD2 family with 5' to 3' exonuclease and flap structure-specific endonuclease activities. HEX1/hExo1 is expressed at low levels in a wide variety of tissues, but at higher levels in fetal liver and adult bone marrow, suggesting HEX1/hExo1 is important for hematopoietic stem cell development. A putative HEX1/hExo1 promoter fragment extending from -6240 to +1600bp exhibits cell-type specific activity in transient transfection assays. This fragment directs high luciferase reporter gene expression in the hematopoietic cell line K562, chronic myelogenous leukemia cells, but low luciferase expression in the non-hematopoietic cell line HeLa, human cervical carcinoma cells. Deletion studies identified a fragment spanning -688 to +1600bp that exhibits full transcriptional activity while a slightly shorter fragment from -658 to +1600bp exhibits significantly decreased promoter activity. In vitro binding assays revealed DNA-binding activities that interact with -687 to -681bp and -665 to -658bp elements. Oligonucleotide competition and antibody disruption studies determined that the transcription factor CREB-1 recognizes the -687 to -681bp element, while transcription factors Sp1 and Sp3 recognize the -665 to -658bp element. Mutation of either the CREB-1 or Sp1/Sp3 binding sites dramatically reduces HEX1/hExo1 promoter activity and elimination of both elements abolishes promoter activity.

Base Sequence↗
PubMed1995

Pathway of promoter melting by Bacillus subtilis RNA polymerase at a stable RNA promoter: effects of temperature, delta protein, and sigma factor mutations.

Bacillus subtilis RNA polymerase (RNAP) contains a catalytic core (beta beta' alpha 2; or E) associated with one of several sigma factors, which determine promoter recognition, and delta protein, which enhances promoter selectivity. We have shown previously that specific mutations in sigma A region 2.3, or addition of delta, decrease the ability of RNAP to melt the ilv-leu promoter. Here we extend these studies to a stable RNA promoter, PtmS, which controls transcription of seven tRNA genes. KMnO4 footprinting was used to visualize DNA melting at PtmS as a function of both temperature and the protein composition of the RNAP holoenzyme. We propose that the pathway leading to productive initiation includes several intermediates: a closed complex (RPc), a complex in which DNA melting has nucleated within the conserved TATA element (RPn), and an open complex in which DNA-melting extends to at least -4 (RPo1). RNAP reconstituted with either of two mutant sigma A proteins, Y189A and W192A, was defective for both the nucleation and propagation of the transcription bubble while a third sigma A mutant, W193A, allows normal nucleation of DNA-melting, but does not efficiently propagate the melted region downstream.

Amino Acid Sequence↗
PubMed2000

Studies of promoter recognition and start site selection by T7 RNA polymerase using a comprehensive collection of promoter variants.

We have examined the behavior of T7 RNA polymerase (RNAP) at a set of promoter variants having all possible single base pair (bp) substitutions. The polymerase exhibits an absolute requirement for initiation with a purine and a strong preference for initiation with GTP vs ATP. Promoter variants that would require initiation at the normal start site (+1) with CTP or UTP result in a shift in initiation to +2 (with GTP). However, the choice of start site is little affected by base substitutions elsewhere in the initiation region. Furthermore, when the initiation region is shifted either one nucleotide (nt) closer or 1 nt further away from the binding region, transcription still begins the same distance downstream. These results indicate that the sequence around the start site is of little importance in start site selection and that initiation is directed a minimum distance of 5 nt downstream from the binding region. At promoters that initiate with +1 GGG, T7 RNAP synthesizes a ladder of poly(G) products as a result of slippage of the transcript on the three C residues in the template strand from +1 to +3. At promoter variants in which there is an opportunity to form a longer RNA-DNA hybrid, this G-ladder is enhanced and extended. This observation is not in agreement with recent suggestions that the RNA-DNA hybrid in the initiation complex cannot extend further than 3 bps upstream from the active site [Cheetham, G., Jeruzalmi, D., and Steitz, T. A. (1999) Nature 399, 80-83].

Bacteriophage T7↗
PubMed2003

Elevation of transgene expression level by flanking matrix attachment regions (MAR) is promoter dependent: a study of the interactions of six promoters with the RB7 3' MAR.

We have analyzed effects of a matrix attachment region (MAR) from the tobacco RB7 gene on transgene expression from six different promoters in stably transformed tobacco cell cultures. The presence of MARs flanking the transgene increased expression of constructs based on the constitutive CaMV 35S, NOS, and OCS promoters. Expression from an induced heat shock promoter was also increased and MARs did not cause expression in the absence of heat shock. There was also no effect of MARs on the pea ferredoxin promoter, which is not normally expressed in this cell line. Importantly, most transgenes flanked by RB7 MAR elements showed a large reduction in the number of low expressing GUS transformants relative to control constructs without MARs.

3' Untranslated Regions↗
PubMed2003

The nectary-specific pattern of expression of the tobacco Nectarin I promoter is regulated by multiple promoter elements.

The major protein secreted into the nectar of tobacco plants (Nectarin I) is a germin-like protein that has superoxide dismutase activity. We have isolated the gene encoding Nectarin I (NECI) and analyzed the expression of a chloramphenicol acetyl transferase (CAT) marker gene driven by its promoter in transgenic plants. Transgenic plant lines that expressed the CAT gene under control of the full-length NECI promoter showed high levels of CAT expression in mature floral nectaries. Tissue specificity of NECI-CAT expression demonstrated that the construct was expressed uniquely in nectaries with a small level of expression in ovary. Further, analysis of its temporal expression showed that the construct is expressed uniquely during those times when nectar is actively being secreted from flowers. An examination of the transcription start site verified that the initiation site of the NECI-CAT mRNA in transgenic plants is identical with that of the native gene in vivo. Two promoter deletion constructs were also prepared and analyzed. Analysis in transgenic plants revealed that the nectary-specific expression is the result of multiple promoter elements and suggests that nectar secretion and flower opening may be coordinately regulated.

Base Sequence↗
PubMed2001

Promoter I of the ovine acetyl-CoA carboxylase-alpha gene: an E-box motif at -114 in the proximal promoter binds upstream stimulatory factor (USF)-1 and USF-2 and acts as an insulin-response sequence in differentiating adipocytes.

Acetyl-CoA carboxylase-alpha (ACC-alpha) plays a central role in co-ordinating de novo fatty acid synthesis in animal tissues. We have characterized the regulatory region of the ovine ACC-alpha gene. Three promoters, PI, PII and PIII, are dispersed throughout 50 kb of genomic DNA. Expression from PI is limited to adipose tissue and liver. Sequence comparison of the proximal promoters of ovine and mouse PIs demonstrates high nucleotide identity and that they are characterized by a TATA box at -29, C/EBP (CCAAT enhancer-binding protein)-binding motifs and multiple E-box motifs. A 4.3 kb ovine PI-luciferase reporter construct is insulin-responsive when transfected into differentiated ovine adipocytes, whereas when this construct is transfected into ovine preadipocytes and HepG2 cells the construct is inactive and is not inducible by insulin. By contrast, transfection of a construct corresponding to 132 bp of the proximal promoter linked to a luciferase reporter is active and inducible by insulin in all three cell systems. Insulin signalling to the -132 bp construct in differentiated ovine adipocytes involves, in part, an E-box motif at -114. Upstream stimulatory factor (USF)-1 and USF-2, but not sterol regulatory element-binding protein 1 (SREBP-1), are major components of protein complexes that bind this E-box motif. Activation of the 4.3 kb PI construct in differentiated ovine adipocytes is associated with endogenous expression of PI transcripts throughout differentiation; PI transcripts are not detectable by RNase-protection assay in ovine preadipocytes, HepG2 cells or 3T3-F442A adipocytes. These data indicate the presence of repressor motifs in PI that are required to be de-repressed during adipocyte differentiation to allow induction of the promoter by insulin.

3T3 Cells↗
PubMed1997

Promoter elements required for phloem-specific gene expression from the RTBV promoter in rice.

Previous studies indicated that a DNA fragment comprising nucleotides (nt) -164 to +45 of the RTBV promoter is sufficient to drive phloem-specific expression of a reporter gene in transgenic rice plants. In addition, two potential cis elements, Box I (nt -3 to +5) and Box II (nt -53 to -39) were identified by DNA-protein interaction assays. In this study, the results of further in vivo studies involving mutagenesis of selected DNA sequences and analysis of expression of a reporter gene in transgenic rice plants revealed that, in addition to Box I and Box II, other elements are required for phloem-specific gene expression, among which are a direct repeat (ASL Box, nt -98 to -79) and a GATA motif (nt -143 to -135). All the these elements bind rice nuclear factors specifically, and mutations of the elements were identified that resulted in loss-of-competition in electrophoretic mobility shift assays. A DNA fragment comprising nt -164 to -32, which contains Box II, the ASL Box and the GATA motif, conferred phloem-specific reporter gene expression independent of Box I when fused to a heterologous CaMV 35S minimal promoter and introduced to transgenic rice plants. Studies that introduced point mutations suggested that in the context of the -103 to +45 promoter fragment, Box II and the ASL Box act synergistically to confer tissue-specific gene expression. Similar studies in the -164 to +45 promoter fragment indicated that the -164 to -103 region, which includes the GATA motif, contains sequences that are functionally redundant with those in the -103 to -32 region, including the ASL Box and Box II. It is concluded that these regions act additively to direct phloem-specific gene expression.

Badnavirus↗
PubMed2001

A novel RNA polymerase binding site upstream of the galactose promoter in Escherichia coli exhibits promoter-like activity.

RNA polymerase is known to bind and utilize the overlapping promoters P1 and P2 in Escherichia coli galactose operon. We have identified an additional specific site upstream of P2, where RNA polymerase binds in a heparin-resistant manner. Binding of polymerase to this site, termed P3, occurs simultaneous to its binding at P1/P2. We have located this P3 site by DNase I footprinting. A 63 base pair region centered around position - 100 with respect to galP1 is protected by polymerase. Interestingly, a Pribnow box TATAAT is present within this protected region (-103 to -108). We have shown that transcription occurs from P3 in vitro. Primer extension analysis provides direct evidence that P3 is transcribed in vivo. The start site of transcription has been mapped at -96 position relative to galP1. beta-galactosidase assays with different gal promoter constructs reveal that while P3 alone functions as a weak in vivo promoter, it has a synergistic effect on transcription from the gal operon, since deletion of P3 or specifically mutating its -10 region result in a substantial reduction in the gal promoter activity.

Binding Sites↗
PubMed2001

Health promotion-who, us? Developing health promoting emergency departments.

This paper describes the initiation of Victoria's Health Promoting Emergency Departments (HPEDs) Program, involving seven of Melbourne's Emergency Departments (EDs). The Program aims to integrate health promotion into the function of EDs, to complement the clinical and curative emergency care that they already provide. A development phase identified the means of achieving this and involved establishing a multi-disciplinary Steering Committee and engaging health promotion specialists to record the opportunities for health promotion within the ED context. A literature review, and consultations with ED staff (focus groups and surveys) and other stakeholders were conducted. A concept paper was published and the development phase findings were documented, including recommendations on the future of the Program.

Emergency Service, Hospital↗
PubMed2000

The specificity loop of T7 RNA polymerase interacts first with the promoter and then with the elongating transcript, suggesting a mechanism for promoter clearance.

During the early stages of transcription, T7 RNA polymerase forms an unstable initiation complex that synthesizes and releases transcripts 2-8 nt in length before disengaging from the promoter and isomerizing to a stable elongation complex. In this study, we used RNA small middle dotprotein and RNA small middle dotDNA crosslinking methods to probe the location of newly synthesized RNA in halted elongation complexes. The results indicate that the RNA in an elongation complex remains in an RNA small middle dotDNA hybrid for about 8 nt from the site of nucleotide addition and emerges to the surface of the enzyme about 12 nt from the addition site. Strikingly, as the transcript leaves its hybrid with the template, the crosslinks it forms with the RNA polymerase involve a portion of a hairpin loop (the specificity loop) that makes specific contacts with the binding region of the promoter during initiation. This observation suggests that the specificity loop may have a dual role in transcription, binding first to the promoter and subsequently interacting with the RNA product. It seems likely that association of the nascent RNA with the specificity loop facilitates disengagement from the promoter and is an important part of the process that leads to a stable elongation complex.

Bacteriophage T7↗