Sp1, a CAAT-binding factor, and the adenovirus major late promoter transcription factor interact with functional regions of the gamma-fibrinogen promoter

1988 ◽  
Vol 8 (6) ◽  
pp. 2628-2637
Author(s):  
J G Morgan ◽  
G Courtois ◽  
G Fourel ◽  
L A Chodosh ◽  
L Campbell ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Transcription Initiation ◽  
Initiation Site ◽  
Internal Deletion ◽  
Developmentally Regulated ◽  
Mrna Accumulation ◽  
Deletion Mutations ◽  
Functional Regions ◽  
Binding Factor

To study the factors which influence the coordinately and developmentally regulated expression of the three adjacent fibrinogen genes, we have defined the functional regions of the gamma-fibrinogen promoter and the proteins which bind to them. Using a series of 5' and internal deletion mutations, we found that sequences between 88 and 43 base pairs (bp) upstream of the gamma-fibrinogen transcription initiation site functioned in cis to direct properly initiated mRNA accumulation in transfected hepatocytes. The efficient function of these sequences was highly distance dependent, since transcriptional activity decreased by 92% when they were moved 32 bp upstream of the TATA box. We demonstrated that two known and one putative transcriptional factors interacted with this 47-bp sequence. The transcription factor Sp1 interacted with sequences between -51 and -46 as demonstrated by protection from DNase I digestion with the purified protein. Directly adjacent to the Sp1 site, between nucleotides -66 and -53, there was a sequence which bound a CAAT-binding factor. Finally, sequences just 5' to the CAAT factor-binding site interacted with the adenovirus major late transcriptional factor as previously demonstrated. Internal deletion mutations which disrupt these interactions diminished the activity of the promoter in vivo. One consequence of the interaction of these proteins is that a bend is placed in the DNA at or near their sites of interaction.

scholarly journals Sp1, a CAAT-binding factor, and the adenovirus major late promoter transcription factor interact with functional regions of the gamma-fibrinogen promoter.

1988 ◽  
Vol 8 (6) ◽  
pp. 2628-2637 ◽  
Author(s):  
J G Morgan ◽  
G Courtois ◽  
G Fourel ◽  
L A Chodosh ◽  
L Campbell ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Transcription Initiation ◽  
Initiation Site ◽  
Internal Deletion ◽  
Developmentally Regulated ◽  
Mrna Accumulation ◽  
Deletion Mutations ◽  
Functional Regions ◽  
Binding Factor

To study the factors which influence the coordinately and developmentally regulated expression of the three adjacent fibrinogen genes, we have defined the functional regions of the gamma-fibrinogen promoter and the proteins which bind to them. Using a series of 5' and internal deletion mutations, we found that sequences between 88 and 43 base pairs (bp) upstream of the gamma-fibrinogen transcription initiation site functioned in cis to direct properly initiated mRNA accumulation in transfected hepatocytes. The efficient function of these sequences was highly distance dependent, since transcriptional activity decreased by 92% when they were moved 32 bp upstream of the TATA box. We demonstrated that two known and one putative transcriptional factors interacted with this 47-bp sequence. The transcription factor Sp1 interacted with sequences between -51 and -46 as demonstrated by protection from DNase I digestion with the purified protein. Directly adjacent to the Sp1 site, between nucleotides -66 and -53, there was a sequence which bound a CAAT-binding factor. Finally, sequences just 5' to the CAAT factor-binding site interacted with the adenovirus major late transcriptional factor as previously demonstrated. Internal deletion mutations which disrupt these interactions diminished the activity of the promoter in vivo. One consequence of the interaction of these proteins is that a bend is placed in the DNA at or near their sites of interaction.

scholarly journals Sporulation Phenotype of a Bacillus subtilis Mutant Expressing an Unprocessable but Active σE Transcription Factor

2004 ◽  
Vol 186 (7) ◽  
pp. 1999-2005 ◽  
Author(s):  
Shonna McBride ◽  
W. G. Haldenwang
Keyword(s):  
Amino Acids ◽  
Transcription Factor ◽  
Bacillus Subtilis ◽  
Transcription Initiation ◽  
Stage Iii ◽  
Wild Type ◽  
Developmentally Regulated ◽  
Proprotein Processing ◽  
Residual Inhibition

ABSTRACT σE, a sporulation-specific sigma factor of Bacillus subtilis, is formed from an inactive precursor (pro-σE) by a developmentally regulated processing reaction that removes 27 amino acids from the proprotein's amino terminus. A sigE variant (sigE335) lacking 15 amino acids of the prosequence is not processed into mature σE but is active without processing. In the present work, we investigated the sporulation defect in sigE335-expressing B. subtilis, asking whether it is the bypass of proprotein processing or a residual inhibition of σE activity that is responsible. Fluorescence microscopy demonstrated that sigE335-expressing B. subtilis progresses further into sporulation (stage III) than do strains lacking σE activity (stage II). Consistent with its stage III phenotype, and a defect in σE activity rather than its timing, the sigE335 allele did not disturb early sporulation gene expression but did inhibit the expression of late sporulation genes (gerE and sspE). The Spo− phenotype of sigE335 was found to be recessive to wild-type sigE. In vivo assays of σE activity in sigE, sigE335, and merodiploid strains indicate that the residual prosequence on σE335, still impairs its activity to function as a transcription factor. The data suggest that the 11-amino-acid extension on σE335 allows it to bind RNA polymerase and direct the resulting holoenzyme to σE-dependent promoters but reduces the enzyme's ability to initiate transcription initiation and/or exit from the promoter.

SPT15, the gene encoding the yeast TATA binding factor TFIID, is required for normal transcription initiation in vivo

Cell ◽  
1989 ◽  
Vol 58 (6) ◽  
pp. 1183-1191 ◽  
Author(s):  
David M. Eisenmann ◽  
Catherine Dollard ◽  
Fred Winston
Keyword(s):  
Transcription Initiation ◽  
Gene Encoding ◽  
Binding Factor

Analysis of Saccharomyces cerevisiae his3 transcription in vitro: biochemical support for multiple mechanisms of transcription

1990 ◽  
Vol 10 (6) ◽  
pp. 2832-2839
Author(s):  
A S Ponticelli ◽  
K Struhl
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Transcriptional Activation ◽  
Transcription Initiation ◽  
Molecular Mechanisms ◽  
Initiation Site ◽  
Activator Proteins ◽  
Nuclear Extracts ◽  
Transcription In Vitro

The promoter region of the Saccharomyces cerevisiae his3 gene contains two TATA elements, TC and TR, that direct transcription initiation to two sites designated +1 and +13. On the basis of differences between their nucleotide sequences and their responsiveness to upstream promoter elements, it has previously been proposed that TC and TR promote transcription by different molecular mechanisms. To begin a study of his3 transcription in vitro, we used S. cerevisiae nuclear extracts together with various DNA templates and transcriptional activator proteins that have been characterized in vivo. We demonstrated accurate transcription initiation in vitro at the sites used in vivo, transcriptional activation by GCN4, and activation by a GAL4 derivative on various gal-his3 hybrid promoters. In all cases, transcription stimulation was dependent on the presence of an acidic activation region in the activator protein. In addition, analysis of promoters containing a variety of TR derivatives indicated that the level of transcription in vitro was directly related to the level achieved in vivo. The results demonstrated that the in vitro system accurately reproduced all known aspects of in vivo his3 transcription that depend on the TR element. However, in striking contrast to his3 transcription in vivo, transcription in vitro yielded approximately 20 times more of the +13 transcript than the +1 transcript. This result was not due to inability of the +1 initiation site to be efficiently utilized in vitro, but rather it reflects the lack of TC function in vitro. The results support the idea that TC and TR mediate transcription from the wild-type promoter by distinct mechanisms.

Identification of promoter elements necessary for transcriptional regulation of a human histone H4 gene in vitro

1985 ◽  
Vol 5 (2) ◽  
pp. 380-389
Author(s):  
S M Hanly ◽  
G C Bleecker ◽  
N Heintz
Keyword(s):  
Transcription Factor ◽  
Transcription Initiation ◽  
Initiation Site ◽  
S Phase ◽  
Histone H4 ◽  
Promoter Elements ◽  
Nuclear Extracts ◽  
Distal Promoter ◽  
Histone H4 Gene

We have examined the nucleotide sequences necessary for transcription of a human histone H4 gene in vitro. Maximal transcription of the H4 promoter requires, in addition to the TATA box and cap site, promoter elements between 70 and 110 nucleotides upstream from the transcription initiation site. These distal promoter elements are recognized preferentially in extracts from synchronized S-phase HeLa cells. The inability of non-S-phase nuclear extracts to recognize the H4 upstream sequences reflects a specific lack of a transcription factor which interacts with those sequences. These results indicate that the cell cycle regulation of human histone gene expression involves both a specific transcription factor and distal transcription signals in the H4 promoter.

scholarly journals Transcription of Drosophila Troponin I Gene Is Regulated by Two Conserved, Functionally Identical, Synergistic Elements

2004 ◽  
Vol 15 (3) ◽  
pp. 1185-1196 ◽  
Author(s):  
María-Cruz Marín ◽  
José-Rodrigo Rodríguez ◽  
Alberto Ferrús
Keyword(s):  
Transcription Initiation ◽  
Troponin I ◽  
Regulatory Element ◽  
Expression Patterns ◽  
In Silico Analysis ◽  
Initiation Site ◽  
Transcription Initiation Site ◽  
Upstream Regulatory Element ◽  
Transgenic Lines

The Drosophila wings-up A gene encodes Troponin I. Two regions, located upstream of the transcription initiation site (upstream regulatory element) and in the first intron (intron regulatory element), regulate gene expression in specific developmental and muscle type domains. Based on LacZ reporter expression in transgenic lines, upstream regulatory element and intron regulatory element yield identical expression patterns. Both elements are required for full expression levels in vivo as indicated by quantitative reverse transcription-polymerase chain reaction assays. Three myocyte enhancer factor-2 binding sites have been functionally characterized in each regulatory element. Using exon specific probes, we show that transvection is based on transcriptional changes in the homologous chromosome and that Zeste and Suppressor of Zeste 3 gene products act as repressors for wings-up A. Critical regions for transvection and for Zeste effects are defined near the transcription initiation site. After in silico analysis in insects (Anopheles and Drosophila pseudoobscura) and vertebrates (Ratus and Coturnix), the regulatory organization of Drosophila seems to be conserved. Troponin I (TnI) is expressed before muscle progenitors begin to fuse, and sarcomere morphogenesis is affected by TnI depletion as Z discs fail to form, revealing a novel developmental role for the protein or its transcripts. Also, abnormal stoichiometry among TnI isoforms, rather than their absolute levels, seems to cause the functional muscle defects.

scholarly journals Interactions of the Antizyme AtoC with Regulatory Elements of the Escherichia coli atoDAEB Operon

2007 ◽  
Vol 189 (17) ◽  
pp. 6324-6332 ◽  
Author(s):  
Meropi K. Matta ◽  
Efthimia E. Lioliou ◽  
Cynthia H. Panagiotidis ◽  
Dimitrios A. Kyriakidis ◽  
Christos A. Panagiotidis
Keyword(s):  
Escherichia Coli ◽  
Transcription Initiation ◽  
Response Regulator ◽  
Regulatory Elements ◽  
Initiation Site ◽  
Integration Host Factor ◽  
Chemical Mutagenesis ◽  
Dual Function

ABSTRACT AtoC has a dual function as both an antizyme, the posttranslational inhibitor of polyamine biosynthetic enzymes, and the transcriptional regulator of genes involved in short-chain fatty acid catabolism (the atoDAEB operon). We have previously shown that AtoC is the response regulator of the AtoS-AtoC two-component signal transduction system that activates atoDAEB when Escherichia coli is exposed to acetoacetate. Here, we show that the same cis elements control both promoter inducibility and AtoC binding. Chromatin immunoprecipitation experiments confirmed the acetoacetate-inducible binding of AtoC to the predicted DNA region in vivo. DNase I protection footprinting analysis revealed that AtoC binds two 20-bp stretches, constituting an inverted palindrome, that are located at −146 to −107 relative to the transcription initiation site. Analyses of promoter mutants obtained by in vitro chemical mutagenesis of the atoDAEB promoter verified both the importance of AtoC binding for the inducibility of the promoter by acetoacetate and the σ54 dependence of atoDAEB expression. The integration host factor was also identified as a critical component of the AtoC-mediated induction of atoDAEB.

scholarly journals Interaction of the Vaccinia Virus RNA Polymerase-Associated 94-Kilodalton Protein with the Early Transcription Factor

2009 ◽  
Vol 83 (23) ◽  
pp. 12018-12026 ◽  
Author(s):  
Zhilong Yang ◽  
Bernard Moss
Keyword(s):  
Transcription Factor ◽  
Rna Polymerase ◽  
Vaccinia Virus ◽  
Transcription Initiation ◽  
Elongation Factor ◽  
Small Subunit ◽  
Functional Link ◽  
Early Transcription

ABSTRACT A multisubunit RNA polymerase (RPO) encoded by vaccinia virus (VACV), in conjunction with specific factors, transcribes early, intermediate, and late viral genes. However, an additional virus-encoded polypeptide referred to as the RPO-associated protein of 94 kDa (RAP94) is tightly bound to the RPO for the transcription of early genes. Unlike the eight RPO core subunits, RAP94 is synthesized exclusively at late times after infection. Furthermore, RAP94 is necessary for the packaging of RPO and other components needed for early transcription in assembling virus particles. The direct association of RAP94 with NPH I, a DNA-dependent ATPase required for transcription termination, and the multifunctional poly(A) polymerase small subunit/2′-O-methyltransferase/elongation factor was previously demonstrated. That RAP94 provides a structural and functional link between the core RPO and the VACV early transcription factor (VETF) has been suspected but not previously demonstrated. Using VACV recombinants that constitutively or inducibly express VETF subunits and RAP94 with affinity tags, we showed that (i) VETF associates only with RPO containing RAP94 in vivo and in vitro, (ii) the association of RAP94 with VETF requires both subunits of the latter, (iii) neither viral DNA nor other virus-encoded late proteins are required for the interaction of RAP94 with VETF and core RPO subunits, (iv) different domains of RAP94 bind VETF and core subunits of RPO, and (v) NPH I and VETF bind independently and possibly simultaneously to the N-terminal region of RAP94. Thus, RAP94 provides the bridge between the RPO and proteins needed for transcription initiation, elongation, and termination.

scholarly journals Identification of a functional hypoxia-responsive element that regulates the expression of the egl nine homologue 3 (egln3/phd3) gene

2005 ◽  
Vol 390 (1) ◽  
pp. 189-197 ◽  
Author(s):  
Nuria Pescador ◽  
Yolanda Cuevas ◽  
Salvador Naranjo ◽  
Marisa Alcaide ◽  
Diego Villar ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcription Initiation ◽  
Initiation Site ◽  
Egg Laying ◽  
Dependent Manner ◽  
Low Oxygen ◽  
Consensus Sequences ◽  
Homologous Protein ◽  
Responsive Element

Low oxygen levels induce an adaptive response in cells through the activation of HIFs (hypoxia-inducible factors). These transcription factors are mainly regulated by a group of proline hydroxylases that, in the presence of oxygen, target HIF for degradation. The expression of two such enzymes, EGLN1 [EGL nine homologous protein 1, where EGL stands for egg laying defective (Caenorhabditis elegans gene)] and EGLN3, is induced by hypoxia through a negative feedback loop, and we have demonstrated recently that hypoxic induction of EGLN expression is HIF-dependent. In the present study, we have identified an HRE (hypoxia response element) in the region of the EGLN3 gene using a combination of bioinformatics and biological approaches. Initially, we isolated a number of HRE consensus sequences in a region of 40 kb around the human EGLN3 gene and studied their evolutionary conservation. Subsequently, we examined the functionality of the conserved HRE sequences in reporter and chromatin precipitation assays. One of the HREs, located within a conserved region of the first intron of the EGLN3 gene 12 kb downstream of the transcription initiation site, bound HIF in vivo. Furthermore, this sequence was able to drive reporter gene expression under conditions of hypoxia in an HRE-dependent manner. Indeed, we were able to demonstrate that HIF was necessary and sufficient to induce gene expression from this enhancer sequence.

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