scholarly journals Roles for Non-TATA Core Promoter Sequences in Transcription and Factor Binding

2000 ◽  
Vol 20 (10) ◽  
pp. 3608-3615 ◽  
Author(s):  
Branden S. Wolner ◽  
Jay D. Gralla
Keyword(s):  
Rna Polymerase Ii ◽  
Core Promoter ◽  
Core Region ◽  
Promoter Activation ◽  
Promoter Sequences ◽  
The Core ◽  
General Transcription Factors ◽  
Specific Effects ◽  
Transcription In Vitro

ABSTRACT Sequence blocks within the core region were swapped among RNA polymerase II promoters to explore effects on transcription in vitro. The pair of blocks flanking TATA strongly influenced general transcription, with an additional effect on promoter activation. These flanking elements induced a change in the ratio of activated to basal transcription, whereas swapping TATA and initiator sequences only altered general transcription levels. Swapping the flanking blocks influenced binding by general transcription factors TBP and TFIIB. The results suggest that the architecture of the extended core sequence is important in determining promoter-specific effects on both general transcription levels and the tightness of regulation.

scholarly journals Selective Promoter Recognition by Chlamydial σ28 Holoenzyme

2006 ◽  
Vol 188 (21) ◽  
pp. 7364-7377 ◽  
Author(s):  
Li Shen ◽  
Xiaogeng Feng ◽  
Yuan Yuan ◽  
Xudong Luo ◽  
Thomas P. Hatch ◽  
...  
Keyword(s):  
Core Promoter ◽  
Sigma Factors ◽  
Promoter Sequences ◽  
E Coli ◽  
The Core ◽  
Recognition Specificity ◽  
Promoter Recognition ◽  
Transcription In Vitro

ABSTRACT The σ transcription factor confers the promoter recognition specificity of RNA polymerase (RNAP) in eubacteria. Chlamydia trachomatis has three known sigma factors, σ66, σ54, and σ28. We developed two methods to facilitate the characterization of promoter sequences recognized by C. trachomatis σ28 (σ28 Ct). One involved the arabinose-induced expression of plasmid-encoded σ28 Ct in a strain of Escherichia coli defective in the σ28 structural gene, fliA. The second was an analysis of transcription in vitro with a hybrid holoenzyme reconstituted with E. coli RNAP core and recombinant σ28 Ct. These approaches were used to investigate the interactions of σ28 Ct with the σ28 Ct-dependent hctB promoter and selected E. coli σ28 (σ28 Ec)-dependent promoters, in parallel, compared with the promoter recognition properties of σ28 EC. Our results indicate that RNAP containing σ28 Ct has at least three characteristics: (i) it is capable of recognizing some but not all σ28 EC-dependent promoters; (ii) it can distinguish different promoter structures, preferentially activating promoters with upstream AT-rich sequences; and (iii) it possesses a greater flexibility than σ28 EC in recognizing variants with different spacing lengths separating the −35 and −10 elements of the core promoter.

scholarly journals Mutational Analysis of the phoD Promoter in Bacillus subtilis: Implications for PhoP Binding and Promoter Activation of Pho Regulon Promoters

1999 ◽  
Vol 181 (7) ◽  
pp. 2017-2025 ◽  
Author(s):  
Steve Eder ◽  
Wei Liu ◽  
F. Marion Hulett
Keyword(s):  
Transcriptional Activation ◽  
Mutational Analysis ◽  
Core Promoter ◽  
Regulatory Region ◽  
Core Region ◽  
Phosphate Deficiency ◽  
Pho Regulon ◽  
Promoter Activation ◽  
Binding Region ◽  
The Core

ABSTRACT The PhoP-PhoR two-component regulatory system controls the phosphate deficiency response in B. subtilis. A number of Pho regulon genes which require PhoP∼P for activation or repression have been identified. The studies reported here were initiated to understand the PhoP-DNA interaction necessary for Pho promoter regulation. The regulatory region of phoD was characterized in detail using oligo-directed mutagenesis, DNase I footprinting, and in vivo transcription assays. These data reveal basic principles of PhoP binding relevant to PhoP’s interaction with other Pho regulon promoters. Our results show that: (i) a dimer of PhoP∼P is able to bind two consensus repeats in a stable fashion; (ii) PhoP binding is highly cooperative within the core promoter region, which is located from −66 to −17 on the coding strand and contains four TT(A/T/C)ACA-like repeats; (iii) specific bases comprising the TT(A/T/C)ACA consensus are essential for transcriptional activation, but the specific base pairs of the intervening sequences separating the consensus repeats are not important for either PhoP binding or promoter activation; (iv) the spacing between two consensus repeats within a putative dimer binding site in the core region is important for both PhoP binding and promoter activation; (v) the exact spacing between two dimer binding sites within the core region is important for promoter activation but less so for PhoP binding affinity, as long as the repeats are on the same face of the helix; and (vi) the 5′ secondary binding region is important for coordinated PhoP binding to the core binding region, making it nearly essential for promoter activation.

scholarly journals Analysis of Core Promoter Sequences Located Downstream from the TATA Element in the hsp70 Promoter from Drosophila melanogaster

2001 ◽  
Vol 21 (5) ◽  
pp. 1593-1602 ◽  
Author(s):  
Chwen-Huey Wu ◽  
Lakshmi Madabusi ◽  
Hokuto Nishioka ◽  
Peter Emanuel ◽  
Michael Sypes ◽  
...  
Keyword(s):  
Core Promoter ◽  
P Element ◽  
Multiple Sequence ◽  
Promoter Sequences ◽  
Tata Element ◽  
Sequence Elements ◽  
Cross Links ◽  
Transcription In Vitro

ABSTRACT TFIID recognizes multiple sequence elements in thehsp70 promoter of Drosophila. Here, we investigate the function of sequences downstream from the TATA element. A mutation in the initiator was identified that caused an eightfold reduction in binding of TFIID and a fourfold reduction in transcription in vitro. Another mutation in the +24 to +29 region was somewhat less inhibitory, but a mutation in the +14 to +19 region had essentially no effect. The normal promoter and the mutants in the initiator and the +24 to +29 region were transformed into flies by P element-mediated transformation. The initiator mutation reduced expression an average of twofold in adult flies, whereas the mutation in the +24 to +29 region had essentially no effect. In contrast, a promoter combining the two mutations was expressed an average of sixfold less than the wild type. The results suggest that the initiator and the +24 to +29 region could serve overlapping functions in vivo. Protein-DNA cross-linking was used to identify which subunits of TFIID contact the +24 to +29 region and the initiator. No specific subunits were found to cross-link to the +24 to +29 region. In contrast, the initiator cross-linked exclusively to dTAF230. Remarkably, dTAF230 cross-links approximately 10 times more efficiently to the nontranscribed strand than to the transcribed strand at the initiator.

scholarly journals Intron-driven gene expression in the absence of a core promoter in Arabidopsis thaliana

2016 ◽  
Author(s):  
Jenna E Gallegos ◽  
Alan B Rose
Keyword(s):  
Gene Expression ◽  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
Core Promoter ◽  
Regulatory Elements ◽  
Intron Position ◽  
Mrna Accumulation ◽  
Practical Applications ◽  
Promoter Sequences ◽  
The Core

AbstractIn diverse eukaryotes, certain introns increase mRNA accumulation through the poorly understood mechanism of intron-mediated enhancement (IME). A distinguishing feature of IME is that these introns have no effect from upstream or more than 1 Kb downstream of the transcription start site (TSS). To more precisely define the intron position requirements for IME in Arabidopsis, we tested the effect of the UBQ10 intron on gene expression from 6 different positions surrounding the TSS of a TRP1:GUS fusion. The intron strongly increased expression from all transcribed positions, but had no effect when 204 nt or more upstream of the 5’-most TSS. When the intron was located in the 5’ UTR, the TSS unexpectedly changed, resulting in longer transcripts. Remarkably, deleting 303 nt of the core promoter, including all known TSS’s and all but 18 nt of the 5’ UTR, had virtually no effect on the level of gene expression as long as a stimulating intron was included in the gene. When the core promoter was deleted, transcription initiated in normally untranscribed sequences the same distance upstream of the intron as when the promoter was intact. Together, these results suggest that certain introns play unexpectedly large roles in directing transcription initiation and represent a previously unrecognized type of downstream regulatory elements for genes transcribed by RNA polymerase II. This study also demonstrates considerable flexibility in the sequences surrounding the TSS, indicating that the TSS is not determined by promoter sequences alone. These findings are relevant in practical applications where introns are used to increase gene expression and contribute to our general understanding of gene structure and regulation in eukaryotes.

scholarly journals Investigation of the Wilson gene ATP7B transcriptional start site and the effect of core promoter alterations

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Clemens Höflich ◽  
Angela Brieger ◽  
Stefan Zeuzem ◽  
Guido Plotz
Keyword(s):  
Wilson Disease ◽  
Transcription Initiation ◽  
Transcriptional Activity ◽  
Core Promoter ◽  
Transcriptional Start Site ◽  
Copper Metabolism ◽  
Biologically Relevant ◽  
The Core ◽  
Translational Start

AbstractPathogenic genetic variants in the ATP7B gene cause Wilson disease, a recessive disorder of copper metabolism showing a significant variability in clinical phenotype. Promoter mutations have been rarely reported, and controversial data exist on the site of transcription initiation (the core promoter). We quantitatively investigated transcription initiation and found it to be located in immediate proximity of the translational start. The effects human single-nucleotide alterations of conserved bases in the core promoter on transcriptional activity were moderate, explaining why clearly pathogenic mutations within the core promoter have not been reported. Furthermore, the core promoter contains two frequent polymorphisms (rs148013251 and rs2277448) that could contribute to phenotypical variability in Wilson disease patients with incompletely inactivating mutations. However, neither polymorphism significantly modulated ATP7B expression in vitro, nor were copper household parameters in healthy probands affected. In summary, the investigations allowed to determine the biologically relevant site of ATP7B transcription initiation and demonstrated that genetic variations in this site, although being the focus of transcriptional activity, do not contribute significantly to Wilson disease pathogenesis.

scholarly journals Activation of yeast polymerase II transcription by herpesvirus VP16 and GAL4 derivatives in vitro.

1989 ◽  
Vol 9 (11) ◽  
pp. 4746-4749 ◽  
Author(s):  
D I Chasman ◽  
J Leatherwood ◽  
M Carey ◽  
M Ptashne ◽  
R D Kornberg
Keyword(s):  
Rna Polymerase Ii ◽  
Fusion Proteins ◽  
In Vitro System ◽  
Transcription System ◽  
Natural Mechanism ◽  
Transcription In Vitro ◽  
Rna Polymerase Ii Transcription ◽  
Fold Stimulation

Fusion proteins known to activate transcription in vivo were tested for the ability to stimulate transcription in vitro in a recently developed Saccharomyces cerevisiae RNA polymerase II transcription system. One fusion protein, whose activation domain was derived from the herpesvirus transcriptional activator VP16, gave more than 100-fold stimulation in the in vitro system. The order of effects of the various proteins was the same for transcription in vitro and in vivo, suggesting that the natural mechanism of activation is preserved in vitro.

BAF is required for emerin assembly into the reforming nuclear envelope

2001 ◽  
Vol 114 (24) ◽  
pp. 4575-4585 ◽  
Author(s):  
Tokuko Haraguchi ◽  
Takako Koujin ◽  
Miriam Segura-Totten ◽  
Kenneth K. Lee ◽  
Yosuke Matsuoka ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Hela Cells ◽  
Core Region ◽  
Lamin B ◽  
The Core ◽  
Double Stranded Dna ◽  
Recessive Form ◽  
Nuclear Envelope Assembly

Mutations in emerin cause the X-linked recessive form of Emery-Dreifuss muscular dystrophy (EDMD). Emerin localizes at the inner membrane of the nuclear envelope (NE) during interphase, and diffuses into the ER when the NE disassembles during mitosis. We analyzed the recruitment of wildtype and mutant GFP-tagged emerin proteins during nuclear envelope assembly in living HeLa cells. During telophase, emerin accumulates briefly at the ‘core’ region of telophase chromosomes, and later distributes over the entire nuclear rim. Barrier-to-autointegration factor (BAF), a protein that binds nonspecifically to double-stranded DNA in vitro, co-localized with emerin at the ‘core’ region of chromosomes during telophase. An emerin mutant defective for binding to BAF in vitro failed to localize at the ‘core’ in vivo, and subsequently failed to localize at the reformed NE. In HeLa cells that expressed BAF mutant G25E, which did not show ‘core’ localization, the endogenous emerin proteins failed to localize at the ‘core’ region during telophase, and did not assemble into the NE during the subsequent interphase. BAF mutant G25E also dominantly dislocalized LAP2β and lamin A from the NE, but had no effect on the localization of lamin B. We conclude that BAF is required for the assembly of emerin and A-type lamins at the reforming NE during telophase, and may mediate their stability in the subsequent interphase.

Sign in / Sign up

Export Citation Format

Share Document