scholarly journals Characterization of Binding Sequences for Butyrolactone Autoregulator Receptors in Streptomycetes

1999 ◽  
Vol 181 (16) ◽  
pp. 5075-5080 ◽  
Author(s):  
Hiroshi Kinoshita ◽  
Tomohiro Tsuji ◽  
Hiroomi Ipposhi ◽  
Takuya Nihira ◽  
Yasuhiro Yamada
Keyword(s):  
Transcription Start Site ◽  
Dna Sequences ◽  
Target Genes ◽  
Start Codon ◽  
Receptor Protein ◽  
Upstream Region ◽  
Start Site ◽  
Transcription Start ◽  
S1 Nuclease ◽  
Nuclease Mapping

ABSTRACT BarA of Streptomyces virginiae is a specific receptor protein for a member of butyrolactone autoregulators which binds to an upstream region of target genes to control transcription, leading to the production of the antibiotic virginiamycin M1 and S. BarA-binding DNA sequences (BarA-responsive elements [BAREs]), to which BarA binds for transcriptional control, were restricted to 26 to 29-nucleotide (nt) sequences on barA and barBupstream regions by the surface plasmon resonance technique, gel shift assay, and DNase I footprint analysis. Two BAREs (BARE-1 and BARE-2) on the barB upstream region were located 57 to 29 bp (BARE-1) and 268 to 241 bp (BARE-2) upstream from the barBtranslational start codon. The BARE located on the barAupstream region (BARE-3) was found 101 to 76 bp upstream of thebarA start codon. High-resolution S1 nuclease mapping analysis revealed that BARE-1 covered the barBtranscription start site and BARE-3 covered an autoregulator-dependent transcription start site of the barA gene. Deletion and mutation analysis of BARE-2 demonstrated that at least a 19-nt sequence was required for sufficient BarA binding, and A or T residues at the edge as well as internal conserved nucleotides were indispensable. The identified binding sequences for autoregulator receptor proteins were found to be highly conserved among Streptomyces species.

scholarly journals In Vitro Analysis of the Butyrolactone Autoregulator Receptor Protein (FarA) of Streptomyces lavendulae FRI-5 Reveals that FarA Acts as a DNA-Binding Transcriptional Regulator That Controls Its Own Synthesis

1999 ◽  
Vol 181 (16) ◽  
pp. 5081-5084 ◽  
Author(s):  
Shigeru Kitani ◽  
Hiroshi Kinoshita ◽  
Takuya Nihira ◽  
Yasuhiro Yamada
Keyword(s):  
Transcription Initiation ◽  
Start Codon ◽  
Receptor Protein ◽  
Blue Pigment ◽  
Upstream Region ◽  
Start Site ◽  
Streptomyces Lavendulae ◽  
Nuclease Mapping ◽  
Binding Sequence

ABSTRACT FarA of Streptomyces lavendulae FRI-5 is a specific receptor protein for IM-2, a butyrolactone autoregulator that controls the production of a blue pigment and the nucleoside antibiotics showdomycin and minimycin. Gel shift assays demonstrated that FarA binds to the farA upstream region and that this binding is abolished in the presence of IM-2. The FarA binding sequence was localized by DNase I footprinting to a 28-bp sequence located approximately 70 bp upstream of the farA translational start site. High-resolution S1 nuclease mapping of farAtranscripts revealed a putative transcription start site, located at an A residue positioned 64 bp upstream from the farAtranslation start codon and 4 bp downstream from an Escherichia coli ς70-like −10 recognition region. The FarA-binding sequence overlaps this −10 region and contains thefarA transcription initiation site, suggesting that FarA acts as a repressor that, in the absence of IM-2, represses transcription of farA.

scholarly journals Preferential Reduction of the Thermodynamically Less Favorable Electron Acceptor, Sulfate, by a Nitrate-Reducing Strain of the Sulfate-Reducing Bacterium Desulfovibrio desulfuricans 27774

2008 ◽  
Vol 191 (3) ◽  
pp. 882-889 ◽  
Author(s):  
Angeliki Marietou ◽  
Lesley Griffiths ◽  
Jeff Cole
Keyword(s):  
Transcription Start Site ◽  
Electron Acceptor ◽  
Nitrate Reduction ◽  
Desulfovibrio Desulfuricans ◽  
Start Codon ◽  
Start Site ◽  
Transcription Start ◽  
Sulfate Reducing ◽  
Reverse Transcription Pcr ◽  
Nitrate Induction

ABSTRACT Desulfovibrio desulfuricans strain 27774 is one of a relative small group of sulfate-reducing bacteria that can also grow with nitrate as an alternative electron acceptor, but how nitrate reduction is regulated in any sulfate-reducing bacterium is controversial. Strain 27774 grew more rapidly and to higher yields of biomass with nitrate than with sulfate or nitrite as the only electron acceptor. In the presence of both sulfate and nitrate, sulfate was used preferentially, even when cultures were continuously gassed with nitrogen and carbon dioxide to prevent sulfide inhibition of nitrate reduction. The napC transcription start site was identified 112 bases upstream of the first base of the translation start codon. Transcripts initiated at the napC promoter that were extended across the napM-napA boundary were detected by reverse transcription-PCR, confirming that the six nap genes can be cotranscribed as a single operon. Real-time PCR experiments confirmed that nap operon expression is regulated at the level of mRNA transcription by at least two mechanisms: nitrate induction and sulfate repression. We speculate that three almost perfect inverted-repeat sequences located upstream of the transcription start site might be binding sites for one or more proteins of the CRP/FNR family of transcription factors that mediate nitrate induction and sulfate repression of nitrate reduction by D. desulfuricans.

scholarly journals Genetic Features of MCR-1-Producing Colistin-Resistant Escherichia coli Isolates in South Africa

2016 ◽  
Vol 60 (7) ◽  
pp. 4394-4397 ◽  
Author(s):  
Laurent Poirel ◽  
Nicolas Kieffer ◽  
Adrian Brink ◽  
Jennifer Coetze ◽  
Aurélie Jayol ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
South Africa ◽  
Transcription Start Site ◽  
Dna Sequences ◽  
Start Site ◽  
Transcription Start ◽  
Content Type ◽  
Promoter Sequences ◽  
Genetic Features ◽  
Community Patients

ABSTRACTA series of colistin-resistantEscherichia coliclinical isolates was recovered from hospitalized and community patients in South Africa. Seven clonally unrelated isolates harbored themcr-1gene located on different plasmid backbones. Two distinct plasmids were fully sequenced, and identical 2,600-bp-long DNA sequences defining amcr-1cassette were identified. Promoter sequences responsible for the expression ofmcr-1, deduced from the precise identification of the +1 transcription start site formcr-1, were characterized.

scholarly journals Isolation and characterization of a TATA-less promoter for the human beta 3 integrin gene

Blood ◽  
1994 ◽  
Vol 83 (3) ◽  
pp. 668-676 ◽  
Author(s):  
M Villa-Garcia ◽  
L Li ◽  
G Riely ◽  
PF Bray
Keyword(s):  
Transcription Start Site ◽  
Phorbol Esters ◽  
K562 Cells ◽  
Start Codon ◽  
Dependent Manner ◽  
Start Site ◽  
Transcription Start ◽  
Specific Expression ◽  
Rnase Protection ◽  
Isolation And Characterization

Proper expression of the human platelet fibrinogen receptor is necessary for the maintenance of normal hemostasis. This receptor is formed by the heterodimer alpha IIb beta 3, a prototypic member of the integrin family of adhesive molecules. beta 3 is also expressed in other tissues with alpha v as the vitronectin receptor. It was not possible to study the basis for tissue-specific expression of this gene, because the beta 3 gene promoter had not been isolated previously. We have now isolated a 6.0-kb human genomic DNA fragment containing 2.0 kb of sequence 5′ to the beta 3 ATG start codon. This clone also contains sequence encoding the signal peptide of the immature beta 3 protein and 3.0 kb of 3′ intronic sequence. Primer extension and RNase protection studies of poly A+ RNA from a human erythroleukemia (HEL) cell line indicated a major transcription start site 30 bp upstream of the ATG start codon. In an orientation-dependent manner, a 584-bp fragment 5′ to the start codon promotes expression of the chloramphenicol acetyl transferase (CAT) reporter gene in K562 cells. CAT expression from this beta 3 promoter is fivefold above expression from a “promoter-less” control CAT construct. This beta 3 promoter lacks TATA and CAAT cis-acting elements, but there are two Sp1 sites flanking the transcription start site. Other potential transcription factor binding sites are also identified. Phorbol esters (TPA), which increase beta 3 transcription in K562 cells, stimulated transcription from the 584-bp 5′ beta 3 region. The isolation of this beta 3 promoter region should permit a more detailed analysis of its transcriptional regulation.

scholarly journals Genomic Characterization of Human DSPG3

1999 ◽  
Vol 9 (5) ◽  
pp. 449-456
Author(s):  
Michelle Deere ◽  
Jose L. Dieguez ◽  
Sung-Joo Kim Yoon ◽  
David Hewett-Emmett ◽  
Albert de la Chapelle ◽  
...  
Keyword(s):  
Transcription Start Site ◽  
Sequence Data ◽  
Stop Codon ◽  
Genomic Structure ◽  
Start Codon ◽  
Start Site ◽  
Transcription Start ◽  
Exon 2 ◽  
Ancestral Gene ◽  
Link Type

DSPG3, the human homolog to chick PG-Lb, is a member of the small leucine-rich repeat proteoglycan (SLRP) family, including decorin, biglycan, fibromodulin, and lumican. In contrast to the tissue distribution of the other SLRPs, DSPG3 is predominantly expressed in cartilage. In this study, we have determined that the human DSPG3 gene is composed of seven exons: Exon 2 ofDSPG3 includes the start codon, exons 4–7 code for the leucine-rich repeats, exons 3 and 7 contain the potential glycosaminoglycan attachment sites, and exon 7 contains the potential N-glycosylation sites and the stop codon. We have identified two polymorphic variations, an insertion/deletion composed of 19 nucleotides in intron 1 and a tetranucleotide (TATT)n repeat in intron 5. Analysis of 1.6 kb of upstream promoter sequence ofDSPG3 reveals three TATA boxes, one of which is 20 nucleotides before the transcription start site. The transcription start site precedes the translation start site by 98 nucleotides. There are 14 potential binding sites for SOX9, a transcription factor present in cartilage, in the promoter, and in the first intron of DSPG3. We have examined the evolution of the SLRP gene family and found that gene products clustered together in the evolutionary tree are encoded by genes with similarities in genomic structure. Hence, it appears that the majority of the introns in the SLRP genes were inserted after the differentiation of the SLRP genes from an ancestral gene that was most likely composed of 2–3 exons.[The sequence data described in this paper have been submitted to GenBank under accession nos.AF031658 and U63814.]

scholarly journals Identification of a 119-bp Promoter of the Maize Sulfite Oxidase Gene (ZmSO) That Confers High-Level Gene Expression and ABA or Drought Inducibility in Transgenic Plants

2019 ◽  
Vol 20 (13) ◽  
pp. 3326 ◽  
Author(s):  
Ziwei Xu ◽  
Meiping Wang ◽  
Ziting Guo ◽  
Xianfeng Zhu ◽  
Zongliang Xia
Keyword(s):  
Drought Stress ◽  
Transgenic Plants ◽  
Transcription Start Site ◽  
Sulfite Oxidase ◽  
Upstream Region ◽  
Regulation Mechanism ◽  
Start Site ◽  
Transcription Start ◽  
Mutant Analysis ◽  
High Level

Drought adversely affects crop growth and yields. The cloning and characterization of drought- or abscisic acid (ABA)-inducible promoters is of great significance for their utilization in the genetic improvement of crop resistance. Our previous studies have shown that maize sulfite oxidase (SO) has a sulfite-oxidizing function and is involved in the drought stress response. However, the promoter of the maize SO gene has not yet been characterized. In this study, the promoter (ZmSOPro, 1194 bp upstream region of the translation initiation site) was isolated from the maize genome. The in-silico analysis of the ZmSOPro promoter identified several cis-elements responsive to the phytohormone ABA and drought stress such as ABA-responsive element (ABRE) and MYB binding site (MBS), besides a number of core cis-acting elements, such as TATA-box and CAAT-box. A 5′ RACE (rapid amplification of cDNA ends) assay identified an adenine residue as the transcription start site of the ZmSO. The ZmSOPro activity was detected by β-glucuronidase (GUS) staining at nearly all developmental stages and in most plant organs, except for the roots in transgenic Arabidopsis. Moreover, its activity was significantly induced by ABA and drought stress. The 5′-deletion mutant analysis of the ZmSOPro in tobacco plants revealed that a 119-bp fragment in the ZmSOPro (upstream of the transcription start site) is a minimal region, which is required for its high-level expression. Moreover, the minimal ZmSOPro was significantly activated by ABA or drought stress in transgenic plants. Further mutant analysis indicated that the MBS element in the minimal ZmSOPro region (119 bp upstream of the transcription start site) is responsible for ABA and drought-stress induced expression. These results improve our understanding of the transcriptional regulation mechanism of the ZmSO gene, and the characterized 119-bp promoter fragment could be an ideal candidate for drought-tolerant gene engineering in both monocot and dicot crops.

Role of the Sulfolobus shibatae viral T6 initiator in conferring promoter strength and in influencing transcription start site selection

2006 ◽  
Vol 52 (11) ◽  
pp. 1136-1140 ◽  
Author(s):  
Sohail A Qureshi
Keyword(s):  
Transcription Start Site ◽  
Dna Sequences ◽  
Site Selection ◽  
Core Promoter ◽  
Promoter Strength ◽  
Start Site ◽  
Transcription Start ◽  
Recognition Element ◽  
Sulfolobus Shibatae

Archaeal promoters contain a TATA-box, an adjacent upstream TFB-recognition element (BRE), and a downstream initiator (INR) region from which transcription originates. While the contribution of A-box and BRE to promoter strength is well established, the role of DNA sequences within the INR region and its vicinity on transcription efficiency and start site selection remains unclear. Here, I demonstrate using the strong Sulfolobus shibatae viral T6 promoter that either substitution of its natural sequence from –17 and beyond with plasmid DNA or introduction of point transversion mutations at +3, –2, –4, and –5 positions reduce promoter strength dramatically, whereas +1, –1, and –2 mutations influence the transcription start site. These data therefore reveal that the INR region plays a role as important as the BRE and the A-box in T6 gene transcription. Key words: Archaea, transcription, initiator (INR), Sulfolobus shibatae, core promoter.

The human prointerleukin 1 beta gene requires DNA sequences both proximal and distal to the transcription start site for tissue-specific induction

1993 ◽  
Vol 13 (3) ◽  
pp. 1332-1344 ◽  
Author(s):  
F Shirakawa ◽  
K Saito ◽  
C A Bonagura ◽  
D L Galson ◽  
M J Fenton ◽  
...  
Keyword(s):  
Transcription Start Site ◽  
Phorbol Myristate Acetate ◽  
Dna Sequences ◽  
Hela Cells ◽  
Start Site ◽  
Myristate Acetate ◽  
Binding Studies ◽  
Transcription Start ◽  
Specific Induction ◽  
Beta Gene

In these studies, we have identified DNA sequences and specific protein interactions necessary for transcriptional regulation of the human prointerleukin 1 beta (proIL-1 beta) gene. A cell-type-independent lipopolysaccharide (LPS)-responsive enhancer element located between -3757 and -2729 bp upstream from the transcription start site (cap site) consisted of at least six discrete subregions which were essential to the maximal induction by LPS in transfected monocytes. The enhancer also appeared to mediate phorbol myristate acetate induction in monocytes and IL-1 responsiveness in fibroblasts. Deletion and base substitution mutations along with DNA binding studies demonstrated that the enhancer contained a minimum of three functional protein binding sequences, two of which appeared to be important for gene induction. One of the essential proteins which bound to the enhancer was similar or identical to members of the C/EBP family of transcription factors required for both IL-1- and LPS-specific induction of the IL-6 gene (i.e., the NF-IL6 proteins). When ligated to the proIL-1 beta cap site-proximal region (located between -131 to +12), both the proIL-1 beta and the simian virus 40 enhancer elements functioned more efficiently in monocytes than in HeLa cells, which are not normally competent for IL-1 beta expression. When ligated to the murine c-fos promoter, however, the proIL-1 beta enhancer was inducible in phorbol myristate acetate-stimulated HeLa cells, suggesting the existence of a proIL-1 beta promoter-proximal requirement for tissue specificity.

scholarly journals The human prointerleukin 1 beta gene requires DNA sequences both proximal and distal to the transcription start site for tissue-specific induction.

1993 ◽  
Vol 13 (3) ◽  
pp. 1332-1344 ◽  
Author(s):  
F Shirakawa ◽  
K Saito ◽  
C A Bonagura ◽  
D L Galson ◽  
M J Fenton ◽  
...  
Keyword(s):  
Transcription Start Site ◽  
Phorbol Myristate Acetate ◽  
Dna Sequences ◽  
Hela Cells ◽  
Start Site ◽  
Myristate Acetate ◽  
Binding Studies ◽  
Transcription Start ◽  
Specific Induction ◽  
Beta Gene

In these studies, we have identified DNA sequences and specific protein interactions necessary for transcriptional regulation of the human prointerleukin 1 beta (proIL-1 beta) gene. A cell-type-independent lipopolysaccharide (LPS)-responsive enhancer element located between -3757 and -2729 bp upstream from the transcription start site (cap site) consisted of at least six discrete subregions which were essential to the maximal induction by LPS in transfected monocytes. The enhancer also appeared to mediate phorbol myristate acetate induction in monocytes and IL-1 responsiveness in fibroblasts. Deletion and base substitution mutations along with DNA binding studies demonstrated that the enhancer contained a minimum of three functional protein binding sequences, two of which appeared to be important for gene induction. One of the essential proteins which bound to the enhancer was similar or identical to members of the C/EBP family of transcription factors required for both IL-1- and LPS-specific induction of the IL-6 gene (i.e., the NF-IL6 proteins). When ligated to the proIL-1 beta cap site-proximal region (located between -131 to +12), both the proIL-1 beta and the simian virus 40 enhancer elements functioned more efficiently in monocytes than in HeLa cells, which are not normally competent for IL-1 beta expression. When ligated to the murine c-fos promoter, however, the proIL-1 beta enhancer was inducible in phorbol myristate acetate-stimulated HeLa cells, suggesting the existence of a proIL-1 beta promoter-proximal requirement for tissue specificity.

scholarly journals Promoter Characterization and Constitutive Expression of the Escherichia coli gcvR Gene

1998 ◽  
Vol 180 (7) ◽  
pp. 1803-1807 ◽  
Author(s):  
Angela C. Ghrist ◽  
George V. Stauffer
Keyword(s):  
Escherichia Coli ◽  
Transcription Start Site ◽  
Promoter Region ◽  
Start Codon ◽  
Start Site ◽  
Transcription Start ◽  
Translation Start Codon ◽  
Translation Start ◽  
Glycine Cleavage ◽  
Extension Analysis

ABSTRACT The Escherichia coli glycine cleavage repressor protein (GcvR) negatively regulates expression of the glycine cleavage operon (gcv). In this study, the gcvR translational start site was determined by N-terminal amino acid sequence analysis of a GcvR-LacZ fusion protein. Primer extension analysis of thegcvR promoter region identified a primary transcription start site 27 bp upstream of the UUG translation start site and a minor transcription start site approximately 100 bp upstream of the translation start codon. The -10 and -35 promoter regions upstream of the primary transcription start site were defined by mutational analysis. Expression of a gcvR-lacZ fusion was unaltered in the presence of glycine or inosine, molecules known to induce or repress expression of gcv, respectively. In addition, it was shown that gcvR-lacZ expression is neither regulated by the glycine cleavage activator protein (GcvA) nor autogenously regulated by GcvR. From DNA sequence analysis, it was predicted that the translation start codon of the downstream bcp gene overlaps the gcvR stop codon, suggesting that these genes may form an operon. However, a down mutation in the -10 promoter region of gcvR had no effect on the expression of a downstreambcp-lacZ fusion, and primer extension analysis of thebcp promoter region demonstrated that bcp has its own promoter within the gcvR coding sequence. These results show that gcvR and bcp do not form an operon. Furthermore, the deletion of bcp from the chromosome had no effect on gcv-lacZ expression.

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