scholarly journals The Divergently Transcribed Streptococcus parasanguis Virulence-Associated fimA Operon Encoding an Mn2+-Responsive Metal Transporter and pepO Encoding a Zinc Metallopeptidase Are Not Coordinately Regulated

2002 ◽  
Vol 70 (10) ◽  
pp. 5706-5714 ◽  
Author(s):  
Joyce Oetjen ◽  
Paula Fives-Taylor ◽  
Eunice H. Froeliger
Keyword(s):  
Transcriptional Start Site ◽  
Regulatory Region ◽  
Regulatory Elements ◽  
Divalent Metal Ions ◽  
Metal Transporters ◽  
Metal Transporter ◽  
Gram Positive Bacteria ◽  
Growth Deficiency ◽  
Domain Mapping ◽  
Transcriptional Start Sites

ABSTRACT The study of how bacteria respond to and obtain divalent metal ions provides insight into the regulation of virulence factors in the host environment. Regulation of metal permease operons in gram-positive bacteria may involve the binding of metal-responsive repressors to palindromic domains in their control regions. The Streptococcus parasanguis fimA operon, which encodes an ATP-binding cassette (ABC) transporter system with sequence homology to the LraI family of metal transporters, possesses a palindromic regulatory region with high homology to that of the Streptococcus gordonii ScaR binding domain. Mapping of the promoter and regulatory regions of fimA and the divergently transcribed pepO gene, which encodes a zinc metalloendopeptidase, indicated that their promoter and regulatory elements overlap. fimA had one transcriptional start site, whereas pepO had three. Analysis of truncated versions of the pepO promoter suggested that all three transcriptional start sites are functional. Analysis of promoter activity under various environmental conditions indicated that the fimA operon promoter and the pepO promoter are not coordinately regulated. The fimA operon is responsive to changes in Mn2+ concentration, but the pepO promoter is not. A S. parasanguis fimA mutant showed a growth deficiency under conditions of limiting Mn2+. This deficiency was not alleviated by compensation with either Mg2+ or Fe3+. Wild-type S. parasanguis could take up Mn2+ and Fe3+, while the fimA mutant showed a marked reduction in this ability. These data suggested that FimA is a component of a metal transporter system capable of transporting both Mn2+ and Fe3+. FimA expression itself was shown to be responsive to Mn2+ concentration, but not to availability of Fe3+ or Mg2+.

scholarly journals Expression System for High Levels of GAG Lyase Gene Expression and Study of the hepA Upstream Region in Flavobacterium heparinum

2002 ◽  
Vol 184 (12) ◽  
pp. 3242-3252 ◽  
Author(s):  
Françoise Blain ◽  
A. Lydia Tkalec ◽  
Zhongqi Shao ◽  
Catherine Poulin ◽  
Marc Pedneault ◽  
...  
Keyword(s):  
Transcriptional Start Site ◽  
Expression System ◽  
Regulatory Region ◽  
Start Codon ◽  
Upstream Region ◽  
Wild Type ◽  
Heparinase I ◽  
High Level ◽  
Flavobacterium Heparinum ◽  
Transcriptional Start Sites

ABSTRACT A system for high-level expression of heparinase I, heparinase II, heparinase III, chondroitinase AC, and chondroitinase B in Flavobacterium heparinum is described. hepA, along with its regulatory region, as well as hepB, hepC, cslA, and cslB, cloned downstream of the hepA regulatory region, was integrated in the chromosome to yield stable transconjugant strains. The level of heparinase I and II expression from the transconjugant strains was approximately fivefold higher, while heparinase III expression was 10-fold higher than in wild-type F. heparinum grown in heparin-only medium. The chondroitinase AC and B transconjugant strains, grown in heparin-only medium, yielded 20- and 13-fold increases, respectively, in chondroitinase AC and B expression, compared to wild-type F. heparinum grown in chondroitin sulfate A-only medium. The hepA upstream region was also studied using cslA as a reporter gene, and the transcriptional start site was determined to be 26 bp upstream of the start codon in the chondroitinase AC transconjugant strain. The transcriptional start sites were determined for hepA in both the wild-type F. heparinum and heparinase I transconjugant strains and were shown to be the same as in the chondroitinase AC transconjugant strain. The five GAG lyases were purified from these transconjugant strains and shown to be identical to their wild-type counterparts.

scholarly journals Repression and activation of the Drosophila dopa decarboxylase gene in glia.

1992 ◽  
Vol 12 (12) ◽  
pp. 5659-5666 ◽  
Author(s):  
G S Mastick ◽  
S B Scholnick
Keyword(s):  
Germ Line ◽  
Transcriptional Start Site ◽  
Regulatory Region ◽  
Regulatory Elements ◽  
Dopa Decarboxylase ◽  
In Vitro Mutagenesis ◽  
Wild Type ◽  
Cell Activator ◽  
Germ Line Transformation

Glial expression of the Drosophila dopa decarboxylase gene (Ddc) is repressed by a regulatory region located approximately 1 kb upstream of the transcriptional start site. We have used in vitro mutagenesis and germ line transformation to determine which elements within the Ddc promoter mediate repression. Our evidence suggests that the hypodermal cell activator elements IIA and IIB play a major role in the transcriptional regulation of Ddc in glial cells. A variety of mutations demonstrate that element IIA is a strong glial activator element and that element IIB is necessary for glial repression. Although these two regulatory elements are nearly identical in sequence, our data suggest that they are not redundant. Altering the wild-type number and spacing of elements IIA and IIB indicates that the wild-type arrangement of this repeat is critical for repression. We conclude that these key elements of the Ddc promoter regulate both activation and repression in glia.

scholarly journals Cloning and Functional characterization of seed-specific LEC1A promoter from peanut (Arachis hypogaea L.)

2020 ◽  
Author(s):  
Guiying Tang ◽  
Pingli Xu ◽  
Pengxiang Li ◽  
Jieqiong Zhu ◽  
Guangxia Chen ◽  
...  
Keyword(s):  
Transcriptional Start Site ◽  
Functional Characterization ◽  
Regulatory Region ◽  
Regulatory Elements ◽  
Histochemical Staining ◽  
Expression Vectors ◽  
Upstream Sequence ◽  
Functional Region ◽  
Chromosomal Walking ◽  
Storage Reserve

AbstractLEAFY COTYLEDON1 (LEC1) is a HAP3 subunit of CCAAT-binding transcription factor, which controls several aspects of embryo and postembryo development, including embryo morphogenesis, storage reserve accumulation and skotomorphogenesis. Herein, using the method of chromosomal walking, a 2707bp upstream sequence from the ATG initiation codon site of AhLEC1A which is a homolog of Arabidopsis LEC1 was isolated in peanut. Its transcriptional start site confirmed by 5’ RACE was located at 82 nt from 5’ upstream of ATG. The bioinformatics analysis revealed that there existed many tissue-specific elements and light responsive motifs in its promoter. To identify the functional region of the AhLEC1A promoter, seven plant expression vectors expressing the GUS (β-glucuronidase) gene, driven by 5’ terminal series deleted fragments of AhLEC1A promoter, were constructed and transformed into Arabidopsis. Results of GUS histochemical staining showed that the regulatory region containing 82bp of 5’ UTR and 2228bp promoter could facilitate GUS to express preferentially in the embryos at different development periods of Arabidopsis. Taken together, it was inferred that the expression of AhLEC1A during seed development of peanut might be controlled positively by several seed-specific regulatory elements, as well as negatively by some other regulatory elements inhibiting its expression in other organs. Moreover, the GUS expression pattern of transgenic seedlings in darkness and in light was relevant to the light-responsive elements scattered in AhLEC1A promoter segment, implying that these light-responsive elements harbored in the AhLEC1A promoter regulate skotomorphogenesis of peanut seeds, and AhLEC1A expression was inhibited after the germinated seedlings were transferred from darkness to light.

Repression and activation of the Drosophila dopa decarboxylase gene in glia

1992 ◽  
Vol 12 (12) ◽  
pp. 5659-5666
Author(s):  
G S Mastick ◽  
S B Scholnick
Keyword(s):  
Germ Line ◽  
Transcriptional Start Site ◽  
Regulatory Region ◽  
Regulatory Elements ◽  
Dopa Decarboxylase ◽  
In Vitro Mutagenesis ◽  
Wild Type ◽  
Cell Activator ◽  
Germ Line Transformation

Glial expression of the Drosophila dopa decarboxylase gene (Ddc) is repressed by a regulatory region located approximately 1 kb upstream of the transcriptional start site. We have used in vitro mutagenesis and germ line transformation to determine which elements within the Ddc promoter mediate repression. Our evidence suggests that the hypodermal cell activator elements IIA and IIB play a major role in the transcriptional regulation of Ddc in glial cells. A variety of mutations demonstrate that element IIA is a strong glial activator element and that element IIB is necessary for glial repression. Although these two regulatory elements are nearly identical in sequence, our data suggest that they are not redundant. Altering the wild-type number and spacing of elements IIA and IIB indicates that the wild-type arrangement of this repeat is critical for repression. We conclude that these key elements of the Ddc promoter regulate both activation and repression in glia.

scholarly journals Cloning and functional characterization of seed-specific LEC1A promoter from peanut (Arachis hypogaea L.)

PLoS ONE ◽  
2021 ◽  
Vol 16 (3) ◽  
pp. e0242949
Author(s):  
Guiying Tang ◽  
Pingli Xu ◽  
Pengxiang Li ◽  
Jieqiong Zhu ◽  
Guangxia Chen ◽  
...  
Keyword(s):  
Transcriptional Start Site ◽  
Functional Characterization ◽  
Regulatory Region ◽  
Regulatory Elements ◽  
Histochemical Staining ◽  
Expression Vectors ◽  
Upstream Sequence ◽  
Functional Region ◽  
Chromosomal Walking ◽  
Storage Reserve

LEAFY COTYLEDON1 (LEC1) is a HAP3 subunit of CCAAT-binding transcription factor, which controls several aspects of embryo and postembryo development, including embryo morphogenesis, storage reserve accumulation and skotomorphogenesis. Herein, using the method of chromosomal walking, a 2707bp upstream sequence from the ATG initiation codon site of AhLEC1A which is a homolog of Arabidopsis LEC1 was isolated in peanut. Its transcriptional start site confirmed by 5’ RACE was located at 82 nt from 5’ upstream of ATG. The bioinformatics analysis revealed that there existed many tissue-specific elements and light responsive motifs in its promoter. To identify the functional region of the AhLEC1A promoter, seven plant expression vectors expressing the GUS (β-glucuronidase) gene, driven by 5’ terminal series deleted fragments of AhLEC1A promoter, were constructed and transformed into Arabidopsis. Results of GUS histochemical staining showed that the regulatory region containing 82bp of 5’ UTR and 2228bp promoter could facilitate GUS to express preferentially in the embryos at different development periods of Arabidopsis. Taken together, it was inferred that the expression of AhLEC1A during seed development of peanut might be controlled positively by several seed-specific regulatory elements, as well as negatively by some other regulatory elements inhibiting its expression in other organs. Moreover, the GUS expression pattern of transgenic seedlings in darkness and in light was relevant to the light-responsive elements scattered in AhLEC1A promoter segment, implying that these light-responsive elements harbored in the AhLEC1A promoter regulate skotomorphogenesis of peanut seeds, and AhLEC1A expression was inhibited after the germinated seedlings were transferred from darkness to light.

scholarly journals Identification of the Ω4499 Regulatory Region Controlling Developmental Expression of a Myxococcus xanthus Cytochrome P-450 System

1999 ◽  
Vol 181 (17) ◽  
pp. 5467-5475 ◽  
Author(s):  
Makda Fisseha ◽  
Dvora Biran ◽  
Lee Kroos
Keyword(s):  
Transcriptional Start Site ◽  
Consensus Sequence ◽  
Myxococcus Xanthus ◽  
Regulatory Region ◽  
Regulatory Elements ◽  
Open Reading Frame ◽  
Developmental Expression ◽  
Reading Frame ◽  
Dna Elements ◽  
Cytochrome P 450

ABSTRACT Ω4499 is the site of a Tn5 lac insertion in theMyxococcus xanthus chromosome that fuses lacZexpression to a developmentally regulated promoter. Cell-cell interactions that occur during development, including C signaling, are required for normal expression of Tn5 lac Ω4499. The DNA upstream of the Ω4499 insertion has been cloned, and the promoter has been localized. Analysis of the DNA sequence downstream of the promoter revealed one complete open reading frame and a second partial open reading frame that is interrupted by Tn5 lac Ω4499. The predicted products of these open reading frames are highly similar to reductase and oxidase components of bacterial cytochrome P-450 systems, which allow catabolism or anabolism of unusual compounds. However, the function of the gene products of the Ω4499 locus remains unclear because M. xanthus containing Tn5 lac Ω4499 exhibits no apparent defect in growth, developmental aggregation, fruiting body formation, or sporulation. Deletion analysis of the Ω4499 regulatory region showed that multiple DNA elements spanning more than 500 bp upstream of the transcriptional start site contribute to developmental promoter activity. At least two DNA elements, one downstream of −49 bp and one between −49 and −218 bp, boosted activity of the promoter in response to intercellular C signaling. Three sequences in the Ω4499 promoter region, centered at −55, −33, and −1 bp, nearly match a 7-bp sequence found in other C signal-dependent promoters. We propose that these sequences, matching the consensus sequence 5′-CAYYCCY-3′, be called C box sequences, and we speculate that these sequences arecis-acting regulatory elements important for the expression of M. xanthus genes that depend upon intercellular C signaling during development.

scholarly journals Deletion of a non-canonical regulatory sequence causes loss of Scn1a expression and epileptic phenotypes in mice

2021 ◽  
Vol 13 (1) ◽  
Author(s):  
Jessica L. Haigh ◽  
Anna Adhikari ◽  
Nycole A. Copping ◽  
Tyler Stradleigh ◽  
A. Ayanna Wade ◽  
...  
Keyword(s):  
Regulatory Element ◽  
Transcriptional Start Site ◽  
Regulatory Region ◽  
Regulatory Elements ◽  
Dravet Syndrome ◽  
Regulatory Sequence ◽  
Functional Requirements ◽  
Transgenic Mouse Line ◽  
Gene And Protein Expression ◽  
Promoter Mutations

Abstract Background Genes with multiple co-active promoters appear common in brain, yet little is known about functional requirements for these potentially redundant genomic regulatory elements. SCN1A, which encodes the NaV1.1 sodium channel alpha subunit, is one such gene with two co-active promoters. Mutations in SCN1A are associated with epilepsy, including Dravet syndrome (DS). The majority of DS patients harbor coding mutations causing SCN1A haploinsufficiency; however, putative causal non-coding promoter mutations have been identified. Methods To determine the functional role of one of these potentially redundant Scn1a promoters, we focused on the non-coding Scn1a 1b regulatory region, previously described as a non-canonical alternative transcriptional start site. We generated a transgenic mouse line with deletion of the extended evolutionarily conserved 1b non-coding interval and characterized changes in gene and protein expression, and assessed seizure activity and alterations in behavior. Results Mice harboring a deletion of the 1b non-coding interval exhibited surprisingly severe reductions of Scn1a and NaV1.1 expression throughout the brain. This was accompanied by electroencephalographic and thermal-evoked seizures, and behavioral deficits. Conclusions This work contributes to functional dissection of the regulatory wiring of a major epilepsy risk gene, SCN1A. We identified the 1b region as a critical disease-relevant regulatory element and provide evidence that non-canonical and seemingly redundant promoters can have essential function.

scholarly journals Identification of Trophoblast-Specific Regulatory Elements in the Mouse Placental Lactogen II Gene

1998 ◽  
Vol 12 (3) ◽  
pp. 418-427 ◽  
Author(s):  
Jiandie Lin ◽  
Daniel I. H. Linzer
Keyword(s):  
Giant Cell ◽  
Regulatory Element ◽  
Transcriptional Start Site ◽  
Regulatory Region ◽  
Giant Cells ◽  
Regulatory Elements ◽  
Positive Element ◽  
Placental Lactogen ◽  
Specific Gene ◽  
Trophoblast Giant Cells

Abstract Placental lactogen II, the major ligand for the PRL receptor during the second half of gestation in rodents, is synthesized specifically by placental trophoblast giant cells. A transient transgenic analysis has been used to localize the giant cell-specific regulatory region within the mouse placental lactogen II gene to sequences between −1340 and −2019 upstream of the transcriptional start site. More precise mapping of the regulatory elements has been accomplished by transfection of promoter constructs into Rcho-1 trophoblast cells, resulting in the characterization of two positive regulatory elements in the −1471 to −1340 region; two other regulatory elements have been implicated but not further characterized, a negative regulatory element between −2019 and −1778 and another positive element within the region from −1340 to −569. Both of the characterized positive regulatory elements are recognized by factors that are enriched in differentiated giant cells compared with proliferative trophoblasts, and these factors are either absent or at low levels in fibroblasts. The complexes that form on the two elements are distinct and neither element competes with the other for factor binding, thus implicating at least two different regulatory elements in late-gestational trophoblast giant cell-specific gene expression.

scholarly journals Deletion of a non-canonical promoter regulatory element causes loss ofScn1aexpression and epileptic phenotypes in mice

2019 ◽  
Author(s):  
Jessica L. Haigh ◽  
Anna Adhikari ◽  
Nycole A. Copping ◽  
Tyler Stradleigh ◽  
A. Ayanna Wade ◽  
...  
Keyword(s):  
Regulatory Element ◽  
Transcriptional Start Site ◽  
Regulatory Region ◽  
Regulatory Elements ◽  
Dravet Syndrome ◽  
Functional Requirements ◽  
Behavioral Deficits ◽  
Evolutionarily Conserved ◽  
Severe Reduction ◽  
Promoter Mutations

AbstractGenes with multiple co-active promoters appear common in brain, yet little is known about functional requirements for these potentially redundant genomic regulatory elements.SCN1A,which encodes the NaV1.1 sodium channel alpha subunit, is one such gene with two co-active promoters. Mutations inSCN1Aare associated with epilepsy, including Dravet Syndrome (DS). The majority of DS patients harbor coding mutations causingSCN1Ahaploinsufficiency, however putative causal non-coding promoter mutations have been identified. To model the functional role of potentially redundantScn1apromoters, we focused on the non-codingScn1a1b regulatory region, previously described as a non-canonical alternative transcriptional start site. Mice harboring deletion of the extended evolutionarily-conserved 1b non-coding interval exhibited surprisingly severe reduction ofScn1aand NaV1.1 expression in brain with accompanying seizures and behavioral deficits. This identified the 1b region as a critical disease-relevant regulatory element and provides evidence that non-canonical and apparently redundant promoters can have essential function.

scholarly journals Identification of the Ω4400 Regulatory Region, a Developmental Promoter of Myxococcus xanthus

1998 ◽  
Vol 180 (8) ◽  
pp. 1995-2004 ◽  
Author(s):  
Janine P. Brandner ◽  
Lee Kroos
Keyword(s):  
Rna Polymerase ◽  
Transcriptional Start Site ◽  
Myxococcus Xanthus ◽  
Regulatory Region ◽  
Cell Interaction ◽  
Regulatory Elements ◽  
Open Reading Frame ◽  
Start Site ◽  
Reading Frame ◽  
Cell Cell

ABSTRACT Ω4400 is the site of a Tn5 lac insertion in theMyxococcus xanthus genome that fuses lacZexpression to a developmentally regulated promoter. Cell-cell interactions that occur during development, including C signaling, are required for normal expression of Tn5 lac Ω4400. The DNA upstream of the Ω4400 insertion has been cloned, the promoter has been localized, and a partial open reading frame has been identified. From the deduced amino acid sequence of the partial open reading frame, the gene disrupted by Tn5 lac Ω4400 may encode a protein with an ATP- or GTP-binding site. Expression of the gene begins 6 to 12 h after starvation initiates development, as measured by β-galactosidase production in cells containing Tn5 lacΩ4400. The putative transcriptional start site was mapped, and deletion analysis has shown that DNA downstream of −101 bp is sufficient for C-signal-dependent, developmental activation of this promoter. A deletion to −76 bp eliminated promoter activity, suggesting the involvement of an upstream activator protein. The promoter may be transcribed by RNA polymerase containing a novel sigma factor, since a mutation in the M. xanthus sigB orsigC gene did not affect Tn5 lac Ω4400 expression and the DNA sequence upstream of the transcriptional start site did not match the sequence of any M. xanthus promoter transcribed by a known form of RNA polymerase. However, the Ω4400 promoter does contain the sequence 5′-CATCCCT-3′ centered at −49 and the C-signal-dependent Ω4403 promoter also contains this sequence at the same position. Moreover, the two promoters match at five of six positions in the −10 regions, suggesting that these promoters may share one or more transcription factors. These results begin to define the cis-acting regulatory elements important for cell-cell interaction-dependent gene expression during the development of a multicellular prokaryote.

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