scholarly journals Characterization of cis-acting elements regulating transcription from the promoter of a constitutively active rice actin gene.

1992 ◽  
Vol 12 (8) ◽  
pp. 3399-3406 ◽  
Author(s):  
Y Wang ◽  
W Zhang ◽  
J Cao ◽  
D McElroy ◽  
R Wu
Keyword(s):  
Transient Expression ◽  
Plant Cell ◽  
Transcription Initiation ◽  
Regulatory Elements ◽  
Initiation Site ◽  
Gus Expression ◽  
Negative Regulator ◽  
Cis Acting ◽  
Highly Active ◽  
Constitutively Active

The promoter of the constitutively expressed rice (Oryza sativa) actin 1 gene (Act1) is highly active in transformed rice plants (W. Zhang, D. McElroy, and R. Wu, Plant Cell 3:1150-1160, 1991). A region 834 bp upstream of the Act1 transcription initiation site contains all the regulatory elements necessary for maximal gene expression in transformed rice protoplasts (D. McElroy, W. Zhang, J. Cao, and R. Wu, Plant Cell 2:163-171, 1990). We have constructed a series of Act1 promoter deletions fused to a bacterial beta-glucuronidase reporter sequence (Gus). Transient expression assays in transformed rice protoplasts, as well as transformed maize cells and tissues, identified two distinct cis-acting regulatory elements in the Act1 promoter. A 38-bp poly(dA-dT) region was found to be a positive regulator of Act1 promoter activity. Deletion of the poly(dA-dT) element lowered Gus expression by at least threefold compared with expression produced by the full-length Act1 promoter. By gel retardation and footprinting, we identified a ubiquitous rice protein which specifically recognizes this poly(dA-dT) element in the constitutively active Act1 promoter. A CCCAA pentamer repeat-containing region was found to be a negative regulator of the Act1 promoter in transformed rice protoplasts. Transient expression assays in different maize cells and tissues with use of the Act1 deletion constructs suggested that the CCCAA pentamer repeat region functions in a complex tissue-specific manner. A CCCAA-binding protein was detected only in root extracts.

Characterization of cis-acting elements regulating transcription from the promoter of a constitutively active rice actin gene

1992 ◽  
Vol 12 (8) ◽  
pp. 3399-3406
Author(s):  
Y Wang ◽  
W Zhang ◽  
J Cao ◽  
D McElroy ◽  
R Wu
Keyword(s):  
Transient Expression ◽  
Plant Cell ◽  
Transcription Initiation ◽  
Regulatory Elements ◽  
Initiation Site ◽  
Gus Expression ◽  
Negative Regulator ◽  
Cis Acting ◽  
Highly Active ◽  
Constitutively Active

The promoter of the constitutively expressed rice (Oryza sativa) actin 1 gene (Act1) is highly active in transformed rice plants (W. Zhang, D. McElroy, and R. Wu, Plant Cell 3:1150-1160, 1991). A region 834 bp upstream of the Act1 transcription initiation site contains all the regulatory elements necessary for maximal gene expression in transformed rice protoplasts (D. McElroy, W. Zhang, J. Cao, and R. Wu, Plant Cell 2:163-171, 1990). We have constructed a series of Act1 promoter deletions fused to a bacterial beta-glucuronidase reporter sequence (Gus). Transient expression assays in transformed rice protoplasts, as well as transformed maize cells and tissues, identified two distinct cis-acting regulatory elements in the Act1 promoter. A 38-bp poly(dA-dT) region was found to be a positive regulator of Act1 promoter activity. Deletion of the poly(dA-dT) element lowered Gus expression by at least threefold compared with expression produced by the full-length Act1 promoter. By gel retardation and footprinting, we identified a ubiquitous rice protein which specifically recognizes this poly(dA-dT) element in the constitutively active Act1 promoter. A CCCAA pentamer repeat-containing region was found to be a negative regulator of the Act1 promoter in transformed rice protoplasts. Transient expression assays in different maize cells and tissues with use of the Act1 deletion constructs suggested that the CCCAA pentamer repeat region functions in a complex tissue-specific manner. A CCCAA-binding protein was detected only in root extracts.

Identification of upstream and intragenic regulatory elements that confer cell-type-restricted and differentiation-specific expression on the muscle creatine kinase gene

1988 ◽  
Vol 8 (7) ◽  
pp. 2896-2909 ◽  
Author(s):  
E A Sternberg ◽  
G Spizz ◽  
W M Perry ◽  
D Vizard ◽  
T Weil ◽  
...  
Keyword(s):  
Transcription Initiation ◽  
Regulatory Element ◽  
Simian Virus 40 ◽  
Muscle Cells ◽  
Regulatory Elements ◽  
Initiation Site ◽  
Simian Virus ◽  
Cell Type ◽  
Specific Expression ◽  
Developing Muscle

Terminal differentiation of skeletal myoblasts is accompanied by induction of a series of tissue-specific gene products, which includes the muscle isoenzyme of creatine kinase (MCK). To begin to define the sequences and signals involved in MCK regulation in developing muscle cells, the mouse MCK gene has been isolated. Sequence analysis of 4,147 bases of DNA surrounding the transcription initiation site revealed several interesting structural features, some of which are common to other muscle-specific genes and to cellular and viral enhancers. To test for sequences required for regulated expression, a region upstream of the MCK gene from -4800 to +1 base pairs, relative to the transcription initiation site, was linked to the coding sequences of the bacterial chloramphenicol acetyltransferase (CAT) gene. Introduction of this MCK-CAT fusion gene into C2 muscle cells resulted in high-level expression of CAT activity in differentiated myotubes and no detectable expression in proliferating undifferentiated myoblasts or in nonmyogenic cell lines. Deletion mutagenesis of sequences between -4800 and the transcription start site showed that the region between -1351 and -1050 was sufficient to confer cell type-specific and developmentally regulated expression on the MCK promoter. This upstream regulatory element functioned independently of position, orientation, or distance from the promoter and therefore exhibited the properties of a classical enhancer. This upstream enhancer also was able to confer muscle-specific regulation on the simian virus 40 promoter, although it exhibited a 3- to 5-fold preference for its own promoter. In contrast to the cell type- and differentiation-specific expression of the upstream enhancer, the MCK promoter was able to function in myoblasts and myotubes and in nonmyogenic cell lines when combined with the simian virus 40 enhancer. An additional positive regulatory element was identified within the first intron of the MCK gene. Like the upstream enhancer, this intragenic element functioned independently of position, orientation, and distance with respect to the MCK promoter and was active in differentiated myotubes but not in myoblasts. These results demonstrate that expression of the MCK gene in developing muscle cells is controlled by complex interactions among multiple upstream and intragenic regulatory elements that are functional only in the appropriate cellular context.

scholarly journals Transcriptional enhancers in the HLA-DQ subregion.

1987 ◽  
Vol 7 (9) ◽  
pp. 3315-3319 ◽  
Author(s):  
K E Sullivan ◽  
B M Peterlin
Keyword(s):  
Major Histocompatibility Complex ◽  
Transient Expression ◽  
Regulatory Elements ◽  
Human Major Histocompatibility Complex ◽  
Cis Acting ◽  
Transcriptional Enhancers ◽  
Histocompatibility Complex ◽  
Hla Dq ◽  
Alpha Gene ◽  
Beta Gene

Using transient expression assays, the HLA-DQ alpha and HLA-DQ beta genes of the human major histocompatibility complex were screened for cis-acting regulatory elements. Two regions in the HLA-DQ alpha gene and one in the HLA-DQ beta gene were identified which fulfilled the criteria for transcriptional enhancers.

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 Genomic organization and transcriptional analysis of the human l-glutaminase gene

2003 ◽  
Vol 370 (3) ◽  
pp. 771-784 ◽  
Author(s):  
Cristina PÉREZ-GÓMEZ ◽  
José M. MATÉS ◽  
Pedro M. GÓMEZ-FABRE ◽  
Antonio del CASTILLO-OLIVARES ◽  
Francisco J. ALONSO ◽  
...  
Keyword(s):  
Transcription Initiation ◽  
Genomic Organization ◽  
Core Promoter ◽  
Regulatory Elements ◽  
Initiation Site ◽  
Genome Project ◽  
Transcriptional Analysis ◽  
Cdna Clones ◽  
Mobility Shift ◽  
Specific Expression

In mammals, glutaminase (GA) is expressed in most tissues, but the regulation of organ-specific expression is largely unknown. Therefore, as an essential step towards studying the regulation of GA expression, the human liver-type GA (hLGA) gene has been characterized. LGA genomic sequences were isolated using the genome walking technique. Analysis and comparison of these sequences with two LGA cDNA clones and the Human Genome Project database, allowed the determination of the genomic organization of the LGA gene. The gene has 18 exons and is approx. 18kb long. All exon/intron junction sequences conform to the GT/AG rule. Progressive deletion analysis of LGA promoter—luciferase constructs indicated that the core promoter is located between nt −141 and +410, with several potential regulatory elements: CAAT, GC, TATA-like, Ras-responsive element binding protein and specificity protein 1 (Sp1) sites. The minimal promoter was mapped within +107 and +410, where only an Sp1 binding site is present. Mutation experiments suggested that two CAAT recognition elements near the transcription-initiation site (-138 and −87), play a crucial role for optimal promoter activity. Electrophoretic mobility-shift assays confirmed the importance of CAAT- and TATA-like boxes to enhance basal transcription, and demonstrated that HNF-1 motif is a significant distal element for transcriptional regulation of the hLGA gene.

scholarly journals Sp1-Mediated Transcription of the Werner Helicase Gene Is Modulated by Rb and p53

1998 ◽  
Vol 18 (11) ◽  
pp. 6191-6200 ◽  
Author(s):  
Yukako Yamabe ◽  
Akira Shimamoto ◽  
Makoto Goto ◽  
Jun Yokota ◽  
Minoru Sugawara ◽  
...  
Keyword(s):  
Transcription Initiation ◽  
Promoter Activity ◽  
Transcriptional Control ◽  
Housekeeping Genes ◽  
Regulatory Elements ◽  
Initiation Site ◽  
Luciferase Reporter ◽  
Upstream Region ◽  
Control Element ◽  
Mrna Levels

ABSTRACT The regulation of Werner’s syndrome gene (WRN) expression was studied by characterizing the cis-regulatory elements in the promoter region and the trans-activating factors that bind to them. First, we defined the transcription initiation sites and the sequence of the 5′ upstream region (2.8 kb) ofWRN that contains a number of cis-regulatory elements, including 7 Sp1, 9 retinoblastoma control element (RCE), and 14 AP2 motifs. A region consisting of nucleotides −67 to +160 was identified as the principal promoter of WRN by reporter gene assays in HeLa cells, using a series of WRNpromoter-luciferase reporter (WRN-Luc) plasmids that contained the 5′-truncated or mutated WRN upstream regions. In particular, two Sp1 elements proximal to the transcription initiation site are indispensable for WRN promoter activity and bind specifically to Sp1 proteins. The RCE enhances WRN promoter activity. Coexpression of the WRN-Luc plasmids with various dosages of plasmids expressing Rb or p53 in Saos2 cells lacking active Rb and p53 proteins showed that the introduced Rb upregulates WRN promoter activity a maximum of 2.5-fold, while p53 downregulates it a maximum of 7-fold, both dose dependently. Consistently, the overexpressed Rb and p53 proteins also affected the endogenous WRN mRNA levels in Saos2 cells, resulting in an increase with Rb and a decrease with p53. These findings suggest that WRN expression, like that of other housekeeping genes, is directed mainly by the Sp1 transcriptional control system but is also further modulated by transcription factors, including Rb and p53, that are implicated in the cell cycle, cell senescence, and genomic instability.

scholarly journals Isolation, Cloning and Characterization of a Constitutive Plant from Potato Aquaporin Gene

2020 ◽  
Vol 63 (3) ◽  
pp. 169-178
Author(s):  
Hira Mubeen ◽  
Rubab Zahra Naqvi ◽  
Ammara Masood ◽  
Mushtaq A. Saleem ◽  
Aftab Bashir ◽  
...  
Keyword(s):  
Genomic Sequence ◽  
Crop Improvement ◽  
Regulatory Elements ◽  
Gus Expression ◽  
Cis Acting ◽  
Transcription Start Sites ◽  
Expression Studies ◽  
Aquaporin Gene ◽  
Highly Expressed Genes

Plasma membrane intrinsic proteins (PIP1) are the most common integral membrane proteins belong to a larger family of intrinsic aquaporin proteins. They are member of aquaporin gene family and have gained importance as highly expressed genes in plants. In this study, the promoter of aquaporin PIP1 gene was identified, analyzed and retrieved from high throughput genomic sequence (HTGS) database. The cis-acting regulatory elements, transcription start sites and transcription factor binding sites of selected promoter were identified through different bio-informatics tools. Many light responsive, phytohormone, stress and defense related cis-regulatory elements were detected in PIP1 promoter region indicating its role as a constitutive promoter. The PIP1 promoter was isolated from Solanum tuberosum. It was initially cloned in TA vector (pTZ57R/T) and later transferred to plant expression binary vectors, pGR1 and pGA482 for transient and stable expression studies in tobacco. The GUS expression results of PIP1 promoter in different tobacco tissues showed its functional importance in regulating gene expression in a constitutive manner. Further, it was concluded that the PIP1 aquaporin promoter is constitutively expressed with a strength equivalent to CaMV 2x35S promoter. These findings indicated the significance of isolated promoter for genetic engineering of plants for crop improvement.  

scholarly journals cis-acting sequences of the rat troponin I slow gene confer tissue- and development-specific transcription in cultured muscle cells as well as fiber type specificity in transgenic mice.

1993 ◽  
Vol 13 (11) ◽  
pp. 7019-7028 ◽  
Author(s):  
S Banerjee-Basu ◽  
A Buonanno
Keyword(s):  
Skeletal Muscle ◽  
Transgenic Mice ◽  
Transcription Initiation ◽  
Troponin I ◽  
Regulatory Elements ◽  
Initiation Site ◽  
C2c12 Cells ◽  
Fiber Types ◽  
Tibialis Muscle ◽  
Flanking Sequences

Transcription of the genes coding for troponin I slow (TnIslow) and other contractile proteins is activated during skeletal muscle differentiation, and their expression is later restricted to specific fiber types during maturation. We have isolated and characterized the rat TnIslow gene in order to begin elucidating its regulation during myogenesis. Transcriptional regulatory regions were delineated by using constructs, containing TnIslow gene sequences driving the expression of the chloramphenicol acetyltransferase (CAT) reporter gene, that were transiently transfected into undifferentiated and differentiated C2C12 cells. TnIslow 5'-flanking sequences directed transcription specifically in differentiated cells. However, transcription rates were approximately 10-fold higher in myotubes transfected with constructs containing the 5'-flanking sequences plus the intragenic region residing upstream of the translation initiation site (introns 1 and 2), indicative of interactions between elements residing upstream and in the introns of the gene. Deletion analysis of the 5' region of the TnIslow gene showed that the 200 bp upstream of the transcription initiation site is sufficient to confer differentiation-specific transcription in C2C12 myocytes. MyoD consensus binding sites were found both in the upstream 200-bp region and in a region residing in the second intron that is highly homologous to the quail TnIfast enhancer. Transactivation experiments using transfected NIH 3T3 fibroblasts with TnI-CAT constructs containing intragenic and/or upstream sequences and with the myogenic factors MyoD, myogenin, and MRF4 showed different potentials of these factors to induce transcription. Transgenic mice harboring the rat TnI-CAT fusion gene expressed the reporter specifically in the skeletal muscle. Furthermore, CAT levels were approximately 50-fold higher in the soleus than in the extensor digitorum longus, gastrocnemius, or tibialis muscle, indicating that the regulatory elements that restrict TnI transcription to slow-twitch myofibers reside in the sequences we have analyzed.

Transcriptional enhancers in the HLA-DQ subregion

1987 ◽  
Vol 7 (9) ◽  
pp. 3315-3319
Author(s):  
K E Sullivan ◽  
B M Peterlin
Keyword(s):  
Major Histocompatibility Complex ◽  
Transient Expression ◽  
Regulatory Elements ◽  
Human Major Histocompatibility Complex ◽  
Cis Acting ◽  
Transcriptional Enhancers ◽  
Histocompatibility Complex ◽  
Hla Dq ◽  
Alpha Gene ◽  
Beta Gene

Using transient expression assays, the HLA-DQ alpha and HLA-DQ beta genes of the human major histocompatibility complex were screened for cis-acting regulatory elements. Two regions in the HLA-DQ alpha gene and one in the HLA-DQ beta gene were identified which fulfilled the criteria for transcriptional enhancers.

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