scholarly journals Mutational Analysis of Regulatory cis-acting Elements for the Transcriptional Activation of the dmsCBA Operon in Rhodobactersphaeroides f. sp. denitrificans

2001 ◽  
Vol 42 (7) ◽  
pp. 703-709 ◽  
Author(s):  
Isamu Yamamoto ◽  
Takeshi Ujiiye ◽  
Yoshinori Ohshima ◽  
Toshio Satoh
Keyword(s):  
Transcriptional Activation ◽  
Mutational Analysis ◽  
Cis Acting

A gene product needed for induction of allantoin system genes in Saccharomyces cerevisiae but not for their transcriptional activation

1989 ◽  
Vol 9 (9) ◽  
pp. 3869-3877
Author(s):  
P A Bricmont ◽  
T G Cooper
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Product ◽  
Transcriptional Activation ◽  
Nitrogen Catabolite Repression ◽  
Wild Type ◽  
Product Analysis ◽  
Cis Acting ◽  
Induction Sequence ◽  
Basal Level Expression ◽  
Activation Sequence

The allantoin-degradative pathway of Saccharomyces cerevisiae consists of several genes whose expression is highly induced by the presence of allophanic acid. Induced expression requires a functional DAL81 gene product. Analysis of these genes has demonstrated the presence of three cis-acting elements in the upstream regions: (i) an upstream activation sequence (UAS) required for transcriptional activation in an inducer-independent fashion, (ii) an upstream repression sequence (URS) that mediates inhibition of this transcriptional activation, and (iii) an upstream induction sequence (UIS) needed for a response to inducer. The UIS element mediates inhibition of URS-mediated function when inducer is present. We cloned and characterized the DAL81 gene and identified the element with which it was associated. The gene was found to encode a rare 3.2-kilobase-pair mRNA. The amount of DAL81-specific RNA responded neither to induction nor to nitrogen catabolite repression. Deletion of the DAL81 gene resulted in loss of induction but did not significantly affect basal level expression of the DAL7 and DUR1,2 genes or the UAS and URS functions present in plasmid constructions. These data suggest that (i) transcriptional activation of the DAL genes and their responses to inducer are mediated by different factors and cis-acting sequences and (ii) the UIS functions only when a wild-type DAL81 gene product is available.

The Aspergillus nidulans abaA gene encodes a transcriptional activator that acts as a genetic switch to control development

1994 ◽  
Vol 14 (4) ◽  
pp. 2503-2515
Author(s):  
A Andrianopoulos ◽  
W E Timberlake
Keyword(s):  
Aspergillus Nidulans ◽  
Transcriptional Activation ◽  
Expression System ◽  
Regulatory Gene ◽  
Binding Motif ◽  
Mobility Shift ◽  
Genetic Switch ◽  
Regulatory Regions ◽  
Cis Acting ◽  
Gel Mobility Shift

The Aspergillus nidulans abaA gene encodes a protein containing an ATTS DNA-binding motif and is required for the terminal stages of conidiophore development. Results from gel mobility shift and protection, missing-contact, and interference footprint assays showed that AbaA binds to the sequence 5'-CATTCY-3', where Y is a pyrimidine, making both major- and minor-groove contacts. Multiple AbaA binding sites are present in the cis-acting regulatory regions of several developmentally controlled structural genes as well as those of the upstream regulatory gene brlA, the downstream regulatory gene wetA, and abaA itself. These cis-acting regulatory regions confer AbaA-dependent transcriptional activation in a heterologous Saccharomyces cerevisiae gene expression system. From these observations, we propose that the AbaA transcription factor establishes a novel set of feedback regulatory loops responsible for determination of conidiophore development.

Analysis of a tissue-specific enhancer: ARF6 regulates adipogenic gene expression

1992 ◽  
Vol 12 (3) ◽  
pp. 1202-1208
Author(s):  
R A Graves ◽  
P Tontonoz ◽  
B M Spiegelman
Keyword(s):  
Gene Expression ◽  
Cultured Cells ◽  
Mutational Analysis ◽  
Cell Types ◽  
Specific Gene ◽  
Enhancer Activity ◽  
Cis Acting ◽  
Specific Gene Expression ◽  
Nuclear Extracts ◽  
Adipogenic Gene

The molecular basis of adipocyte-specific gene expression is not well understood. We have previously identified a 518-bp enhancer from the adipocyte P2 gene that stimulates adipose-specific gene expression in both cultured cells and transgenic mice. In this analysis of the enhancer, we have defined and characterized a 122-bp DNA fragment that directs differentiation-dependent gene expression in cultured preadipocytes and adipocytes. Several cis-acting elements have been identified and shown by mutational analysis to be important for full enhancer activity. One pair of sequences, ARE2 and ARE4, binds a nuclear factor (ARF2) present in extracts derived from many cell types. Multiple copies of these elements stimulate gene expression from a minimal promoter in preadipocytes, adipocytes, and several other cultured cell lines. A second pair of elements, ARE6 and ARE7, binds a separate factor (ARF6) that is detected only in nuclear extracts derived from adipocytes. The ability of multimers of ARE6 or ARE7 to stimulate promoter activity is strictly adipocyte specific. Mutations in the ARE6 sequence greatly reduce the activity of the 518-bp enhancer. These data demonstrate that several cis- and trans-acting components contribute to the activity of the adipocyte P2 enhancer and suggest that ARF6, a novel differentiation-dependent factor, may be a key regulator of adipogenic gene expression.

Insights into Enhancer RNAs: Biogenesis and Emerging Role in Brain Diseases

2021 ◽  
pp. 107385842110468
Author(s):  
Yuxin Shen ◽  
Zhengyi Huang ◽  
Ruiqing Yang ◽  
Yunlong Chen ◽  
Qiang Wang ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Small Proportion ◽  
Target Genes ◽  
Noncoding Rnas ◽  
Histone Mark ◽  
Brain Diseases ◽  
Enhancer Activity ◽  
Cis Acting ◽  
High Level ◽  
Classification Function

Enhancers are cis-acting elements that control the transcription of target genes and are transcribed into a class of noncoding RNAs (ncRNAs) termed enhancer RNAs (eRNAs). eRNAs have shorter half-lives than mRNAs and long noncoding RNAs; however, the frequency of transcription of eRNAs is close to that of mRNAs. eRNA expression is associated with a high level of histone mark H3K27ac and a low level of H3K27me3. Although eRNAs only account for a small proportion of ncRNAs, their functions are important. eRNAs can not only increase enhancer activity by promoting the formation of enhancer-promoter loops but also regulate transcriptional activation. Increasing numbers of studies have found that eRNAs play an important role in the occurrence and development of brain diseases; however, further research into eRNAs is required. This review discusses the concept, characteristics, classification, function, and potential roles of eRNAs in brain diseases.

Mechanism of RSV-induced IL-8 gene expression in A549 cells before viral replication

1996 ◽  
Vol 271 (6) ◽  
pp. L963-L971 ◽  
Author(s):  
M. A. Fiedler ◽  
K. Wernke-Dollries ◽  
J. M. Stark
Keyword(s):  
Gene Expression ◽  
Transcription Factor ◽  
Viral Replication ◽  
Transcriptional Activation ◽  
Mutational Analysis ◽  
A549 Cells ◽  
Nf Kappa B ◽  
Mobility Shift ◽  
Syncytial Virus ◽  
Time Point

Previous studies demonstrated that respiratory syncytial virus (RSV) infection of A549 cells induced interleukin (IL)-8 gene expression and protein release from the cells as early as 2 h after treatment [M. A. Fiedler, K. Wernke-Dollries, and J. M. Stark. Am. J. Physiol. 269 (Lung Cell. Mol. Physiol. 13): L865-L872, 1995; J. G. Mastronarde, M. M. Monick, and G. W. Hunninghake. Am. J. Respir. Cell Mol. Biol. 13: 237-244, 1995]. Furthermore, the effects of RSV at the 2-h time point were not dependent on viral replication. The studies reported here were designed to test the hypothesis that active and inactive RSV induce IL-8 gene expression in A549 cells at the 2-h time point by a mechanism dependent on the activation of the nuclear transcription factor NF-kappa B Northern blot analysis indicated that IL-8 gene expression occurred independent of protein synthesis 2 h after A549 cells were treated with RSV. Analysis of nuclear extracts from RSV-treated A549 cells by electrophoretic mobility shift assays demonstrated that NF-kappa B was activated as early as 15 min after RSV was added to the cells and remained activated for at least 90 min. In contrast, baseline levels of NF-IL-6 and activator protein-1 (AP-1) did not change over this period of time. Deoxyribonuclease footprint analysis of a portion of the 5'-flanking region of the IL-8 gene demonstrated two potential regions for transcription factor binding, which corresponded to the potential AP-1 binding site, and potential NF-IL-6 and NF-kappa B binding sites. Mutational analysis of the 200-bp 5'-untranslated region of the IL-8 gene demonstrated that activation of NF-kappa B and NF-IL-6 were required for RSV-induced transcriptional activation of the IL-8 gene.

scholarly journals Transcriptional control of the factor IX gene: analysis of five cis- acting elements and the deleterious effects of naturally occurring hemophilia B Leyden mutations

Blood ◽  
1994 ◽  
Vol 84 (9) ◽  
pp. 2992-3000 ◽  
Author(s):  
DJ Picketts ◽  
CR Mueller ◽  
D Lillicrap
Keyword(s):  
Dna Binding ◽  
Protein Binding ◽  
Transcriptional Activation ◽  
In Vitro Transcription ◽  
Factor Ix ◽  
Hemophilia B ◽  
Transcription Assay ◽  
Cis Acting ◽  
Nuclear Extracts

Abstract Hemophilia B Leyden is a rare form of inherited factor IX deficiency in which patients experience spontaneous postpubertal recovery of factor IX levels. The mutations resulting in this disorder are localized in a 40-nucleotide region encompassing the major transcriptional start site for factor IX. Here we report the further characterization of five cis- acting elements in the factor IX promoter and the effects on protein binding and transcriptional activation of five Leyden mutations (at nucleotides +13, -5, -6, -20, and -26) that occur within the proximal three elements (sites 1 through 3). Bandshift studies using nuclear extracts from four different rat tissues have shown that at least some of the proteins binding to each of the five sites are ubiquitous in nature. The pattern of DNA binding at site 1 suggests that this element plays an important role in mediating the liver-specific expression of factor IX. Additional studies with liver nuclear extracts obtained at several different points in development have shown an increase in DNA binding at sites 1, 4, and 5 between 1 day and 1 week. Using DNase I footprint analysis and competition bandshift studies, we have shown that the binding of nuclear proteins to each of the mutant sites is disrupted to a variable extent. There appears to be some, although reduced, protein binding to all of the mutant oligonucleotides apart from the -26 mutant. In vitro transcription assays have shown that each of the mutations reduces the global proximal promoter activity by approximately 40%. Two double mutant promoters did not show any additional downregulation in the in vitro transcription assay. In experiments designed to assess the relative transcriptional activity mediated from each of the five sites independently, we have tested artificial homopolymer promoters of each site in the in vitro transcription assay. These studies show that sites 4 and 5 are the strongest activators and that transactivation from site 5 is further enhanced by the albumin D site-binding protein. In summary, these investigations show deleterious effects of each of the Leyden mutations tested on the binding of trans-acting factors and also show disruption of transcriptional activation in a functional in vitro transcription assay. Our results also show that cis-acting elements 4 and 5 are the principal activators of this locus.

scholarly journals Participation of the yeast activator Abf1 in meiosis-specific expression of the HOP1 gene.

1996 ◽  
Vol 16 (6) ◽  
pp. 2777-2786 ◽  
Author(s):  
V Gailus-Durner ◽  
J Xie ◽  
C Chintamaneni ◽  
A K Vershon
Keyword(s):  
Transcriptional Activation ◽  
Mutational Analysis ◽  
Consensus Sequence ◽  
Promoter Sequence ◽  
Regulatory Elements ◽  
Specific Gene ◽  
Specific Expression ◽  
Autonomously Replicating Sequence

The meiosis-specific gene HOP1, which encodes a component of the synaptonemal complex, is controlled through two regulatory elements, UASH and URS1H. Sites similar to URS1H have been identified in the promoter region of virtually every early meiosis-specific gene, as well as in many promoters of nonmeiotic genes, and it has been shown that the proteins that bind to this site function to regulate meiotic and nonmeiotic transcription. Sites similar to the UASH site have been found in a number of meiotic and nonmeiotic genes as well. Since it has been shown that UASH functions as an activator site in vegetative haploid cells, it seemed likely that the factors binding to this site regulate both meiotic and nonmeiotic transcription. We purified the factor binding to the UASH element of the HOP1 promoter. Sequence analysis identified the protein as Abf1 (autonomously replicating sequence-binding factor 1), a multifunctional protein involved in DNA replication, silencing, and transcriptional regulation. We show by mutational analysis of the UASH site, that positions outside of the proposed UASH consensus sequence (TNTGN[A/T]GT) are required for DNA binding in vitro and transcriptional activation in vivo. A new UASH consensus sequence derived from this mutational analysis closely matches a consensus Abf1 binding site. We also show that an Abf1 site from a nonmeiotic gene can replace the function of the UASH site in the HOP1 promoter. Taken together, these results show that Abf1 functions to regulate meiotic gene expression.

scholarly journals Transcriptional Activation of the Decidual/Trophoblast Prolactin-Related Protein Gene1

Endocrinology ◽  
1999 ◽  
Vol 140 (9) ◽  
pp. 4032-4039 ◽  
Author(s):  
Kyle E. Orwig ◽  
Michael J. Soares
Keyword(s):  
Transcriptional Activation ◽  
Promoter Activity ◽  
Mutational Analysis ◽  
Regulatory Element ◽  
Gene Activation ◽  
Cell Formation ◽  
Regulatory Elements ◽  
Related Protein ◽  
Trophoblast Cells ◽  
Decidual Cells

Abstract The decidual/trophoblast PRL-related protein (d/tPRP) is dually expressed by decidual and trophoblast cells during pregnancy. We have characterized the proximal d/tPRP promoter responsible for directing d/tPRP expression in decidual and trophoblast cells. We have demonstrated that the proximal 93 bp of d/tPRP 5′-flanking DNA are sufficient to direct luciferase gene expression in primary decidual and Rcho-1 trophoblast cells, but not in fibroblast, undifferentiated uterine stromal cells or trophoblast cells of a labyrinthine lineage. The 93-bp d/tPRP promoter was also sufficient to direct differentiation-dependent expression in trophoblast giant cells. Mutational analysis demonstrated the differential importance of activating protein-1 and Ets regulatory elements (located within the proximal 93 bp of d/tPRP 5′-flanking DNA) for activation of the d/tPRP promoter in decidual vs. trophoblast cells. Disruption of the activating protein-1 regulatory element inhibited d/tPRP promoter activity by more than 95% in decidual cells, and approximately 80% trophoblast cells. Disruption of the Ets regulatory element reduced d/tPRP promoter activity by approximately 50% in decidual cells, while inactivating the d/tPRP promoter in trophoblast cells. Protein interactions with the trophoblast Ets regulatory element were shown to be cell type specific and to change during trophoblast giant cell formation. In conclusion, a 93-bp region of the d/tPRP promoter is shown to contain regulatory elements sufficient for gene activation in decidual and trophoblast cells.

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